01 April 2024, Volume 38 Issue 4

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  • Review
    Giuseppe Marano, Francesco Maria Lisci, Gianandrea Traversi, Caterina Brisi, Domenico De Berardis, Eleonora Gaetani, Paola Aceto, Carlo Lai, Giovanni Martinotti, Osvaldo Mazza, Marianna Mazza
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2693-2712. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.212
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    Background: Recent achievements in neurosciences have shown a growing interest in the role of endocannabinoid system (ECS) in the regulation of neuroprotection and neurodevelopment, with growing interest in the regulation of psychological processes. As well as neurotransmitters, endocannabinoids are molecules produced in our organisms and used as messengers in order to transmit signals between cells and activate responses, by interacting with several receptors. Recent studies have highlighted the involvement of the ECS in the physiopathology of several neurodegenerative disorders, such as Parkinson's Disease (PD), Alzheimer's Disease (AD) and Huntington Disease (HD).

    Methods: We searched the PubMed/Medline, Scopus, Embase, and ScienceDirect databases on 1st September 2023, focusing on recent research regarding the role of endocannabinoids on the physiopathogenesis of neurodegenerative diseases with neuropsychiatric symptoms. We explored the role of ECS in the development of neurodegenerative disorders and the link to the onset of neuropsychiatric symptoms and future therapeutic strategies that implicate ECS activity in relation to these diseases.

    Results: There is a strong involvement of endocannabinoids in neurodegenerative diseases and psychiatric conditions, with growing evidences on efficacy of Cannabidiol (CBD)-based treatments.

    Conclusions: Recent ECS-mediated pathways have been found to be involved in neurodegenerative diseases and psychiatric conditions. CBD-based treatments may be promising neuropsychopharmacological interventions for psychiatric symptoms on AD, PD and HD patients, worthy of being further explored.

  • Review
    Iqra Khurram, Muhammad Umer Khan, Ayman Saleem, Saooda Ibrahim, Muhammad Usman Ghani, Muhammad Shahid, Iram Amin, Daniela Calina, Javad Sharifi-Rad
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2713-2732. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.213
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    Background: Cancer, characterized by the rapid and abnormal growth of cells affecting any part of the body, stands as the leading cause of death worldwide. Cell-free DNA (cfDNA) has garnered significant attention as a non-invasive liquid biopsy approach for disease detection, therapy evaluation, and prognosis. This review aims to provide a comprehensive exploration of cfDNA within the realm of oncology. It encompasses its diagnostic and therapeutic applications while identifying areas necessitating standardization and optimization.

    Methods: We reviewed existing literature to delve into the biological properties of cfDNA, exploring genetic and epigenetic aberrations found in various bodily fluids. Additionally, we explored its correlation with circulating tumor DNA (ctDNA). The review also encompasses preanalytical procedures and emerging technologies geared towards maximizing the complete potential of cfDNA.

    Results: Genetic and epigenetic markers have been identified in cfDNA across plasma, serum, and urine, presenting promising diagnostic and prognostic applications. Furthermore, ctDNA preserves genomic profiles akin to those found in corresponding tumor tissues, enabling a nuanced evaluation of tumor heterogeneity and mutation burdens. However, despite its potential, current methodologies suffer from a lack of standardization and optimization, thereby restraining the complete clinical utility of cfDNA.

    Conclusions: Although cfDNA stands as a compelling avenue for non-invasive early cancer diagnosis and therapy evaluation, unlocking its maximum potential requires methodological refinement and a deeper understanding of its biological characteristics. This review advocates for targeted research to standardize and optimize cfDNA analytical techniques, thereby enhancing its role in oncology.

  • Review
    Rakesh Bhaskar, Kannan Badri Narayanan, Aarti Nagayach, Soon Mo Choi, Sung Soo Han
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2733-2749. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.214
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    Neurodegenerative disorders pose significant societal and individual burdens, marked by progressive degeneration of the nervous system. The limited efficacies of current treatments underscore the pressing need for innovative therapeutic approaches. Cannabigerol (CBG) is a cannabinoid derivative found in the plant genus Cannabis that is viewed as a molecule of therapeutic interest due to its unique pharmacological profile and interactions with the endocannabinoid system (ECS). This comprehensive review explores the therapeutic potential of CBG for the treatment of disorders of the nervous system, such as Alzheimer's and Parkinson's disease. Preclinical investigations have revealed that the neuroprotective properties of CBG involve ECS modulation, promoting neuronal survival, and mitigating neuroinflammation and oxidative stress. However, it should be acknowledged that much of the research on the therapeutic potential of CBG is still in its early stages and primarily consists of in vitro and animal studies. Further clinical trials and research are needed to determine the efficacy and safety of CBG and the optimal dosing strategies required to treat neurodegenerative diseases. This review also explores the transformative potential of nanocarrier systems in CBG-centered therapies targeting neurodegenerative disorders and discusses the merits and demerits of various nanocarrier systems. This dual focus on the therapeutic potential of CBG and innovative distribution systems underscores the extent of the investigative work required to advance neurodegenerative disease management.

  • Review
    Jelena Marković Filipović, Ivana Ivelja, Jelena Vukčević, Maja Milanović
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2751-2767. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.215
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    Humans are heavily exposed to acrylamide (AA), a Maillard reaction product present in everyday diet. However, only recently, the European Commission set recommendations for monitoring AA levels in certain foods. Owing to their quick metabolism after intake, both AA and its adducts induce oxidative stress and inflammatory processes, as well as, histopathological alterations in the liver. To date, the exact concentration and the length of exposure associated with AA adverse effects remain unclear. A diverse number of natural products have been investigated in attempts to identify their protective effects against liver disorder. This review aims to highlight the significance of several phytochemicals and phytochemical mixtures in ameliorating AA-induced hepatotoxicity. Among phytochemicals, quercetin, morin, thymoquinone, allicin, alpha-lipoic acid, ellagic acid, curcumin, cysteine and carvacrol, revealed anti-inflammatory, antioxidant, hepatoprotective, antiapoptotic and DNA-protective effects. On the other hand, among phytochemical mixtures, Carica papaya, wine polyphenols, pomegranate juice, anthocyanins and propolis, showed anti-inflammatory, antioxidant, hepatoprotective and antiapoptotic properties against AA-induced hepatotoxicity. Further studies are needed in order to elucidate the major natural products, as well as, the molecular mechanisms underlying their hepatoprotective effects in AA-related hepatoxicity. Additionally, the safety of their application should be investigated. Having in mind that AA levels in food cannot be completely controlled, well-designed clinical trials could enable the development and application of new pharmaceutical products aiming to prevent and alleviate AA-induced hepatotoxicity.

  • Systematic Review
    Sonakashee Deshmukh, Amol Somaji Patil, Sonakshi Sharma
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2769-2780. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.216
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    Background: Recently, numerous attempts have been made to expedite orthodontic tooth movement (OTM) to reduce treatment time. Platelet-rich concentrates have emerged as a potential intervention to accelerate OTM. This study aimed to comprehensively examine the existing available literature on the effect of platelet-rich plasma (PRP) on the rate of OTM.

    Methods: This systematic review adhered to Preferred Reporting Items for Systematic Review and Meta-analysis (PRISMA) statement guidelines, and the protocol was registered in Prospective Registration of Systematic Review (PROSPERO) (CDR42022355356). Electronic databases were searched to retrieve relevant randomized controlled trials (RCTs) assessing the rate of orthodontic tooth movement as the primary outcome, and secondary outcomes included pain (visual analog scale (VAS) score), loss of anchorage, and molar rotation. Risk of bias (ROB) was assessed using Cochrane's ROB-2, and Grading of Recommendations, Assessment, Development and Evaluation (GRADE) quality assessment was employed. Standardized mean difference (SMD) served as a summary statistic measure, with p < 0.05 indicating statistical significance. Publication bias was explored through a funnel plot.

    Results: Eight studies met the eligibility criteria for the systematic review, and a meta-analysis was conducted on five studies. The ROB for all studies ranged from moderate to high, while the quality of evidence was moderate. The pooled estimate, as indicated by SMD, demonstrated a significant increase in the rate of orthodontic tooth movement during the first month after the intervention. Meta-analysis indicated the absence of publication bias.

    Conclusions: PRP demonstrated the potential to accelerate the rate of OTM during the first month. Further studies conducted with a rigorous protocol extended duration are recommended.

  • Article
    Spiros Gerostathis, Alexandros Athanasopoulos, Katerina N. Panagiotaki, Zili Sideratou, Dimitris Tsiourvas, Vicky Sophianopoulou
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2781-2794. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.217
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    Background: Drug resistance is the major problem for all of the currently available antifungals and, therefore, a completely novel class of antifungal compounds is an urgent need. The aim of this study is to investigate whether appropriately functionalized cationic dendritic polymers that have extensively been studied as drug delivery systems and/or antibacterial agents exhibit fungicidal activity and thus could be used as antifungals.

    Methods: In this work, hyperbranched polyethyleneimine and its end-group functionalized guanidinylated or quaternized derivatives were synthesized and their antifungal activity was evaluated against the opportunistic pathogen Aspergillus nidulans (A. nidulans) as a model eukaryotic microorganism. Specifically, their subcellular localization in germlings originating from fungal conidiospores, their effect on fungal growth and their ability to cause cell death in both hyphae and quiescent conidia, were studied using confocal laser and epifluorescent microscopy, the colony forming unit (CFU) assay and the propidium iodide (PI) staining assay.

    Results: It was found that polyethyleneimine (PEI) was the most effective compound causing complete growth inhibition at relatively lower minimum inhibitory concentration (MIC) of ca. 2 μg/mL compared to the guanidinylated (GPEI) (3 μg/mL) or quaternized (QPEI) (5 μg/mL) derivatives. More importantly, the compounds prevent quiescent conidiospore germination and, therefore, the formation of hyphae and mycelium. Employing the fluorescently-labeled derivatives, it was evident that the compounds are initially localized in the plasma membrane of both quiescent conidiospores and hyphae, while subsequently enter the cytoplasm, without showing localization in specific subcellular organelles. Appropriate staining with propidium iodide (PI) revealed that the membrane of cells, grown in the presence of all tested polymers at concentrations above the minimum inhibitory concentration (MIC), exhibits increased permeability to the dye, suggesting that the primary cause of cell death is the disruption of the plasma membrane's integrity. Furthermore, by employing the CFU assay it was evident that a significant percentage of fungal conidiospores exhibited inability to recover growth following treatment with the examined compounds and that their survival was reduced by up to 70% (p < 0.001).

    Conclusions: Our results showed that the cationic hyperbranched polymeric compounds PEI and its derivatives GPEI and QPEI exhibited apparent fungicidal activity on A. nidulans cells. In addition, all compounds showed negligible in vitro cytotoxicity against three mammalian cell lines in this concentration range. Overall, our results suggest that the investigated polymeric compounds are promising antifungals and their activity deserves further investigation especially against drug-resistant fungal cells.

  • Article
    Tae-Kyeong Lee, Myoung Cheol Shin, Joon Ha Park, Ji Hyeon Ahn, Choong-Hyun Lee, Yang Hee Kim, Jun Hwi Cho, Moo-Ho Won, Dae Won Kim, Seongkweon Hong
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2795-2806. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.218
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    Background: Whole-body ischemia and reperfusion (IR) injury following the return of spontaneous circulation (ROSC) after cardiac arrest (CA) can cause multiple organ dysfunction syndrome accompanied by adverse outcomes, including serious mortality. Renal IR injury following ROSC after CA is associated with complicated pathological processes. Renal IR injury is commonly observed in inpatients with heart failure and is associated with high mortality. Risperidone (Risp), an atypical antipsychotic drug, has been reported to exert beneficial effects against IR. This study investigated whether treatment with risperidone enhanced survival rates and reduced kidney failure following CA-induced whole-body IR injury.

    Methods: Rats were subjected to asphyxial CA for five minutes followed by ROSC induction, and risperidone (10 mg/kg) or saline was intravenously injected. Survival rate was evaluated using Kaplan-Meier survival analysis was used to evaluate the survival rate. Changes in serum levels of creatinine, lactate dehydrogenase (LDH), and blood urea nitrogen (BUN) were assessed. We carried out hematoxylin and eosin staining and immunohistochemistry for 4-hydroxy-2-nonenal (4-HNE), 8-hydroxy-2′-deoxyguanosine (8-OHdG), superoxide dismutase 1 (SOD-1), and superoxide dismutase 2 (SOD-2).

    Results: Treatment with risperidone significantly improved the survival rate after CA and attenuated LDH, creatinine, and BUN levels. Histopathological injury was remarkably improved in the renal cortex following risperidone treatment including a significant attenuation of the immunoreactivities of 4-HNE and 8-OHdG. Conversely, immunoreactivities of SODs were significantly improved by risperidone treatment.

    Conclusions: The current results demonstrated that treatment with risperidone reduced the CA-induced renal injury by reducing oxidative stress and retaining antioxidant enzymes, which significantly influenced the improvement of survival rate after CA.

  • Article
    Blanka Maria Borowiec, Wiesława Kranc, Magdalena Kulus, Patrycja Sujka-Kordowska, Aneta Konwerska, Piotr Celichowski, Małgorzata Józkowiak, Jakub Kulus, Małgorzata Blatkiewicz, Michal Jeseta, Paul Mozdziak, Bartosz Kempisty
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2807-2824. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.219
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    Background: A frequent reason for in vitro fertilization failure is insufficient oocyte competence and difficulties in mimicking optimal in vivo conditions at the stage of in vitro maturation. This study aimed to identify the expression profile of genes associated with “vasculature development”, “circulatory system process”, “hemopoiesis”, “patterning of blood vessels” and “hemopoietic or lymphoid organ development” ontological groups during in vitro maturation of domestic pig oocytes. The goal was also to describe new genetic markers that can be used in assisted reproduction techniques.

    Methods: Porcine oocytes were subjected to in vitro maturation, and the expression of selected genes was compared before and after maturation. Microarray and reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis were performed on total RNA isolated from the samples. Additionally, immunocytochemical staining was performed and the histological morphology of ovaries was examined.

    Results: Regardless of the individual processes, the gene expression pattern remained the same and all 32 gene ontology biological process term-genes were represented in down-regulated gene sets. The greatest change in expression was shown by the Vascular Endothelial Growth Factor A (VEGFA) and the smallest by myoferlin (MYOF). Moreover, the expression of VEGFA was confirmed by immunohistochemical analysis of ovaries.

    Conclusions: The potential contribution of chosen genes to oocyte performance during in vitro maturation can enhance comprehension of the underlying processes in oocytes and their microenvironment. It may also improve the effectiveness of in vitro fertilization.

  • Article
    Haina Tao, Chengcai Zhang
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2825-2836. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.220
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    Background: Gastric cancer presents a significant global health burden, with existing therapies exhibiting limited efficacy and potential side effects. Molecular-targeted therapy has emerged as a promising approach. This study aimed to explore the role of guanylate binding protein 2 (GBP2) in programmed death 1 ligand (PD-L1)-related immune response in gastric cancer and to elucidate the underlying molecular regulatory mechanisms.

    Methods: The gastric adenocarcinoma cell line AGS, the human immortalized gastric epithelium cell line GES-1, and the peripheral blood mononuclear cells (PBMCs) obtained from healthy donors were utilized. Lentiviral plasmids, small interfering (si)RNAs, and the signal transducer and activator of transcription 3 (STAT3) inhibitor BP-1-102 were used to modulate targeted protein expression. Differential expression of GBP2 in gastric cancer cells was assessed, and the impact of GBP2 levels on STAT3, nuclear factor-kappa B (NF-κB), CD8, PD-L1, T-cell response-related cytokines of interleukin-2 (IL-2), interferon-γ (IFN-γ), tumor necrosis factor-alpha (TNF-α), and cell viability was determined. Immunofluorescence and gene expression analyses were conducted using Enzyme-Linked Immunosorbent Assay (ELISA), the RT-qPCR, Western Blot, and flow cytometry.

    Results: GBP2 was upregulated in gastric cancer cells, positively regulating PD-L1 expression and activating the STAT3-NF-κB pathway while negatively regulating CD8 expression. GBP2 overexpression enhanced the STAT3-NF-κB pathway activation and PD-L1 expression, which was reversed upon suppressing of the STAT3-NF-κB signaling pathway. Moreover, GBP2 knockdown in AGS cells cultured alone or with activated T-cells resulted in increased cell proliferation and invasion, and reduced cell apoptosis. Additionally, the increased/decreased GBP2 inhibited/promoted the expressions of interleukin-2 (IL-2), interferon-γ (IFN-γ) and tumor necrosis factor-alpha (TNF-α).

    Conclusion: GBP2 positively regulates PD-L1 expression through the STAT3-NF-κB signaling pathway and subsequently causes a suppression on T-cell immune responses, thus suggesting GBP2 as a potential immunotherapeutic target in gastric cancer.

  • Article
    Tao Xie, Xue-xin He, Qian Sun, Zhuo-feng Mao, Xiao-peng Wang
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2837-2843. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.221
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    Background: Epilepsy is a common disease that often causes cognitive impairment in patients. Ginsenoside Rb1 has been shown to have a protective effect on hippocampal neurons, but its role in epilepsy is unclear. Herein, we investigate the protective effects of ginsenoside Rb1 in pentylenetetrazole (PTZ)-kindled rats.

    Methods: The 40 Sprague-Dawley rats were randomised into 4 groups (n = 10/group): control, PTZ, PTZ + ginsenoside and ginsenoside groups. Rats in the control group were subjected to saline every other day for a total of 13 injections. Similarly, rats in the PTZ group were injected every other day with PTZ (dose, 35 mg/kg). Rats in the PTZ + ginsenoside group were additionally treated with ginsenoside (dose, 100 mg/kg) in addition to receiving PTZ. Rats in the ginsenoside group received injections of ginsenoside at a dose of 100 mg/kg. Following completion of the final injection, the Morris water maze test and relevant behavioural experiments were performed 24 h later on all rats in the four groups. The malondialdehyde (MDA) and glutathione (GSH) content, as well as Nissl staining, were subsequently assessed in rats.

    Results: This study found that ginsenoside Rb1 pre-administration decreased the mean seizure duration and improved cognitive function compared to the PTZ group (p < 0.05). The behavioural test results indicated that the latency of the PTZ group during all sessions was longer compared with the control group (p < 0.05), and ginsenoside Rb1 alleviated this poorer performance. Additionally, ginsenoside Rb1 pretreatment diminished the MDA content and enhanced the GSH content compared to the PTZ group (p < 0.05). Furthermore, pretreatment with ginsenoside Rb1 observably reduced seizure-induced neuronal loss (p < 0.001).

    Conclusion: Ginsenoside Rb1 can inhibit seizures and improve PTZ-induced cognitive dysfunction by alleviating oxidative stress damage and neuronal loss in kindled rats.

  • Article
    Dhinesh Kumar Murugesan, Kalirajan Rajagopal, Arumugam R. Vijayakumar, Gothandam Sundararajan, Kannan Raman, Gowramma Byran, Esakkimuthukumar Murugesan, Jeetendra Kumar Gupta, Rani S. Kankate, Firzan Nainu, Rashu Barua, Sheikh F. Ahmad, Talha Bin Emran
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2845-2859. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.222
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    Background: Breast cancer, a common and potentially fatal disease, has several subtypes, with the frequent being hormone receptor-positive breast cancer. This study addresses the critical need to tackle this subtype by focusing on the estrogen receptor alpha (ERα), a crucial factor in its development and progression.

    Methods: We used computational approaches to create and improve new pyrazole-substituted 9-anilinoacridine compounds with ERα inhibition potential. This included ligand preparation, receptor grid generation, Glide molecular docking and Prime/ Molecular Mechanics/Generalized Born Surface Area (MM-GBSA) binding free energy calculations. We tested the ligand–protein interactions and binding affinities using these approaches. The lead compounds were tested for anti-breast cancer efficacy in vitro by utilising MDA-MB-231 cell lines.

    Results: Our computational research showed numerous chemicals with strong ERα receptor binding sites as evidenced by favourable interactions involving hydrogen bonding, hydrophobic interactions and van der Waals forces. These molecules were subsequently synthesised and structurally characterised. Compound 1z displayed considerable cytotoxicity in in-vitro cytotoxicity studies using the MDA-MB-231 cell line, with concentration required for 50% inhibition of cell growth (IC50) value of 23.86 g/mL indicating a good potential against breast cancer. Despite having high docking scores and binding free energy, compound 1x had a high IC50 value (36.08 g/mL). This emphasises the importance of experimental validation in supplementing computer predictions.

    Conclusions: Thus, pyrazole-substituted 9-anilinoacridine derivatives show promise as possible ERα inhibitors for breast cancer therapy. While our computational analysis provides a solid foundation, more research is needed to ensure their safety and efficacy as prospective therapies for ERα breast cancer, including in-vivo evaluations and Absorption, Distribution, Metabolism, Excretion and Toxicity (ADMET) profiling.

  • Article
    Qiang Gao, Jie Li
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2861-2874. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.223
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    Background: Lung adenocarcinoma (LUAD) is a severe type of lung cancer, characterized by high prevalence and mortality rate. Adipocyte enhancer binding protein 1 (AEBP1) is a critical cancer-promoting factor in several types of cancer. Nevertheless, the hidden mechanism underlying the effect of AEBP1 on LUAD has not been investigated.

    Methods: The mRNA and protein expressions of AEBP1 and Yin Yang 1 (YY1) were detected by reverse transcription-quantitative PCR (RT-qPCR) and western blot. By means of Cell Counting Kit-8 (CCK-8), 5-ethynyl-2′-deoxyuridine (EdU) staining, colony formation assay, transwell and wound healing assays, A549 cell viability, cell proliferation, invasion and migration were appraised. The angiogenesis of human umbilical vein endothelial cells (HUVECs) was evaluated with tube formation assay. Western blot was used to detect proteins associated with epithelial-mesenchymal transition (EMT) and ferroptosis. Thiobarbituric acid-reactive substances (TBARS) production rate and Fe2+ levels were measured using the corresponding kits. Reactive oxygen species (ROS) activity was detected by 4,4-difluoro-5-(4-phenyl-1,3-butadienyl)-4-bora-3a,4a-diaza-s-indacene-3-undecanoic acid (C11-BODIPY(581/591)). Luciferase reporter assay was used to detect AEBP1 promoter activity and chromatin immunoprecipitation was performed to estimate the binding ability of YY1 and AEBP1 promoter.

    Results: AEBP1 silencing inhibited the proliferation, metastasis and angiogenesis whereas induced ferroptosis in LUAD cells (p < 0.001), which were all reversed by YY1 (p < 0.05 or p < 0.01 or p < 0.001). Besides, YY1 was found to promote AEBP1 transcription (p < 0.001).

    Conclusion: Collectively, YY1-regulated AEBP1 could facilitate LUAD advancement and inhibit ferroptosis.

  • Article
    Yunxia Jiang, Da Li, Wenting Chen, Yiting Zhou, Zhongmin Yang, An Wu
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2875-2886. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.224
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    Background: Intracranial aneurysm (IA) rupture poses a significant threat to health. Thrombospondin 1 (THBS1) has been implicated in the prevention of IA rupture. This study aimed to elucidate the mechanistic role of THBS1 in IA rupture.

    Methods: An IA model was induced in Sprague-Dawley (SD) rats by ligating the right common carotid artery, left external carotid artery, and left pterygopalatine artery. Vascular smooth muscle cells (VSMCs) were isolated from SD rats and stimulated with tumor necrosis factor-alpha (TNF-α; 20 ng/μL) to induce a proinflammatory phenotype. Cerebral vessel morphology was assessed using hematoxylin-eosin (HE) staining. Levels of alpha-smooth muscle actin (α-SMA), smooth muscle 22 alpha (SM22α), matrix metalloproteinase 2 (MMP2), THBS1, thrombospondin receptor CD36, and transforming growth factor-beta 1 (TGF-β1) were evaluated via western blot. The transfection efficiency of THBS1 plasmid and short hairpin CD36 (ShCD36) was determined using quantitative real-time polymerase chain reaction (qRT-PCR). Cell viability, proliferation, and apoptosis were assessed using the cell counting kit-8 (CCK-8), 5-ethynyl-2′-deoxyuridine (EdU) assay, and Hoechst assay, respectively.

    Results: IA model rats exhibited reduced levels of α-SMA, SM22α, THBS1, and CD36, accompanied by increased MMP2 levels (p < 0.01). THBS1 overexpression reversed the effects of TNF-α on VSMCs viability, proliferation, apoptosis, and the expression levels of TGF-β1, α-SMA, SM22α, MMP2, CD36, and THBS1 (p < 0.05). Silencing CD36 reversed the effects of THBS1 overexpression on TNF-α-treated VSMCs (p < 0.05).

    Conclusions: Our findings demonstrate that THBS1 promotes VSMC proliferation and inhibits phenotypic transformation to reduce IA rupture.

  • Article
    Yanhua Lu, Qiang Hou, Qinghua Li
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2887-2901. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.225
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    Background: Although advanced therapeutic approaches have been used to improve the outcomes of burn patients, burn injuries continue to pose a life-threatening challenge. This study aimed to identify the innovative biomarkers and potential key gene-related drug candidates for burns.

    Method: We identified differentially expressed genes (DEGs) in the GSE182616 dataset, comparing control individuals with burn patients across various time points within 24 h. Co-DEGs were identified at specific intervals (0–2 h, 2–4 h, 4–8 h, 8–12 h, and 12–24 h) through overlapping analysis. Furthermore, key genes were found before and after DJK-5 treatment in the GSE210640 dataset. Based on these key genes, we screened key gene-related sets and subsequently predicted potential gene inhibitors through molecular docking.

    Result: Based on overlapping analysis of the GSE182616 dataset, 21 co-DEGs were screened, identifying 6 key genes, including integrin beta2 (ITGB2), GTPase of immunity-associated protein 6 (GIMAP6), tripartite motif-containing 22 (TRIM22), IFN regulatory factor 7 (IRF7), ubiquitin carboxyl-terminal hydrolase 15 (USP15), and mitogen-activated protein kinase 14 (MAPK14) with significant changes before and after treatment. These key genes demonstrated positive associations with inflammatory-related pathways such as the janus kinase-transducer and activator of transcription (JAK-STAT) signaling pathway, nuclear factor kappa-B (NF-κB) signaling, Toll-like receptor signaling, B cell receptor signaling, and T cell receptor signaling pathways. Additionally, these key genes were linked to immune cells including activated CD8 T cells, activated dendritic cells, mast cells, monocytes, natural killer cells, and neutrophils. Molecular docking demonstrated that compound DB08059 was a potential inhibitor of GIMAP6 and USP15. Moreover, compounds DB05442, DB14059, DB08437, and DB07186 acted as inhibitors of IRF7, ITGB2, MAPK14, and TRIM22 proteins, respectively.

    Conclusions: This study provided novel ideas for the underlying mechanism of key genes in burns and shared promising strategies for the treatment of burn injuries.

  • Article
    Xiaoli Xu, Xueju Xu, Yuan Zhang, Wenhao Yan, Yufeng Liu
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2903-2911. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.226
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    Background: Although advances in treatment have increased the five-year overall survival of pediatric patients with T-cell acute lymphoblastic leukemia (T-ALL) to >80%, relapse, resulting in poor prognosis, remains a considerable challenge for all age groups. Therefore, further improvements in T-ALL treatment are required. This study aimed to determine the roles of miR-96-5p and spleen tyrosine kinase (SYK) as novel therapeutic targets that regulate T-ALL cells.

    Methods: The expression levels of miR-96-5p and SYK were evaluated from patient samples, and the correlation between the two was determined. The effects of miR-96-5p on the proliferation of T-ALL cells were detected using the cell counting kit-8 (CCK-8) and 5-ethynyl-2′-deoxyuridine (EdU) assays. The effect of miR-96-5p on T-ALL cell apoptosis was evaluated by flow cytometry. In addition, bioinformatics analysis, luciferase reporter assays, and rescue assays were used to verify whether SYK was a downstream target of miR-96-5p. The regulatory relationship between miR-96-5p and SYK was also analyzed.

    Results: The results showed that the expression of miR-96-5p was lower in patients with T-ALL than in healthy volunteers (p < 0.05). Additionally, miR-96-5p enhanced proliferation and inhibited apoptosis of T-ALL cells in vitro. Furthermore, the expression of SYK and miR-96-5p in T-ALL patients showed an opposite trend, and this negative correlation was also reflected in the effect on cells.

    Conclusions: This study demonstrated that miR-96-5p and SYK may be potential therapeutic targets for regulating T-ALL cell proliferation and apoptosis at the cellular level.

  • Article
    Yang Zhang, Xueqing Zhou, Jinpeng Zhang, Mingfei Zhang, Danhong Zeng, Jie Zhou, Baohu Zhang, Li Zhang, Shucai Yang
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2913-2926. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.227
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    Background: Non-small cell lung cancer (NSCLC) represents the predominant pathological subtype of lung cancer in China. Amidst the advent of precision medicine, immunotherapy has emerged as a pivotal approach in managing malignant neoplasms, substantially improving patient prognosis and survival rates. However, the efficacy of immunotherapy remains limited, primarily attributed to the development of resistance among advanced-stage patients. This study used bioinformatics methodologies to analyze and identify potential key genes governing immune resistance in NSCLC, offering novel insights into therapeutic avenues.

    Methods: Gene expression datasets (GSE126044 and GSE135222) encompassing NSCLC cases with immunotherapy resistance and control groups were retrieved from the Gene Expression Omnibus (GEO) repository. Differential gene expression analysis was conducted using Gene Expression Omnibus 2 R (GEO2R) with criteria set at |log FC (fold change)| ≥1 and p < 0.05. Subsequent analyses involved Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, Gene Ontology (GO) functional annotation, Protein-Protein Interaction (PPI) network construction, and Gene Set Enrichment Analysis (GSEA). The findings were visualized through volcano plots and box plots in the R program. Candidate genes were cross-validated with Genecard database entries and scrutinized against existing literature for clinical relevance. The association between key genes, immune cells, and immune-associated gene expressions was analyzed using the Tumor Immune Estimation Resource (TIMER) database. Immunohistochemistry assays were employed to assess the differentially expressed genes (DEGs) in lung cancer tissues.

    Results: Sixty-four upregulated DEGs were obtained from datasets GSE126044 and GSE135222. PPI network analysis identified one cluster and twelve candidate genes, further corroborated through module examination of common DEGs. Integration with Genecard database entries and literature confirmed Fas Ligand (FASLG) as a pivotal gene. KEGG and GSEA pathway analyses unveiled potential mechanisms predominantly related to the interaction between immune cell functions and cytokines, especially T cells. Analysis in the TIMER database revealed a significant positive correlation between FASLG expression and six types of infiltrating immune cells, as well as specific immune cell subsets, alongside three immune checkpoint-associated molecules: Cluster of Differentiation 274 (CD274), Cytotoxic T-Lymphocyte-Associated protein 4 (CTLA-4), and Programmed Cell Death Protein 1 (PDCD1) (p-value < 0.05). Furthermore, in The Cancer Genome Atlas (TCGA) database, FASLG was strongly associated with T cell gene markers and regulatory factors associated with T cell exhaustion, demonstrating statistical significance (p-value < 0.05). Immunohistochemical results preliminarily confirmed the significant upregulation of FASLG in lung cancer tissues.

    Conclusion: The identification of key genes and associated signaling cascades deepens our understanding of the molecular mechanisms governing immunotherapy resistance in NSCLC. Notably, FASLG is a potential facilitator of immune escape in NSCLC tumor cells by promoting T cell exhaustion, highlighting NSCLC as a viable target for anticancer interventions.

  • Article
    Lin Zhang, Xingmin Zhang, Juan Du, Xiaobing Chen
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2927-2932. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.228
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    Background: Morphine and oxycodone are effective analgesics for treating both acute and chronic pain. However, previous studies have primarily focused on their oral efficacy and potential side effects. This study aims to assess the effectiveness and adverse reactions of oxycodone and morphine in the treatment of cancer-related pain.

    Methods: A total of 39 patients admitted to our hospital from April 5, 2022 to April 5, 2023 received oxycodone (n = 19) or morphine (n = 20). Their treatment data were retrospectively analyzed. These patients received either oxycodone or morphine orally for five consecutive days. Pain intensity was evaluated using the Numeric Rating Scale for Pain (NRS Pain) from day 0 to day 5. The study also recorded pain relief rates, incidence of adverse reactions, and treatment satisfaction.

    Results: After receiving treatment with either oxycodone or morphine, all subjects experienced a reduction in cancer pain grade from moderate or severe to mild. On the third day of drug administration, there was a statistically significant decrease in NRS Pain scores (p < 0.05). Furthermore, by the fifth day of treatment, although pain symptoms were notably alleviated in both groups, satisfaction was significantly higher in the morphine group (p < 0.05). Both groups exhibited adverse effects such as vomiting, nausea, and constipation. However, the incidence of adverse effects and the number of patients experiencing vomiting were significantly lower in the oxycodone group compared to the morphine group. The incidence of adverse effects increased after the administration of high doses of oxycodone or morphine but remained lower in the oxycodone group than in the morphine group.

    Conclusions: When compared to morphine, oxycodone exhibits a slightly shorter onset of action, a more potent short-term analgesic effect, and fewer adverse effects. This study provides clinical evidence to refine drug regimens in managing cancer pain. Further basic research is expected to elucidate the different mechanisms underlying the effects of oxycodone and morphine.

  • Article
    Rui Wang, Bing Wang, Jiali Li, Xixi Xiang, Xiaoqi Wang, Shijie Yang, Xi Zhang, Li Gao
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2933-2943. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.229
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    Background: The inhibitory effect of dermal mesenchymal stromal cells (DMSCs) on graft versus host disease (GVHD) has been demonstrated in mice, but the underlying mechanism remains unclear. Semaphorin-3A (Sema3A) is an effective regulator in all stages of immune response. The purpose of this manuscript is to investigate the role of Sema3A in the immunosuppressive effect of human DMSCs (hDMSCs).

    Methods: Coculture systems of hDMSCs expressing varying levels of Sema3A and T cells were established in vitro. The effects of hDMSCs expressing different Sema3A levels on T-cell proliferation, apoptosis, cell cycle, subsets, and cytokine secretion were examined.

    Results: Among the hDMSC types, ov3A-hDMSCs exhibited the most significant inhibition of T-cell proliferation. si3A-hDMSCs also suppressed T-cell proliferation, albeit with noticeably weaker effects compared to ov3A-hDMSCs. Sema3A arrested T cells in the G0/G1 phase, with ov3A-hDMSCs inducing S-phase arrest in T cells more potently than si3A-hDMSCs. This effect was associated with the inhibition of cyclin dependent kinase 4 (CDK4) and cyclin D1 and the promotion of p27 expression by Sema3A secreted by hDMSCs. Further investigation revealed that Sema3A secreted by hDMSCs suppressed the phosphorylation of extracellular regulated protein kinases (ERK) and zeta-chain-associated protein of 70 kDa (ZAP70), thus influencing the expression levels of downstream factors cyclin D1 and CDK4, thereby inhibiting T-cell proliferation. Additionally, Sema3A secreted by hDMSCs promoted Th2 and Treg cell differentiation while inhibiting Th1 and Th17 differentiation, thereby impacting cytokine secretion by these cells.

    Conclusions: Sema3A serves as the mediator for the negative regulatory effect of hDMSCs on T lymphocyte responses.

  • Article
    Bárbara Guerra-Carvalho, Ivana Jarak, Rui A. Carvalho, Soraia Pinto, Sónia C. Correia, Alberto Barros, Marco G. Alves, Pedro F. Oliveira
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2945-2958. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.230
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    Background: Aging is the progressive decline in an organism's structural and functional characteristics. Aging has also been linked to a reduction in male reproductive potential. In contemporary societies where delayed parenthood is prevalent, there is a burgeoning interest in comprehending the age-related decline in male fertility and its implications for offspring. This study delves into investigating the metabolome of human seminal fluid, aiming to unveil potential metabolite biomarkers of male reproductive potential during the aging process, employing a Nuclear Magnetic Resonance (NMR)-based metabolomics approach.

    Methods: Seminal samples from 31 healthy normozoospermic men (age range from 25 to 50 years old) undergoing fertility treatment were examined. Routine semen analyses were performed on fresh semen samples to determine the sperm parameters according to the World Health Organization guidelines. Seminal fluid was separated from spermatozoa and the metabolic profile was analyzed through 1H-Nuclear Magnetic Resonance (1H-NMR). The samples were separated into two groups based on age (<35 years and ≥35 years) and multivariate analysis was performed on the quantified metabolites.

    Results: Employing this NMR-based metabolomics approach, we identified 37 small metabolites across all seminal fluid samples. Partial least squares-discriminant analysis unveiled a clustering trend between age groups. Notably, a significant age-associated increase in the amino acids serine and aspartate was observed. Moreover, a propensity for increased levels of various amino acids in seminal fluid with advancing age was observed. Conversely, seminal levels of malate exhibited a decline in older individuals.

    Conclusions: Our findings provide compelling evidence of age exerting a discernible influence on the comprehensive seminal metabolome. These results underscore the potential for future omics studies, focusing on unraveling the intricate molecular mechanisms that underlie the age-associated decline in male fertility.

  • Article
    Ya Cheng, Mengjiao Zhu, Xiangdong Qu, Pan Wang, Ling Zhang
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2959-2969. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.231
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    Background: Fibroblast-derived Thymic Stromal Lymphopoietin (TSLP), an inflammatory cytokine, plays a significant role in regulating the tumor microenvironment and immune responses. This study aimed to investigate the role of TSLP in ovarian cancer (OVCA) metastasis and its underlying mechanisms that modify the immune microenvironment and drive type 2 immunity.

    Methods: The role of TSLP in OVCA metastasis was evaluated using both the in vivo and in vitro experimental models. The expression levels of TSLP, its associated immune cell infiltration and type 2 immune-related factors were evaluated in non-metastatic and metastatic OVCA employing flow cytometry, Quantitative Reverse Transcription Polymerase Chain Reaction (qRT-PCR), and Western blot analysis.

    Results: We found that TSLP was upregulated in fibroblasts and the TSLP receptor (TSLPR) was significantly upregulated in immune cells in the metastatic microenvironment (p < 0.01). Moreover, inhibition of TSLP substantially decreased the migration of both T cells and eosinophils. Furthermore, inhibition of TSLP reduced the formation of metastatic foci and modulated immune response within these foci (p < 0.01). Additionally, both in vivo and in vitro experimental results showed that TSLP promoted OVCA cell migration and invasion, and enhanced type 2 immune response of immune cells (p < 0.01).

    Conclusion: Our findings indicate that fibroblast-derived TSLP plays a crucial role in promoting OVCA metastasis by altering the immune microenvironment and stimulating type 2 immune response. This study provides a theoretical basis for devising novel therapeutic strategies and targeted treatment of TSLP.

  • Article
    Yuxiang Chen, Keke Yang, Jinsuo Yang, Guanmei Xu, Feng Zhou
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2971-2990. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.232
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    Background: Methylation-induced alterations of mucin 3A (MUC3A) expression have been observed in various malignancies, yet few studies have investigated its role in gastric cancer (GC). Herein, we elucidate the role of MUC3A in GC progression and its associated mechanisms.

    Methods: GC cells were treated with 5-Aza-2′-Deoxycytidine (5-Aza) and the phosphatidylinositol 3-kinase (PI3K)/AKT signaling pathway inhibitor LY294002, or transfected with short hairpin RNA targeting MUC3A (shRNA-MUC3A), MUC3A overexpression, and DNA methyltransferase 2 (DNMT2) overexpression constructs. Quantification of MUC3A was conducted using quantitative reverse transcription-PCR (qRT-PCR), while the methylation level of MUC3A was evaluated through methylation-specific PCR. Cell proliferation, migration, invasion, and apoptosis were assessed using 5-Ethynyl-2′-Deoxyuridine (EdU) incorporation assay, wound healing assay, Transwell assay, and flow cytometry, respectively. Additionally, protein expressions of programmed cell death-ligand 1 (PD-L1), phospho-AKT (p-AKT), and total AKT were determined through Western blot analysis.

    Results: Upregulation of MUC3A was observed in GC tissues (p < 0.001), while methylation of MUC3A was nearly absent in tumor tissues. Treatment with 5-Aza promoted proliferation, migration, invasion, and PD-L1 expression while inhibiting apoptosis in GC cells (p < 0.05). Conversely, shRNA-MUC3A demonstrated opposite effects (p < 0.05). Moreover, the effects induced by 5-Aza on GC cells could be reversed by MUC3A knockdown (p < 0.05). Additionally, overexpression of MUC3A enhanced the malignant behaviors of GC cells (p < 0.01), but this effect was reversed by pre-treatment with LY294002, a PI3K/AKT pathway inhibitor (p < 0.05). Furthermore, DNMT2 was found to bind to the MUC3A CpG island site, and overexpression of DNMT2 weakened the impact of MUC3A overexpression on accelerating the progression of GC (p < 0.05).

    Conclusion: Hypomethylation of MUC3A can promote the malignant behaviors of GC cells through the regulation of PD-L1 and the PI3K/AKT pathway, suggesting its potential as a biomarker in gastric carcinogenesis.

  • Article
    Peng Zhao, Tenghua Yu, Lifei Deng, Zhaokun Liu, Yanxiao Huang, Xinhua Zhu, Lixiang Peng
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 2991-3000. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.233
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    Background: Hypopharyngeal cancer (HPC) is a prevalent malignancy commonly occurring in the head and neck region, with a five-year survival rate ranging from 30–50%. Consequently, there is an urgent need to delve into the underlying molecular mechanisms and to identify novel biomarkers that can serve as therapeutic targets for treating HPC.

    Methods: The GSE2379 dataset and HPC data in The Cancer Genome Atlas (TCGA) database were utilized for combination bioinformatics analysis. The mRNA and protein expression levels of thyroid hormone receptor interactor 13 (TRIP13) were evaluated both in vitro and in vivo, as well as in clinical samples using quantitative real-time polymerase chain reaction (qRT-PCR) and western blot (WB) analysis. Furthermore, the malignant phenotypes of HPC cells were assessed employing the Transwell, wound healing, and flow cytometry assays. A xenograft tumor nude mice model was established to observe the tumor growth and lung metastases. Moreover, the role of TRIP13 in regulating the mitogen-activated protein kinase (MAPK) pathway was assessed using bioinformatic analysis.

    Results: Our analysis revealed the participation of 66 common up-regulated differential expressed genes (DEGs) in the epithelial-mesenchymal transition (EMT) related signaling pathway. Furthermore, TRIP13 level was elevated in FaDu cells, xenograft tumor nude mice, and clinical HPC samples. The knocking down of TRIP13 suppressed the invasion, migration, and cell cycle of FaDu cells in vitro, and inhibited tumor growth, lung metastases, and pathological conditions in vivo.

    Conclusion: This study reveals the role of TRIP13, as a potential oncogene, in promoting the development of HPC.

  • Article
    Xiaoning Li, Yiran Meng, Zhongkui Wang, Lihua Sun, Yuanyuan Zhang, Hui Li, Wenqian Yu, Liyan Pan
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3001-3009. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.234
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    Backgrounds: Atherosclerosis is a chronic and progressive vascular disease. Adiponectin, a hormone produced by adipose tissue, has been implicated in the development of atherosclerosis. Therefore, this study aims to explore the biological functions of Adiponectin in vascular cells and its impact on the progression of atherosclerosis.

    Methods: Initially, atherosclerosis-related cell models were established in human coronary artery endothelial cells (HCAEC), human mononuclear cell line (THP-1), and vascular smooth muscle cells (VSMC) by stimulating them with Oxidized Low-Density Lipoprotein (oxLDL). The impact of adiponectin on the proliferation and migration capabilities of oxLDL-stimulated HCAEC, THP-1, and VSMC cells was assessed using the Cell Counting Kit-8 (CCK-8) method and the Transwell assay. The influence of adiponectin on the expression levels of inflammatory factors as well as the levels of Protein Kinase B (AKT) and phosphorylated AKT was determined in oxLDL-stimulated HCAEC, THP-1, and VSMC cells employing Quantitative Reverse Transcription Polymerase Chain Reaction (qRT-PCR) and western blot analysis.

    Results: Adiponectin inhibited the proliferation and migration capabilities of oxLDL-stimulated HCAEC, THP-1, and VSMC cells (p < 0.01, and p < 0.001). Additionally, adiponectin reduced inflammation by modulating the mRNA expression of inflammatory factors in HCAEC, THP-1, and VSMC cells (p < 0.05, p < 0.01, and p < 0.001). Furthermore, Adiponectin decreased the phosphorylation levels of AKT in oxLDL-induced HCAEC, THP-1, and VSMC cells (p < 0.01, and p < 0.001).

    Conclusions: This study suggests that adiponectin may play a role in attenuating atherosclerosis by inhibiting AKT phosphorylation, impeding the migration and proliferation of vascular cells, and reducing inflammation.

  • Article
    Noopur Srivastava, Shiwani Singh, A. Muthukumar, Padmaa M Paarakh, Saad Alobid, Kuntal Das, Ali Ibrahim Almoteer, Moneer E. Almadani, Fuzail Ahmad, Syed Imam Rabbani, Syed Mohammed Basheeruddin Asdaq
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3011-3021. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.235
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    Background: Oxidative stress, which is tightly linked to free radical generation, is cited as contributing to depression. Cichorium intybus L. (CI) is widely known as a viable source of abundant substances with antioxidant properties. In this study, we evaluated the possible antidepressant properties of CI in mice subjected to chronic unpredictable mild stress (CUMS).

    Method: Swiss albino mice were subjected to CUMS induction procedure for 14 days and then treated with imipramine (15 mg/kg) and CI (250 and 500 mg/kg) for 10 days. The treatment was followed by several behavioral parameter evaluations, such as sucrose preference test, forced swim test, open field test, and tail suspension test. Antioxidant enzyme levels, lipid peroxidation degree, and brain serotonin content of the sacrificed animals were assessed.

    Results: The neurochemical analysis of the mice exposed to CUMS revealed significantly decreased serotonin and antioxidant enzyme levels as well as heightened lipid peroxidation (p < 0.001). The mice with CUMS underwent significant behavioral changes, marked by protracted immobility, decreased exploratory behavior, and decreased preference for sucrose solution (p < 0.001). However, these behavioral changes could be reversed by administering CI, which also resulted in elevated antioxidant enzyme levels, diminished lipid peroxidation, and ultimately increased serotonin levels in the brain (p < 0.001).

    Conclusion: Our findings demonstrated that CI holds immense promise as a therapeutic agent for depression by strengthening the antioxidant defense system to mitigate oxidative stress, decrease lipid peroxidation, and preserve serotonin levels in the brain.

  • Article
    Zhengwen Sun, Xueyan Liu, Dong Han, Yinghui Sui, Qiaoling Zhu
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3023-3033. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.236
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    Background: Periosteal stem cells (PSCs) are pivotal in bone tissue repair and regeneration. This study explores the effect of Loxoprofen (Lox) on PSCs’ osteogenic differentiation, focusing on the Wingless/Integrated/Extracellular Signal-Regulated Kinase (Wnt/ERK) signaling pathway and mitochondrial dynamics.

    Methods: Initially, we induced osteogenic differentiation in PSCs and exposed them to varying concentrations of Lox (5 μg/mL, 10 μg/mL, 20 μg/mL). Employing a murine fracture healing model and Hematoxylin and Eosin (H&E) staining, we assessed Lox's in vivo impact on fracture healing in mice. PSCs' proliferation, adhesion, and migration effects under Lox influence were scrutinized using Cell Counting Kit-8 (CCK-8), cell adhesion experiments, and Transwell assays. Furthermore, Lox's effects on Alkaline Phosphatase (ALP) activity, mineralized nodules, osteogenic markers, and Wnt/ERK pathway-associated proteins during PSCs' osteogenic differentiation were analyzed via Enzyme-Linked Immunosorbent Assay (ELISA), ALP staining, Alizarin Red staining, western blot, and Quantitative Real-Time Polymerase Chain Reaction (qRT-PCR). Microscopy was utilized to explore Lox's impact on mitochondrial morphology and cytoskeleton in PSCs post-osteogenic differentiation. Additionally, we measured Lox's effects on adenosine triphosphate (ATP) levels, reactive oxygen species (ROS) levels, and mitochondrial membrane potential in PSCs after osteogenic differentiation with ELISA and flow cytometry. Finally, we demonstrated the inhibitory effect of Wnt/ERK pathway inhibitors (Dickkopf-1 (DKK1)/SCH772984) on Lox-induced Wnt/ERK signaling pathway activation using western blot.

    Results: Our findings confirm that Lox, at low, medium, and high doses, increases the proliferation, adhesion, and migration capacities of PSCs (p < 0.05). Lox demonstrates a pronounced elevation in the osteogenic potential of PSCs while significantly promoting mitochondrial activity, fusion, and progress of fracture healing (p < 0.05). Additionally, Lox treatment leads to significant enhancement in the activation of the Wnt/ERK signaling pathway (p < 0.05). The addition of Wnt/ERK pathway inhibitors significantly suppressed the effect of Lox on the Wnt/ERK signaling pathway (p < 0.05).

    Conclusions: Lox facilitates PSCs’ osteogenic differentiation by enhancing the Wnt and ERK signaling pathways and mitochondrial dynamics, contributing to improved fracture healing.

  • Article
    Ziyuan Zhang, Haijuan Hu, Demin Liu, Xue Geng, Huimin Sun, Wei Cui
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3035-3045. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.237
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    Background: DL-3-n-Butylphthalide (NBP) sodium chloride injection is widely used in treating cerebral ischemia stroke, yet it may lead to adverse effects, including a reduction in heart rate (HR). However, the specific component responsible for affecting cardiac function remains unclear.

    Aim: This study aimed to elucidate the component of NBP that affects cardiac function and delineate its mechanisms of action.

    Methods: The Langendorff perfusion system was employed to investigate the effects of NBP and its principal components on the functional parameters of isolated rat hearts. The apoptosis/necrosis assay kit evaluated the impact of β-cyclodextrin (β-CD) on cardiomyocyte apoptosis and necrosis. Furthermore, Di-8-ANEPPS staining was performed to visualize T-tubule disruptions. Finally, patch clamp techniques were employed to examine alterations in L-type calcium channel function.

    Results: β-CD, rather than butylphthalide, impaired cardiac function. Higher concentrations or longer perfusion times of β-CD resulted in significant reductions in HR (p < 0.05). Moreover, increased concentrations or longer perfusion time of β-CD led to elevated left ventricular systolic pressure (LVSP) and left ventricular end-diastolic pressure (LVEDP) (p < 0.05). β-CD perfusion also decreased the maximal left ventricular pressure rising rate (dp/dt max) and -dp/dt max (p < 0.05). Treatment with different concentrations of β-CD significantly decreased B-cell lymphoma-2 associated X protein (Bax) expression levels and increased cardiomyocyte apoptosis and necrosis rates (p < 0.01). Pre-treatment and post-treatment with cholesterol significantly mitigated β-CD-induced cardiomyocyte necrosis, with post-treatment exhibiting a stronger effect than pre-treatment (p < 0.05). Di-8-ANEPPS staining revealed the stronger concentration-dependent disruption of T-tubule integrity and continuity following β-CD treatment. Furthermore, paracellular administration of β-CD did not alter the current density or activation curve trend of L-type calcium channels but caused a rightward shift in the activation curve.

    Conclusion: β-CD, a principal component of NBP, induces a reduction in HR, which depends on the concentration and duration of β-CD treatment. The mechanism underlying this effect may involve cardiomyocyte necrosis, T-tubule damage, and modulation of L-type calcium channel function.

  • Article
    Jin-Zhu Zhuang, Jun Zhang, Yu-E Zhang, Di Zhang, Hui Sun, Yun Su, Shu-Feng Li
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3047-3055. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.238
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    Background: Retinal ganglion cells (RGCs) affected by ischemia cause irreversible visual impairment. Hydrogen sulfide (H2S) may protect RGC-5 from oxidative damage by increasing glutathione levels. This study aimed to evaluate the ability of H2S to reduce oxygen-glucose deprivation (OGD)-induced apoptosis in RGC cells by activating ATP-sensitive potassium (KATP) channels.

    Methods: First, purified primary RGCs were cultured and treated with OGD or OGD + NaSH. Cell mortality was observed through Hoechst-33258 staining and enzymatic immunoassay. Then, the RGCs were treated with pinacidil or glibenclamide (ATP-sensitive potassium channel agonist or inhibitor) to observe whether H2S protects against OGD-induced cell damage through activation of ATP-sensitive potassium channels using patch clamp, immunoblot and calcium imaging techniques.

    Results: Under OGD, apoptotic cell death was effectively prevented by NaSH. Simultaneous treatment with 300 μM NaSH and OGD exposure delayed cell death. H2S protected RGCs at an early stage after OGD and delayed the start of OGD injury. Resting potential (RP) was significantly hyperpolarized after NaSH or pinacidil exposure compared to vehicle (p < 0.05). Hyperpolarization due to NaSH was induced by the activation of KATP channels. RGCs apoptosis induced by OGD was suppressed by H2S through the activation of the KATP channel.

    Conclusion: H2S reduced calcium efflux and prevented OGD-induced neuronal death in primary RGCs, and the neuroprotective effect of H2S was associated with the activation of KATP channels.

  • Article
    Arshad Iqbal, Ayaz Ali Sher, Syed Lal Badshah, Muhammad Adil, Sher Wali, Ghallab Alotaibi, Muhammad Ayaz, Nada M. Mostafa
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3057-3072. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.239
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    Background: Withania somnifera (L.) Dunal is traditionally used as laxative, diuretic and sedative remedy. The current study was aimed to perform phytochemical analysis, antioxidant, cholinesterase inhibitory, laxative, prokinetic and spasmolytic potentials of the plant using in-vitro, in-vivo and in-silico approaches.

    Methods: Phytochemical analysis was performed using Gas Chromatography-Mass Spectrometry (GC-MS). In-vitro cholinesterase inhibitory and antioxidant studies were performed using standard procedures. The laxative effect was evaluated using pigeons and rabbits whereas the spasmolytic study was performed using rabbits jejunal slices. In-silico study was performed against the identified compounds using Discovery studio version 4.1.

    Results: The GC-MS revealed the presence of several bioactive compounds. Methanolic extract (MEWS) and n-butanol fraction (NBFWS) showed antioxidant potential using 2,2′-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS) and 2,2-Diphenyl-1-picrylhydrazyl (DPPH) assays with half maximal inhibitory concentration (IC50) values of (90 μg mL-1, 180 μg mL-1) and (170 μg mL-1, 169 μg mL-1) respectively. Furthermore, they inhibited acetylcholinesterase (AChE) with IC50 values of 50 and 150 μg mL-1 respectively. Both samples exhibited prokinetic effects in metronidazole and loperamide hydrochloride-induced constipation models. Metronidazole was found to be effective by reducing the percentage of wet feces as compared to loperamide hydrochloride. Both samples promoted intestinal transit (propulsion) compared to distilled water. The NBFWS showed significant spasmolytic activity with an effective concentration (EC50 = 0.85 ± 0.31 mg mL-1) in rabbit jejunum when compared with MEWS (EC50 = 4.3 ± 0.5 mg mL-1). Similarly, NBFWS exhibited significant relaxing activity in potassium chloride (KCl)-induced (80 mM) contraction (EC50 = 3.41 ± 0.67 mg mL-1) compared to MEWS (EC50 of 9.6 ± 0.4 mg mL-1). In-silico molecular docking study against the active site of AChE enzyme revealed that 8-cis Octadecenoic acid methyl ester exhibited C-docker energy of –50.0022 Kcal/mol relative to the co-crystallized ligand (–43.6427 Kcal/mol).

    Conclusions: The purpose of the current study was to find a natural remedy which can overcome the problem associated with constipation and to regulate the GIT motility. The results validated the use of Withania somnifera(W. somnifera) in the treatment of gastrointestinal disorders like constipation and spasms, yet further work is warranted to isolate and therapeutically validate its bioactive phytochemicals.

  • Article
    Min Kong, Lishan Jiang, Tong Wu, Huadong Ni, Xu Shen, Xuyan Zhou
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3073-3078. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.240
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    Background: Neurotensin (NT), an endogenous neuropeptide, plays a crucial role in modulating dopaminergic transmission. Dopaminergic drugs may serve as an effective therapy against chronic pain, a frequent adverse symptom observed in spinal cord injury (SCI) patients. Therefore, we aimed to investigate whether the mechanism of NT for chronic pain after SCI involves the modulation of dopaminergic neuronal activity.

    Methods: The SCI rat model was used with or without NT administration based on the experimental design. Ultrasonic vocalizations (USVs) and von Frey filaments were performed to assess the effect of NT on pain behavior in SCI rats. Dopaminergic neuron firing and bursting within the ventral tegmental area (VTA) were examined using electrophysiology. Furthermore, the dopamine D2 receptor (D2DR) was quantified using quantitative real-time polymerase chain reaction and immunohistochemistry.

    Results: Significant spontaneous and induced pain could be observed in SCI rats along with a decrease in dopaminergic neuron firing and bursting activities as well as upregulation of D2DR expression (p < 0.001). Importantly, NT treatment significantly alleviated pain symptoms and promoted dopaminergic neuron firing and bursting activities in SCI rats (p < 0.001). Additionally, NT treatment diminished D2DR mRNA level and positive expression in SCI rats (p < 0.001).

    Conclusion: NT alleviates expression level of D2DR and activates dopaminergic firing and bursting activities in the VTA region, thereby relieving SCI-induced chronic pain.

  • Article
    Shihai Zhou, Fuxin Zheng, Zheng Zheng, Jinlong Yu
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3079-3090. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.241
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    Objectives: Previous studies have demonstrated that tumor-associated macrophages (TAMs) have a significant impact on the initiation and progression of cancer. Interactions between cells within the tumor microenvironment (TME) are mediated by extracellular vesicles (EVs), which transport various functional molecules such as mRNA, miRNA, proteins, and lipids. However, the specific molecular mechanisms that regulate the interaction between EVs derived from tumors and TAMs in the context of liver metastasis of colorectal cancer liver metastasis (CRLM) remain unclear. Our goal is to show that EVs originating from colorectal cancer cells stimulate the M2 polarization of macrophages.

    Methods: We examined the relationship between macrophages and the potential impact of EVs derived from colorectal cancer cells in this study. Transmission electron microscopy (TEM) and differential ultracentrifugation were utilized to validate the presence of EVs. Several methods, including dual-luciferase reporter assay, quantitative polymerase chain reaction (qPCR), and western blotting, were employed to elucidate the mechanism by which EVs miR-28-5p regulate the interaction between colorectal cancer (CRC) cells and macrophages.

    Results: Initially, we observed an increased expression of miR-28-5p in CRC patients, specifically in CRLM patients. Analysis of data from The Cancer Genome Atlas (TCGA) and The Encyclopedia of RNA Interactomes (ENCORI) databases revealed its association with poor prognosis in CRC patients. Subsequently, we verified that miR-28-5p, derived from EVs of colorectal cancer cells, induced the polarization of macrophages by inhibiting phosphatase and tensin homolog deleted on chromosome ten (PTEN) and activating phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT). Additionally, we identified that M2 macrophages polarized by CRC cell-derived EV-miR-28-5p promoted epithelial-mesenchymal transition (EMT) and facilitated CRLM.

    Conclusions: In summary, our findings show that miR-28-5p in EVs derived from CRC can promote CRLM by regulating TAM polarization. Furthermore, we have uncovered another mechanism, mediated by tumor-derived EVs, that influences CRLM.

  • Article
    Chunyu Li, Shuo Yang, Haochen Xia, Yufeng Jin
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3091-3099. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.242
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    Background: The disorders of bone resorption and bone formation processes may contribute to the progression of osteoporosis (OP). Moreover, the human bone marrow mesenchymal stem cells (hBM-MSCs) are precursor cells of osteoblasts. Exploring the differentiation mechanism of hBM-MSCs holds immense significance for the prevention and treatment of OP. Furthermore, the steroid receptor coactivator-3 (SRC-3), an important co-activator of estrogen receptor and estrogen-related receptor α, may serve as an important regulator of bone metabolism. However, there are limited studies available on the role as well as the molecular mechanism of SRC-3 in postmenopausal OP (PMOP). Therefore, this study investigated the role of SRC-3 in hBM-MSCs osteogenic differentiation (OD) and delved into its underlying regulatory mechanism.

    Methods: A total of 60 study subjects, including 30 PMOP patients (OP group) and 30 healthy menopausal women (non-OP (NOP) group) were recruited in this study. Serum SRC-3 levels were assessed using enzyme-linked immunosorbent assay (ELISA). Furthermore, 10 nmol/L of dexamethasone, 10 mmol/L of β-glycerolphosphate, and 50 μg/mL of ascorbic acid were added to the culture medium to induce OD in hBM-MSCs. The expression level of SRC-3 was evaluated employing real-time quantitative polymerase chain reaction (RT-qPCR) and Western blot analysis following OD induction. The impact of SRC-3 on the OD and mineralization of hBM-MSCs was assessed by analyzing alkaline phosphatase (ALP) activity and alizarin red staining (ARS) procedure. Furthermore, changes in the expression levels of OD indicators and Wnt3a/β-catenin were determined by RT-qPCR and Western blot analysis.

    Results: The expression level of SRC-3 was significantly reduced in the serum of PMOP patients (p < 0.0001). However, OD was successfully induced (p < 0.01, p < 0.001, p < 0.0001) and the expression of SRC-3 was upregulated by SRC-3 overexpression vector (p < 0.01, p < 0.001). Furthermore, SRC-3 overexpression promoted OD of hBM-MSCs (p < 0.01, p < 0.001, p < 0.0001) as well as up-regulated the expression of Wnt3a and β-catenin (p < 0.05, p < 0.01, p < 0.001, p < 0.0001). Moreover, the inhibition of Wnt3a or β-catenin was found to reverse the promotion of SRC-3 on OD of hBM-MSCs (p < 0.05, p < 0.001, p < 0.0001).

    Conclusion: In summary, we observed significantly reduced serum SRC-3 level in PMOP patients. Furthermore, SRC-3 overexpression promoted OD and mineralization of hBM-MSCs through the activation of the Wnt/β-catenin signaling pathway, thereby ameliorating OP.

  • Article
    Xiangzhi Zhang, Haiyan Zhang, Xinna Li
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3101-3109. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.243
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    Background and Objective: Vemurafenib (VEM) is commonly used to treat papillary thyroid cancer (PTC) with the B-Raf proto-oncogene, serine/threonine kinase (BRAF)V600E mutation. However, a major challenge during treatment is the emergence of cellular resistance to vemurafenib. Furthermore, FKBP Prolyl Isomerase 10 (FKBP10) has been associated with the occurrence and progression of various cancer types. Therefore, the current study aimed to investigate the role of FKBP10 in the emergence of cellular resistance to vemurafenib within PTC patients.

    Methods: The potential of FKBP10 in the proliferation of VEM-resistant cells was assessed in vitro using the Cell Counting Kit-8 (CCK-8), colony formation, and cell migration assays. Western blotting, Co-Immunoprecipitation, Quantitative Reverse Transcription Polymerase Chain Reaction (qRT-PCR), and confocal immunofluorescence labeling assay were employed to investigate the underlying molecular mechanism of FKBP10 overexpression in VEM-resistant cells.

    Results: Compared to Nthy-ori 3-1, expression levels of FKBP10 were substantially increased in PTC cells. Furthermore, the knockdown of FKBP10 significantly increased their drug sensitivity. However, resistant cell lines exhibited a significant reduction in cell survival, migration, and colony formation (p < 0.05) and elevated apoptotic rate. Additionally, silencing FKBP10 (sh-FKBP10) significantly decreased the expression levels of p-Protein kinase B (AKT) and p-Phosphoinositide 3-kinase (pI3K) in human thyroid cancer papillary cells (p < 0.05). Similarly, treatment with PI3K inhibitor LY294002 and AKT inhibitor MK-2206 substantially decreased the expression levels of the p-AKT and p-pI3K proteins (p < 0.05) and hence reduced cell survival. Furthermore, Co-Immunoprecipitation (Co-IP) and confocal microscopy experiments revealed a significant interaction between the PI3K/AKT signaling pathway and FKBP10.

    Conclusions: The present study shows that vemurafenib resistance can be reversed through FKBP10 knockdown.

  • Article
    Jia-lu Li, Ju-qun Xi
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3111-3121. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.244
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    Background: Lung cancer is the leading cause of cancer-related deaths, where non-small cell lung cancer (NSCLC) accounts for approximately 85% of all lung cancer cases. The potential of apatinib, a small-molecule anti-angiogenic drug, and salidroside, the Chinese herbal extract, as anti-tumor agents has been recognized. Therefore, this study aims to investigate the synergistic effects of apatinib and salidroside on NSCLC and provide insights into this combination to evaluate their combined potential in treating NSCLC.

    Methods: 3-(4,5-dimethylthiazol)-2,5-diphenyl-tetrazolium bromide (MTT) assay was employed to determine the inhibitory effects of different concentrations of apatinib, salidroside and their combination on NSCLC cell proliferation. The levels of reactive oxygen species (ROS) in NSCLC cells were assessed in treated cells. Furthermore, flow cytometry, wound healing assay, and tube formation assay were used to determine the apoptosis rate, migration ability, and angiogenesis of cells, respectively. Additionally, western blot analysis was utilized to evaluate the levels of apoptosis-related proteins and mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK)/vascular endothelial growth factor (VEGF) axis-related proteins.

    Results: Compared to using either apatinib or salidroside for treatment alone, apatinib in combination with salidroside (Sal) significantly inhibited NSCLC cell proliferation, migration, and tube formation (p < 0.01). Meanwhile, this combination ((apatinib) Apa+Sal) induced ROS production and promoted apoptosis by up-regulating Bcl-2 associated X (Bax) expression and down-regulating B cell lymphoma-2 (Bcl-2) expression (p < 0.01). Western blot analysis revealed a substantial increase in the phosphor (p)-p38 expression, and decreased p-ERK, VEGF, and VEGF receptor (VEGFR) levels in the combined treated group compared to the single treatment group (p < 0.05). Combined treatment coupled with MAPK/ERK inhibitor (SB203580) or activator (anisomycin) significantly enhanced or decreased the inhibitory effect of apatinib and salidroside on human umbilical vein endothelial cells (HUVEC) tube formation, respectively.

    Conclusion: Apatinib in combination with salidroside exerts a synergistic anticancer effect in NSCLC by down-regulating the MAPK/ERK/VEGF signaling axis, which suggests the potential of this combined therapy in treating NSCLC.

  • Article
    Sara Hosseinchi Gharehaghaji, Fatemeh Sadat Abtahi, Emad Khalilzadeh, Sina Pakkhesal, AmirMohammad Sharafi, Ali Ahmadalipour
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3123-3133. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.245
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    Background: Major Depressive Disorder (MDD) is a disabling multifactorial neuropsychiatric disorder in which several neurobiological systems and signaling pathways are involved. Physical activity improves mood and displays antidepressant effects. However, the exact mechanism underlying these effects from different exercise sessions has not been comprehensively studied. The aim of this study was to investigate the effects of short-term exercise on depressive behaviors and also its possible modulatory effects on three critical pathways, which are dysregulated in depressive pathophysiology, including serotonergic tryptophan hydroxylase 2 (TPH2), mitogen-activated protein kinase (MAPK) including p38 and c-Jun N-terminal kinases (JNK), and cannabinoid receptor 1 (CB1) systems in the stress-induced rat model of depression.

    Methods: Following seven weeks of chronic unpredictable mild stress (CUMS) protocol, male Wistar rats were exposed to 60 minutes of treadmill-running exercise (10 min preparatory; 50 min training session, 10 m/min) for five consecutive days. Then, the anti-depressive effects were evaluated by the forced swimming test (FST) and the open field test (OFT), and finally, rats were decapitated, and their prefrontal cortex (PFC) was extracted for further analysis.

    Results: The results showed that CUMS induced a significant increase in immobility time in the FST (p < 0.01), which was reversed by short-term exercise (p < 0.01), and a slight decrease in horizontal distance in the OFT. Moreover, CUMS increased the protein expressions of CB1 and p-JNK/JNK in PFC (p < 0.001), and no effect of exercise was observed on these markers (p > 0.05). Interestingly, we found that the CUMS-induced p-p38/p38 ratio increment was reversed by the training session (p < 0.001). Meanwhile, short-term exercise also significantly upregulated TPH2, whose level was diminished in CUMS rats.

    Conclusions: Our results reveal that short-term exercise alleviates depression behavior induced by chronic stress and improves the 5-hydroxytryptamine (5-HT) neurotransmission regulating enzyme, TPH2. In addition, the observed reduction of p38 phosphorylation may play a key modulatory role in the anti-depressive-like properties of short-term exercise, which needs further study to approve the exact mechanisms.

  • Article
    Shunfu Deng, Hui Hu, Jinke Lv, Kaixi Liu, Zhaoqian Lin, Zhenyu Wang, Boru Shen, Xuhui Wu, Minqian Huang, Jiaqing Wang, Changshan Song
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3135-3146. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.246
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    Objective: This study aimed to investigate the molecular insights by which microRNA (miR-522-3p) in cancer-associated fibroblast (CAF) derived exosomes regulate lung cancer cell growth, proliferation, and invasive capabilities.

    Method: CAFs and normal fibroblasts (NFs) were isolated from lung cancer and adjacent non-cancerous/normal tissues and identified. Exosomes secreted from CAFs and NFs were isolated to analyze their effects on lung cancer cell proliferation, migration, and invasion by Cell Counting Kit-8 (CCK-8), colony formation, and Transwell assays. Next-generation sequencing (NGS) technology was applied to analyze microRNAs (miRNA) expression within the exosomes, specificaly measuring miR-522-3p. Additionally, a dual-luciferase reporter gene assay was utilized to validate the binding of zinc-finger RAN-binding domain-containing protein 2 (ZRANB2) to miR-522-3p.

    Results: The proliferation ability of cells in the CAF-exosomes (exo) groups significantly increased compared to NF-exo group (p < 0.05). Up-regulated miR-522-3p expression level was found in CAFs-exo (p < 0.001). Functional experiments demonstrated that miR-522-3p inhibitor can suppress lung cancer cell proliferation, migration, and invasion compared to the control inhibitor-exo groups (p < 0.05). ZRANB2 was a target gene of miR-522-3p.

    Conclusions: The present study highlights the role of CAFs in promoting lung cancer progression and metastasis via exosomal miR-522-3p, suggesting that the inhibition of CAF-derived miR-522-3p is an alternative modality for lung cancer treatment. These outcomes also provide a new perspective for further research on the lung cancer microenvironment and the development of treatment strategies.

  • Article
    Yang Zhang, Rong Chen, Juan Wang, Jingsong Zhu, Xu Han, Zhonglu Jian, Jian Tang, Linghao Wang, Huiyuan Ming, Songjiang Tang, Baojun Min
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3147-3153. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.247
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    Background: Cognitive dysfunction resulting from sleep deprivation has received widespread attention. However, the impact of postpartum sleep disorders (PSD) arising from childbirth-related factors on maternal cognitive function remains unclear. Butorphanol, a potential pharmacological agent, may play a crucial role in improving cognitive function in postpartum sleep disorders.

    Objective: This study aimed to investigate the impact of butorphanol on cognitive function among rats with postpartum sleep disorders and elucidate the underlying molecular mechanisms.

    Methods: A PSD rat model was established 24 hours after natural delivery. The rats were divided into three groups: the control group, the model group, and the butorphanol group. Furthermore, neurological function, brain tissue damage, apoptosis rate, expression levels of inflammatory factors, and related signaling pathways were assessed employing Enzyme-Linked Immunosorbent Assay (ELISA), Quantitative Reverse Transcription Polymerase Chain Reaction (qRT-PCR), and Western blot analysis. The Morris water maze test was used to evaluate cognitive function in rats.

    Results: The butorphanol-treated group of rats exhibited significant improvement in sleep, with reduced wakefulness compared to the model group (p < 0.05). In the Morris water maze test, rats in the butorphanol-treated group showed significantly decreased escape latency and increased platform entries (p < 0.05), indicating improved cognitive function after treatment. Furthermore, butorphanol reduced the expression levels of inflammatory factors (Tumor Necrosis Factor-alpha (TNF-α), Interleukin (IL)-1β, and IL-6) by modulating the Toll-Like Receptor 4/Nuclear Factor Kappa B (TLR4/NF-κB) signaling pathway (p < 0.05), thereby exerting anti-inflammatory and anti-apoptotic effects.

    Conclusion: Butorphanol can enhance cognitive function in postpartum sleep-disordered rats by regulating the TLR4/NF-κB signaling pathway, providing a theoretical basis for its clinical application in treating postpartum sleep disorders.

  • Article
    Zhongwei Ai, Fan Yang, Xue Wu, Wei Dou
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3155-3165. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.248
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    Objective: The study aims to investigate the role of Glucocorticoid-induced tumor necrosis factor (TNF) receptor superfamily 18 (TNFRSF18, also called GITR) in the proliferation and migration of breast carcinoma cells.

    Methods: A total of 94 patients with breast carcinoma who underwent surgical excision were enrolled in this study. Cancer tissues and para-cancer tissues were obtained during the operation. Human breast carcinoma cell lines MCF-7 plus MDA-MB-231 were cultivated and transfected. The mRNA or protein levels of TNFRSF18, ubiquitinated-like protein 3 (UBL3), CyclinD1, and matrix metalloproteinase-2 (MMP-2) in breast cancer, para-cancer tissues, MCF-7 cells, and MDA-MB-231 cell lines were quantified using quantitative real-time polymerase chain reaction (qRT-PCR) or western blot analysis. Cell proliferation activity was assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. The interaction between TNFRSF18 and UBL3 was verified by immunoluciferase reporter assay.

    Results: The levels of TNFRSF18 in breast cancer samples were remarkably higher than those in para-cancer samples (p < 0.001). Conversely, the mRNA and protein levels of UBL3 in cancer tissues were significantly lower than those in normal samples (p < 0.001). Both TNFRSF18 and UBL3 mRNA levels were found to be correlated with clinical stage, histological grade, as well as expression levels of the nuclear protein Ki67 and Bromodeoxyuridine (BrdU), respectively. TNFRSF18 overexpression was observed to enhance the proliferation of MDA-MB-231 and MCF-7 cell lines (p < 0.05). Conversely, inhibition of TNFRSF18 or overexpression of UBL3 led to a decrease in the proliferation of MDA-MB-231 and MCF-7 cell lines. Furthermore, an immunoluciferase reporter assay confirmed that UBL3 was a target of TNFRSF18 and was negatively regulated by TNFRSF18. The inhibitory effect of TNFRSF18 downregulation on MDA-MB-231 and MCF-7 cell proliferation was reversed by UBL3 knockdown.

    Conclusions: The expression of TNFRSF18 is upregulated, while UBL3 expression is downregulated in breast carcinoma samples. TNFRSF18 promotes the proliferation of breast cancer by suppressing UBL3.

  • Article
    Shuming Yuan, Guang Li, Feifei Feng
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3167-3176. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.249
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    Background: Dry eye disease (DED) is closely coupled with ocular surface inflammation. Irigenin, an active ingredient in traditional Chinese medicine Belamcanda chinensis (L.) DC, possesses various pharmacological effects including anti-inflammation. This study aims to reveal the impact of irigenin on DED.

    Methods: DED model cells in vitro were constructed by 24-h hyperosmolarity intervention (90 mM sodium chloride (NaCl)) and treated with different concentrations (10, 20 and 40 μM) of irigenin in the same time. Cell counting kit-8, 5-ethynyl-2′-deoxyuridine, Transwell, enzyme-linked immunosorbent assays, and flow cytometry, were used to determine the effect and mechanism of irigenin on DED. The expressions of pyroptosis-related proteins were measured by Western blot. Hyaluronan synthase 2 (HAS2) and HAS3 expressions were quantified by quantitative reverse transcription polymerase chain reaction.

    Results: Irigenin exerted no cytotoxicity on HCE-2 cells at concentrations of 10, 20 or 40 μM. Irigenin enhanced the viability, proliferation, migration, and inhibited the apoptosis of hyperosmolarity-induced HCE-2 cells, but these effects were reversed by interleukin 18 (IL-18) overexpression. Irigenin decreased the level of IL-1β, tumor necrosis factor alpha (TNF-α) as well as the expressions of Gasdermin-D (GSDMD), IL-18 and caspase-1, but increased the expressions of HAS2 and HAS3 in hyperosmolarity-induced HCE-2 cells. These effects of irigenin on inhibiting inflammation and pyroptosis as well as on promoting hyaluronic acid synthesis in hyperosmolarity-induced HCE-2 cells were reversed by IL-18 overexpression.

    Conclusion: Irigenin protected HCE-2 cells against hyperosmolarity-induced inflammation and dysfunction by reducing IL-18-mediated pyroptosis. Irigenin has potential in developing therapeutic agents for DED.

  • Article
    Rashita Makkar, Aayush Sehgal, Sukhbir Singh, Neelam Sharma, Ravi Rawat, Monica Gulati, Celia Vargas-De-La-Cruz, Nemat Ali, Havagiray R Chitme, Tapan Behl, Simona Gabriela Bungau
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3177-3187. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.250
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    Background: Rheumatoid arthritis is a chronic inflammatory autoimmune disorder characterized with destruction and degradation of synovial joints. The current study aims to evaluate the physical and hematological effects of gallic acid (GA) isolated from Tecoma stans leaf extract and further synthesis of its gallic acid phytosome (GAP) to evaluate antiarthritic activity in Complete Freund's Adjuvant induced rheumatoid arthritis in Wistar rats.

    Methods: The experimental rats were categorized into nine groups (n = 6 each group) namely normal control, disease control, methotrexate (0.75 mg/kg) treated group, GA 50 mg/kg treated group, GA 100 mg/kg treated group, GAP 50 mg/kg treated group, GAP 100 mg/kg treated group, GA 100 mg/kg treated group + methotrexate (0.75 mg/kg), GAP 100 mg/kg treated group + methotrexate (0.75 mg/kg) for 21 days. All the parameters were assessed at the end of the study. To assess the role of GA in mediating anti-arthritic activity through inhibition of NLR family pyrin domain containing 3 (NLRP3) inflammasome, an additional Molecular Dynamics (MD) simulation was also conducted.

    Results: GA and GAP significantly decreased arthritic score (p < 0.01, p < 0.05, and p < 0.001), paw volume, pain latency (p < 0.01, p < 0.05, and p < 0.001) and increased body weight (p < 0.01, p < 0.05, and p < 0.001) in a dose-dependent manner in arthritic rats. The level of Haemoglobin (Hb) and number of red blood cells (RBCs) were significantly increased while the levels of white blood cell (WBC), platelet count and erythrocyte sedimentation rate (ESR) were drastically dropped in treatment groups compared to disease control group (p < 0.001, p < 0.01 and p < 0.05). During the MD simulation, GA demonstrated the formation of 2–3 stable hydrogen bond contacts with the NLR family pyrin domain containing 3 (NLRP3, previously known as NACHT domain), primarily involving the residues Threonine439 (THR439), Single Amino Acid of Gα16 Alanine228 (ALA228), and Arginine578 (ARG578).

    Conclusion: The results produced by GAP were higher than GA indicating the benefit of formulation in providing better efficacy and suggesting its potential role in rheumatoid arthritis. The MD simulation result also suggested that GA has the potential to function as an inhibitor of NLRP3 protein and thus expressing anti-arthritic activity.

  • Article
    Ling Jiang, Lifang Duan, Ru Wang
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3189-3199. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.251
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    Background: The NOD-like receptor protein 3 (NLRP3) inflammasome plays a crucial role in the immune microenvironment. Therefore, we investigated the involvement of NLRP3 in the inflammatory response associated with term and preterm birth, as well as the potential of N-acetylcysteine (NAC) to mitigate this inflammatory response.

    Methods: First, we induced infectious preterm birth in mice using Lipopolysaccharide (LPS) and treated them with NAC. Next, LPS was used to trigger inflammation in uterine smooth muscle cells (USMC), which were then treated with NAC. The levels of cleaved caspase-4/5/11, NLRP3, and Apoptosis-associated speck-like protein containing CARD (ASC) proteins in both the myometrium and USMC were determined by western blot. Enzyme-linked immunosorbent assay (ELISA) was used to quantify the levels of caspase-1, interleukin-1β (IL-1β), and IL-18 in the myometrium and USMC. Subsequently, siRNA targeting NLRP3 was transfected into LPS-treated USMC, and the expression levels of NLRP3, oxytocin receptor (OTR), ASC, connexin 43 (CX43), and inflammatory cytokines were evaluated using western blotting and quantitative reverse transcription polymerase chain reaction (qRT-PCR).

    Results: The NLRP3 inflammasome, cleaved caspase-4/5/11, and ASC were found to be upregulated in USMC and myometrium tissues treated with LPS (p < 0.01). Administration of NAC not only prolonged the duration of pregnancy but also suppressed the activation of NLRP3 and the noncanonical pyroptosis pathway (p < 0.01 and p < 0.001). Deletion of NLRP3 under LPS conditions led to a decrease in the expressions of OTR and CX43 (p < 0.05), along with a reduction in the levels of associated inflammatory factors (p < 0.05).

    Conclusions: NAC inhibits immune inflammation and promotes uterine quiescence by suppressing the noncanonical pyroptosis pathway and NLRP3 inflammasome activation.

  • Article
    Manwen Yang, Yuluo Huang, Di Zhang, Xiuhai Lu, Xianwen Wang, Tengchuan Jin, Lei Shi
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3201-3214. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.252
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    Background: Fungal keratitis (FK) is characterized by corneal inflammation induced by fungal infection. Thymic stromal lymphopoietin (TSLP) is crucial in FK pathogenesis. This study aimed to explore the impact of TSLP on FK and elucidate the mechanism of TSLP signaling on macrophage-mediated inflammation.

    Methods: Transformed human corneal epithelial cells (THCEs) were stimulated with heat-killed fragments of Aspergillus fumigatus hyphae. TSLP secretion was assessed using quantitative polymerase chain reaction (qPCR), enzyme-linked immunosorbent assay (ELISA), and western blotting (WB). THCEs stimulated with hyphae or recombinant thymic stromal lymphopoietin (rTSLP) were co-cultured with tohoku hospital pediatrics-1 (THP-1) cell-derived macrophages. The inflammatory activity of rTSLP was evaluated in THP-1 macrophages through real-time PCR, ELISA, WB, and flow cytometry. Additionally, high-throughput transcriptome sequencing was employed to elucidate the underlying mechanisms.

    Results: Secretion of TSLP by A. fumigatus-stimulated THCEs was correlated with increased inflammation in THP-1 macrophage. Treatment with rTSLP resulted in elevated expression of proinflammatory markers associated with macrophage type 1 (M1) phenotype, including tumor necrosis factor-alpha (TNF-α) (p < 0.01), interleukin-1 beta (IL-1β) (p < 0.01), inducible nitric oxide synthase (iNOS) (p < 0.001), and CD86 (p < 0.001) at 24 hours. Exposure of THP-1 macrophages to rTSLP induced activation of the nuclear factor kappa B (NF-κB) signaling pathway. The improved level of phosphorylated NF-κB and inflammatory factors (iNOS, IL-1β, TNF-α) were attenuated by a specific NF-κB inhibitor.

    Conclusions: In A. fumigatus keratitis, TSLP secreted by THCEs promoted TNF-α, IL-1β, and iNOS secretion in THP-1-derived macrophages by upregulating the NF-κB signaling pathway. This led to dysregulation of the pro-inflammatory (M1)/alternative (M2) macrophage ratio and polarization of THP-1 macrophages towards the pro-inflammatory phenotype.

  • Article
    Yanghui Tan, Jianhui Tan, Yu Hong
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3215-3223. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.253
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    Objective: Galectin-3 (Gal-3), is a crucial protein involved in regulating cell adhesion, inflammation, and fibrosis, thereby playing a pivotal role in the occurrence and progression of Atherosclerosis (AS). Using the rat carotid artery AS model, this study aimed to investigate the regulatory effects of Galectin-3 expression on lipid accumulation and AS lesion.

    Methods: Thirty male Wistar rats, aged 10 weeks, were randomly assigned to three groups: the control, model, and treatment groups, each comprising 10 rats. In the control group, dissection was limited to the left common carotid artery. Moreover, the carotid atherosclerosis model was induced in both the model and treatment groups. The treatment group was administered with 10% Modified citrus pectin (MCP) concurrently one week before surgery. Blood and tissue samples were collected on the day after the operation, and the atherosclerosis modeling was observed using Hematoxylin-Eosin (H&E) staining. The levels of triglyceride, total cholesterol, low-density lipoprotein, and high-density lipoprotein were assessed in these three rat groups. Oil red O staining was used to analyze the lipid deposition in the intima of arteries. Furthermore, the expression levels of Cholesterol ester hydrolase (CEH) and Gal-3 in the carotid artery were determined by quantitative Real-time Polymerase Chain Reaction (qRT-PCR) and Western blot analysis. Enzyme-Linked immunosorbent assay (ELISA) was employed to assess the expression levels of Gal-3, tumor necrosis factor-α (TNF-α), vascular endothelial growth factor (VEGF), Interleukin 6 (IL-6), and Interleukin 8 (IL-8). Additionally, flow cytometry was utilized to evaluate mitochondrial membrane potential (MMP) and the expression of iron death-related proteins (Glutathione Peroxidase 4 and Nicotinamide Adenine Dinucleotide Phosphate (NADPH) oxidase 4 (NOX4)) was determined using Western blot analysis.

    Results: Gal-3 was highly expressed in atherosclerotic rats. After the inhibition of Gal-3, a significant reduction was observed in the levels of triglyceride, total cholesterol, and low-density lipoprotein cholesterol in tissues. The degree of lesions, the levels of inflammatory factors and iron death were significantly alleviated, while the expression of CEH was increased.

    Conclusion: Inhibition of Galectin-3 can slow down the lesion and lipid accumulation in carotid atherosclerotic rats.

  • Article
    Pingping Dong, Xiaoyan Liang, Feng Wang, Guoqing Gu, Fengyun Su, Haimin Wang
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3225-3232. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.254
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    Background: Preterm birth is a major cause of neonatal mortality and morbidity, with inflammation being a crucial contributor to its occurrence. The nuclear factor-kappa B (NF-κB) pathway serves as an important regulator of inflammatory responses and is strongly associated with preterm birth. However, lentinan has been recognized for its immunomodulatory and anti-inflammatory effects. Therefore, this study aimed to delve into the potential impact as well as the underlying mechanism of lentinan on preterm birth.

    Methods: In this study, we used a lipopolysaccharide (LPS)-induced preterm birth model in mice to evaluate the effect of lentinan on preterm birth and its influence on the NF-κB inflammatory pathway. Mice were randomly assigned to the control, LPS, and LPS+Lentinan intervention groups. The levels of inflammatory factors and expression of NF-κB pathway-related proteins in uterine tissues were assessed using enzyme-linked immunosorbent assay (ELISA), western blot analysis, and immunohistochemistry techniques.

    Results: Compared to the LPS group, the LPS+Lentinan intervention group showed a significant reduction in the rate of preterm birth (p < 0.01), as well as improvements in offspring survival and birth weight (p < 0.01). Moreover, lentinan inhibited the expression of inflammation-related proteins (such as jun proto-oncogene (C-JUN) and phosphorylated(p)-P65) in LPS-induced uterine smooth muscle (p < 0.01), while reducing the upregulation of inflammatory factors (such as tumor necrosis factor (TNF)-α, interleukin (IL)-6; p < 0.01). These results suggest that lentinan may exert anti-inflammatory effects by inhibiting the activation of the NF-κB pathway.

    Conclusion: Lentinan effectively prevents LPS-induced preterm birth in mice and improves offspring survival. Its mechanism may be related to the inhibition of the activation of NF-κB inflammatory pathway in uterine smooth muscle. This finding provides a potential natural drug candidate for the development of new interventions to prevent preterm birth, thereby reducing neonatal mortality and associated complications.

  • Article
    Yayun Chen, Yuxin Sun, Wenjie Cai, Kaixin Li, Bingzong Gao, Runzhi Mao
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3233-3243. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.255
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    Background: Overexpression of the transcription factor Dp-1 (TFDP1) has been demonstrated in breast cancer (BC), indicating a possible involvement in the progression of BC. This study explored the downstream mechanism of TFDP1 in BC.

    Methods: Quantification of TFDP1, anti-silencing function 1B histone chaperone (ASF1B) and CDC28 protein kinase regulatory subunit 1B (CKS1B) in BC cells was realized through quantitative real-time polymerase chain reaction (qRT-PCR). The relationship among TFDP1, ASF1B and CKS1B was predicted by GEPIA and verified by qRT-PCR and western blot. Through loss- and gain-of-function assays and rescue assays, the effects of TFDP1 and TFDP1/ASF1B/CKS1B axis upon viability, proliferation, migration, and invasion of BC cells were studied. Proliferating cell nuclear antigen (PCNA), matrix metalloproteinases (MMPs) and vimentin protein levels were determined by western blot.

    Results: TFDP1, ASF1B and CKS1B were highly expressed in BC and were positively correlated in pairs (p < 0.001). TFDP1 overexpression and ASF1B overexpression enhanced while TFDP1/CKS1B/ASF1B silencing suppressed the viability, migration, and invasion of BC cells, and TFDP1 silencing, CKS1B silencing or ASF1B silencing suppressed BC cell viability, migration, and invasion (p < 0.05). TFDP1 silencing suppressed CKS1B and ASF1B expressions (p < 0.05). ASF1B overexpression increased CKS1B, PCNA, MMP-2, MMP-9 and vimentin expression, while these effects were reversed by CKS1B silencing (p < 0.001).

    Conclusion: TFDP1 facilitates the malignant biological behaviors of BC cells by promoting ASF1B-mediated CKS1B upregulation.

  • Article
    Xidong Zhang, Chao Xu
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3245-3256. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.256
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    Background: Osteosarcoma (OS) is the most prevalent primary malignant bone tumor among adolescents, yet the efficacy of current medications is limited by a lack of understanding of the molecular mechanisms underpinning OS growth and metastasis. In previous studies, cantharidin (CTD), an effective component found in several clinical traditional Chinese medicine formulations, has demonstrated promising pharmacological efficacy against various malignancies. However, the precise biological functions and regulatory mechanisms of CTD in OS, particularly its interaction with the Forkhead box O3A (FOXO3A)/Methyltransferase Like 14 (METTL14)/Type III Intermediate Filament Protein (Vimentin) pathway, remain incompletely understood, warranting further investigation.

    Methods: Eighteen female BALB/c nude mice (four-week-old; 15 ± 2 g) were utilized in this study and divided into three groups (n = 6/group): Normal control, Control, and CTD groups. Mice in the Normal control group received saline injections, while those in the other groups were injected with U-2 OS cells at the proximal end of the left tibia to establish an osteosarcoma model. Following successful model establishment, mice in the Control group received a daily intraperitoneal injection of saline (1 mg/kg), while those in the CTD group received an intraperitoneal injection of CTD (2.5 mg/kg). The inhibitory effect of CTD on tumor growth was evaluated after 28 days of continuous treatment. Additionally, the effects of CTD on OS cells were assessed using a Cell Counting Kit-8 (CCK-8), colony formation, and Transwell assays. Western blotting and quantitative real-time PCR (qRT-PCR) were employed to quantify FOXO3A/METTL14 expression levels.

    Results: Lower levels of FOXO3A expression were observed in OS tissues (p < 0.01), with these expression levels significantly correlating with OS patient prognosis (p < 0.01). Compared to controls, CTD significantly suppressed osteosarcoma cell growth and metastasis as well as tumor growth (p < 0.001) by activating the FOXO3A/METTL14 pathway, consequently downregulating Vimentin expression (p < 0.001).

    Conclusion: Our findings indicate that cantharidin restricts osteosarcoma cell proliferation and metastasis through modulation of the FOXO3A/METTL14/Vimentin pathway.

  • Article
    Xinxin Li, Wei Zhou, Liangliang Zhou, Yingbin Li, Xufeng Wu, Jianjun Chen
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3257-3267. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.257
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    Background: Cold-inducible RNA-binding protein (CIRP) promotes inflammatory responses. Activator protein-1 (AP-1) potentially acts as a transcriptional regulator of CIRP. Consequently, this study aims to investigate the impact of the AP-1-CIRP axis on microglial activation to mitigate neural injury in sepsis-associated encephalopathy (SAE).

    Methods: We employed a Streptococcus pneumoniae-induced mouse SAE model in vivo and lipopolysaccharide (LPS)-induced BV-2 cells for in vitro experiments. The spatial exploration ability of mice was assessed using the Morris water maze assay. Levels of inflammatory factors were determined via enzyme-linked immunosorbent assay. Microglial activation in the cerebral cortex and hippocampus was determined through immunohistochemistry or immunofluorescence. Degenerated neurons were identified using Fluoro-Jade C staining. Quantifications of mRNA and protein levels in tissues, cells, and supernatants were conducted through real-time quantitative polymerase chain reaction and Western blot, respectively. The binding relationship between AP-1 and CIRP was analyzed using JASPAR and chromatin immunoprecipitation. Reactive oxygen species (ROS) levels were measured via 2,7-dichlorodihydrofluorescein diacetate (DCFH-DA) assay.

    Results: Silencing of AP-1 led to reduced levels of tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-1β levels, as well as a decrease in Iba1+ cells in SAE mice. Cognitive impairment and neuronal degeneration in the brains of SAE mice were partially alleviated by AP-1 silencing. Furthermore, elevated levels of FOS Like 1 (Fosl1), JunB Proto-Oncogene (Junb), Jun Proto-Oncogene (Jun), and CIRP in SAE mice were partially attenuated by AP-1 silencing. We observed that AP-1 is bound to the CIRP promoter region. Inhibition of AP-1 or CIRP reduced levels of inflammatory factors, CD11b+ cells, ROS, pro-apoptosis-related protein, and extracellular CIRP. Conversely, AP-1 overexpression exhibited the opposite effects reversed by CIRP silencing in LPS-induced cells.

    Conclusion: Inhibition of AP-1 ameliorates neuronal injury in SAE by suppressing microglial activation through transcriptional regulation of CIRP.

  • Article
    Wanglin Zhao, Xiyun Li, Heng Bi, Fanbing Li
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3269-3279. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.258
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    Objective: This work aimed to evaluate the therapeutic effects of icariin (ICA) on postmenopausal osteoporosis (PMOP) and explore its association with the fatty acid-binding protein 4 (FABP4) gene, providing novel insights into the mechanisms of osteoporosis.

    Materials and Methods: Forty-five healthy rats were randomly divided into three groups: Group S1 (icariin treatment), Group S2 (model), and Group S3 (sham surgery). Measurements included body weight, spatial structural parameters of tibia and lumbar vertebrae tissue, FABP4 gene expression, and Bone Gla protein (BGP) levels.

    Results: The body weight of rats in Group S1 was slightly lower than that of Group S2 at weeks 7 and 14, but the difference was not significant (p > 0.05). Lumbar vertebrae and femoral neck bone mineral density (BMD) were also slightly lower than those in Group S3, but the difference was not significant (p > 0.05). However, the spatial structural parameters of tibia and lumbar vertebrae tissue, including bone volume (BV), total bone volume (TV), trabecular thickness (Tb.Th), and trabecular connectivity density (Conn.D), were significantly higher in Group S1 than in Groups S2 and S3. FABP4 gene expression and trabecular separation (Tb.Sp) were significantly lower in Group S1 than in Groups S2 and S3 (p < 0.05). Additionally, the maximum bending and compressive loads of the tibia and serum BGP levels were significantly higher in Group S1 than in Groups S2 and S3 (p < 0.05).

    Conclusion: ICA effectively improved the strength of bone tissue in PMOP rats and enhanced the ultrastructural characteristics of trabecular bone. These effects may be associated with ICA's inhibition of FABP4 gene expression, increased serum BGP levels, and improved osteoblast activity. These findings provided important clues for further exploration of the mechanisms of PMOP treatment.

  • Article
    Baoping Li, Shaning Yang
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3281-3288. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.259
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    Background: This study aimed to explore the relevant factors and clinical significance of procalcitonin (PCT) levels in patients diagnosed with persistent atrial fibrillation (AF) and heart failure (HF). However, AF is the most common cardiac arrhythmia, while HF is a clinical syndrome characterized by the heart's inability to pump sufficient blood to meet the body's metabolic demands. While PCT has been widely used for the clinical diagnosis of infectious diseases, its role in non-infectious diseases, particularly cardiovascular diseases, remains unclear.

    Methods: We conducted a cross-sectional observational study involving 286 hospitalized patients diagnosed with persistent AF and HF. Peripheral venous blood samples were collected, and PCT levels were assessed using the chemiluminescence immunoassay method. Moreover, clinical data, including HF severity, ventricular rate, age, and other variables of the study cohort were recorded. Furthermore, nonparametric tests were used to compare PCT levels among different groups, and multiple logistic regression analysis was employed to explore potential factors influencing PCT levels.

    Results: We observed that PCT levels were significantly associated with HF severity, ventricular rate, and age, even after adjusting for other clinical variables. The odds of having a positive PCT level (defined as >0.05 ng/mL) increased by 2.74 times for each increase in HF grade, by 3.21 times for a ventricular rate of ≥110 bpm compared to <110 bpm, and by 1.05 times for each year of age. These findings suggest that PCT could serve as a potential biomarker for the diagnosis and assessment of HF in patients with AF, as well as an indicator of the cardiovascular stress or damage associated with AF.

    Conclusion: This study provides new insights into the potential of PCT in AF and HF, highlighting the need for additional investigations on the causal relationship and underlying biological pathways between PCT and these factors, as well as the diagnostic and prognostic significance of PCT in this population.

  • Article
    Na Wang, Shu Wang, Linqi Lu, Lei Yu, Rui He, Huan Liu, Qiang Zhang, Bing Liu, Chao Zhang
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3289-3297. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.260
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    Background: Acute myocardial infarction (AMI) is a cardiovascular disease induced by acute or persistent hypoxia in cardiomyocytes, resulting in irreversible damage to the heart. Therefore, we aimed to elucidate the role and mechanism of the Brain and muscle Arnt-like protein-1 (Bmal1)/AKT/P53 signaling pathway in mediating cardiomyocyte ferroptosis, oxidative stress, and inflammatory response in AMI.

    Methods: This study utilized H9C2 cardiomyocytes for hypoxia culture to establish an in vitro AMI model. Quantitative polymerase chain reaction (qPCR), Western blot analysis, and Enzyme-Linked Immunosorbent Assay (ELISA) assays were used to assess the expression levels of Bmal1/AKT/P53 signaling pathway-related molecules, Bmal1, p-AKT, and P53, along with other expression levels of associated factors within cardiomyocytes.

    Results: We observed that cardiomyocyte hypoxia promoted cardiomyocyte reactive oxygen species (ROS) production through the Bmal1/AKT/P53 signaling pathway. Furthermore, the expression levels of acyl-CoA synthetase long-chain family member 4 (ACSL4) were significantly increased (p < 0.01), whereas glutathione peroxidase 4 (GPX4) and solute carrier family 7a member 11 (SLC7A11) were significantly decreased. Additionally, the inflammatory response-related factors, including interleukin-1α (IL-1α), IL-1β, IL-2, IL-6, Transforming growth factor β (TGF-β), and TGF-α were significantly increased (p < 0.01).

    Conclusion: This study explored that overexpression of Bmal1 activates AKT phosphorylation and inhibits oxidative stress, ferroptosis, and inflammation caused by cardiomyocyte hypoxia, thereby alleviating acute myocardial infarction.

  • Article
    Wenfeng Liu, Shu Li, Xia Xing, Weiling Wang, Min Zhang, Miao Miao
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3299-3307. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.261
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    Objective: This study aimed to explore the correlation of proliferative cell nuclear antigen-67 (Ki-67) with clinicopathological characteristics and prognosis in endometrial cancer (EC) patients.

    Methods: The study included 216 EC patients admitted from January 2010 to December 2014. The corresponding clinicopathological data of the patients were collected and comprehensively analyzed. Their tissue samples were collected during surgical procedures and the patients were followed up for 3 to 60 months. Receiver operating characteristic curve (ROC) was drawn to determine the critical value of Ki-67 for predicting the prognosis and occurrence of postoperative events. Spearman correlation analysis and Kaplan-Meier survival curve were used for the evaluation of patient prognosis.

    Results: The expression of Ki-67 >67% did not show a significant correlation with patient age, gravidity, menopause, family history, and tumor size, but exhibited a significant correlation with pathological classification, pathological type, depth of myometrial invasion, lymphatic vascular space invasion, lymph node metastasis, International Federation of Gynecology and Obstetrics (FIGO) stage, and histological grade. Furthermore, the expression of Ki-67 >67% in cancer tissue was negatively correlated with the expression of estrogen receptor (ER) and progesterone receptor (PR) (r = –0.818 and –0.842, p < 0.001). Conversely, it indicated a positive correlation with the expression of human epidermal growth factor receptor-2 (HER-2) and tumor suppressor gene (P53) (r = 0.613 and 0.587, p < 0.001). Moreover, the Kaplan-Meier survival curve analysis revealed a substantial association between the expression level of Ki-67 and the 5-year survival rate of EC patients. Concurrently, the 5-year event-free survival rate was significantly lower in patients with Ki-67 >67%+ER-negative and (or)+PR-negative compared to those with Ki-67 >67%+ER-positive and (or)+PR-positive (p < 0.001). Additionally, it was also lower in patients with Ki-67 >67%+HER-2-positive and (or)+P53-positive compared to those with Ki-67 >67%+HER-2-negative and (or)+P53-negative (p < 0.001).

    Conclusion: The elevated expression of Ki-67 exhibits a strong correlation with the poor clinicopathological characteristics of EC patients, suggesting a poor prognosis in EC patients with Ki-67 >67%.

  • Article
    Xiaofang Chen, Shufang Qi, Na Yu, Qing Liu
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3309-3320. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.262
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    Background: Recurrent pregnancy loss (RPL) is characterized by multiple miscarriages before reaching 20 weeks of pregnancy, affecting women in their reproductive age. This study aimed to investigate the effect of interleukin-2 (IL-2)/signal transducer and activator of transcription 5 (STAT5) axis on the RPL and its association with regulatory T cells (Treg) and T helper cells (Th) 17.

    Methods: The female CBA/J mice were used to construct the RPL model by mating with male DBA/2 mice. The IL-2 administration was used to increase the IL-2 content and activate IL-2/STAT5 axis, and the IL-2 plus STAT5 inhibitor was used to increase the IL-2 content while at the same time inactivate the IL-2/STAT5 axis. Moreover, the resorption rate in mice was evaluated. In the mice uteri tissues and intracardiac blood sera, the levels of IL-2 and phosphorylated (p)-STAT5/STAT5, as well as levels of cytokines and factors related to Treg and Th17 cells were assessed using quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot analysis. The inflammation associated cytokines in intracardiac blood sera were detected. Additionally, the peripheral blood mononuclear cells (PBMCs) and the single-cell suspension of the spleen tissues were prepared and analyzed employing flow cytometry to determine the percentage of both Th17 cells and Treg cells.

    Results: IL-2 treatment increased the activation of the IL-2/STAT5 axis and decreased resorption in mice (p < 0.01), which was reversed following STAT5 inhibitor (p < 0.01). Moreover, the IL-2-induced IL-2/STAT5 axis activation promoted the expression of forkhead box P3 (Foxp3), IL-10, and transforming growth factor-beta (TGF-β) while decreasing the expression of retinoid-related orphan receptor gamma t (RORγt), IL-17 and IL-22 (p < 0.001), which was reversed by STAT5 inhibitor (p < 0.001). Furthermore, the activation of the IL-2/STAT5 axis decreased the levels of IL-1β, tumour necrosis factor-alpha (TNF-α), IL-6, Interferon-gamma (IFN-γ), granulocyte-macrophage colony-stimulating factor (GM-CSF), and Th17 percentage while increasing the levels of IL-4 and Treg percentage (p < 0.001), which was reversed with the disruption of the IL-2/STAT5 axis activation (p < 0.01).

    Conclusion: The IL-2/STAT5 axis can activate Treg cells while suppressing the Th17 cells and their associated inflammatory response, leading to effects on protecting pregnant mice from miscarriage. Thus, the IL-2/STAT5 axis could be a potential therapeutic target for treating RPL.

  • Article
    Jinxin Liu, Dapeng Ding, Feiye Liu, Yizhi Chen
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3321-3330. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.263
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    Background: Expression confusion of tripartite motif (TRIM) family proteins has been associated with various cellular processes such as immune disorders, neurological-related diseases, protein degradation, and tumorigenesis. Therefore, we aim to explore the function and clinical significance of TRIM family proteins in chemotherapy resistance in ovarian cancer.

    Methods: Western blot analysis and immunohistochemistry (IHC) assay were conducted to assess the protein expression levels of TRIM37 in both ovarian cancer cell lines and patient tissues. Furthermore, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays were conducted to detect the effect of TRIM37 on 50% inhibitory concentration (IC50). The rate of apoptotic cells was conducted using fluorescence-activated cell sorting (FACS) assays. Moreover, Luciferase assays were performed to detect the luciferase reporter activity. Additionally, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assays were conducted to detect the pro-cancer effect of TRIM37.

    Results: We found that TRIM37 was increased in ovarian cancer tissues, showing partial response to cisplatin chemotherapy (p < 0.01). The high TRIM37 expression was linked to a worse overall survival prognosis and lower disease-free survival rate in our cancer tissue samples and public ovarian cancer databases (p < 0.001, p < 0.01). Functionally, overexpression of TRIM37 endows cisplatin resistance in ovarian cancer cells. However, inhibition of TRIM37 counteracted this resistance in ovarian cancer. Additionally, TRIM37 was found to activate the wnt/β-catenin signaling pathway.

    Conclusions: Our findings indicate that TRIM37 contributes to enhanced cisplatin resistance in ovarian cancer, suggesting its potential as a promising chemotherapy biomarker.

  • Article
    Sohi Kang, Sungmoo Hong, Joong-Sun Kim, Hongbing Wang, Changjong Moon, Taekyun Shin
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3331-3344. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.264
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    Background: Chlorogenic acid (CGA), a phenolic compound, is renowned for its capacity as an antioxidant, anti-inflammatory agent, and scavenger of free radicals. Multiple sclerosis (MS) is a chronic autoimmune condition impacting the central nervous system, characterized by demyelination and neuroinflammation. This research delved into the potential therapeutic effects of CGA in experimental autoimmune encephalomyelitis (EAE), an established animal model mirroring MS pathology.

    Methods: We induced EAE in C57BL/6 mice via immunization with myelin oligodendrocyte glycoprotein 35–55 peptide and conducted daily monitoring to record clinical signs, including paraparesis and body weight changes. We performed histological and molecular analyses on spinal cord tissues to evaluate neurodegeneration and neuroinflammation.

    Results: Intraperitoneal CGA administration significantly reduced the incidence and severity of EAE paraparesis. Histological analysis showed a significant reduction in inflammatory cell infiltration within the spinal cord parenchyma following EAE induction. CGA treatment substantially downregulated mRNA levels of interleukin-1β, interleukin-6, tumor necrosis factor-α, and interferon-gamma in the spinal cord tissues. Additionally, CGA treatment resulted in a significant decrease in astrogliosis and microglial/macrophage activation in the spinal cords of EAE mice.

    Conclusions: Our findings demonstrate that CGA has a therapeutic effect on EAE by modulating pro-inflammatory cytokines. These results position CGA as a promising pharmacotherapeutic candidate for MS treatment.

  • Article
    Shuxiang Duan, Haiyan Meng, Ping Sun, Ning Zhang, Hongmei Niu, Yonghui Ma, Haoyuan Li, Hongqiang Zhang
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3345-3353. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.265
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    Background: Atherosclerosis is one of the primary causes of cardiovascular diseases. Previous studies have demonstrated the significant role of the Found in Inflammatory Zone 1 (FIZZ1) in the development of atherosclerosis. Therefore, this study aims to explore the potential mechanisms of FIZZ1 in atherosclerosis.

    Methods: The mouse aortic endothelial cells were utilized in the present study. The quantitative reverse transcription polymerase chain reaction (RT-qPCR) and RNA sequencing approaches were employed to assess the impact of FIZZ1 on vascular generation-related genes as well as the expression of differentially expressed genes (DEGs) in endothelial cells. Furthermore, gene functional annotation and pathway enrichment analyses were conducted using the Gene Ontology (GO) and the Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis for FIZZ1-induced differentially expressed genes (DEGs). Additionally, Cell Counting Kit-8 and 5-ethynyl-20-deoxyuridine (EdU) assays were utilized to investigate the effect of FIZZ1 on cell proliferation. Transwell assay was used to evaluate cell migration, and angiogenesis assay was used to assess cell angiogenesis.

    Results: FIZZ1 treatment significantly downregulated the expression levels of angiogenesis-inhibiting genes such as Collagen Type I Alpha 1 Chain (COL1A1) and Endothelin Receptor Type A (EDNRA) while upregulated the expressions levels of angiogenesis-promoting genes including Fms Related Receptor Tyrosine Kinase 4 (FLT4) and Integrin Subunit Alpha 3 (ITGA3) (p < 0.05, and p < 0.01). Furthermore, FIZZ1 treatment substantially increased the proliferation, migration, and angiogenesis of aortic endothelial cells (p < 0.05, p < 0.01, and p < 0.001). Additionally, FIZZ1-induced DEGs were associated with crucial cellular functions which positively participate in cell proliferation, migration, and angiogenesis.

    Conclusion: In summary, FIZZ1 treatment can promote the proliferation, migration, and angiogenesis of aortic endothelial cells. Furthermore, it can induce numerous differentially expressed genes involved in angiogenesis-related functions or pathways. Consequently, FIZZ1 promotes the angiogenesis of aortic endothelial cells, thereby contributing to the progression of atherosclerosis. Therefore, FIZZ1 can serve as a potential therapeutic target for ameliorating atherosclerosis.

  • Article
    Qin Chen, Songrui Wei, Mei Yang, Jungang Zhao, Fei Hong, Feng Liu, Guoji Zhu
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3355-3363. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.266
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    Background & Aims: Resveratrol (RES), as a natural compound, has demonstrated potential therapeutic effects in sepsis and anti-immune suppression properties in various diseases. However, during the development of sepsis, immune suppression is primarily caused by T-cell dysfunction. Therefore, this study aims to explore the specific mechanisms by which RES regulates the immunosuppression of T lymphocytes in septic rats.

    Method: Initially, we simulated the pathological state of sepsis in rats using the cecal ligation and puncture (CLP) model and treated them with RES. The therapeutic effect of RES was assessed by observing the survival rate of CLP rats. Subsequently, we determined the protein expression levels of programmed death-ligand 1 (PD-L1) and Programmed death-1 (PD-1) in blood, as well as the number of PD-L1-positive neutrophils. We also assessed the histopathological morphology and apoptosis in the spleen and thymus, along with the levels of inflammatory factors. Finally, neutrophils were isolated from both the Sham and CLP groups, co-cultured with T lymphocytes, and the apoptosis rate of T lymphocytes was examined.

    Results: We observed that RES boosted the survival rate of CLP rats. Additionally, RES inhibited the expression of PD-L1 on neutrophils and exhibited a mitigating effect on immunosuppression of T lymphocytes. This was demonstrated by the improvement in the histopathological structure of the spleen and thymus in CLP rats, along with the inhibition of apoptosis in T lymphocyte by RES.

    Conclusion: RES alleviates immunosuppression of T lymphocytes in septic rats by inhibiting PD-L1 expression on neutrophils. Consequently, RES enhances the survival rate of CLP rats.

  • Article
    Juan Ma, Wenjing Yang
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3365-3374. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.267
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    Background: Diabetic nephropathy (DN) is the leading cause of end-stage renal disease throughout the world. Currently, hemodialysis (HD) and peritoneal dialysis (PD) are widely applied in treating patients with DN.

    Objective: The present research is dedicated to investigating the impact of hemodialysis on heart function and structure, oxidative stress, and inflammatory factor levels of DN patients.

    Methods: The data of 100 cases of DN patients (between 2021 and 2022 in our hospital) with normal left ventricular systolic function were collected. These patients were randomly divided into two groups: the observation group (48 cases of DN patients that underwent HD) and the control group (52 cases of those subjected to PD). Echocardiography was employed to evaluate the changes in cardiac function and structure of DN patients in both groups. The enzyme-linked immunosorbent assay (ELISA) and oxidative stress commercial assay kits were used to measure the levels of inflammatory factors and oxidative stress markers in the serum of both groups of patients. Additionally, the nutritional status and occurrence of complications during HD were observed and recorded in both groups. The Hamilton Anxiety Scale (HAMA) and Hamilton Depression Rating Scale (HAMD) were used to evaluate the social-psychological status of patients. The simplified comfort state scale and the 36-item short-form health survey (SF-36) were used to assess the comfort and quality of life of patients, respectively, before and after hemodialysis care.

    Results: Compared to the control group, the observation group of patients with DN showed more significant cardiac structural and functional impairments, as well as elevated levels of inflammatory markers and oxidative stress. Meanwhile, compared to the control group, the observation group of patients exhibited significantly higher levels of anxiety and depression. Moreover, the patients reported significantly decreased comfort and quality of life (p < 0.05).

    Conclusions: Compared to peritoneal dialysis, hemodialysis reduces the quality of life for DN patients and may not be the optimal choice for DN patients.

  • Article
    Sipho Mapfumari, Kokoette Bassey
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3375-3384. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.268
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    Background: Cancer describes a vast category of diseases our body suffers from. The total global oncology spending was USD186bn as of 2021 and was estimated to rise to USD208.9bn in 2022. The adverse effects of managing cancer with orthodox medications includes death among other dire consequences. Several Viscum species including Viscum album L, Viscum angulatum, and Viscum articulatum Burm f. from other countries have been reported for their anticancer potentials.

    Methods: Four (n = 4) mistletoe extracts namely n-hexane, dichloromethane, acetone, and methanol were tested for their anti-cancer potemtials against Vero (African green monkey, kidney, non-cancerous), BJ-5ta (Human skin fibroblast), A549 (Human non-small cell lung carcinoma), Michigan Cancer Foundation (MCF7) human breast cancer cell lines. The percentage cancer cells survival rate and selectivity index for each extract were calculated against positive controls (untreated cells) and blank dimethyl sulfoside (DMSO) solutions, and the compounds with anticancer potentials were identified to be present in the extracts using gas chromatography-mass spectrometry analysis.

    Results: All four extracts showed concentration dependent anti-cancer activity as measured from the percentage average cell viability or cytotoxicity. The cytotoxicity to the lung (A549) was recorded as 1.25, 10, 0.5, 1.25 mg/mL for the hexane, dichloromethane, acetone and methanol extracts respectively. Those for the breast (MCF7) cancer cell lines appeared as as 1.25, 0.5, 0.5 and 10 mg/mL for the hexane, dichloromethane, acetone and methanol extracts in that order. In terms of the selectivity index (SI), that is, which extracts is cytotoxic to a specific cancer cell line, all the extracts were highly selective (SI >2). However, the methanol extract was selectively more toxic to the lung cancer cell (A459) with SI of 6.08 and Half maximal Inhibitory Concentration (IC50) value of 0.251 ± 3.96. The other three extracts were highly selective against breast cancer cell (MCF7) with SI of 5.90 (IC50 = 0.38), 9.06 (IC50 = 0.04) and 8.15 (IC50 = 0.02) for n-hexane, dichloromethane (DCM), and acetone extracts, respectively.

    Conclusions: Eucalyptol, 9(E),11(E)-conjugated linoleic acid, ester and 9, 12, 15-octadecatrienoic acid detected in South African mistletoe by gas chromatoghraphy-mass spectrometry (GC-MS) analysis is proposed to be responsible for the anticancer potentials of the extracts.

  • Article
    Xin Luo, Jian Zhao
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3385-3394. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.269
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    Background: Gestational diabetes mellitus (GDM) elevates the risk of complications in pregnant women and fetuses. The follistatin (FST) expression is decreased during GDM. However, its precise impact and the underlying mechanisms in human umbilical vein endothelial cell (HUVEC) are not fully understood. Therefore, this study aimed to delve into the impact of FST on HUVEC and to elucidate the underlying molecular mechanisms.

    Methods: FST level in GDM was bioinformatically analyzed using two datasets, GSE49524 and GSE87295. Human extravillous trophoblast cells (HTR-8/SVneo) were cultured in a high glucose (HG) medium and subsequently transfected with lentiviral vectors overexpressing FST or its negative control. Moreover, HUVEC was grown in HTR-8/SVneo cell culture media replete with Compound C (CC, an inhibitor of adenosine monophosphate (AMP)-activated protein kinase (AMPK)), and FST level was observed using enzyme linked immunosorbent assay (ELISA). Furthermore, the expression levels of FST, glucose-6-phosphatase (G6Pase), proliferating cell nuclear antigen (PCNA), marker of proliferation Ki-67 (Ki67), phosphorylated AMPK (pAMPK), AMPK, histone deacetylase 4 (HDAC4), and phosphorylated histone deacetylase 4 (pHDAC4) were assessed in the treated HUVECs utilizing quantitative real time polymerase chain reaction (qRT-PCR) and western blot analysis. Additionally, the viability and proliferation of HUVECs were determined through 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) and 5-ethynyl-2′-deoxyuridine (EdU) staining assays.

    Results: HG treatment inhibited FST level in HTR-8/SVneo cells as well as in their cell culture medium (p < 0.01). Furthermore, overexpression of FST promoted the viability and proliferation of HUVECs, increased the levels of Ki67, PCNA, and pAMPK, and decreased G6Pase and pHDAC4 levels in HG-induced HUVECs (p < 0.05). Additionally, CC treatment further counteracted the influences of overexpressed FST on pAMPK/AMPK and pHDAC4/HDAC4 ratios, G6Pase, Ki67, and PCNA levels, and the viability and proliferation in HUVECs (p < 0.05).

    Conclusion: In summary, HTR-8/SVneo-derived FST promoted the viability and proliferation of HUVEC by activating the AMPK signaling pathway.

  • Article
    Yayun Sun, Quanbin Zha, Yongbin Ma, Dan Zhou, Junhua Zhang
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3395-3405. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.270
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    Background: α-Humulene (Hum) can alleviate cerebral ischemia/reperfusion (I/R) injury. However, the mechanism of Hum in microglial activation induced by cerebral I/R injury remains unclear. This study aims to investigate the role of Hum in microglial activation resulting from cerebral I/R injury.

    Methods: Several cerebral I/R injury models were established in adult Sprague-Dawley (SD) rats treated with 25 and 50 mg/kg Hum, and in primary microglia extracted from newborn SD rats treated with 0.25, 0.5, and 1.0 mM Hum. Neurological deficits were assessed using a modified neurological severity score. Cerebral infarction was assessed by triphenyltetrazolium chloride staining. Immunofluorescence assay was applied to measure the levels of potassium voltage-gated channel subfamily A member 3 (KCNA3) and ionized calcium-binding adapter molecule 1 (Iba-1). Gene expression levels of KCNA3, NOD-like receptor thermal protein domain-associated protein 3 (NLRP3), and brain-derived neurotrophic factor (BDNF) were measured by western blot and quantitative real-time polymerase chain reaction (qRT-PCR). The levels of tumor necrosis factor (TNF)-α and interleukin (IL)-6 were measured using enzyme-linked immune sorbent assay (ELISA). Additionally, a rescue assay was used to investigate whether the inhibitory effect of Hum on microglial activation was mediated through KCNA3 regulation.

    Results: Hum significantly reduced neurological deficits and cerebral infarction in rats with cerebral I/R injury (p < 0.001). Both in vivo and in vitro, Hum reduced the levels of Iba-1, KCNA3, and NLRP3, yet promoted the level of BDNF in cerebral I/R injury models (p < 0.01). Moreover, Hum suppressed the levels of TNF-α and IL-6 in oxygen-glucose deprivation/reperfusion (OGD/R)-exposed microglia (p < 0.001). Notably, KCNA3 overexpression reversed the effects of Hum on Iba-1, TNF-α, IL-6, NLRP3, and BDNF in OGD/R-exposed microglia (p < 0.001).

    Conclusion: Hum improves cerebral I/R injury by blocking KCNA3 to inhibit microglial activation.

  • Article
    Yan Zhang, Rujia Fan, Danting Wu, Kangning Zhu, Yue Wang
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3407-3417. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.271
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    Background: The incidence and mortality of cervical cancer are increasing, which seriously threatens the life and health of women. Lymph node metastasis is a critical factor involved in the survival rate of patients. Research has shown that the immune system plays a key role in suppressing tumor development and metastasis. Thus, to study the relationship between immunity and tumor metastasis, we explored the mechanism by which Janus kinase-signal transducer and activator of transcription (JAK-STAT) signaling pathway promotes lymph node metastasis in cervical cancer based on CD8+T cell senescence.

    Methods: The level of JAK-STAT protein was detected by collecting cervical cancer tissues and adjacent normal tissues. JAK-STAT protein levels were compared in patients with different Federation International of Gynecology and Obstetrics (FIGO) stages and lymph node metastasis. To analyze the diagnostic value of JAK-STAT level in cervical cancer lymph node metastasis, cervical cancer cell lines were infected with JAK-STAT pathway inhibitors to detect cell viability, migration, and invasion. Concurrently, CD8+ T cells were co-cultured with cervical cancer cell lines to assess CD8+ T cell count, surface molecule expression (CD28, CD27, and CD57), and inflammatory factor levels. The lymphoid metastasis model of cervical cancer in nude mice was constructed to observe changes in the general state and lymph node metastasis of nude mice. Cervical cancer infiltration in lymph nodes was certified by pathological method, and the expression of JAK-STAT in lymph tissues was detected. The expressions of CD8+ T cells and surface molecules (CD28, CD27, and CD57), as well as inflammatory factors in lymph nodes were examined.

    Result: JAK-STAT signaling pathway was activated in cervical cancer tissues, correlated with different FIGO stages and lymph node metastasis, and exhibited a certain degree of clinical diagnostic value (p < 0.05). After JAK-STAT inhibition, cervical cancer cell viability decreased, and migration and invasion ability weakened (p < 0.05). The co-cultivation of CD8+ T cells with cervical cancer cell lines revealed a time-dependent fluctuation in the cell vitality, initially increasing and subsequently decreasing (p < 0.05). Moreover, the expressions of CD28 and CD27 decreased (p < 0.05), and the expression of CD57 increased (p < 0.05). While the cell vitality in the JAK-STAT inhibition + co-culture group initially increased and then slowly decreased (p < 0.05), with an increase in the expressions of CD28 and CD27 and a decrease in the expression of CD57 (p < 0.05). The lymphatic metastasis model of cervical cancer in nude mice revealed that after JAK-STAT inhibition, the lymph nodes were fewer and smaller, and the cervical invasion in the lymph nodes was reduced (p < 0.05). Podoplanin and p-cytokeratin (p-CK) confirmed cervical cancer metastasis in lymph nodes. The number and function of CD8+ T cells in vitro showed that the expressions of CD28, CD27, and CD57 tended to be normal in the blank and the solvent groups. Nevertheless, inflammatory factors Interleukin (IL)-6, IL-1β, and Tumor Necrosis Factor (TNF)-α exhibited an increase, and inhibiting JAK-STAT led to elevated expressions of CD28 and CD27 (p < 0.05). Conversely, CD57 expressions and levels of inflammatory cytokines IL-6, IL-1β, and TNF-α demonstrated a significant reduction (p < 0.05).

    Conclusions: The JAK-STAT signaling pathway is activated in cervical cancer, contributing to the growth and metastasis of the disease, and influencing the senescence of CD8+ T cells. Inhibition of the JAK-STAT pathway can delay the senescence of CD8+ T cells, weaken the biological behavior of cervical cancer cells, and inhibit tumor metastasis.

  • Article
    Yiwei Zhang, Jun Qu, Zhen Wang, Weifeng Liu, Wenhao Xu
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3419-3433. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.272
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    Background: Myocardial ischemia-reperfusion (I/R) injury (MIRI) often occurs as a complication following myocardial infarction, resulting in inflammation and apoptosis of myocardial cells, thereby worsening cardiac dysfunction. Leukotriene B4 (LTB4) serves as an inflammatory mediator known for its regulation of various cellular functions through its receptor, leukotriene B4 receptor 2 (BLT2). This study delved into the precise role of LTB4 in MIRI and the underlying mechanisms.

    Methods: This study utilized LTB4-deficient mouse models and an in vitro model of cardiomyocytes overexpressing LTB4 subjected to H/R (hypoxia/reoxygenation). LTB4 expression levels were analyzed using enzyme linked immunosorbent assay (ELISA), while activation of the BLT2/janus kinase 1 (JAK1)/signal transducer and activator of transcription 1 (STAT1) pathway was assessed via western blotting. Cardiac function in mice was evaluated through echocardiographic and hemodynamic assessments. Myocardial injury was determined by hematoxylin-eosin (H&E) staining, as well as measurement of lactate dehydrogenase (LDH), creatine phosphokinase (CPK), cardiac troponin-I (CTNI), cardiac troponin-T (CTNT), and creatine kinase MB (CK-MB) levels. Inflammation was assessed by real-time reverse transcriptase-polymerase chain reaction (RT-qPCR) to determine the expression of inflammation-related genes. Terminal deoxynucleotidyl transferase (TDT) dUTP Nick-End Labeling (TUNEL) staining and analysis of apoptosis-related protein levels were employed to assess apoptosis. Additionally, selective BLT2 agonists and antagonists were utilized in targeting experiments to demonstrate that BLT2 acts upstream of JAK1/STAT1.

    Results: Our findings revealed a significant elevation in LTB4 expression during MIRI (p < 0.01). LTB4-deficient mice demonstrated enhanced cardiac function and markedly reduced activation of the BLT2/JAK1/STAT1 pathway, myocardial injury, inflammation, and cell apoptosis following I/R injury compared to wild-type mice (p < 0.05). Moreover, in vitro experiments indicated that LTB4 overexpression heightened activation of the BLT2/JAK1/STAT1 pathway, exacerbating inflammation and apoptosis in H/R-stimulated cardiomyocytes by activating the BLT2/JAK1/STAT1 pathway (p < 0.05).

    Conclusion: This study reaffirms the crucial involvement of LTB4 through the BLT2/JAK1/STAT1 signaling pathway in MIRI, potentially by promoting inflammation and apoptosis of myocardial cells. These results offer valuable insights for the exploration of novel therapeutic approaches. Future research should explore interventions aimed at modulating the LTB4/BLT2/JAK1/STAT1 signaling pathway to mitigate MIRI and enhance cardiac function.

  • Article
    Liang Xiang, Bei Wang, Hailin Deng, Shuping Zhou
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3435-3447. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.273
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    Background: Osteoarthritis (OA) is a chronic progressive joint disease with a complex pathogenesis in which inflammation plays a vital role in its development. This study aims to investigate the function of the Kappa Opioid Receptor (KOR) in the Hedgehog (Hh) signaling pathway, as well as its effects and mechanisms on the development of inflammation in osteoarthritis through the Smoothened (Smo)/Glioma-associated oncogene homolog 1 (Gli1) signaling pathway.

    Methods: The expression of KOR, Smo, Gli1, and Hh was detected by Western Blotting (WB) and Polymerase Chain Reaction (PCR). Interleukin-1 beta (IL-1β), tumor necrosis factor-alpha (TNF-α), and interleukin-6 (IL-6) were monitored by enzyme-linked immunosorbent assay (ELISA). The expression of apoptotic proteins B-cell lymphoma 2 (Bcl-2), Bcl-2-associated X protein (Bax), and cysteine aspartic acid-specific protease-3 (Caspase-3) was detected by WB, and cell growth was assessed using the Cell Counting Kit-8 (CCK-8) assay. The chondrocyte injury was observed by toluidine blue staining, and apoptosis was detected by flow cytometry. A rat OA model was established using the meniscectomy method, and Hematoxylin and Eosin (HE) staining and Modified Safranin-fast green staining were used to observe the histopathological structure of cartilage in OA rats. The histopathological assessment of the severity of cartilage osteoarthritis lesions was made by Mankin scoring. Terminal deoxynucleotidyl transferase dUTP Nick End Labeling (TUNEL) staining was used to identify apoptosis in cartilage tissues.

    Results: In cellular experiments, the osteoarthritis group showed elevated levels of Smo, Gli1, and Hh, reduced expression of KOR, decreased proliferation of cartilage cells, increased apoptosis, and increased inflammatory response (p < 0.05). The addition of JT09 or cyclopamine amplified cell viability and diminished the expression of pro-apoptotic proteins, which was more effective when dual action was applied to osteoarthritis. In animal experiments, concerning the osteoarthritis group, the addition of JT09 or cyclopamine increased the expression of KOR, and reduced the expression of Smo, Gli1, and Hh. It also increased Bcl-2, declined Bax and Caspase-3, and diminished the inflammatory response, and the effect of both simultaneous actions was significant (p < 0.05). These indicated that JT09 synergistic treatment with cyclopamine declined the inflammatory response in rat osteoarthritis.

    Conclusions: KOR mediates Hedgehog (Hh) to promote osteoarthritis inflammation progression and exacerbates bone damage through the Smo/Gli1 pathway.

  • Article
    Hafiz Muhammad Rehman, Wardah Shoaib, Muhammad Naveed Khan, Numan Yousaf, Fareeha Bashir, Hamid Bashir, Qurban Ali, Shiming Han
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3449-3461. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.274
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    Background: Breast cancer remains a pervasive global health concern among women, with conventional therapies such as chemotherapeutic agents, radiography, and surgery lacking selectivity and causing undesirable side effects. In response to this challenge, fusion proteins are emerging as a promising avenue for targeted cancer therapy. This study aimed at in silico design of a chimeric protein by fusing the cell-penetrating peptide, nemo binding domain (NBD) peptide, with the cell-targeting peptide, interleukin 24 (IL-24), using linkers of varying lengths. The selected bifunctional peptide was investigated for its potential of enhanced anti-tumor activity through in silico methods.

    Methods: The 3D structure of the fusion peptide was initially modeled using I-TASSER (Iterative Threading Assembly Refinement) and AlphaFold2. The predicted structures underwent rigorous quality check and validation through Ramachandran plot analysis, ERRAT, and VERIFY 3D. The best model was refined, and physiochemical properties were evaluated. Subsequently, docking analysis was performed using ClusPro 2.0, and molecular dynamic simulations were performed using NAMD with VMD.

    Results: The results demonstrated a valid 3D structure with high model quality. The docked complex exhibited stability with 23 salt bridges, 17 hydrogen bonds, and 236 non-bonded contacts, suggesting a successful interaction of the fusion peptide with its potential receptor, thereby generating an apoptotic signal. Molecular dynamic simulations further confirmed the stability, flexibility, and functionality of the fusion protein, indicated by non-significant fluctuations in the generated graphs.

    Conclusion: In conclusion, our newly designed peptide holds promise as a potential drug candidate against breast cancer, warranting further evaluation through in vitro studies. Computational tools for designing such fusion proteins have proven valuable as an initial step preceding in vitro production.

  • Article
    Tiantian Han, Linzhen Jia, Haisu Wang, Dandan Cui, Hongye Zhao, Yu Xiao, Xue Li
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3463-3473. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.275
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    Background: Hypertension serves as a significant risk factor for various cardiovascular and renal diseases, necessitating a deeper understanding of its pathogenesis, as well as the identification of potential therapeutic targets. This study aims to investigate the molecular mechanisms underlying hypertension, specifically focusing on the regulation of renal outer medullary potassium (ROMK) channels by the mitogen-activated protein kinase (MAPK) signaling pathway and its impact on water-electrolyte metabolism.

    Methods: A mouse hypertension model was constructed, and mouse primary renal tubular epithelial cells were stimulated with Angiotensin II (AngII). Differential gene expression, key signaling pathways, and molecules were analyzed utilizing bioinformatics approaches. Furthermore, Western blot and quantitative polymerase chain reaction (qPCR) analyses were used to assess the expression of the ROMK channel both at mRNA and protein levels as well as the signaling molecule MAPK. Additionally, the MAPK inhibitors were applied to further elucidate the involvement of this signaling pathway in hypertension.

    Results: The contents of sodium, potassium, chloride in mouse blood pressure (BP) and urine were detected. A gradual increase was observed in both mRNA and protein levels of ROMK following AngII stimulation, reaching a peak on day 7. Furthermore, in vitro experiments revealed that following AngII stimulation, ROMK protein and mRNA expression levels were significantly increased compared to the control cells. Similarly, the expression levels of Phospholipase C (PLC), Protein kinase C (PKC), and MAPK proteins were significantly higher. Furthermore, Western blot analysis revealed a significant reduction in the levels of ROMK in renal tubular epithelial cells. Meanwhile, the BP decreased, and the amount of sodium and potassium ions decreased.

    Conclusion: The study demonstrates the potential significance of AngII regulating the upregulation of ROMK channels through MAPK-dependent pathway to promote potassium excretion and affect water-salt balance, leading to the development of hypertension.

  • Article
    Ran Li, Die Zhou, Guanghui Li, Minglei Sun
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3475-3487. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.276
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    Background: Oral squamous cell carcinoma (OSCC) stands as a prevalent malignant tumor worldwide, with its progression and metastasis representing primary contributors to therapeutic resistance and poor prognosis. Tumor necrosis factor receptor-associated factor 2 (TRAF2) plays a pivotal role in various tumors. However, its precise function and underlying mechanisms in OSCC remain unclear. TRAF2 interacts with Signal Transducer and Activator of Transcription 3 (STAT3) in regulating cellular processes, including inflammation and immune responses. This study aimed to investigate the role of TRAF2 in OSCC proliferation and metastasis by mediating the STAT3 signaling pathway and its underlying mechanisms.

    Methods: The impact of TRAF2 overexpression, silencing, and STAT3 inhibition on OSCC cell proliferation, invasion, and morphological alterations was evaluated using Cell Counting Kit-8 (CCK-8) assays, Transwell assays, and cytoskeleton staining, respectively. Real-time quantitative Polymerase Chain Reaction (PCR) and Western blot analysis were employed to investigate the impact of TRAF2 on the expression of key proteins in the STAT3 signaling pathway.

    Results: TRAF2 overexpression upregulated phosphorylated Janus Kinase 2 (JAK2) and STAT3 levels (p < 0.05). Additionally, TRAF2 and STAT3 were co-localized within the cytoplasm. TRAF2 knockout suppressed STAT3 transcriptional processes, with notable reductions in mRNA levels of target genes E2F Transcription Factor 3 (E2F3), LIM Homeobox 1 (LHX1), Kinesin Family Member 18B (KIF18B), Spindle Pole Body Component 25 (SPC25), Nuclear Factor of Activated T cells 2 (NFATC2), Chromatin Accessibility Factor I A (CHAFIA), and Cytoskeleton-Associated Protein 2-Like (CKAP2L) (p < 0.05). Enhanced proliferation and invasion capacities were observed upon TRAF2 overexpression in OSCC cells, whereas TRAF2 silencing or STAT3 inhibition effectively reversed these effects. Additionally, TRAF2 overexpression induced significant alterations in cell morphology, which were reversed upon STAT3 silencing. Collectively, TRAF2 promotes OSCC cell proliferation and invasion by activating the STAT3 signaling pathway.

    Conclusion: This study reveals the pivotal role of TRAF2 in OSCC proliferation and metastasis through modulation of the STAT3 signaling pathway. Elevated TRAF2 and STAT3 expression correlates with enhanced proliferation and invasion capabilities of OSCC cells. The promotion of OSCC development by TRFA2 via STAT3 pathway activation underscores the potential of targeting the TRAF2/STAT3 axis as a therapeutic strategy for OSCC. These findings offer insights into OSCC molecular pathogenesis and lay a theoretical foundation for developing novel treatment approaches.

  • Article
    Yan Huang, Yuanyuan Lin, Shenghua Xu, Xiangxiang Song, Weiming Zhang
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3489-3500. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.277
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    Background: In recent years, there has been a growing interest in understanding the role of RNA binding proteins (RBPs), such as pumilio homolog 2 (PUM2), in the development of diffuse large B-cell lymphoma (DLBCL). Despite this attention, the precise contribution of PUM2 in the progression of DLBCL has not been fully elucidated. This study aimed to examine the role of PUM2 in the development of DLBCL.

    Methods: To determine the expression level and prognostic value of PUM2 in DLBCL, we employed quantitative real-time polymerase chain reaction (qRT-PCR) analysis, immunoblotting, and immunohistochemical (IHC) staining techniques. Moreover, we examined the impact of PUM2 on DLBCL cell proliferation and glucose metabolism through PUM2 overexpression or silencing. In vivo studies were conducted to evaluate the effect of PUM2 on tumor growth post-transplantation. Furthermore, we utilized luciferase reporter assays, RNA immunoprecipitation (RIP), and RNA pull-down experiments to elucidate the molecular mechanism by which PUM2 regulates chromobox protein homolog 3 (CBX3) expression via its 3′UTR.

    Results: Our research revealed that PUM2 was upregulated in clinical samples and DLBCL cell lines (p < 0.001). Additionally, ectopic expression of PUM2 enhanced the proliferation and aerobic glycolysis of DLBCL cells, and this effect was inhibited by silencing PUM2 (p < 0.05). Moreover, PUM2 directly bound to the 3′UTR of CBX3 mRNA and increased its stability. Consequently, PUM2 interacted with and stabilized CBX3 mRNA, thereby promoting the proliferation and glycolytic metabolism of DLBCL cells.

    Conclusions: PUM2 might be a novel regulator of glucose metabolism in DLBCL cells by modulating CBX3 mRNA stability.

  • Article
    Xiaoli Xin, Di Wang, Ruichao Liu, Zhongqi Wang, Zhenye Xu, Yang Guo, Yan Zhang
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3501-3512. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.278
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    Background: Drug resistance in non-small cell lung cancer (NSCLC) cells will affect the therapeutic efficacy of cisplatin (DDP) in treatment. Retinoic acid receptor beta (RARB) has been implicated in the prognosis of NSCLC. This study aimed to investigate the mechanism of RARB in the drug resistance of NSCLC cells.

    Methods: Differential gene expression in drug-resistant NSCLC cells was analyzed using bioinformatics. DDP-resistant NSCLC cells were generated through continuous exposure to DDP over six months. mRNA and protein levels of RARB, nuclear receptor coactivator 3 (NCOA3), and phosphorylated p65 (p-p65) were quantified using quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting. Cell viability was assessed using the cell counting kit-8 (CCK-8) assay, and the half-maximal inhibitory concentration (IC50) of DDP was determined. Cell apoptosis was analyzed via flow cytometry. A rescue experiment was conducted to explore whether the regulatory effect of RARB on DDP-resistant NSCLC cell sensitivity was mediated by NCOA3 regulation.

    Results: RARB expression was downregulated in DDP-resistant NSCLC cells, while NCOA3 and p-p65 expressions were upregulated. RARB overexpression (RARB-OE) inhibited cell viability and enhanced the sensitivity and apoptosis of DDP-resistant NSCLC cells, whereas overexpression of NCOA3 (NCOA3-OE) had the opposite effect. Additionally, RARB-OE reduced NCOA3 and p-p65 levels in DDP-resistant NSCLC cells, while NCOA3-OE increased p-p65 levels. Notably, the impact of RARB-OE and NCOA3-OE on DDP-resistant NSCLC cells was reversed following co-transfection.

    Conclusion: RARB enhances the sensitivity of DDP-resistant NSCLC cells by downregulating NCOA3.

  • Article
    Yang Kong, Bin Fu, Huaye Jiang, Huilin Yang
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3513-3521. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.279
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    Background: Osteoclast formation serves as a triggering factor during osteoarthritis (OA), therefore, our study aimed to elucidate the influence of nuclear receptor corepressor 1 (NCOR1) on osteoclast differentiation and formation.

    Methods: At days 0, 1, 3, or 5 of osteoclast differentiation, osteoclasts were quantified using tartrate-resistant acid phosphatase (TRAP) staining assay, and their apoptosis rate was assessed employing flow cytometry. Moreover, the protein levels of c-Src, β3-integrin, matrix metalloproteinase 9 (MMP9), NCOR1, histone deacetylase 3 (HDAC3), and signal transducer and activator of transcription 3 (STAT3) were measured by western blot analysis. Additionally, the acetylation of STAT3 during the differentiation of osteoclast precursor (OCP) cells was examined through chromatin immunoprecipitation assay. Furthermore, following the transfection of NCOR1 and STAT3 overexpression, the mechanism of NCOR1 in regulating osteoclastogenesis was determined.

    Results: The number of TRAP-positive cells and the protein levels of osteoclast-specific genes (MMP9, c-Src, and β3-integrin) as well as STAT3 and its acetylation were significantly increased during osteoclast differentiation. However, the protein expression levels of NCOR1 and HDAC3 were substantially decreased. Meanwhile, these outcomes were reversed with overexpression of NCOR1. Furthermore, STAT3 up-regulation reversed the hindering effect of NCOR1 on osteoclastogenesis.

    Conclusion: NCOR1 inhibits osteoclastogenesis by suppressing STAT3 acetylation, and this effect can be reversed by STAT3 overexpression.

  • Article
    Xianglian Tang, Yuxiang Zhou, Jiabing Wen, Zhao Huang, Qiangxing Xiang, Denghui Liu
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3523-3533. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.280
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    Background: Neuroblastoma, originating from neural crest, presents a challenging pediatric malignancy. The pituitary tumor-transforming gene 1 (PTTG1) is a recently discovered oncogene implicated in various cancers. This study aimed to elucidate the impact of the PTTG1-tripartite motif 59 (TRIM59) protein complex on the proliferation and metastasis of pediatric neuroblastoma by modulating the phosphorylation status of signal transducer and activator of transcription 3 (STAT3).

    Methods: Utilizing SK-N-AS and SK-N-BE cell lines, we conducted in vitro experiments with gene expression regulation via cell transfection. Migration and invasion capabilities were assessed through matrix gel invasion, wound healing assays, cell activity detection via Cell Counting Kit-8 (CCK-8), and Kaplan-Meier survival analysis. PTTG1 expression in cancer tissue was determined via immunohistochemical techniques. Western blot and co-immunoprecipitation (co-IP) assays were employed to identify PTTG1/STAT3/phosphorylated (p)-STAT3/Bcl-2-associated X (Bax)/B-cell lymphoma-2 (Bcl-2)/TRIM59 expression and interaction. Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) was used to detect the expression levels of PTTG1/Bax/Bcl-2/E-Cadherin/Vimentin/Twist1/Twist2/Snai1/Slug/Zinc finger E-box binding homeobox 1 (ZEB1)/ZEB2.

    Results: PTTG1 knockdown inhibited neuroblastoma cell proliferation, migration, and invasion while promoting apoptosis. It also suppressed STAT3 phosphorylation and epithelial-mesenchymal transition (EMT). Patients exhibiting low PTTG1 expression (p < 0.05) had a significantly better prognosis. qRT-PCR analysis revealed increased expression (p < 0.05) of proapoptotic factors (Bax/E-cadherin) and STAT3-related transcription factors (Bax/Bcl-2/E-cadherin/Vimentin) upon PPTG1 knockdown. Notably, our findings demonstrate the role of the PTTG1-TRIM59 complex in facilitating neuroblastoma progression through the STAT3 pathway.

    Conclusion: The PTTG1-TRIM59 complex and STAT3 represent potential therapeutic targets for pediatric neuroblastoma. Our findings underscore the pivotal role of the PTTG1-TRIM59 complex in stabilizing STAT3 phosphorylation and promoting neuroblastoma growth and metastasis.

  • Article
    Yang Song, Li Pang, Qingjie Li, Shihui Yang, Lan Wei, Shuhan Fan, Chongyong Gao, Jie Yuan
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3535-3545. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.281
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    Background: Chronic atrophic gastritis (CAG) is a prevalent gastrointestinal disorder characterized by gastric inflammation and epithelial cell impairment. Berberine, renowned for its potent anti-inflammatory, antioxidant, and immunomodulatory properties, is frequently utilized in managing gastrointestinal disorders and chronic inflammatory conditions. This study aimed to investigate the protective effects of berberine against CAG in mice and its underlying molecular mechanisms.

    Methods: CAG was induced in mice through Helicobacter pylori (Hp) infection and ethanol administration. Subsequently, mice were treated with varying doses of berberine (10 and 50 mg/kg). The therapeutic efficacy of berberine was evaluated by analyzing histopathological alterations, serum inflammatory markers, and oxidative stress levels. Moreover, the expression levels of stromal cell-derived factor-1 alpha (SDF-1α), C-X-C chemokine receptor type 4 (CXCR4), vascular endothelial growth factor (VEGF), and proteins related to the nuclear factor-kappa B (NF-κB) signaling pathway were examined through Western blot analysis and immunohistochemistry.

    Results: Compared to the CAG group, berberine treatment can significantly improve CAG-related histopathological changes, reduce inflammatory infiltrating cells, and reduce gastric gland atrophy and epithelial cell damage. Berberine administration also reduced the levels of tumor necrosis factor-α (TNF-α), Interleukin-6 (IL-6), Interleukin-1β (IL-1β), and cyclooxygenase-2 (COX2) compared to the CAG group. Additionally, berberine downregulated the expression of SDF-1α, CXCR4, VEGF, and NF-κB p65 in gastric tissue, suggesting its inhibitory effect on these signaling pathways.

    Conclusion: This study demonstrates that berberine exerts protective effects against CAG in mice by suppressing the SDF-1α-CXCR4-VEGF and NF-κB signaling pathways. These findings underscore the potential utility of berberine in managing CAG and provide a basis for further investigation into its clinical applicability.

  • Article
    Shaoxian Liu, Jingjie Luan
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3547-3557. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.282
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    Background: Bone fracture is a common injury, and the healing process involves the intricate regulation of multiple cells and signaling pathways. Bone Morphogenetic Protein-5 (BMP-5) plays a crucial role as a bone morphogenetic protein in the healing of bone fractures. This study aims to investigate the precise mechanism of BMP-5 in fracture healing.

    Methods: Mouse mesenchymal stem cells (MSCs) were used as the research model to explore the role of BMP-5 in bone fracture healing. Models of MSCs with BMP-5 knockdown and BMP-5 overexpression were established. The study investigated the effects of BMP-5 on the osteogenic differentiation ability of MSCs, the activity of the β-catenin and Extracellular Signal-Regulated Kinase (ERK) 1/2 signaling pathways, as well as mitochondrial dynamics using quantitative real-time polymerase chain reaction (qRT-PCR), Western blot, and immunofluorescence staining. Additionally, the impact of BMP-5 on fracture-healing tissues was assessed through histological analysis and tube formation assays.

    Results: The experimental results demonstrated that silencing BMP-5 expression significantly hindered the osteogenic differentiation potential of murine peripheral blood-derived MSCs (mPB-MSCs) (p < 0.05). Conversely, the overexpression of BMP-5 substantially enhanced the osteogenic differentiation ability of mPB-MSCs (p < 0.05). Additionally, BMP-5 overexpression elevated the activity of the β-catenin and ERK1/2 signaling pathways in mPB-MSCs and modulated mitochondrial dynamics (p < 0.05). Histological analysis and tube formation assay results revealed that BMP-5 overexpression in mPB-MSCs promoted angiogenesis and enhanced tube formation capacity in the tissue during bone fracture healing.

    Conclusions: The upregulation of BMP-5 can promote the osteogenic differentiation of mPB-MSCs and angiogenesis in fracture healing by activating the Wnt/β-catenin and ERK1/2 signaling pathways and enhancing mitochondrial dynamics.

  • Article
    Min-Ho Cha, Myong-Min Lee, Jin-Yeul Ma
    Journal of Biological Regulators and Homeostatic Agents. 2024, 38(4): 3559-3568. https://doi.org/10.23812/j.biol.regul.homeost.agents.20243804.283
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    Background: Acer Palmatum Thumb is a traditional medicinal herb used in East Asia, and previous studies performed by our team revealed the anti-inflammatory and neuroprotective effects of KIOM-2015EW, a water extract of Acer Palmatum Thumb leaves. Hence, this study aimed to explore the inhibitory effects of KIOM-2015EW on decreasing proprotein convertase subtilisin/kexin type 9 (PCSK9) expression in a fructose-induced fatty liver model.

    Method: HepG2, a human hepatocyte-derived hepatocellular carcinoma, was exposed to low-glucose Dulbecco's modified Eagle's medium (DMEM) media with or without fructose, statins, and KIOM-2015EW. Cell viability was measured by cell counting kit-8 (CCK-8) analysis, and fat accumulation and low-density lipoprotein (LDL) uptake were, respectively, detected by oil-red O staining and LDL-uptake assay. The mRNA and protein levels of PCSK9, low-density lipoprotein receptor (LDLR), peroxisome proliferator-activated receptor alpha (PPARα), and retinoid X receptor alpha (RXRα) were measured via real-time quantitative polymerase chain reaction (RT-qPCR) and western blot. PCSK9 transcription factors were predicted via chemical-protein interactions using the Stichi database.

    Result: KIOM-2015EW significantly decreased fat accumulation and recovered LDL uptake in fructose-induced cells (p < 0.05). That also decreased PCSK9 levels induced by fructose or fructose/lovastatin (p < 0.05) and recovered LDLR mRNA expression. Chemical-protein interactions showed that eleven transcription factors (TFs) interacted with two major compounds of KIOM-2015EW. Inhibition of PPARα/RXRα and mitogen-activated protein kinase (p38) significantly attenuated PCSK9 expression induced by fructose. KIOM-2015EW decreased PPARα and RXRα expression which was increased by fructose and fructose/statin (p < 0.05).

    Conclusion: This study suggested that KIOM-2015EW may lower serum cholesterol, and it can be used as a natural substance for the prevention or treatment of hypercholesterolemia.