20 November 2023, Volume 37 Issue 11

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  • Review
    Fabio Cardinale, Michele Fiore, Giorgio Piacentini, Alfonso Varricchio
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5745-5755. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.551
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    Recurrent Respiratory Infections (RRIs) in the pediatric population have an important clinical and economic impact, which can be reduced through appropriate preventive measures, including immunomodulating agents, like bacterial lysates. This manuscript aimed to provide an overview of the clinical evidence on the efficacy and safety of bacterial lysates in the prevention of RRIs in children, with a particular focus on OM-85 (Broncho Vaxom®) as well as to shed light on the concomitant use of OM-85 and vaccines as a successful strategy for reducing respiratory morbidity in the pediatric population. In children with RRIs, OM-85 treatment significantly reduce the number and duration of Respiratory Tract Infections (RTIs), antibiotic courses and wheezing respiratory illnesses in young children. Its efficacy seemed to be more pronounced in children with more respiratory infections. Moreover, OM-85 was effective and well tolerated when administered concomitantly with inactivated influenza vaccine. Due to its long-term efficacy and good safety profile, OM-85 can be recommended as a valid prophylactic approach for preventing RTIs in children, especially in cases where the incidence of the disease is high and attempts to eliminate contributing environmental factors have proven unsuccessful.

  • Review
    Bhagyalakshmi Nair, Aparna Ashokan, Veena Gopalakrishnan Prema, Girish Banavara Narasimhamurthy, Rajesh Gopalakrishna, Devika Vadhyaruparambil Padamakumar, Lekshmi R. Nath, Anca Oana Docea, Javad Sharifi-Rad, Daniela Calina
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5757-5768. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.552
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    Background: The global pandemic instigated by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), or the 2019 Novel Coronavirus (2019-nCoV), has intensified the quest for efficacious preventive and therapeutic solutions. Zinc, an essential trace element known for its antioxidant properties, has emerged as a potential player.

    Objective: This review aims to scrutinize the role of Zinc, focusing on its antioxidant capacities, in the prevention and management of coronavirus disease 2019 (COVID-19), elucidating the mechanisms by which it may impede the progression and severity of the disease.

    Methods: Comprehensive data were gleaned from various clinical trials, research publications, and observational studies exploring the interaction between Zinc and SARS-CoV-2. The review evaluates both the direct antiviral effects of Zinc and its impact on host immune responses.

    Results: Initial research indicates that Zinc could interfere with specific phases of the SARS-CoV-2 virus's life cycle, such as its entry into cells and subsequent replication. This effect may be due to Zinc's ability to reduce the activity of certain viral proteins. These include the interaction between the virus's Spike protein and the angiotensin-converting enzyme-2 (ACE-2) receptor on human cells, and the RNA-dependent RNA polymerase enzyme essential for viral replication. Additionally, Zinc's antioxidant properties are pivotal in bolstering the host's immune response, mitigating oxidative stress, and reducing the severity of inflammatory cascades associated with severe COVID-19 presentations.

    Conclusions: The multifaceted roles of Zinc, particularly its antioxidant capabilities, underscore its potential significance in the prophylaxis and therapeutic management of COVID-19. Further rigorous clinical studies are imperative to delineate the optimal dosing, timing, and synergistic interventions that can maximize Zinc's therapeutic benefits against the disease.

  • Review
    Sarika Bano, Muhammad Asim Raza, Shampa Ghosh, Nayaab S. Pandit, Saumya Srivastava, Mohammad Azam, Sanjay Kumar Dey, Sung Soo Han, Jitendra Kumar Sinha, Munindra Ruwali
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5769-5784. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.553
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    Alzheimer's disease (AD) stands as a devastating neurodegenerative disorder, depicted by the relentless erosion of cognitive and memory faculties. Despite relentless research spanning decades, the quest for efficacious disease-modifying therapies remains an ongoing challenge. Recent strides in the realm of molecular neuropharmacology, however, have revealed a number of potential therapeutic targets for AD, instilling a renewed sense of optimism. This comprehensive review navigates the landscape of emerging avenues in the quest to combat AD, shedding light on a diverse array of targets that hold substantial promise. Central to this discourse are novel targets such as beta-secretase (BACE) and gamma-secretase, instrumental in the making of amyloid-beta (Aβ) peptides, the hallmark culprits of AD pathology. Deconstructing the pathological cascade further, the review delves into the intricate involvement of tau protein abnormalities, neuroinflammation cascades, and proteins linked with synaptic dysfunction. Beyond the traditional scope, this review also ventures into the realm of cutting-edge modalities, elucidating the potential of ground-breaking techniques including immunotherapies and microRNA-based interventions. These innovative strategies, harnessing immune modulation and intricate gene regulation, introduce new dimensions to AD therapeutics, providing novel avenues to halt disease progression. A thorough understanding of these emerging therapeutic targets, coupled with in-depth exploration of their underlying mechanisms, sparks a paradigm shift in conceptualizing potent AD interventions. By targeting these prospects, the objective extends beyond mere symptomatic relief, aiming to not just impede disease advancement but also enhance the well-being of patients and caregivers. However, the transition from potential to clinical reality mandates unwavering commitment to rigorous scientific inquiry and meticulous clinical validation. These promising candidates require further scientific scrutiny and systematic trials to establish efficacy, safety, and long-term benefits. Only through such concerted efforts can the transformative potential of these burgeoning therapeutic targets be translated into robust and secure AD treatments.

  • Article
    Minami Kosuda, Midori Yamana, Akiko Nagasawa, Aya Tanaka, Kentaro Watanabe, Hisamitsu Ishihara
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5785-5794. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.554
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    Background: Carbonic anhydrase (CA) catalyzes the formation of bicarbonate ions and protons from carbon dioxide and water, and also the reverse reaction, thus regulating acid-base balance and processes requiring HCO3- in the cell. Previous data from us and others have suggested that CAs participate in the regulation of insulin secretion. The aim of this study was to examine sublines of insulin-secreting MIN6 cells in terms of CA isoform transcript levels, and to analyze their roles in insulin secretion.

    Methods: Differential expression of CA isoforms in MIN6 cells differing in glucose responsiveness were analyzed by reverse transcription-polymerase chain reaction (RT-PCR). Next, we established stable cell lines with a doxycycline-inducible system overexpressing several of the CA isoforms. We also generated MIN6 sublines with shRNA-mediated suppression and studied glucose-stimulated insulin secretion, as well as glucose metabolism, using radiolabeled glucose in these cells.

    Results: We confirmed expression of a mitochondrial CA isoform, CA VB, and a cytosolic isoform, CA XIII, was significantly greater (4.18-fold, p < 0.01 for CA VB; 48.3-fold, p < 0.01 for CA XIII), while expression of CA XI, a cytosolic isoform that lacks catalytic activity, was lower (0.50-fold, p < 0.01), in highly glucose-responsive sublines. CA VB overexpression increased (1.32-fold, p < 0.05), while its knockdown suppressed (0.77-fold, p < 0.05), glucose-stimulated insulin secretion. In contrast, CA XIII overexpression and CA XIII knockdown failed to modulate insulin secretion evoked by glucose. CA XI overexpression produced no changes in insulin secretion. Changes in insulin secretion due to CA VB overexpression or knockdown were not accompanied by changes in [5-3H]glucose and [U-14C]glucose metabolism.

    Conclusions: CA VB serves as a positive regulator of glucose-stimulated insulin secretion in MIN6 cells.

  • Article
    Federica Vaccaro, Roberta Corsaro, Elisabetta Miraldi, Giulia Collodel, Marco Biagi, Cinzia Signorini, Giulia Baini, Lucia Micheli, Rosetta Ponchia, Giorgio Cappellucci, Elena Moretti
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5795-5809. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.555
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    Background: Semen handling and cryopreservation increase reactive oxygen species (ROS) production, exposing spermatozoa to oxidative stress (OS) that can be minimised by antioxidant supplementation. We studied if Olea europaea leaf extract (OE) could have antioxidant and protective activity during sperm manipulation.

    Methods: The extract was characterized by high-performance liquid chromatography with diode array detection (HPLC-DAD) and antioxidant activity was determined by 2,2-diphenyl-1-picrylhydrazyl (DPPH) and oxygen radical absorbance capacity (ORAC) assays. Then, spermatozoa of 35 normozoospermic donors were treated with OE. First, swim-up selected spermatozoa were incubated with OE (1:100–1:400) and sperm motility and DNA integrity (acridine orange test) were assessed. Then, swim-up selected sperm were treated with 100 μM H2O2 to induce OS with and without OE; motility, DNA integrity and F2-Isoprostanes (F2-IsoPs quantified by gas chromatography/negative ion chemical ionization tandem mass spectrometry analysis), an OS marker, were assessed. Finally, sperm were frozen with and without OE. Beside the previous endpoints, acrosome shape was evaluated by Tetramethylrhodamine (TRITC)-conjugated Pisum sativum agglutinin (PSA).

    Results: OE was enriched in polyphenols (0.34%) and triterpenes (0.72%) and the synergistic action of phytocomplex components was responsible for antioxidant activity. Since OE was not toxic for spermatozoa, 1:100 dilution was used for the other experiments. OE protected motility, DNA integrity and reduced F2-IsoPs (p < 0.001) in in vitro experiment with OS induction. OE treated frozen-thawed spermatozoa showed increased motility, DNA integrity, reduced F2-IsoP concentration (p < 0.001) and normal acrosomes versus non-supplemented samples.

    Conclusions: OE is characterized by high concentration of polyphenols and exhibits protective properties against oxidative damage induced by H2O2 in human ejaculated sperm. After cryopreservation, the samples supplemented with the extract showed increased sperm quality. OE could represent a supplement of culture media during semen handling where OS is exacerbated.

  • Article
    Ramadan S. Hussein, Salman Bin Dayel, Othman Abahussein, Shereen H. Elsayed
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5811-5816. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.556
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    Background: Elderly populations are expanding in both wealthy and developing countries. The aged people have a particular skin disease spectrum that varies geographically. With aging, the skin system experiences structural and functional changes. Geriatric dermatoses provide a tough condition for clinicians in terms of diagnosis, treatment, and follow-up. There is little data on skin illness in Saudi Arabia's elderly community. This research aimed to investigate the prevalence and distribution of cutaneous illnesses in patients in Saudi Arabia's middle region.

    Methods: The retrospective study included patients who attended the dermatology outpatient clinic of Prince Sattam Bin Abdulaziz University Hospital between January 1, 2020, and December 31, 2022. Their genders, ages, and cutaneous diagnoses were collected, and the descriptive analysis was performed using SPSS version 22.

    Results: 1152 patients were present, with a male predominance of 63.45%. The most prevalent diagnosis was skin eczema or dermatitis (39.91%), followed by infections (20.31%), papulosquamous illness (13.37%), and benign neoplasms (12.85%). There were no cancerous tumors.

    Conclusions: Eczema/dermatitis and infections are major health issues for older persons in this region. Most illnesses are preventable; to lessen the prevalence of certain skin disorders, it is crucial to increase public awareness of risk factors and conduct additional research in related fields.

  • Article
    Meiqin Yuan, Jianfei Fang, Li Wang, Rongrong Chen, Haijun Zhong, Hongming Pan, Wangxia Lv
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5817-5827. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.557
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    Background: The most significant prognostic variables for colorectal cancer (CRC) are tumor sidedness and patterns of metastatic spread. However, less is known about the differences in the mutational profile of hepatic and extrahepatic metastases between left-sided and right-sided colorectal cancer. To address this knowledge gap, we sought to analyze the association between different metastatic sites and the gene mutational profiles in each population. We aimed to discover the molecular heterogeneity between these two subtypes of colorectal cancer.

    Methods: We retrospectively analyzed mutations from 185 colorectal cancer patients. The samples for next generation sequencing were primary colon tumor tissues. Formalin-fixed, paraffin-embedded tumor tissues were collected and deep sequencing targeting 1021 genes was conducted to analyze genomic alteration. Correlation analysis was performed by Chi-square or Fisher's exact tests.

    Results: A total of 185 CRC patients (132 left-sided colon cancer (LCC) and 53 right-sided colon cancer (RCC)) were analyzed. These included 60 patients with (32.4%) solitary liver metastasis (SLM) cohort, 43 patients (23.2%) with multiple liver metastases (MLM) cohort, 56 patients (30.3%) with extrahepatic metastases (EHM) cohort, and 26 patients (14.1%) without metastases (control cohort). Tumor protein 53 (TP53, 88% vs. 68%, p < 0.05) and adenomatous polyposis coli (APC, 78% vs. 58%, p < 0.05) were found to be mutated more frequently in LCC than RCC. In contrast, B-raf murine sarcoma viral oncogene homologue B (BRAF, 5% vs. 11%, p < 0.05), phosphatidylinositol-4,5-bisphosphate 3-kinase, catalytic subunit alpha (PIK3CA, 13% vs. 23%, p < 0.05), lysine demethylase 6A (KDM6A, 1% vs. 8%, p < 0.05), FAT atypical cadherin 2 (FAT2, 2% vs. 11%, p < 0.05), and SRY-box transcription factor 9 (SOX9, 7% vs. 19%, p < 0.05) were more common in RCC patients compared with those with LCC. In LCC, APC was more frequently mutated in SLM, MLM, and control cohorts than in the EHM cohort (85% vs. 83% vs. 90% vs. 57%, p < 0.05). In RCC, the mutation frequencies of APC and PIK3CA were more significant in the SLM cohort when compared to the EHM cohort. However, a converse result was observed for the SMAD family member 4 (SMAD4) gene.

    Conclusions: These results demonstrate the molecular heterogeneity between hepatic and extrahepatic metastases from left- and right-sided colorectal cancer. These findings could help us better understand the mechanisms underlying tumor metastasis and reveal disparities in survival among metastatic locations.

  • Article
    Fang Zong, Yingzi Zhao, Yanmin Cao, Qian Geng, Qinying Cao
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5829-5836. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.558
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    Background: Poly (ADP-ribose) polymerase (PARP) 1 and PARP2 deficiency in the uterus results in the loss of pregnancy. Metastasis associated lung adenocarcinoma transcript 1 (MALAT1), a PARP1 binding nucleus-localized long non-coding ribonucleic acid (RNA), is downregulated in patients with recurrent pregnancy loss (RPL). However, the correlation between MALAT1 and PARPs, and their roles in RPL remain unclear.

    Methods: In this study, MALAT1, PARP1, and PARP2 levels were examined in the decidua from a cohort consisting of 35 patients with RPL and 15 healthy controls. The interaction between MALAT1 and PARP2 was identified in primary decidual stromal cells using a ribonucleoprotein immunoprecipitation assay and confirmed by RNA pull-down and immunofluorescence. Deoxyribonucleic acid (DNA) damage, cell viability, and apoptosis were examined by immunoblotting, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay, and flow cytometry in PARP2 knockdown cells.

    Results: MALAT1 and PARP2 levels were downregulated in patients with RPL. The messenger RNA level of PARP2 was positively correlated with MALAT1 levels (r = 0.41, p = 0.013). PARP2 bound MALAT1 close to the 3′ end and colocalizes with it in the cell nucleus. In PARP2 knockdown cells, MALAT1 was re-localized to the nucleus rim and cytoplasm and degraded more quickly. In decidual stromal cells, PARP2 knockdown led to DNA damage, decreased cell viability, and increased apoptosis.

    Conclusions: We identified the interaction between PARP2 and MALAT1 for the first time and constructed a correlation between MALAT1/PARP2 dysregulation and the occurrence of RPL, which provides new clues for RPL treatment.

  • Article
    Feng Wang, Qionghui Huang, Qimin Wu, Lishu Wang, Huanji Zhang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5837-5850. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.559
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    Background: Atherosclerosis is a complex and pervasive chronic disease with a substantial global health impact. Emerging evidence highlights the role of osteogenic transdifferentiation in vascular smooth muscle cells (VSMCs) as a contributor to atherosclerosis development. However, the precise molecular mechanisms involved remain unclear. This study aimed to uncover common genes that link osteogenic transdifferentiation of VSMCs and atherosclerosis.

    Methods: We conducted weighted gene co-expression network analysis (WGCNA) using publicly available atherosclerosis-related datasets, namely GSE43292 and GSE28829. We identified key module genes that exhibited a positive correlation with both osteogenic transdifferentiation and atherosclerosis, designating them as Geneset 1. Functional enrichment analysis of Geneset 1 was conducted using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses. We further screened for upregulated differentially expressed genes in the GSE135626 dataset that intersected with Geneset 1, resulting in the identification of genes with positive associations with osteogenic transdifferentiation of VSMCs and atherosclerosis, designating them as Geneset 2. We validated the expression of Geneset 2 in the osteogenic-induced VSMCs cell line (MOVAS-1) through RT-qPCR and identified hub genes. The hub genes were further validated in mouse atherosclerotic plaque samples using immunohistochemistry (IHC). Moreover, we performed receiver operating characteristic (ROC) analysis to assess the predictive value of the hub genes for atherosclerotic plaque progression and stability. Furthermore, we constructed a network illustrating the interaction between transcription factors (TFs) and hub genes based on our findings.

    Results: Using WGCNA, we identified three significant gene modules and 99 osteogenic-related genes positively associated with atherosclerosis in Geneset 1. These genes were primarily involved in osteoclast differentiation pathways, exosome secretion-related functions, and immunoinflammatory processes according to GO and KEGG analysis. Subsequently, we isolated 11 candidate genes (Geneset 2) that demonstrated a positive linkage between osteogenic transdifferentiation of VSMCs and atherosclerosis. RT-qPCR confirmed the upregulation of four hub genes (branched-chain amino acid transaminase 1 (BCAT1), receptor-interacting protein kinase 3 (RIPK3), haematopoietic cell kinase (HCK), and plasminogen activator urokinase receptor (PLAUR)) from Geneset 2 in osteogenic-induced MOVAS-1 cells and in mouse atherosclerotic plaque samples through IHC. Moreover, the four hub genes exhibited strong predictive values for atherosclerotic plaque progression and stability in ROC analysis. Finally, we identified 26 TFs that target the hub genes, shedding light on potential regulatory mechanisms.

    Conclusions: This study identifies BCAT1, RIPK3, HCK, and PLAUR as shared hub genes linking osteogenic transdifferentiation of VSMCs and atherosclerosis. These findings provide valuable insights for future research.

  • Article
    Gizem Ayna Duran
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5851-5864. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.560
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    Background: Pan-cancer research has recently been conducted to identify genomic differences in multiple cancers compared with those in normal tissues. Our aims were to determine common differentially expressed genes (DEGs) by analysing the genomes of multiple cancers and to identify common cancer biomarkers for both diagnosis and treatment by determining the effect of these commonly changed genes on survival.

    Methods: We performed DEG and survival analyses by considering differences in expression levels in tumour and related normal tissues using the R programme and bioinformatic webtools Gene Expression Profiling Interactive Analysis 2 (GEPIA2), UALCAN, TIMER2.0, KM-plot, ShinyGO 0.77, miRTargetLink, and CancerMIRNome.

    Results: In our study, 5 genes (ribonucleotide reductase regulatory subunit M2 (RRM2), enhancer of the zeste homolog 2 (EZH2), Proliferating cell nuclear antigen (PCNA) clamp associated factor (PCLAF), mitotic checkpoint serine/threonine kinase B (BUB1B), and centromere protein U (CENP-U)) were found to be upregulated in multiple cancers. Among these genes, the RRM2 gene was upregulated in more cancers and datasets. According to the gene enrichment analysis, the EZH2 gene also acted together with the RRM2 gene in terms of biological significance. Furthermore, instead of finding chemical agents that would inhibit the overexpression of both the RRM2 and EZH2 genes, miRNA screening targeting both genes were performed. As a result, hsa-let-7a-5p miRNA was found to have a strong interaction with both genes through the p53- and cell cycle-dependent pathways.

    Conclusion: The commonly upregulated RRM2 and EZH2 genes can be utilised as biomarkers for early diagnosis of multiple cancers. The joint interactions of the RRM2 and EZH2 genes with hsa-let-7a-5p may be a promising early treatment by inhibiting the expression levels of both genes by targeting hsa-let-7a-5p, an miRNA with low expression levels in cancers.

  • Article
    Shiyan Wu, Jie Meng, Wei Liu, Yong Fan, Jing Xu, Aziguli Aji, Peng Xi, Shuli Zhang, Yimei Li, Yan Hao
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5865-5872. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.561
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    Background: Diabetic peripheral neuropathic pain (DPNP) is one of the most common complications of diabetes mellitus (DM). This study aimed to screen and analyze the differential expression of long non-coding RNAs (lncRNAs) and mRNAs in Uygur and Han diabetic peripheral neuropathic pain (DPNP) patients in Xinjiang.

    Methods: From July 2019 to December 2019, we recruited 3 Uyghur patients who were diagnosed with type 2 DM and DPNP. We then selected 3 Han Chinese patients with DPNP from the same hospital and matched them accordingly. The diagnostic criteria for DPNP were in line with the 2018 Expert Consensus on the Diagnosis and Treatment of Diabetic Peripheral Neuralgia. Peripheral blood samples were taken from these patients in each group to analyze the expression profile of lncRNAs through microarray. Differentially expressed lncRNAs and mRNAs were screened through gene chip analysis of peripheral blood from 3 pairs of DPNP patients.

    Results: 275 differentially expressed lncRNAs (171 upregulated, 104 downregulated) and 279 differentially expressed mRNAs (152 upregulated, 127 downregulated) were identified between Uygur and Han DPNP patients. The potential target genes of lncRNAs were found to be concentrated in the toll-like receptor signaling pathway and Wnt signaling pathway. Additionally, the Gene Ontology (GO) analysis showed that the differentially expressed lncRNAs were mainly associated with the activity of transmembrane signal receptors, the response to tumor necrosis factor, and the positive regulation of natural killer cell chemotaxis.

    Conclusions: There were differentially expressed lncRNAs and mRNAs between Xinjiang Uygur and Han DPNP patients. These lncRNAs and mRNAs may play an important role in the occurrence and development of DPNP.

  • Article
    Yifei Wang, Lizhen Liu, Peng Xu, Runsheng Guo, Shanjuan Wang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5873-5878. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.562
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    Objective: This study aims to investigate the effect of partial splenic embolization (PSE) in the treatment of liver cirrhosis with hypersplenism and its influence on portal vein blood flow and T lymphocyte levels.

    Methods: Ninety-four liver cirrhosis patients with hypersplenism admitted to Jiading District Central Hospital from February 2018 to June 2021 were selected and randomly divided into two groups, with 47 cases in each group. The patients in Group A accepted PSE treatment, while patients in Group B accepted traditional splenectomy. Routine blood indices, portal vein hemodynamic indices, liver perfusion level, peripheral blood T lymphocytes and surgical complications were compared between the two groups at different time before and after the operation.

    Results: The levels of red blood cell, white blood cell, and platelet in Group A and Group B were considerably higher than those before the operation at 1 month and 3 months after the operation (p < 0.05). At 3 months after the operation, internal diameter of the portal vein, blood flow velocity of the portal vein, portal vein blood flow and portal vein pressure in Group A were lower than those in Group B (p < 0.05). At 3 months after the operation, hepatic artery perfusion (HAP) values in Group A were higher than those in Group B, and the portal vein perfusion (PVP) and total liver perfusion (TLP) values in Group A were lower than those in Group B (p < 0.05). Three months after the operation, the CD3+ and CD4+ values in Group A were higher than those in Group B (p < 0.05). The complication rate of Group A (8.5%) was lower than that of Group B (23.40%) (p < 0.05).

    Conclusion: The effect of PSE in treating liver cirrhosis with hypersplenism is better than that of traditional splenectomy. It has better effect on improving portal vein blood flow and liver perfusion, promoting the recovery of immune function and causing fewer complications.

  • Article
    Zhihui Hu, Yan Zhang, Yan Li, Yaguang Han
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5879-5886. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.563
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    Background: Osteopontin (OPN) conducts a vital part in the metastasis of non-small cell lung cancer (NSCLC). This research investigates whether OPN modulates the radiosensitivity of NSCLC cells through the JAK (Janus tyrosine Kinase) 2/STAT (Signal Transducer and Activator of Transcription) 3 pathway in NSCLC.

    Methods: NCI-H1299 cells were exposed to different doses of radiation. The cells were divided into 6 subgroups: Control (CON) subgroup, si-non-specific control (NC) subgroup, si-OPN subgroup, CON+8Gray (GY) subgroup, si-NC+8GY subgroup and si-OPN+8GY subgroup. The OPN mRNA level was measured by quantitative real-time polymerase chain reaction (qRT-PCR). The OPN, p-H2A histone family member X (p-H2AX), H2AX, p-STAT3, STAT3, p-JAK2 and JAK2 levels were measured by western blot. DNA damage in cells was measured by comet assay. Cell proliferation was measured by colony formation assay. Scratch healing assay and transwell assay were used to measure the migration and invasion ability of the cells.

    Results: Different doses of irradiation could reduce the mRNA and protein levels of OPN (p < 0.05). Both si-OPN and 8GY rays could reduce OPN, p-STAT3 and p-JAK2 levels in NCI-H1299 cells, boost p-H2AX level, promote DNA damage, reduce cell activity, and restrain cell metastasis (p < 0.05).

    Conclusion: OPN may affect the radiosensitivity and activity of NCI-H1299 cells through the JAK2/STAT3 pathway.

  • Article
    Haci Ömer Osmanlıoğlu, Mustafa Nazıroğlu
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5887-5901. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.564
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    Background: Lipopolysaccharide (LPS) production through the upregulation of transient receptor potential melastatin 2 (TRPM2)-caused Ca2+ influx, cytokine production, intracellular reactive oxygen molecules (iROS), and mitochondrial free reactive oxygen radicals (mROS) generation in microglia induces neurodegenerative diseases. As a synthetic glucocorticoid and anesthetic drug, dexamethasone (DEX) inhibits LPS-induced oxidative stress, cytokine production, and apoptosis in brain cells, including mice microglia (BV-2). We aimed to investigate the protective role of DEX on LPS-induced inflammation, apoptosis, and oxidative cytotoxicity in BV-2 cells by attenuating TRPM2 signaling.

    Methods: We induced five primary groups in the cultured BV-2 cells: control, DEX (100 nM for 24 h), LPS (1 μg/mL for 24 h), LPS plus DEX, and LPS plus TRPM2 blocker (N-(p-amylcinnamoyl) anthranilic acid or carvacrol). The cell death, oxidants, and Ca2+ analyses of the current study were performed using laser scan confocal microscopy (via fluorescent dyes). A spectrophotometer was used for the lipid peroxidation and antioxidant analyses. An automatic plate reader was used for the apoptosis, cytokine, and caspase analyses. The cell number, debris, and cell viability were determined using a cell counter.

    Results: The amount of cell death, apoptosis, oxidants (mROS, iROS, and lipid peroxidation), apoptotic indicators (caspase -3, -8, and -9), cytokines (tumor necrosis factor-alpha (TNF-α), interleukin-1beta (IL-1β), and interleukin-6 (IL-6)), death cell waste (debris), and Ca2+ were upregulated by LPS incubation, although their amounts were diminished by the DEX and TRPM2 blocker treatments (p ≤ 0.05). The DEX treatments enhanced the LPS-mediated declines in cell viability, cell number, glutathione, and glutathione peroxidase values (p ≤ 0.05).

    Conclusion: The treatment of DEX diminished the LPS-caused inflammatory cytokine, apoptosis, and oxidative stress via TRPM2 attenuation in microglia cells. The DEX may be considered a potential therapeutic way to treat LPS-caused microglia oxidative neuronal injury and neurodegenerative diseases.

  • Article
    Yang Li, Chuan Wang, Jiangya Wang, Xiangdan Yang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5903-5911. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.565
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    Background: Lymphoma has emerged as the most common neoplasm among children, nearly half of them being non-Hodgkin lymphoma (NHL). To explore appropriate therapeutic targets and suggest directions for future research, this study aimed to identify the effects of TNF receptor-associated factor 6 (TRAF6) on NHL and its correlation with the phosphatidylinositol 3-kinase and nuclear factor kappa-B (PI3K/NF-κB) pathway.

    Methods: Blood samples were collected from healthy children and those affected with NHL. The cell lines, Raji human B lymphoma (iCell-h178) and U-937 human leukemia (iCell-h267), were maintained on RPMI-1640 culture medium. The expression profile of TRAF6, phosphatidylinositol 3-kinase (PI3K), and nuclear factor kappa-B (NF-κB) was determined by performing the real-time reverse transcriptase-polymerase chain reaction (RT-qPCR) using blood samples from both healthy and affected children. Furthermore, RT-qPCR, Western Blot, Cell Counting Kit-8 (CCK-8) assay, and flow cytometry were used to evaluate the impact of TRAF6 expression levels on NHL cell proliferation and apoptosis, as well as the role of TRAF6 in mediating the PI3K/NF-κB pathway.

    Results: TRAF6, PI3K, and NF-κB were over-expressed in NHL patients (p < 0.05). Decreased/increased TRAF6 expression led to decreased/increased Proliferating cell nuclear antigen (PCNA), Cyclin D1, Bcl-2 protein expression and NHL cell proliferation (p < 0.05), while increasing/decreasing Cleaved Caspase-3, Bax protein expressions and NHL cell apoptosis (p < 0.05). In addition, decreased/increased TRAF6 expression could lead to decreased/increased p-PI3K, p-NF-κB protein expressions (p < 0.05).

    Conclusion: In summary, TRAF6 was over-expressed in NHL patients and NHL cell lines. However, down-regulation of TRAF6 reduced proliferation and induced apoptosis in NHL cells through deactivating PI3K/NF-κB pathway, thus suppressing the progression of NHL.

  • Article
    Yan Yang, Zhong-jun Wang, Chun-xiang Li, Bin Li, Ze-gang Liu, Fu-huan Wang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5913-5928. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.566
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    Background: Breast cancer (BC) is highly prevalent in women and is often fatal. Vasculogenic mimicry (VM) is associated with cancer progression. The expression of Cavolin-2 (CAV-2) affects the occurrence and development of BC, but the specific mechanism of CAV-2 in BC is still poorly understood. The aim of our study was to explore the specific mechanism of the effect of CAV-2 on the occurrence of VM and the malignant biological behavior of BC.

    Methods: First, we detected the gene expression of CAV-2 in BC tissues and cell lines. The overexpressed CAV-2 plasmid (oe-CAV-2) and si-CAV-2 were then transfected into michigan cancer foundation-7 (MCF-7) cells to determine the effects of different expression levels of CAV-2 on BC cells. In addition, to further determine the feedback loop mechanism between CAV-2 and epidermal growth factor receptor (EGFR), the cells were treated with (1) AG1478, which inhibits EGFR phosphorylation, (2) an EGFR phosphorylation activator epidermal growth factor (EGF), (3) a ubiquitination proteasome inhibitor MG-132, and (4) an extracellular regulated protein kinases1/2 (ERK1/2) inhibitor SHR2415. In the cell experiment, VM and the biological behavior of BC cells were mainly observed by tumor spheroid formation, tube formation, Transwell, 5-ethynyl-2'-deoxyuridine (EdU), and scratch tests. In addition, we performed Western blot analysis of epithelial–mesenchymal transition (EMT)-associated proteins and angiogenesis analog markers. Finally, subcutaneous injection of breast cancer cells into nude mice was used to establish an animal model of BC so that the effect of CAV-2 on the progression of BC could be verified in vivo.

    Results: CAV-2 was reduced in BC patient cancer tissue and in human BC cells (p < 0.001). Overexpression of CAV-2 effectively inhibited the progression of BC cells and reduced the formation of tumor spheres and tubules, whereas knockdown of CAV-2 had the opposite effect. CAV-2 and EGFR could interact, and the expression of EGFR was higher after overexpression of CAV-2 (p < 0.001), but P-EGFR expression was lower (p < 0.01). Treatment with the EGFR phosphorylation inhibitor AG1478 further enhanced the effect of oe-CAV-2. Treatment with the EGFR phosphorylation activator EGF reversed the effects of oe-CAV-2. After EGF treatment, the CAV-2 ubiquitination level was increased, but its gene expression decreased (p < 0.01). EGF promoted VM occurrence and malignant biological behavior of BC cells, and the EGF effect was reversed after overexpression of CAV-2. EGF also promoted the upregulation of P-ERK1/2 expression (p < 0.001), and the promotion effect of EGF on the behavior and angiogenic mimicry of BC cells was reversed after treatment with SHR2415, an ERK1/2 inhibitor. Animal experiments have also confirmed that CAV-2 can inhibit tumor growth and the phosphorylation of EGFR and ERK1/2, and this effect can also be reversed by treatment with EGF to a certain extent.

    Conclusion: CAV-2 can inhibit the phosphorylation of ERK1/2 by antagonizing EGFR phosphorylation and downregulating the expression of p-EGFR, thereby inhibiting VM and the malignant biological behavior of BC. In the process of regulating EGFR phosphorylation by CAV-2, p-EGFR can induce CAV-2 ubiquitination, that is, CAV-2 regulates the ERK signaling pathway through an EGFR feedback loop and influences the angiogenic mimicry and malignant biological behavior of BC.

  • Article
    Jun Xue, Chien-Wei Chen, Po-Hua Su, Pang-Yen Chen
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5929-5938. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.567
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    Introduction: Whole transcriptome analysis found substantial variations in butyrophilin-like 9 (BTNL9) expression between osteosarcoma (OS) and healthy bone tissues. The biological involvement of BTNL9 in the physiological regulation of OS cells and its underlying mechanism were investigated herein.

    Methods: Cell proliferation, colony formation, apoptosis, migration, invasion, and angiogenesis were evaluated to determine the influence of BTNL9 on the malignant phenotypes of OS cells. Given that the STRING (https://string-db.org/) database analysis indicated a potential association between BTNL9 and ras homolog family member B (RHOB), co-immunoprecipitation (Co-IP) was performed to confirm their relationship. To verify the role of RHOB, it was knocked down, and the impacts on the above aspects, as well as the phosphatidylinositol-3-kinase/AKT serine/threonine kinase (PI3K/Akt) signaling pathway, were evaluated.

    Results: BTNL9 plays a negative role in OS cells, manifested as inhibition of cell proliferation, colony formation, migration, invasion and angiogenesis and promotion of apoptosis. Co-IP results demonstrated a direct association between BTNL9 and RHOB. RHOB knockdown partially reversed the regulatory effect of BTNL9 on the malignant phenotypes of OS cells and the PI3K/Akt signaling pathway.

    Conclusion: This study highlights that BTNL9 acts as a tumor suppressor in OS and alleviates several malignant phenotypes. BTNL9 collaborates with the recognized tumor suppressor RHOB to block the PI3K/Akt signaling pathway, which is one of the regulatory mechanisms.

  • Article
    Mingtao Tan, Wenxuan Sun, Lijun Xie
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5939-5944. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.568
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    Background: Open fracture of the tibia and fibula is a common fracture. Most patients with this fracture need surgical treatment, but the optimal anesthesia scheme remains inconclusive. Epidural anesthesia with nalbuphine and ropivacaine is used for intraoperative analgesia, but its efficacy in patients undergoing surgery for open fracture of the tibia and fibula remains unclear. This study aims to explore the application of nalbuphine combined with ropivacaine for epidural anesthesia in such patients.

    Methods: The data of 172 patients with open fractures of the tibia and fibula admitted to Zibo Central Hospital from February 2020 to February 2021 were selected for retrospective analysis. Among them, 9 patients failed in anesthesia, and the remaining patients were divided into the experimental group (EG, n = 79) and the control group (CG, n = 84) according to anesthetic drug. All patients underwent internal fracture fixation, patients of the CG were given 0.2 μg/mL of sufentanil and 0.2% of ropivacaine for epidural anesthesia, while patients of the EG received 0.2 mg/mL of nalbuphine and 0.2% of ropivacaine. Hemodynamic indices of patients in both groups were recorded before anesthesia (T1), 15 min after anesthesia (T2), 30 min after anesthesia (T3) and 24 hours (h) after surgery (T4) to compare postoperative pain score, the levels of inflammatory factors and immune function indexes.

    Results: There was no intergroup difference in the hemodynamic indices at T1 and T4 (p > 0.05). While the heart rate and mean arterial pressure in both groups showed a downward trend at T2 and T3, the EG was notably higher than the CG (p < 0.001). The EG showed significantly lower serum levels of Interleukin-10 (IL-10) and tumor necrosis factor-α (TNF-α) than the CG at 3 h, 6 h and 12 h after surgery (p < 0.001). There were significant differences in postoperative pain scores between the two groups at 3 h, 6 h and 12 h after surgery (p < 0.001). Higher levels of cluster of differentiation 4 (CD4+) and CD4+/CD8+ existed in the EG than the CG at the end of surgery, 24 h after surgery and 48 h after surgery (p < 0.001). The severity of adverse events in both groups was grade 1, with no difference in the incidence of nausea and vomiting and urinary retention between the two groups (p > 0.05). However, the incidence of pruritus in the EG was significantly lower than in the CG (p < 0.05).

    Conclusion: Nalbuphine combined with ropivacaine for epidural anesthesia stabilized the hemodynamic indices of patients undergoing surgery for open fracture of the tibia and fibula, improved the levels of inflammatory factors after surgery, exerted a small effect on immune cells, and reduced pain degrees of patients, which is worthy of clinical promotion.

  • Article
    Linchun Wen, Lu Xu, Beibei Cai, Zihao Liu, Lixue Wang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5945-5954. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.569
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    Background: Studies have shown that nucleotide-binding oligomerization domain (NOD)-like receptors family CARD domain containing 5 (NLRC5) is involved in promoting the transformation and invasion of malignant cancer cells. However, the effects and mechanism of action of NLRC5 in lung adenocarcinoma have yet to be elucidated. The aim of this study was therefore to investigate the effect of NLRC5 expression on the invasion and metastasis of lung adenocarcinoma (LUAD) cells and its mechanisms.

    Methods: Tumor and tumor-adjacent normal tissues were acquired from LUAD patients admitted to the affiliated Suqian hospital of Xuzhou medical university from June 2017 to December 2019. A549 and H292 cells were transfected with negative siRNA (si-NC) and NLRC5 siRNA (si-NLRC5), respectively. The phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) pathway agonist 740Y-P was added to cells in the si-NLRC5 group, while cells without any treatment were used as controls. NLRC5 expression was evaluated by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and Western blot. In addition, the techniques of cell colony formation, Transwell assay and scratch tests were employed to assess cell viability, invasion, and migration, respectively. The expressions of genes related to epithelial-mesenchymal transition (EMT) and the PI3K/AKT pathway was also evaluated by Western blot.

    Results: NLRC5 expression in LUAD cell lines and tissues was markedly increased compared to normal human bronchial epithelial (HBE) cells and tumor-adjacent normal tissues (p < 0.05). Knockdown of NLRC5 expression significantly reduced cell viability, invasion and migration, as well as reducing EMT processes and inhibiting activation of the PI3K/AKT pathway (p < 0.05). However, treatment with 740Y-P partially reversed the impact of NLRC5 knockdown in LUAD cells (p < 0.05).

    Conclusion: NLRC5 stimulates the invasion and metastasis of LUAD cells through activating the PI3K/AKT pathway.

  • Article
    Orhan DALKILIC
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5955-5959. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.570
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    Background: Type 2 diabetes mellitus is associated with an increased risk of certain infections, such as pneumonia. Timely diagnosis and treatment of infections is crucial in populations with diabetes. The C-reactive protein to lymphocyte ratio (CLR) has been proposed as a novel marker of inflammation. We studied the role of CLR in subjects with diabetes who had pneumonia.

    Methods: A total of 426 subjects with pneumonia and type 2 diabetes mellitus who presented to the pulmonology clinics of Hisar Intercontinental Hospital were enrolled in the study, and 161 patients with diabetes who had no pneumonia were enrolled as controls. Overall, 176 (66.4%) of the diabetic pneumonia group and 101 (62.7%) of the control subjects were men. C-reactive protein (CRP) was analyzed using Enzyme-Linked Immunosorbent Assay (ELISA) immune assay, and lymphocytes were counted by an automatic analyzer. The CLR levels of patients with diabetes comorbid with pneumonia were compared to those diabetics without pneumonia in the present cross-sectional study.

    Results: Median CLR levels of the patients with diabetes comorbid with pneumonia and control subjects were 28.7 (1.1–116)% and 1.9 (0.04–12.2)%, respectively (p < 0.001). The CLR of the participants was significantly and positively correlated with their white blood cell count (r = 0.21, p < 0.001) and serum creatinine levels (r = 0.2, p < 0.001). Receiver operating characteristic (ROC) analysis revealed that CLR (when higher than 5.73%) had 86% sensitivity and 91% specificity in detecting pneumonia in patients with diabetes (AUC (area under the curve): 0.945, p < 0.001, 95% CI (confidence interval): 0.926–0.964). The sensitivity and specificity of CLR was higher than separate values of C-reactive protein (76% sensitivity and 71% specificity when higher than 7.1 mg/L; AUC: 0.813, p < 0.001, 95% CI: 0.774–0.852) and blood leukocyte count (65% sensitivity and 70% specificity when higher than 7.9 k/mm3; AUC: 0.665, p < 0.001, 95% CI: 0.614–0.716).

    Conclusion: These findings suggest that CLR could be a useful marker in the timely diagnosis of pneumonia in populations with diabetes.

  • Article
    Yu Liang, Jianzhong Cao, Qing Hou, Xin Cao, Ningning Yao, Bochen Sun, Peixin Feng, Wenjuan Zhang, Jinliang Niu
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5961-5969. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.571
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    Background: The lactate dehydrogenase (LDH)-to-lymphocyte ratio (LLR) is a simple, non-invasive prognostic marker. We aimed to reveal the prognostic role that LLR plays in limited-stage small cell lung cancer (LS-SCLC).

    Methodology: This study was a retrospective analysis of 497 patients diagnosed with LS-SCLC from 2015 to 2018, pretreated with LDH, and whose absolute lymphocyte counts were recorded. Patients were categorized into two groups based on their LLR scores: high-risk (LLR ≥182) and low-risk (LLR <182). The Kaplan–Meier test was used to plot the survival curve, and the log-rank test helped evaluate the differences in survival between the high- and low-risk groups. Variables associated with overall survival (OS) and progression-free survival (PFS) were assessed using multivariate and univariate analyses. Additionally, the inverse probability of treatment weight (IPTW) and propensity score matching (PSM) methods helped minimize the selection and confounding bias.

    Results: The high LLR group had 151 patients, and the low LLR group had 346 patients. In the statistical analysis, we found that the patients' PFS and OS were significantly decreased in the high LLR group (PFS: 7.85 vs. 10.41 months, p < 0.0001; OS: 13.4 vs. 18.6 months, p < 0.0001). LLR was an independent prognostic factor for OS (hazard ratio [HR]: 0.719; p = 0.004) and PFS (HR: 0.714; p = 0.002). Similar results were obtained after compensating and matching with the IPTW or PSM methods.

    Conclusion: Pre-treatment LLR is a potential marker against PFS and OS in patients with LS-SCLC.

  • Article
    Yanping Yuan, Junqing Wang, Yunlun Li, Zhijuan Zheng
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5971-5981. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.572
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    Background: Piezo1 is a mechanosensitive Ca2+ channel protein that participates in stem cells fate. Salvianolic acid B (Sal-B) is a polyphenolic compound that promotes endothelial differentiation. This study aims to assess the role of Sal-B in facilitating endothelial differentiation by regulating the activity of Piezo1.

    Methods: The system of human-induced pluripotent stem cells (iPSCs) differentiating into endothelial cells (ECs) in vitro was established to study the role and underlying mechanism of Piezo1 and Sal-B in the regulation of endotheliogenesis. Human iPSCs were divided into four groups, including the normal Sal-B shPiezo1, shPiezo1+Sal-B, shNT and shNT+Sal-B. Fura2-AM was used to record intracellular calcium ion changes. The relative mRNA levels of Piezo1, CD144, endothelial Nitric Oxide Synthase (eNOS), mitogen-activated protein kinase (MAPK) kinase (MEK) and extracellular signal-regulated kinase1/2 (Erk1/2) were identified by Quantitative Reverse Transcription-Polymerase Chain Reaction (RT-qPCR) while the protein levels were measured by Western blotting (WB).

    Results: The addition of 50 μM Sal-B significantly increased the differentiation of ECs. WB and RT-qPCR results showed that Sal-B increased the relative levels of eNOS mRNA, CD144 mRNA, CD144 protein and the tube formation in vitro. Sal-B treatment enhanced Piezo1 expression and Yoda1 activated Ca2+ influx, and amplified MEK/Erk1/2 phosphorylation of endothelial cells (ECs) derived from iPSCs (iPSC-ECs). Silencing of Piezo1 impeded the effect of Sal-B on increasing the rate of endotheliogenesis, as well as the MEK/Erk1/2 phosphorylation of iPSC-ECs.

    Conclusions: The results indicate that Sal-B promotes endothelial differentiation of iPSCs through Piezo1 by triggering the internal flow of Ca2+ and thus further amplifying MAPK/Erk1/2 signaling. Here, we found that Sal-B allows the increase in the number and quality of ECs differentiated from iPSCs. These in-vitro studies constitute a starting point for the future use of this compound in regenerative medicine.

  • Article
    Shadman Ahmad, Khalid Amin, Amin Ullah, Hayat Khan, Izzat Ullah, Iftikhar Ahmed, Alamgir Khan, Mohammed S. Almuhayawi, Soad K. Al Jaouni, Samy Selim, Mutasem Saad Almehayawi, Arshad Farid
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5983-5993. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.573
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    Background: Medicinal plants are abundant reservoirs of antimicrobial compounds. Medicinal plants, which are rich sources of many possible medications, are used therapeutically by people in different countries. Justicia adhatoda, a widespread perennial shrub in the tropical region of Southeast Asia, is known for expectorant and antispasmodic activity. In the present study, various extracts of Justicia adhatoda leaves and extract-mediated silver nanoparticles (AgNPs) were assessed for antimicrobial potency against a panel of thirteen multi-drug-resistant bacterial strains and seven fungal strains.

    Methods: Various solvent extracts of Justicia adhatoda leaves and extract-mediated AgNPs were prepared and dried. Following incubation, extracts were obtained through the evaporation of solvent. The extracts were also evaluated for various parameters such as moisture contents, extractive value, phytochemical and Fourier transform infrared (FT-IR) analysis. The AgNPs were characterized for size and structure using X-rays diffraction (XRD) and scanning electron microscopy (SEM). Solvent extracts and extract-mediated AgNPs were assessed for their antimicrobial activities using a well diffusion assay.

    Results: The FT-IR and phytochemical analysis revealed various types of bioactive compounds and functional groups such as tannins, reducing sugars, flavonoids, steroids, polysterol, terpenoids, saponins, glycosides, phenols, anthraquinones, and alkaloids. The nanoparticles synthesized were round and spherical in shape, with a size range of 35–45 nm, as demonstrated by XRD and SEM analysis. All the extracts demonstrated comparable activity against the bacterial strains; however, the methanolic extract proved relatively more potent. Among the panel of multidrug-resistance (MDR) bacterial strains, Staph aureus, Klebsiella, Citrobacter and Shigella proved relatively more sensitive towards all solvent extracts. Likewise, all fungal strains were susceptible towards all the extracts, though Helminthosporium and Trichoderma were relatively more susceptible. The antimicrobial activity of the AgNPs was also assessed alone and in combination with standard drugs.

    Conclusion: Various solvent extracts of Justicia adhatoda contain numerous bioactive compounds with antimicrobial activities. Both solvent extracts and extract-mediated AgNPs hold broad-spectrum antimicrobial potential. However, the AgNPs synthesized in the aqueous extract were relatively more potent against bacterial and fungal strains. Herein, the potential of Justicia adhatoda as an antimicrobial agent is demonstrated.

  • Article
    Linxiang Yu, Haiwen Zhen, Zhong Guan, Junyi Yang, Changhui Ma, Bin Zhang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 5995-6004. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.574
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    Background: Lung squamous cell carcinoma (LUSC) is one of the familiar types of lung cancer, but the molecular mechanism of LUSC progression remains unclear. The aim of this study is to investigate the biological function of tripartite motif-containing protein 27 (TRIM27) on LUSC progression.

    Methods: Cell immunochemistry, quantitative real-time polymerase chain reaction (qRT-PCR), western blot, and cell transfection were used to detect the expression efficiencies of TRIM27, phosphatidylinositol 3-kinase (PI3K), protein kinase B (AKT), and phosphorylated AKT (pAKT) and the regulatory mechanism of these molecules. Cell Counting Kit-8 (CCK-8) assay, Transwell assay, and flow cytometry were used to detect the viability, metastasis, and apoptosis of LUSC cells.

    Results: Our data indicate that TRIM27 is overexpressed in LUSC. Up-regulation of TRIM27 increases the viability and metastasis of LUSC cells and suppresses apoptosis. The PI3K/AKT pathway is highly active in LUSC. TRIM27 knockdown reduces the expression efficiencies of PI3K, AKT, and pAKT. The addition of v-PI3K plasmid or v-AKT plasmid reverses the silencing of the TRIM27-induced inhibitory effect on LUSC cells.

    Conclusion: Our research demonstrates that TRIM27 promotes LUSC cell viability and metastasis by affecting the PI3K/AKT axis. We conclude that the TRIM27-PI3K/AKT axis plays a crucial role in LUSC.

  • Article
    Aziz Sener, Selin Yildiz, Gulce Kiren, Canan Topcuoglu
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6005-6013. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.575
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    Background: Recently, new inflammatory parameters obtained from complete blood count and biochemical measurements have gained importance. They are easy and inexpensive to perform. These parameters have been shown to have relationships with glycemic control and complications in diabetic patients in various studies. We aimed to compare new inflammatory markers [neutrophil to lymphocyte ratio (NLR), red cell distribution width (RDW), monocyte to lymphocyte ratio (MLR), monocyte to high-density lipoprotein ratio (MHR), platelet to lymphocyte ratio (PLR), and mean platelet volume (MPV)] among well-controlled and poorly controlled diabetic patient groups and also between albuminuria groups in Type 2 Diabetes Mellitus (T2DM) patients.

    Methods: We included 415 T2DM patients and analyzed them retrospectively. We divided the patients into well-controlled diabetics (hemoglobin A1C (HbA1c) ≤7%) and poorly-controlled diabetics (HbA1c >7%) to evaluate diabetic control. We also grouped patients in order to evaluate albuminuria as follows: A1: urine albumin-creatinine ratio (UACR) <30 mg/g, A2: 30 mg/g ≤ UACR ≥ 300 mg/g, A3: UACR >300 mg/g. We compared the groups we formed based on diabetic control and albuminuria levels regarding new inflammatory markers and determined the correlation between these markers and HbA1c and UACR levels.

    Results: MHR showed a more remarkable and noticeable difference than other parameters when comparing A2 and A1 albuminuria group patients. Our study found that MLR was positively correlated with UACR measurements but not superior to other parameters that were positively correlated with UACR (p < 0.01). NLR and PLR differed significantly between the albuminuria groups and were positively correlated with UACR measurements (p < 0.01). RDW, on the other hand, did not show statistical differences between the albuminuria groups but was positively correlated with UACR measurements (p > 0.05, p < 0.01 respectively). MPV, neither statistically differed between the albuminuria groups nor showed correlation with UACR measurements (p > 0.05 for each). There was no difference in the results of new inflammatory markers between diabetes control groups, and there was no correlation with HbA1c measurements (p > 0.05 for each).

    Conclusions: These parameters are useful in assessing albuminuria in patients with T2DM, but not in assessing glycemic control.

  • Article
    Yangyang Yu, Rong Liu, Fei Xia, Yahui Yan
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6015-6027. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.576
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    Background: Fluorosis is a health condition caused by excessive fluoride intake, which can harm the body, especially the teeth. In this study, we investigated whether transmembrane protein 16A (TMEM16A) plays a role in tooth damage caused by fluorine and whether calcium supplementation can mitigate this damage in mice with dental fluorosis.

    Methods: Fluorosis in mice was induced by drinking water containing 100 mg/L of sodium fluoride (NaF) and administering different doses of calcium for 10 weeks. The mice were categorized into four groups: the Model group, the Low-Calcium group, the Medium-Calcium group, and the High-Calcium group. We then assessed the effects of calcium supplementation on body weight, fluoride levels in blood, incisors, and bones. We evaluated tooth development-related proteins (bone morphogenetic protein 2 (BMP-2), transformation growth factor beta (TGF-β), nuclear factor kappa B (NF-κB), BCL-2-associated X protein (BAX), and B-cell lymphoma-2 (BCL-2)), and TMEM16A expression using techniques such as quantitative polymerase chain reaction (qPCR), Western blot, hematoxylin and eosin (HE) staining, and immunohistochemistry. We also examined the effect of the TMEM16A inhibitor T16Ainh-A01 on tooth development-related proteins by supplementing it in the calcium administration group. Various doses of NaF (0 mg/L, 50 mg/L, 100 mg/L, 150 mg/L) were given to evaluate the impact of NaF on dental germ development.

    Results: Compared to the model mice, those receiving medium and high calcium supplementation displayed increased body weight (p < 0.01), reduced fluoride levels (p < 0.01), and decreased apoptotic enamel cells (p < 0.01). TMEM16A, NF-κB, and BAX expression levels were higher in the model group but decreased following calcium treatment. Moreover, calcium treatment significantly boosted BMP-2, TGF-β, and BCL-2 expression levels. When combined with the TMEM16A inhibitor T16Ainh-A01, medium-dose calcium further enhanced BMP-2 and TGF-β expression while decreasing NF-κB expression. As NaF concentration increased, a corresponding rise in fluoride content was observed in blood, tooth germs, and bone tissues. Notably, tooth germ development was delayed and slowed in the NaF-treated group compared to the control group, with TMEM16A expression increasing alongside higher NaF concentration and tooth germ development stage.

    Conclusions: High concentrations of NaF lead to elevated TMEM16A expression, activating the calcium-activated chloride channel and causing tooth damage. Calcium supplementation or inhibition of TMEM16A expression shows promise in alleviating tooth damage caused by fluorosis, contributing to dental repair in fluoride-affected mice. These insights offer potential therapeutic strategies for managing dental fluorosis.

  • Article
    Shakeel Ahmed, Kouadio Ibrahime Sinan, Nilofar, Claudio Ferrante, Ozan Emre Eyupoglu, Ouattara Katinan Etienne, Gokhan Zengin
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6029-6039. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.577
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    Background: Mondia whitei, an indigenous African plant, is widely recognised for its medicinal properties. Throughout ancient times, African communities have employed this remedy to address a wide range of health conditions. However, there is a lack of online High-Performance Liquid Chromatography (HPLC) studies available for this plant. The present investigation aimed to address and alleviate this deficiency by providing a comprehensive analysis.

    Methods: Methanol, combination of water and methanol, ethyl acetate, and water extracts of Mondia whitei (M. whitei) leaves were utilized in this study. The phenolic and flavonoids composition of the extracts determined by Folin-Ciocalteu and Aluminum chloride (AlCl3) assays, respectively. The antioxidant potential of all the extracts was assessed through a range of in vitro assays, including FRAP (Ferric reducing antioxidant power), DPPH (2,2-diphenyl-1-picrylhydrazyl), ABTS [2,2′-Azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid)], CUPRAC (cupric reducing antioxidant capacity), PBD (phosphomolybdenum), and MCA (metal chelating ability). Subsequently, the online HPLC-linked system had established to compare these extracts using four distinct antioxidant analysis methods: FRAP, DPPH, ABTS, and CUPRAC. The enzyme inhibitory properties were also investigated against cholinesterases, tyrosinase, α-amylase and α-glucosidase.

    Results: The methanol-water and methanol extracts had the highest phenolic contents and demonstrated superior efficacy as scavenging/reducing agents and enzyme inhibitors, particularly against tyrosinase and cholinesterase (p < 0.05). Through the utilisation of various online HPLC antioxidant techniques, the identification of four molecules exhibiting antioxidant activity was achieved. These compounds were identified as Coumarin, Vanillin, p-OH benzoic acid, and Chlorogenic acid.

    Conclusions: Mondia whitei leaves can be regarded as a valuable source of natural bioactive compounds, with the potential to serve as a material basis for pharmacological effects in nutraceutical and pharmaceutical applications.

  • Article
    Ning Li, Xiao Liu, Ping Liu, Xiaojiao Meng
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6041-6047. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.578
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    Background: Cyclophilin A (CyPA) is one of the important members of cyclophilin family and is well known for its biological function. It mediates macrophages to express matrix metalloproteinase (MMPS) through ERK1/2 and IkB/NF-kB pathways and provides a basis for inflammatory cell infiltration and migration. This study aims to analyze the value of CyPA combined with double low dose technique and echocardiography in the diagnosis of acute myocardial infarction (AMI).

    Methods: The clinical data of 63 patients with AMI admitted to Zibo Central Hospital from March 2021 to March 2022 were selected as the lesion group (LG) for retrospective analysis. In contrast, 57 healthy people who underwent physical examination in Zibo Central Hospital during the same period were selected as the control group (CG). In total, 120 research subjects were included in this study. All patients in both groups received double low dose of coronary CT angiography (CCTA) and echocardiography examination. The enzyme-linked immunosorbent assay (ELIZA) was adopted to detect serum CyPA level, and the receiver operating characteristic (ROC) curve was used to analyze the application value of serum CyPA. The double low dose of CCTA combined with echocardiography was used for the diagnosis of AMI.

    Results: The LG had remarkably lower left ventricular ejection fraction (LVEF) levels and higher serum CyPA, left ventricular end diastolic diameter (LVEDD), and Gensini scores than the CG (p < 0.001). Results of logistic regression analysis showed that serum CyPA, LVEF, Gensini scores, and LVEDD were independent risk factors of AMI. The area under the curve of ROC (AUC) value and Youden indexes of the combined diagnosis of the three were higher than those of each single diagnosis.

    Conclusions: Serum CyPA combined with double low dose technique and echocardiography had higher clinical values in the diagnosis of AMI, which provides the basis for using it for clinical treatment program.

  • Article
    Peng Sun, Ti Fu, Siyu Wu, Xiangyun Wang, Zhaoquan Shi, Guotai Yao, Xiaoli Wang, Xiaolie Wang, Jianghan Chen, Yi Jin
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6049-6059. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.579
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    Background: Chemo-immune therapy is a common treatment for lung cancer (LC) which possesses serious adverse effects and provides only minimal survival gain. There is an urgent need to find novel biomarkers to refine the chemo-immune therapy application in LC treatment. This prospective study was designed to investigate the association of oral mycobiome features (prior or post treatment) with the prognosis of LC patients receiving chemo-immune therapy.

    Methods: The prognosis was based on the Response evaluation criteria in solid tumors (RECIST) 1.1 criteria, and progressive disease (PD), partial response (PR), and stable disease (SD) in 30 cases were confirmed before and after chemo-immune therapy. The oral mycobiome was characterized based on the Internal Transcribed Spacer (ITS) ribosomal RNA V3~V4 region with the Illumina™ platform.

    Results: There were no differences in α-diversity and β-diversity before or after treatment in LC patients with different prognoses. The fungal composition of the oral cavity changed at almost all levels before and after chemo-immune therapy. Especially at the genus level, the relative abundance of Malassezia was severely elevated within the PD group before treatment in comparison with the PD group after treatment (Pre-Treatment (Pre-Tr)-PD vs. Post-Treatment (Post-Tr)-PD), and in the PR group, the result of Malassezia was conversed (Pre-Tr-PR vs. Post-Tr-PR).

    Conclusions: The abundance of genus Malassezia over the course of chemo-immune therapy treatment was associated with prognosis in LC patients. The mechanisms and therapeutic implications of these findings need to be investigated in future studies.

  • Article
    Arzu Keskin-Aktan, Aslı Nur Bahar, Fatma Gizem Sonugür, Saadet Özen Akarca-Dizakar, Kazime Gonca Akbulut
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6061-6072. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.580
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    Background: Fibrosis induced by transforming growth factor-β1 (TGF-β1) activity and the Wnt/β-catenin pathway is a significant hallmark of progressive kidney disease and kidney aging. We aimed to investigate the effects of pharmacological silent mating type information regulation 2 homolog-2 (SIRT2) inhibition on renal functions, histopathological changes, fibrosis, TGF-β1/β-catenin and klotho signaling, and apoptosis in D-galactose (D-Gal)-induced aging model.

    Methods: The study was conducted with three months old male rats divided into four groups: control (Saline solution (0.9%, 0.5 mL/day) was administered subcutaneously (sc) for ten weeks) (n = 6), D-Gal (D-galactose saline solution (150 mg/kg/day) was administered sc for ten weeks) (n = 8), D-Gal+DMSO (D-galactose (150 mg/kg/day) and 4% dimethyl sulfoxide (DMSO) in phosphate-buffered saline (PBS) (10 μL/bw/day) were administered sc for ten weeks) (n = 8), and D-Gal+acylglycerol kinase (AGK)-2 (D-galactose (150 mg/kg/day) and AGK-2 in 4% DMSO-PBS (10 μM/bw/day) was administered sc for ten weeks) (n = 8). The kidney index was calculated, renal function markers (sodium (Na+), creatinine (Cr), blood urea nitrogen (BUN)) in plasma and urine samples were analyzed, and fractional excretion of sodium (FeNa%) was calculated. Glomerular diameter, fibrosis, and basement membrane thickness were analyzed with histopathological methods. TGF-β1 and β-catenin mRNA expression were determined with quantitative real-time polymerase chain reaction (qRT-PCR), klotho protein levels were determined with the enzyme linked immunosorbent assay (ELISA) method, and SIRT2 protein expression was determined with western blot. The immunohistochemical method was employed to determine the immunoreactivities of β-catenin, klotho, SIRT2, and fibronectin. Apoptosis was determined with the terminal deoxynucleotidyl transferase deoxyuridine triphosphate (dUTP) nick end labeling (TUNEL) method.

    Results: AGK-2 and D-galactose co-administration increased kidney index and decreased plasma and urine Na+ and Cr levels, as well as BUN and FeNa% (p < 0.05). AGK-2 improved the histopathological changes induced by D-galactose, reducing fibrosis and basal membrane thickness (p < 0.05). Furthermore, AGK-2 administration decreased TGF-β1, β-catenin, SIRT2, and fibronectin in the kidney (p < 0.05). AGK-2 and D-galactose co-administration increased klotho protein levels in the kidney; however, the increase was not statistically significant in klotho immunoreactivity (p > 0.05). D-galactose induced apoptosis in the kidney (p < 0.05); however, AGK-2 did not significantly mitigate apoptosis (p > 0.05).

    Conclusion: Our findings suggested that pharmacological SIRT2 inhibition could ameliorate alterations in functional, histopathological, and fibrosis protein pathway activities in the kidney that are associated with aging.

  • Article
    Luping Zhang, Qi Wang, Chao Yang, Jinyun Chen, Wenzhi Chen
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6073-6084. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.581
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    Background: High-intensity focused ultrasound ablation (HIFUa) has been shown to rapidly ablate tumour tissues. However, HIFUa alone often fails to stimulate sufficient and long-lasting antitumour immunity. To overcome this limitation, we investigated the use of non-ablation focused ultrasound (na-FUS), which has potent immunomodulatory effects, combined with HIFUa. In this study, we evaluated the potential of this improved HIFUa method, referred to as “na-FUS+HIFUa", along with the addition of gemcitabine (GEM), to enhance antitumour immunity, inhibit tumour development, and prevent metastasis in murine 4T1 tumours.

    Methods: This study was conducted in three stages. First, Ultrasonic images, TTC (2,3,5-triphenyltetrazolium chloride) and HE (hematoxylin-eosin) were used to prove na-FUS does not lead to coagulation necrosis. Then, we verify the efficacy and related immune mechanism of “na-FUS+HIFUa" in treating 4T1 breast cancer BALB/c mice. In unilateral tumor model mouse, the tumor was removed for ribonucleic acid sequencing (RNA-seq) transcriptome sequencing and immunofluorescence. Flow cytometry was performed use tumor and spleen. In bilateral tumor model mice, the treated tumor, untreated tumor, and spleen were removed for flow cytometry to detect infiltrating lymphocytes. Finally, GEM was combined to further elucidates the effect of “na-FUS+HIFUa" compared to HIFUa. Flow cytometry was performed use tumors and spleen. All mice underwent tumor growth monitoring and survival records, as well as counting the number of lung metastases.

    Results: Through RNA-seq analysis, we observed differential mRNA expression related to the immune microenvironment in the tumours. Gene Set Enrichment Analysis (GSEA) revealed a higher number of differentially expressed genes associated with adaptive immune response in the “na-FUS+HIFUa" group compared to the HIFUa group. Heatmap analysis demonstrated increased expression of marker genes associated with immune response following “na-FUS+HIFUa" treatment. Flow cytometry results further confirmed that “na-FUS+HIFUa" induced higher levels of local intratumoural lymphocyte infiltration, including CD4+ and CD8+ T cells, compared to HIFUa treatment alone (p < 0.05). Moreover, the combination of “na-FUS+HIFUa+GEM" significantly enhanced both local and systemic immune responses, leading to increased lymphocyte infiltration in the treated tumours, untreated tumours, and spleen. This combination approach also inhibited the development of lung metastases and improved overall survival.

    Conclusion: We have successfully developed a novel, non-invasive and feasible method for combining na-FUS with HIFUa. The pretreatment of na-FUS improved the efficacy of HIFUa and promoted the infiltration of CD4+T lymphocytes and CD8+T lymphocytes within the tumor. Combining GEM can further enhance the therapeutic effect of “na-FUS+HIFUa".

  • Article
    Qionghui Huang, Xinxin Lu, Zhaohui He, Chao Zeng, Zhenwei Chen, Tiecheng Yang, Lang-Jing Zhu
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6085-6099. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.582
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    Background: Rubicon is highly expressed in aging kidneys, but its role in chronic kidney disease (CKD) has not been investigated. This study aimed to explore the altered expression of Rubicon in the kidneys, tubulointerstitia, and glomeruli of CKD patients. Additionally, this study analyzed glomerular Rubicon expression based on different disease types and patient sex and investigated potential mechanisms of glomerular Rubicon through bioinformatic methods.

    Methods: This study analyzed sequencing and clinical data of 61 kidney samples, 199 glomerulus samples, and 201 tubulointerstitium samples from the Nephroseq database. The aim was to investigate the correlation between Rubicon and the sex-based estimated glomerular filtration rate (eGFR) and the predictive value of Rubicon for CKD. Glomerular Rubicon alternation was validated through immunohistochemical analysis of 56 kidney samples collected from CKD patients, as well as the correlation between Rubicon expression and eGFR. Furthermore, correlation analysis and weighted gene co-expression network analysis were performed to identify genes associated with Rubicon, followed by pathway enrichment analysis. Immune cell types associated with Rubicon were identified via correlation analysis between glomerular Rubicon and immune cell proportion counted through the microenvironment cell populations counter (MCP-counter) algorithm.

    Results: Rubicon expression was found to be upregulated in the kidneys of CKD patients, particularly in the glomeruli. Moreover, baseline Rubicon expression in glomeruli was higher in females than in males. In males, glomerular Rubicon expression was affected by and inversely correlated with the eGFR. A receiver operating characteristic curve analysis revealed that glomerular Rubicon expression might have predictive value for male patients with CKD. Results of immunohistochemistry (IHC) analysis and correlation analysis validated glomerular Rubicon upregulation in kidneys of CKD patients and a negative correlation between glomerular Rubicon and eGFR in males. Further bioinformatic analysis showed that Rubicon expression was associated with immune and inflammatory pathways, particularly those involved in immune cell activation and migration. Rubicon expression was closely correlated with monocyte/macrophage and dendritic cell infiltration scores.

    Conclusions: Rubicon is a protein that is upregulated in the glomeruli of patients with CKD. In males, its expression is negatively correlated with eGFR. Rubicon is closely related to immune and inflammatory pathways, especially those involving monocyte/macrophage and dendritic cell infiltration.

  • Article
    Shenghua Wu, Shuofan Wang, Yongbiao Meng
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6101-6111. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.583
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    Background & aims: Osteoporosis, a widespread bone disorder primarily affecting postmenopausal women, poses a significant public health challenge. Recent research has linked ferroptosis to the development of osteoporosis. This study explored the possible anti-osteoporosis benefits of quercetin. Additionally, the underlying mechanisms were studied, with a special emphasis on the modulation of ferroptosis.

    Method: In this study, ovariectomy (OVX)-induced osteoporotic rats were employed. They were subjected to treatment with Quercetin and Deferoxamine (DFO, a ferroptosis inhibitor) to investigate the therapeutic effects of Quercetin and ferroptosis inhibition on osteoporosis. Various parameters were assessed, including the morphological characteristics of femoral bone tissue and bone quality (femoral bone mineral density and bone mineral salt content). To evaluate the influence of Quercetin on ferroptosis, hepatic iron deposition, apoptosis rates of bone tissue cells and the expression levels of ferroptosis-associated proteins Xc-cystine (xCT), Voltage-Dependent Anion-Selective Channel Protein 2 (VDAC2), transferrin receptor 1 (TFR1), and glutathione peroxidase 4 (GPX4) were quantified.

    Results: In ovariectomized rats, we found a significant decrease in bone mineral density and impaired morphological characteristics of femoral bone tissue, which is typical of osteoporosis (p < 0.05). Notably, quercetin therapy effectively ameliorated these negative effects and increased bone health in the osteoporotic rat model (p < 0.05). The ferroptosis pathway was significantly suppressed by quercetin administration, evidenced by altered expression levels of key ferroptosis-related components such as GPX4 and transferrin receptor 1 (TFR1) (p < 0.05).

    Conclusion: Quercetin suppresses ferroptosis, which contributes to enhanced bone density and bone mineral content in ovariectomized rats. It additionally corresponds to a decrease in bone turnover, indicating that it might address bone-related disorders.

  • Article
    Fatima Zahrae Radi, Noureddine Bencheikh, Mohamed Bouhrim, Omar M. Noman, Ramzi A. Mothana, Mansour N. Ibrahim, Mohamed Bnouham, Touriya Zair
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6113-6130. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.584
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    Background: Aromatic and medicinal plants are a source of biologically active secondary metabolites, such as polyphenols. These substances have several biological properties, such as antioxidant, antimicrobial, and antihyperglycemic activities. This study aims to evaluate the phenolic compound content and antioxidant, antibacterial, antifungal, and antihyperglycemic activities of extracts from two aromatic and medicinal plant species growing wild in the Middle Atlas region of Morocco: Thymus zygis L. and Thymus willdenowii Bois..

    Method: Phytochemical screening was carried out using colorimetric and precipitation reactions. Total polyphenols were determined using the Folin-Ciocalteu reagent, flavonoids by aluminum trichloride (AlCl3), and condensed tannins by vanillin in an acidic medium. Polyphenols were identified by High-Performance Liquid Chromatography (HPLC) and ultraviolet-visible (UV/VIS) detection. Antioxidant activity was assessed by 1,1-diphenyl-2-picrylhydrazyl (DPPH) and Ferric Reducing Antioxidant Power (FRAP) methods. Antimicrobial activity was assessed on six bacterial strains (Escherichia coli (E. coli), Staphylococcus aureus (S. aureus), Acinetobacter baumanii (A. baumanii), Shigella dysenteria (Sh. dysenteria), Salmonelle Typhi (S. Typhi), and Enterobacter cloacae (E. cloacae)) by disk diffusion, Minimum Inhibitory Concentration (MIC), and Minimum Bactericidal Concentration (MBC) tests. The oral glucose tolerance test and the α-amylase inhibition test assessed antihyperglycimiant activity.

    Results: The analysis of the aqueous extracts of these plants revealed the existence of polyphenolic compounds, sterols and triterpenes, saponosides, and polyuronides. Furthermore, the quantification of total polyphenols, flavonoids, and tannins showed that the extracts of Thymus zygis (T. zygis) and Thymus willdenowii (T. willdenowii) are rich in these compounds. The DPPH and FRAP antioxidant activity test revealed that our extracts have antioxidant activities against DPPH with an median inhibitory concentration (IC50) of 6.98 ± 0.03 μg/mL and 11.82 ± 0.30 μg/mL, and against FRAP with IC50 of 5.95 ± 0.03 mg/mL and 11.90 ± 0.03 mg/mL for T. zygis and T. willdenowii, respectively. The area under the curve (AUC) of antihyperglycemic activity was significantly lower (p < 0.01) in rats treated with plant extracts 54.76 g/L/h for T. zygis, and 53.21 g/L/h for T. willdenowii, respectively. The antimicrobial activity assay revealed that only T. zygis extract was active starting at a concentration of 10 mg/mL. Acute toxicity results showed that administering these extracts up to the dose limit (2 g/kg) induced non-toxicity or mortality in mice.

    Conclusions: The results show that T. zygis and T. willdenowii decocts have exceptional antioxidant and antihyperglycemic properties to fight against the harmful elements that can weaken the body.

  • Article
    Chunhe Sha, Xiaodong Liu, Hua Wang, Guohui Liu, Zhiqing Wang, Haitao Liu, Kai Huang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6131-6140. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.585
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    Background: Spinal cord injury is a condition where the spinal cord is damaged by external trauma or other types of injuries. The spinal cord is part of the central nervous system, located within the spine and responsible for transmitting neural signals to control the body's motor and sensory functions. This study aimed to analyze the mechanism and pathway of nuclear factor erythroid-2-related factor 2 (Nrf2) in spinal cord injury (SCI).

    Methods: Twenty SD rats were randomized to sham, model, blank, and intervention groups, with a surgical incision made alone in the sham group and an SCI model established in the other three groups. Rats in the intervention group received intramyelin injections of vectors targeted to increase Nrf2 expression, while those in the blank group were injected with corresponding empty vectors. Morris water maze (MWM) tests were carried out on each group of rats, and the escape latency and swimming speed were recorded. Subsequently, the rats were killed to collect the spinal cord tissue for Hematoxylin-Eosin (HE), toluidine blue and Terminal deoxynucleotidyl transferase dUTP Nick End Labeling (TUNEL) staining, as well as the determination of Nrf2 fluorescence intensity using immunofluorescence. Finally, the levels of oxidative stress, inflammatory factors and Glyceraldehyde-3-Phosphate Dehydrogenase (GAPDH)/Seven in Absentia Homolog 1 (Siah1) pathway proteins were detected.

    Results: The Nrf2 fluorescence intensity was higher in the model, blank and intervention groups compared with the sham group (p < 0.05). In the MWM test, the escape latency of the intervention group was lower compared with the model and blank groups, and was higher compared with the sham group, while the swimming speed of the intervention group was lower versus the sham group (p < 0.05). According to spinal cord tissue staining, the model and blank groups had severe nerve injury and a large number of cells apoptosis, which were significantly ameliorated in the intervention group (p < 0.05). Milder oxidative stress and inflammatory reactions in the intervention group were identified compared with the model and blank groups, which were still higher as compared to the sham group (p < 0.05). Finally, the GAPDH/Siah1 axis was found to be activated in the model and blank groups, but it was inhibited in the intervention group (p < 0.05).

    Conclusion: Upregulation of Nrf2 expression can alleviate the pathological injury of SCI, and its mechanism is related to the inhibition of GAPDH/Siah1 axis.

  • Article
    Xiangyang Mo, Meiling He, Man Chen, Peng Huang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6141-6150. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.586
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    Background: Acute cerebral infarction (ACI) is an acute cerebrovascular disease with high mortality. Intravenous thrombolysis using alteplase is the primarily available treatment method for ACI, but it has limitations. Studies have shown that using tirofiban following alteplase can improve the stroke scale score in ACI patients. Therefore, we aimed to observe the effects of alteplase in combination with tirofiban on the risk factors of thrombosis in ACI.

    Methods: The disease model of ACI was established and divided into control, sham, model, alteplase, and combined (alteplase+tirofiban) groups. Bederson's method was applied to assess the neurobehavioral score of each group before and after treatment. The percentage of cerebral infarction was determined using the 2,3,5-triphenyltetrazolium chloride (TTC) staining method. Hematoxylin-Eosin (HE) staining approach was used to observe the pathological changes in the morphology of brain tissue. Furthermore, enzyme-linked immunosorbent assay (ELISA) was utilized to evaluate the levels of soluble P-selectin (sP-sel), von Willeophilic factor (vWF), endothelin-1 (ET-1), and interleukin-1β (IL-1β). A positive expression rate of intercellular adhesion molecule 1 (ICAM-1) was detected using immunohistochemistry methods.

    Results: There was no neurological impairment in the control and sham groups, and the model group showed no prominent change in neurobehavioral score before and after treatment (p > 0.05). However, in the alteplase and combined groups, the neurobehavioral scores were significantly reduced after treatment than before treatment (p < 0.05). When compared to the control group, the sham group did not exhibit any significant changes in cerebral infarction, while it was increased in the model group. However, compared with the model group, the area of cerebral infarction was found to be reduced in the alteplase and combined groups. Furthermore, normal brain cell morphology was observed in the control and sham groups, whereas it was disorderly arranged in the model group. Moreover, some brain cells in the alteplase group and a small number of brain cells in the combined group were also found to be disorganized. Compared to the control group, the levels of sP-sel and IL-1β were elevated in the sham and model groups. However, these levels were reduced in the alteplase and combined groups (p < 0.05). There was no significant difference in vWF and ET-1 levels, as well as the positive expression rate of ICAM-1, between the control and sham groups. However, they were significantly increased in the model group compared to the sham group. However, the contents of vWF and ET-1, as well as the expression of ICAM-1 in the alteplase and combined groups were decreased compared to the model group (p < 0.05).

    Conclusion: Alteplase in combination with tirofiban in ACI treatment reduced the area of cerebral infarction, down-regulated the expression of thrombosis risk factors, and alleviated arterial endothelial injury and inflammatory response, making it a method of choice rather than alteplase treatment alone.

  • Article
    Lu Yuan, Daohuan Kang, Weizhong Gu, Qing Chen, Zhen Meng, Yunyan Zheng, Bin Lu
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6151-6158. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.587
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    Background: The widespread use of light-emitting diode (LED) lamp has raised concerns about their potential health effects. This study was designed to explore the effects of light-emitting diode (LED) and electrodeless light on the morphology and thickness of guinea pig retinas.

    Methods: Guinea pigs were randomly divided into three groups: normal control, LED lamp, and electrodeless lamp. The guinea pigs in the normal control group were exposed to natural light, while those in the other two groups were exposed to LED light (wavelength: 450–460 nm) and electrodeless light (frequency: 220–200 mHz) for 8 hours daily, respectively. After 5 days, morphological changes of retinal were assessed using hematoxylin-eosin staining. Retinal cell apoptosis was evaluated through TdT-mediated dUTP nick-end labeling (TUNEL) staining. Levels of apoptosis-related factors and heat shock protein 70 (HSP70) in retinal cells were analyzed by Western blot.

    Results: In the normal control group, guinea pigs exhibited distinct inner structures and outer nuclear layers with well-defined boundaries and organized inner and outer retinal segments. However, the retinal structure was unclear in the LED lamp and electrodeless lamp groups, and the inner layer was noticeably thinner. The cell count and thickness in the electrodeless lamp and LED groups differed from those in the normal control group, with the LED group showing fewer cell numbers than the normal control group (p < 0.01). Both LED and electrodeless lamp exposure led to retinal cell death, elevated protein levels of HSP70, Bcl-2-associated X protein (Bax) and Cleaved caspase-3, while diminishing the level of B-cell lymphoma-2 (Bcl-2) (p < 0.01).

    Conclusion: Exposure to electrodeless lamp and LED lamp results in damage to the retinas of guinea pigs.

  • Article
    Ling Li, Jun Zhang, Qiang Hou
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6159-6168. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.588
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    Background: Systemic Lupus Erythematosus (SLE) is a chronic autoimmune disease that affects multiple organs and is characterised by various symptoms such as fatigue, fever, joint pain and swelling, skin rashes, and inflammation of the kidneys or brain. Children with SLE may experience additional symptoms, such as delayed puberty. The main objective of the study is to perform an integrated bioinformatics analysis of the interferon regulatory factor 5 (IRF5) signalling pathway in children with SLE.

    Methods: We obtained two microarray datasets from the Gene Expression Omnibus (GEO) database. The Gene Set Enrichment (GSE17926) dataset, based on the GPL9059 platform, consists of 13 and 37 IRF5 samples. The GSE81622 dataset was based on the GPL10558 platform and consisted of 30 SLE patients and 25 controls. Data normalisation and relevant plots were produced in R software. We visualized the protein-protein interaction (PPI) networks using Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) and performed gene ontology and pathway enrichment analysis using ShinyGO.

    Results: We identified 357 differentially expressed genes (DEGs), of which 190 were downregulated while 167 were upregulated. Using the STRING tool, we constructed a PPI network and identified 10 hub genes (interleukin 1 receptor associated kinase 1 (IRAK1), Cyclic adenosine monophosphate-response element binding protein (CREBBP), signal transducer and activator of transcription 1 (STAT1), myeloid differentiation factor 88 (MyD88), interferon simulated gene 15 (ISG15), signal transducer and activator of transcription 2 (STAT2), interferon beta 1 (IFNB1), tumour necrosis receptor-associated factor 6 (TRAF6), rel-associated protein (RELA), and interferon regulatory factor 3 (IRF3) associated with IRF5 signalling in SLE. We also identified three Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways involved in IRF5 and the hub genes: the Janus Kinase-signal transducer and activator of transcription (JAK-STAT) signalling pathway, the Toll-like receptor signalling pathway, and the cellular differentiation of T helper cells. Furthermore, we found that IRF5 signalling was enriched in pathways such as response to peptidoglycans, muramyl dipeptide and regulation of interferons.

    Conclusions: The dysregulation of the IRF5 signalling pathway can contribute to the development of SLE by promoting inflammation and activating the immune system. This can lead to tissue damage and the production of autoantibodies.

  • Article
    Yi Liu, Xiaobao Qian, Dandan Li, Zhensheng Qin
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6169-6178. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.589
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    Background: Cuproptosis, a recently reported novel form of cell death that is dependent on abnormal Cu (II) accumulation, has been reported to be mediated by the ferredoxin 1 (FDX1) and lipoylation of dihydrolipoamide S-acetyltransferase (DLAT) and dihydrolipoamide S-succinyltransferase (DLST) proteins. Erianin is extracted from traditional herbs and reported as a potential anti-tumor agent. In this study, we aimed to delve to whether erianin induces cuproptosis of colorectal cancer (CRC) cells and to explore the underlying mechanisms.

    Methods: CRC cells were treated with the cuproptosis inducer elesclomol (Ele) and erianin (ERA). Then cell proliferation was measured with a cell counting kit 8 (CCK-8) assay and colony formation. The in vivo growth of CRC cells was measured by a xenograft tumor model. The intracellular levels of Cu, pyruvate acid (PA), and α-ketoglutarate dehydrogenase (α-KG) were measured to determine the relationship between cuproptosis and the tricarboxylic acid (TCA) cycle. The lipoylated proteins levels were measured by western blot. The oligomerization of the proteins was observed using immunofluorescence. Knockdown of FDX1 was conducted using siRNA and then cell growth and cuproptosis were measured.

    Results: Elesclomol suppressed CRC cell growth in vitro and in vivo, induced accumulation of Cu and decreased the production of PA and α-KG, suggesting the induction of cuproptosis. Administration of erianin enhanced the anti-tumor effects of elesclomol. Erianin treatment increased the expression of FDX1 and induced the lipoylation of DLAT and DLST protein, whereas depletion of FDX1 suppressed the erianin-induced CRC cell cuproptosis and protein lipoylation.

    Conclusions: Erianin induced cuproptosis of CRC cells to suppress CRC cell growth. Erianin may increase FDX1 expression and protein lipoylation to induce ferroptosis.

  • Article
    José-Luis Rodríguez, Zoyla-Mirella Clavo, Mariella Ramos-Gonzalez, Max De La Cruz, Luis Inostroza-Ruiz, Wendy Bada-Laura, Olga Palomino, Maria S. Fernández-Alfonso, Raquel Mateos, Luis Goya
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6179-6192. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.590
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    Background: Oxidative stress is a potential cause of cardiovascular pathologies, so the protection of endothelial cells and vascular tissues is essential to avoid such conditions, perhaps with the use of natural compounds rich in phenolic compounds with a proven high antioxidant capacity. The present study was designed to show the cytoprotective capacity of phenolic extracts from the plant Brunfelsia grandiflora, as well as describe its antioxidant defense mechanisms and the expression of some molecular markers involved in cellular protection.

    Methods: Human EA.hy926 cells were exposed to Brunfelsia grandiflora (B. grandiflora) extract (1, 10, 25, 50, 100, and 200 μg/mL) in co-treatment (22 h with 100 μM tert-Butyl hydroperoxide (t-BOOH) and B. grandiflora concentrations) and pre-treatment (18 h of B. grandiflora concentrations and then 200 μM t-BOOH for 4 h). Cell viability, reactive oxygen species (ROS) production, nitric oxide (NO) levels, caspase 3/7 activity, malondialdehyde (MDA) concentration, and reduction in reduced glutathione (GSH) levels, glutathione peroxidase (GPx), and glutathione reductase (GR) activity were measured, and real-time PCR molecular assays superoxide dismutase (SOD2), nuclear factor E2-related factor (NRF2), BCL-2-associated X protein (BAX), and B-cell lymphoma 2 (BCL2) were performed. Data were analyzed via one-way ANOVA followed by Tukey's post hoc test.

    Results: B. grandiflora bark extract, mainly at concentrations of 25, 50, 100, and 200 μg/mL, significantly (p < 0.05) reversed (pre-treatment and co-treatment) the deleterious effects of t-BOOH on EA.hy926 endothelial cells, which were significantly decreased cell viability, GSH activity, and SOD2 and NRF2 expression (p < 0.05); and significantly increased levels of ROS, NO, MDA, caspase-3/7 activity, GPx activity, GR activity, and the BAX/BCL2 ratio (p < 0.05).

    Conclusions: B. grandiflora extract was able to reduce the deleterious effects of t-BOOH on EA.hy926 endothelial cells, which may indicate its potential phytotherapeutic benefit against cytotoxic damage caused by chemical agents.

  • Article
    Ping Xing, Hua-Rong Luo, Cheng Xu, Yong Liang, Feilin Cao
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6193-6203. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.591
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    Background: The immunological relationship and the molecular mechanisms underlying the co-occurrence of papillary thyroid carcinoma (PTC) and Hashimoto's thyroiditis (HT) are yet to be known. This study aimed to explore the immunological and clinical association between papillary thyroid carcinoma (PTC) and Hashimoto's thyroiditis (HT).

    Method: 680 PTC patients were evaluated for clinicopathological factors and serum thyroid-function level. Among 680 PTC patients, 129 PTC patients with or without HT were investigated the immune-related cytokine levels such as Interleukin, tumor necrosis factor and immune cell percentages (cluster of differentiation (CD)4, CD8, CD3) in the serum using the immunofluorescence-flow cytometry technique. We explored the characteristics of tumor immune microenvironment and the levels of immune cell infiltration of CD4, CD8, CD3, CD20 in tissues using Immunohistochemistry.

    Results: It was found that the patients with co-existing PTC and HT (PTC+HT) had lower levels of Interleukin-2 (IL-2) (p = 0.023) and Interferon-γ (IFN-γ) (p = 0.003) than those with PTC alone. There was no significant difference in the proportions of CD4, CD8, and CD3 cells in the two groups (p > 0.05). Additionally, the proportions of CD4 and CD3 cells were significantly lower in the PTC+HT group than those with HT alone (p < 0.05). Furthermore, the PTC+HT group had lower lymph node metastasis rates (p = 0.001) and showed no significant difference in the stratification of postoperative recurrence risk (p < 0.05) than those with PTC alone. Moreover, it was observed that the patients with HT had higher levels of CD4+ and CD3+ cell infiltration in their intact thyroid tissues than those with PTC+HT. The level of CD8+ cells was significantly higher in the PTC+HT group of patients than those with PTC alone. However, the level of CD4+ cells was found to be lower in the PTC+HT group of patients. The PTC+HT group of patients with negative lymph node status had more prominent CD4+ T cells intratumoral infiltration than those with positive lymph node metastasis (LNM) (p < 0.05). Moreover, PTC+HT and PTC patients with LNM+ status had lower CD8+ T cell infiltration levels than those with LNM-status (p < 0.05).

    Conclusion: HT does not attenuate the aggressive factor of lymph node metastasis in PTC patients. Patients with PTC+HT may have different anti-tumor immune mechanisms impacting lymph node metastasis and lower levels of CD4+Th1 cellular immunity than patients with PTC.

  • Article
    Yan Zhang, Jing Cui, Ze Wang, Ying Zhu, Ningkun Zhang, Yu Chen
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6205-6216. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.592
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    Background: Apelin, an emerging endogenous peptide ligand, has been associated with insulin resistance, but the precise mechanism of its influence remains unclear. This study aims to determine the effect of Apelin on insulin resistance and glucose metabolism in 3T3-L1 adipocytes.

    Methods: Insulin resistance was induced in 3T3-L1 adipocytes through tumor necrosis factor-α treatment, followed by infection with Apelin-containing lentivirus particles. Apelin gene expression was confirmed by fluorescence microscope, reverse transcription polymerase chain reaction (RT-PCR), and western blot analysis. Glucose utilization was measured by a glucose uptake assay. The expressions of protein kinase B (AKt) and endothelial nitric oxide synthase (eNOS) were detected by RT-PCR and western blot. The RT-PCR evaluated the inflammatory cytokines interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α), and anti-inflammatory factors adiponectin.

    Results: The translocation of glucose transporter 4 (GLUT4) was detected by flow cytometry. Apelin overexpression in insulin-resistant 3T3-L1 adipocytes resulted in a 35% (p < 0.01) increase in glucose uptake when stimulated with insulin. Apelin overexpression facilitated the translocation of GLUT4 from the cytoplasm to the plasma membrane and elevated AKt and eNOS levels. Furthermore, Apelin overexpression led to an up-regulation of adiponectin mRNA and reduced TNF-α and IL-6 mRNA expressions.

    Conclusions: This study shows that Apelin stimulates GLUT4 translocation and improves glucose utilization through the AKt and eNOS pathways. Apelin can potentially enhance the mechanism of insulin resistance by mediating the inflammatory response in 3T3-L1 adipocytes.

  • Article
    Lu Sun, Beige Zong, Fei Li, Yina Qiao, Ting Jin, Yihua Wang, Yingzi Zhang, Yudi Jin, Fanli Qu, Shengchun Liu
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6217-6229. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.593
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    Background: Tamoxifen is an adjuvant endocrine drug used as first-line therapy for treating breast cancer (BC) with positive estrogen receptor (ER) status. However, its effectiveness is limited by the emergence of tamoxifen resistance (TR) caused by changes in autophagy. The aim of this research was to evaluate the role of the cyclic adenosine monophosphate (cAMP)-dependent protein kinase inhibitor-β (PKIB) in the emergence of TR in BC, together with its involvement in autophagy.

    Methods: PKIB expression was assessed in tamoxifen-sensitive and -resistant BC cell lines and clinical specimens using RT-qPCR, Western blotting, and immunohistochemistry. Overexpression and knockdown of PKIB were employed to evaluate the effects on autophagy and on the response to tamoxifen in vitro. In addition, the mechanism of PKIB-mediated TR was explored using immunofluorescence and Western blot analysis. Furthermore, the activation of cAMP-responsive element-binding protein (CREB)/Autophagy related gene 7 (ATG7) signaling after knockdown of PKIB was evaluated in MCF7 and T47D cells.

    Results: PKIB was found to be upregulated in tamoxifen-sensitive cell lines (MCF7 and T47D) and in primary tumor tissues. Knockdown of PKIB promoted TR, both in vivo and in vitro, and markedly increased autophagy levels in tamoxifen-sensitive cells. In contrast, the overexpression of PKIB suppressed autophagy and restored tamoxifen sensitivity in TR cells. Furthermore, the protein levels of ATG7 and pCREB were upregulated in MCF7/si2-PKIB and T47D/si2-PKIB cells. Lastly, Kaplan-Meier survival analysis showed that upregulation of PKIB was associated with favourable prognosis of BC patients.

    Conclusions: This study demonstrated that PKIB inhibits autophagy via the CREB/ATG7 axis, thereby leading to TR in BC.

  • Article
    Hayat Bilal, Mubarak Ali Khan, Syed Wadood Ali Shah, Mehreen Ghias, Haya Hussain, Huma Ali, Mehboob Ur Rahman, Muhammad Zahoor, Riaz Ullah, Amal Alotaibi
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6231-6245. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.594
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    Background: Helicteres isora is a highly valued medicinal plant whose fruits have been traditionally used throughout Asia, Africa, and other parts of the world in treating a variety of diseases, including diabetes mellitus and memory-related health problems. This study aimed to evaluate the in-vitro/in-vivo toxicity and anti-amnesiac profile of Helicteres isora (H. isora) extract.

    Methods: Initially, experimental animals were subjected to acute, sub-chronic, and chronic toxicity evaluation to establish the safety profile of the H. isora-crude (Hi-Crd) extract of selected plant fruit extract. In the first phase, oral administrations of the graded doses of 500, 1000, and 1500 mg/kg body weight (b.w.) were administered to experimental rats for 14 days. In the second phase, higher doses (3000, 4000, and 5000 mg/kg b.w.) were administered to assess acute toxicity. On the 28th day, blood was obtained and subjected to serum biochemical analysis and lipid profiling to evaluate chronic toxicity. To further evaluate the toxicity in deep, histopathology studies of the kidney, liver, and pancreas were performed. Acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) inhibitory activities of the extract were also performed, and IC50 values were calculated. The scopolamine-induced amnesic mice were utilized in behavioral studies (Y-maze and Novel Object Recognition Test (NORT)), where doses of 75 and 150 mg/kg b.w. of Hi-Crd were administered to experimental animals.

    Results: In the acute toxicity test, no symptoms of morbidity or toxicity were observed. Hi-Crd increased the relative liver and kidney weight at the end of the sub-chronic toxicity test when given at a dose of 4000 mg/kg b.w.. During the chronic toxicity test, the extract showed substantial effects on several biochemical parameters. Although a rise in the lipid profile was noted, the overall effect on other biochemical parameters was negligible. At day 28, there was no significant effect on the liver enzyme markers, and thus the extract was considered to be safe at tested doses. Hi-Crd has a safe profile in terms of uric acid, bilirubin, and blood urea while there was a slight rise in the serum creatinine level. The H.isora-chloroform fraction (Hi-Chl) showed maximum activity with IC50 values of 54.29 ± 1.17 μg/mL against AChE and 105.66 ± 2.91 μg/mL against BuChE. In behavioral studies, the chloroform extract considerably enhanced spontaneous alteration performance as measured through the Y-maze test (p < 0.001), at tested doses. Its counterpart, the ethyl acetate fraction, also showed noteworthy outcomes with good significance levels; p < 0.01 and p < 0.001 at 75 and 150 mg/kg b.w., respectively. In addition, the chloroform fraction significantly improved the discrimination index (DI) in experimental mice to 67.08% as compared to the amnesic group (29.87%) mice results.

    Conclusions: Helicteres isora Linn was found to be well tolerated during the performed set of tests and could be considered an effective treatment to enhance memory impairment in short- and long-term therapy with an adequate margin of safety. However, the study found that it may have adverse effects to an imperceptible level on kidney and liver functions. Therefore, further studies on other animal models are necessary to confirm its future use as a drug.

  • Article
    Wenjun Pan, Wulong Wang, Guodai Hong, Shengyang Chen, An Luo, Wenbin Gao
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6247-6256. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.595
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    Background: Early diagnosis of lung cancer helps to extend the patient's survival time. Our previous study demonstrated that tumor-derived exosomal microRNA-141 (miR-141) promotes lung cancer angiogenesis and malignant progression by targeting the gene transcript for growth arrest-specific homeobox (GAX). Here, we evaluated miR-141's prognosis value to lung cancer diagnosis.

    Methods: MiR-141 was steady overexpressed in A549 cell by lentivirus vector. The GAX was expressed in miR-141+ cells by pcDNA 3.1 (+) plasmid. The protein expression levels were assessed using western blot analysis. The functional significance of miR-141 and GAX was further investigated in vivo models. MiR-141 level in cells and serum were detected by real-time quantitative polymerase chain reaction (RT-qPCR) assay.

    Results: The overexpression of miR-141 has been shown to enhance cellular proliferation and migration. Overexpression of miR-141 accompanied with rescue overexpression of GAX in cells could restore cell proliferation and migration to control. We induced constitutive overexpression of miR-141 in the A549 cell line (A549-miR-141) and found that the tumor growth rate of A549-miR-141 was faster than that of the control group in a nude xenograft mouse model. The tumors in the A549-miR-141 group were significantly enriched in blood vessels relative to the control group. Correlation of clinical information and miR-141 expression levels within the serum exosomes from 98 lung cancer patients showed that serum miR-141 levels were unrelated to gender, age, operation, chemotherapy, or radiotherapy. However, the mRNA level of miR-141 was correlated with clinical stage and lymph node metastasis (p < 0.001). In addition, there was strong correlation between overall survival and the level of miR-141, with longer overall survival in patients with lower miR-141 expression level.

    Conclusions: Our study suggests that exosomal miR-141 derived from lung cancer cells promotes angiogenesis and facilitates malignant progression. The expression level of serum exosome miR-141 exhibits a significant correlation with the clinical stage and lymph node metastasis, thereby offering a novel prognostic marker and potential therapeutic target for lung cancer management.

  • Article
    Chunli Ma, Lili Yang, Tianying Xu, Lei Yang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6257-6267. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.596
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    Background: Ischemic is one of the major types of strokes and is regarded as the third important cause of mortality and the leading factor of permanent disability. Especially, the ischemia-reperfusion (I/R) injury always induces traumatic tissue damage and irreversible loss of neurons, making it crucial to explore novel therapeutic strategies for treating I/R injury. This study investigated the function of microglial exosomes in I/R injury and explored the association of underlying mechanisms with Notch1 and signal transducer and activator of transcription 3 (STAT3)/nuclear factor kappa B (NF-κB) signaling pathway.

    Methods: The immortalized murine microglial cell line BV2 was purchased and the oxygen-glucose deprivation (OGD)-treated neurons were established in this study. In addition, real-time reverse transcriptase-polymerase chain reaction (RT-qPCR), western blot, transmission electron microscope, particle size analysis, cell counting kit-8 (CCK-8), the terminal deoxynucleotidyl transferase (TdT) dUTP nick-end labeling (TUNEL) assay, and lactate dehydrogenase (LDH) assay were utilized for mechanistic studies.

    Results: BV2 cells were successfully polarized (M2) and exosomes were successfully isolated in this study. OGD treatment resulted in increased apoptosis and decreased viability of neuronal cells (p < 0.05), which could be reversed by exosomes of polarized BV2 cells (p < 0.05). Decreased Notch1 expression was associated with the attenuation of OGD-induced neuronal apoptosis (p < 0.05). Decreased Notch1 expression could activate STAT3/NF-κB pathway, and activated STAT3/NF-κB pathway was associated with less cell apoptosis and more cell viability (p < 0.05).

    Conclusion: The present study demonstrates that the down-regulated Notch1 in exosomes secreted by M2 microglia attenuates the ischemia-reperfusion brain injury by targeting the downstream STAT3/NF-κB pathway.

  • Article
    Zhihui Ma, Rongjun Cui, Fusheng Zhao, Pei Liu, Jinli Ji, Xingyun Wang, Zheyao Song, Ying Jiang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6269-6281. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.597
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    Background: Dysregulation of ADAMTS9-AS1 is engaged in the carcinogenesis and metastasis of different cancers. However, whether ADAMTS9-AS1 dysregulation may have an impact on the prognosis and tumor lymphocytes infiltration in kidney cancer remains unclear.

    Methods: Data from the cancer genome atlas (TCGA) were used to assess ADAMTS9-AS1 expression pattern in different cancers. Survival rates of patients with different tumors (with a focus on kidney renal clear cell carcinoma (KIRC) and kidney renal papillary cell carcinoma (KIRP)), and ADAMTS9-AS1 expression level were subjected to Cox regression and Kaplan-Meier curve analysis. Associations of ADAMTS9-AS1 expression with clinicopathological parameters were determined by Wilcoxon singed-rank test. ADAMTS9-AS1 expression prognostic potential was evaluated with univariable and multivariable Cox analyses. Gene set enrichment analysis (GSEA) and pathway enrichment analysis were used to determine pathways and gene sets associated with ADAMTS9-AS1 expression level. Tumor microenvironment and immune cell infiltration were assessed using Estimate and cell-type identification by estimating relative subsets of RNA transcript (CIBERSORT) methods.

    Results: ADAMTS9-AS1 was downregulated in KIRC, and was associated with a poor survival. Low ADAMTS9-AS1 expression level in KIRC was associated with more advanced clinical stages, larger tumor sizes, more metastases and higher histological grades. In contrast, ADAMTS9-AS1 was overexpressed in KIRP which was associated to better survival rate. Low ADAMTS9-AS1 level in KIRP was associated with more advanced clinical stages, larger tumor sizes, and more lymph node metastases and distant metastases. GSEA analysis showed that histidine metabolism and autophagy pathway regulation were enriched in ADAMTS9-AS1 high expression group of KIRC and KIRP. In addition, regulatory T cell (Treg) markers, including Forkhead Box O3 (FOXO3) and transforming growth factor-β1 (TGFβ1), were negatively correlated with ADAMTS9-AS1 level. Low ADAMTS9-AS1 expression was associated with macrophage and Treg enrichment in KIRC and KIRP.

    Conclusion: ADAMTS9-AS1 may be used as a potential biomarker to indicate the infiltration of immunosuppressive cells in KIRC and KIRP.

  • Article
    Tongjun Liu, Zhiwei Sun, Rui Yang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6283-6294. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.598
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    Objective: The macrophage polarization is predominantly involved in the progression of peri-implantitis. However, the matrine has been shown to potentially inhibit M1 macrophage polarization. Therefore, this study aimed to explore the in vivo effects of matrine on M1 macrophage polarization in peri-implantitis.

    Methods: RAW264.7 cells were treated with lipopolysaccharide (LPS) and then incubated with different concentrations (0, 20, 50 μM) of matrine. The cells were transfected with si-matrix metallopeptidase 9 (si-MMP9) and si-nuclear factor-kappa B (si-NF-κB). Furthermore, Sprague-Dawley (SD) rats were used to establish peri-implantitis models by surgically implanting titanium alloy screws into the left upper first molar. The rats were divided into four groups, each containing five rates: a control group, a model group, a saline group, and a matrine group. The rats in the matrine group were administered matrine (100 mg/kg) every day for four weeks. However, in the saline group, the rats were injected with an equivalent dose of physiological saline. After this treatment, the rats were anesthetized with 4% to 5% isoflurane, and their maxillae were dissected. The levels of matrix metallopeptidase 9 (MMP9) and nuclear factor-kappa B (NF-κB), both at the mRNA and protein levels, were determined in cell lines as well as in rat models using quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting methods. Furthermore, the level of inflammatory factors was also evaluated. The cellular viability and apoptosis were analyzed using cell counting kit-8 (CCK-8) assay and flow cytometry. Additionally, macrophage polarization was observed by immunofluorescence, and the inflammatory factor infiltration was assessed using histopathological examination.

    Results: It was observed that LPS successfully induced M1 macrophages (p < 0.001), while matrine significantly reduced the activity of M1 macrophages (p < 0.001). Moreover, matrine also induced apoptosis (p < 0.001) and promoted the polarization of M2 macrophages (p < 0.001), thereby inhibiting the expression of inflammatory factors tumor necrosis factor (TNF) (p < 0.001), interleukin (IL)-1β (p < 0.001), IL-6 (p < 0.001), and increasing the levels of IL-10 (p < 0.001) and transforming growth factor (TGF) (p < 0.001) by decreasing the levels of MMP9 (p < 0.01) and NF-κB (p < 0.01). Additionally, the rat model of peri-implantitis was successfully constructed. Matrine treatment significantly inhibited inflammatory infiltration (p < 0.01) and successfully inhibited the polarization of M1 macrophages (p < 0.001) by regulating the MMP9/NF-κB signaling pathway.

    Conclusion: Matrine effectively decreases the inflammatory infiltration in peri-implantitis by regulating the MMP9/NF-κB signaling pathway, inhibiting M1 macrophage polarization and inducing M2 phenotype.

  • Article
    Rukayat Abiola Abdulsalam, Jamiu Olaseni Aribisala, Saheed Sabiu
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6295-6318. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.599
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    Background: The relevance of aminoglycosides has decreased in recent times due to the prevalence of their modifying enzymes, such as aminoglycoside N-acetyltransferases (AAC) [e.g., AAC (6')-ly], aminoglycoside O-nucleotidyltransferases (ANT) [e.g., ANT (2”)] and aminoglycoside O-phosphotransferases (APH) [e.g., APH (3')-IIa)], which inactivate them. However, the recent development of plazomicin, which resists the modifying actions of some subclasses of AAC, ANT, and APH, has sparked renewed interest in aminoglycosides. This study aims to identify computationally potent and less toxic modulators of AAC (6')-ly, ANT (2”), and APH (3')-IIa from Gnaphalium affine (G. affine) that could be used in combinatory therapy with aminoglycosides.

    Methods: Potent and less toxic modulators of the three enzymes were identified using molecular docking, quantitative structure-activity relationship (QSAR) study, pharmacokinetics properties, and a 100 ns molecular dynamic simulation for stability and energy refinement.

    Results: The results show that plazomicin is a potent aminoglycoside-like antibiotic (probability of being active (Pa): 0.803) with significant binding free energies of –51.01, –53.01, and –81.86 kcal/mol against ANT (2”), APH (3')-IIa, and AAC (6')-ly, respectively. The high binding free energy of plazomicin against the three enzymes correlated with its predicted high bioactivity against them. Interestingly, bioprospecting the metabolites of G. affine against the three enzymes revealed that the top-ranked compounds in each case formed thermodynamically stable complexes (root mean square deviation (RMSD): 1.43–2.37 Å) with pharmacokinetic properties that potentiated them as less toxic enzyme inhibitors than plazomicin.

    Conclusions: Overall, the thermodynamic binding free energy and the QSAR analyses identified alpha-cedrol as a putative inhibitor of the three enzymes. The binding free energy of alpha-cedrol ranged from –46.26 to –53.27 kcal/mol, which is comparable to that of plazomicin against ANT (2”) and APH (3')-IIa. Further validation of alpha-cedrol as a novel inhibitor of the investigated enzymes is highly recommended. Both in vitro and in vivo studies should be conducted to confirm this efficacy.

  • Article
    Xin’na Li, Xiangzhi Zhang, Haiyan Zhang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6319-6327. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.600
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    Background: Papillary thyroid carcinoma (PTC), being one of the malignant tumors, ranks as the most common thyroid tumor. Sorafenib (SFB), Zinc finger E-box binding homeobox1 (ZEB1), and the phosphatidylinositol-3-kinase and protein kinase B (PI3K/AKT) signaling pathway all play critical roles in cancer development and are involved in various biological processes within cancer cells. However, the precise mechanism of PTC remains unclear. Therefore, the objective of this study was to investigate the regulatory role of SFB/ZEB1 in the process of epithelial-mesenchymal transition (EMT) in PTC, with a focus on the PI3K/AKT signaling pathway.

    Methods: Quantitative Real-time polymerase chain reaction (qRT-PCR) and Western blot analysis were employed to examine the expression levels of ZEB1, phosphatidylinositol-3-kinase (PI3K), protein kinase B (AKT), pAKT, E-Cadherin (E-Ca), N-cadherin (N-Ca), and Vimentin, while regulatory mechanisms controlling these molecules were also investigated. Additionally, PTC cell viability and metastasis were assessed using scratch assays, Transwell assays, and 3-45-dimethylthiahiazo-z-y1-35-di-phenytetrazoliumromide (MTT) assays.

    Results: We observed that SFB effectively reduced the viability of PTC cells in a concentration-dependent manner, with an inhibitory concentration 50% (IC50) of 9.56 μmol/L. Furthermore, SFB demonstrated significant inhibitory effects on the metastasis of PTC cells when compared to the control group (p < 0.05). ZEB1 exhibited an upregulation in PTC cells, and the overexpression of ZEB1 led to a noteworthy enhancement in the viability, metastatic potential, and induction of epithelial-mesenchymal transition (EMT) in PTC cells compared to the overexpression-negative control (v-NC) group (p < 0.05). Notably, the upregulation of ZEB1 was associated with elevated protein levels of PI3K, AKT, and pAKT. However, SFB effectively reduced the expression level of ZEB1 and suppressed the promoting effects on PTC induced by ZEB1 overexpression (p < 0.05).

    Conclusions: In this study, our findings suggest that ZEB1 enhances the viability, metastatic potential, and EMT process of PTC cells by modulating the PI3K/AKT signaling pathway. SFB is effective in impeding the progression of PTC by inhibiting the biological effects of ZEB1 in PTC.

  • Article
    Tan Wang, Kang-lun Jiang, Da-quan Wu, Lei Zhou, Na Shen, Ben-yu Nan, Xin-sheng Huang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6329-6340. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.601
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    Background: Cisplatin (Cis) is a common chemotherapy medication which is widely applied to treat various types of tumors. But ototoxicity caused by Cis is the major concern of its application. Salidroside (Sal) is the primary active component of Rhodiola rosea, which has multiple pharmacological effects, including anti-inflammatory and antioxidant. This study was performed to evaluate the protective effect and understand the potential mechanisms of Sal on Cis-induced ototoxicity.

    Methods: Neonatal cochlear explants and HEI-OC1 cells were adopted to construct the Cis damage model, and treated with Sal. The effects of Sal on HEI-OC1 cells viability were detected using Cell Counting Kit-8 (CCK-8) and Adenosine Triphosphate (ATP) assay. The mitochondrial membrane potentials (MMPs) and the cellular and mitochondrial reactive oxygen species (ROS) levels in HEI-OC1 cells were evaluated. The hair cells in neonatal cochlear explants were labeled using immunofluorescence. The protein related to phosphatase and tensin homolog deleted on chromosome ten (PTEN) induced putative kinase 1 (PINK1)/Parkin pathway was detected by western blot. The neonatal cochlear explants and HEI-OC1 cells were incubated with mitochondrial division inhibitor 1 (Mdivi-1). The changes in cell viability and ROS level in HEI-OC1 cells and the number of hair cells in neonatal cochlear explants were examined.

    Results: Sal increased the cell viability and MMPs, and inhibited the cellular and mitochondrial ROS levels in Cis-induced HEI-OC1 cells. Besides, Sal increased the number of hair cells, and enhanced the PINK1/Parkin pathway-mediated mitophagy in neonatal cochlear explants induced by Cis. Mdivi-1 reversed the protective effect of Sal on ototoxicity caused by Cis.

    Conclusions: Sal promotes mitochondrial autophagy by activating the PINK1/Parkin pathway. It protects hair cells from Cis-induced ototoxicity by scavenging ROS accumulation.

  • Article
    Chunmiao Chen, Pingping Liu, Guozhe Zhang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6341-6349. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.602
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    Background: Diabetic nephropathy (DN) is a common complication of diabetes with intricate mechanisms. Our objective was to investigate the potential improvement effects of astragaloside IV (ASV) in a DN cell model and its underlying molecular mechanisms.

    Methods: High glucose (HG) was employed to induce DN cell model in HK-2 cells, a human renal tubular epithelial cell line. In the initial in vitro experiment, aimed at assessing the impact of ASV on the DN cell model, HK-2 cells were subjected to ASV treatment following HG stimulation. The experimental groups were comprised of Control group, HG group, and HG + ASV treatment group. In the second in vitro experiment, five groups were studied, including Control group, HG group, HG + ASV treatment group, HG + ASV + pcDNA group, and HG + ASV + nuclear factor of activated T cells 5 (NFAT5) group, to elucidate the mechanisms underlying ASV's influence on DN progression. To evaluate the levels of NFAT5, reverse transcription-quantitative PCR (RT-qPCR) was employed. Cell proliferation and apoptosis were measured using flow cytometry, Cell Counting Kit-8 (CCK-8), and 5-Ethynyl-2'-deoxyuridine (EdU) assays. Western blot analysis was performed to assess the expression of Collagen Type I (Col. I), α-Smooth Muscle Actin (α-SMA), Fibronectin (FN), Notch1, E-cadherin, Transforming Growth Factor-beta 1 (TGF-β1), NFAT5, Vimentin, Hes1, Jagged1, and Snail.

    Results: ASV treatment effectively alleviated HG-induced cell damage by enhancing the proliferative capacity, inhibiting cell apoptosis, and suppressing epithelial-mesenchymal transition (EMT) and fibrosis in HK-2 cells (p < 0.05 or p < 0.01). Following HG stimulation, NFAT5 level was increased, and the Notch/Snail pathway was activated (p < 0.01). These effects were significantly ameliorated by ASV treatment (p < 0.01). ASV also demonstrated a direct ability to reduce NFAT5 expression (p < 0.01). Moreover, the positive impact of ASV on cell proliferation and its inhibitory effects on cell apoptosis, EMT, and fibrosis were compromised by NFAT5 overexpression in HK-2 cells after HG stimulation (p < 0.01). Additionally, ASV treatment restrained HG-induced activation of the Notch/Snail pathway (p < 0.01), which was counteracted by NFAT5 overexpression (p < 0.01).

    Conclusion: ASV demonstrated the protective effects against cell damage induced by HG in HK-2 cells through downregulating NFAT5 and inactivating the Notch/Snail pathway.

  • Article
    Jing-han Yang, Wen Gao
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6351-6360. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.603
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    Background: Circular RNA (circRNA) plays a crucial role in several cerebral cardiovascular disorders; however, the mechanism of coronary heart disease (CHD) is still not fully understood. Circ_0000003 is one of the circRNAs which affects the cell proliferation or apoptosis and may have a regulatory function in CHD. Thus, the aim of this study is to explore the role of circ_0000003 in CHD and its mechanism.

    Methods: The blood samples of clinical CHD patients (CHD group, n = 20) and healthy volunteers (normal group, n = 20) were collected. The expression level of circ_0000003 in the blood was observed by quantitative real-time polymerase chain reaction (qRT-PCR). To mimic CHD in vitro, H9C2 rat cardiomyocytes were induced with 50 μmol/L H2O2, and the expression levels of circ_0000003 in the resulting cardiomyocytes were recorded. The circ_0000003 overexpression vector (circ_0000003) and negative control (vector), as well as the shRNA circ_0000003 vector (sh-circ_0000003) and shRNA negative control (sh-NC) were transfected into H2O2-induced H9C2 cells. Cell counting kit-8 (CCK-8) assay and propidium iodide (PI)/fluorescein isothiocyanate (FITC)-Annexin V staining were performed to determine the cell proliferation and apoptosis levels, as well as the levels of inflammation-related factors interleukin (IL)-6, IL-1β, tumor necrosis factor (TNF)-α and oxidative stress-related factors malondialdehyde (MDA), superoxide dismutase (SOD), and reactive oxygen species (ROS) in each group. The expression levels of toll-like receptor 4 (TLR4)/nuclear factor kappa-B (NF-κB) p65 pathway-related proteins (TLR4, phosphorylation-nuclear factor kappa-B (p-NF-κB), NF-κB) in each group of cells were observed by western blotting.

    Results: The expression levels of circ_0000003 were significantly reduced in CHD patients and CHD cell models (p < 0.01). When circ_0000003 was overexpressed in the CHD cell model, it greatly promoted cell proliferation and inhibited apoptosis (p < 0.01). On the other hand, the overexpression decreased the levels of IL-6, IL-1β, TNF-α, MDA, and ROS, but increased SOD levels (p < 0.01). In addition, TLR4 expression and p-NF-κB/NF-κB ratio were lowered by circ_0000003 overexpression in the CHD cell model (p < 0.01). More importantly, the knockdown of circ_0000003 showed the opposite results to those by overexpression of circ_0000003.

    Conclusions: Circ_0000003 inhibits H2O2-induced inflammatory and oxidative stress responses in H9C2 cells, thereby promoting cell proliferation and inhibiting apoptosis, which is associated with blocking the TLR4/NF-κB p65 pathway.

  • Article
    Sahar Aldosari, Chukwuebuka Egbuna, Hind Albadrani, Alaa Alshareeda, Kingsley C. Patrick-Iwuanyanwu, Adedokun A. Kamoru, Sikiru O. Imodoye, Eugene N. Onyeike, Sadaf Jahan, Johra Khan
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6361-6374. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.604
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    Background: The Janus kinase 2-Signal Transducer and Activator of the Transcription 5 pathway (JAK2-STAT5 pathway) is a signaling pathway that plays a role in a variety of cellular processes, including cell growth, proliferation, and survival. Dysregulation of the JAK2-STAT5 pathway has been linked to several diseases, including cancer and its relapse. This study performed molecular docking simulation and Absorption, Distribution, Metabolism, Excretion, and Toxicity (ADMET) investigations on selected bioactive compounds against JAK2-STAT5 proteins for cancer drug discovery.

    Methods: JAK2 protein (Protein Data Bank (PDB) ID: 3Q32), STAT5 protein (PDB ID: 1Y1U), and 313 bioactive compounds, including standard anticancer drugs, were used for molecular docking simulation. The proteins' crystal structures were retrieved from PDB and prepared using BIOVIA Discovery Studio. The chemical structures of the compounds were retrieved from the National Center for Biotechnology Information (NCBI) PubChem database and prepared using the Open Babel and VConf software. PyRx equipped with AutoDock vina was used to perform molecular docking. The ADMET parameters for the top-performing compounds were determined using the SwissADME and pkCMS web servers.

    Results: The result from this study suggests that the following top-performing compounds, hypericin, withanolide, tomatidine, silymarin, baicalin, and diosmin, had the best binding energies with the potentials to serve as scaffolds or leads for new drug discovery against cancer. The ADMET study revealed that hypericin, baicalin, and diosmin violated more than one Lipinski's rule of 5 (RO5), which implies poor oral bioavailability, while withanolide, tomatidine, silymarin, ailanthone, elliptinium, ellipticine, etc. did not violate the rule which implies that they would be the best oral drug candidates based on the RO5 rules.

    Conclusions: The top-performing compounds, hypericin, withanolide, tomatidine, silymarin, baicalin, and diosmin, could serve as potential inhibitors/therapeutics against cancer caused by JAK2-STAT5 dysregulation.

  • Article
    Yan Ma, Yingge Peng, Xin Li, Xiaoguang Huo, Wenzhe Xu, Wei Zhao, Yingnan Wang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6375-6388. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.605
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    Background: Breast cancer (BC) is one of the most common malignant tumors in women. There is an urgent need to explore the key genes responsible for the prognosis of BC. The aim of this study was to screen out differentially expressed genes (DEGs) in BC and to provide reliable prognosis prediction. DEGs related to prognosis of BC were identified and verified as markers for predicting BC prognosis.

    Methods: Two data sets GSE22820 and GSE29431 were selected from the Gene Expression Omnibus (GEO) database and the DEGs were screened by the online tool of GEO2R and visualized by the volcano maps. The bioinformatics software of database for annotation, visualization, and integrated discovery (DAVID) was used to annotate associated functions of DEGs. Search tool for the retrieval of interacting genes/proteins (STRING) was used to construct the protein-protein interaction (PPI) networks which were visualized by Cytoscape software. Afterward, the data from The Cancer Genome Atlas (TCGA) were used for multivariate Cox regression analysis and Kaplan-Meier (K-M) survival analysis was used to identify prognosis-related key genes. The immortalized breast epithelial cell line MCF10A and BC cell line MCF7 were purchased and exploited to verify the differential expression of selected hub genes through real-time reverse transcriptase-polymerase chain reaction (RT-qPCR) and western blot. The expressions of prognostic-related genes were regulated through transfecting MCF7 cells, followed by exploring the effects of prognosis-related genes. The cell proliferation was determined by assays of colony formation. The cell counting kit (CCK)-8 and the cell migration were evaluated by the tests of transwell and cell scratch. The flow cytometry of Annexin V-PE and 7-AAD double stained cells was used for measuring cell apoptosis.

    Results: The bioinformatics analysis indicated that 969 DEGs were found in GSE22820, out of which 421 genes were over-expressed and 548 genes were under-expressed. In GSE29431, 724 DEGs were found, out of which 109 genes were over-expressed and 615 genes were under-expressed. GSE22820, GSE29431, and Co-DEGs are involved in mitotic nuclear division, regulation of angiogenesis, connective tissue development, muscle organ development, glucose metabolic process, and hexose metabolic process. Genes of peroxisome proliferator-activated receptor alpha (PPARA), peroxisome proliferator-activated receptor gamma (PPARG), lipoprotein lipase (LPL), leptin (LEP), insulin like growth factor 1 (IGF1), type I collagen α1 (COL1A1), Fibronectin (FN)1, and anillin (ANLN) were identified as hub genes, and ANLN, PPARA, PPARG, and LPL were verified for having effect on prognosis of BC. The upregulation of PPARA and PPARG, and the downregulation of ANLN and LPL restrained both proliferation and migration and induced apoptosis of BC cells.

    Conclusions: PPARA, PPARG, LPL, LEP, IGF1, COL1A1, FN1, and ANLN were identified as hub genes and ANLN, PPARA, PPARG, and LPL could affect the development of BC by inhibiting proliferation and migration and promoting apoptosis of cancer cells.

  • Article
    Jizheng Zhang, Yi Li, Xiaohua Sun, Wanlu Ren
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6389-6395. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.606
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    Background: Dexmedetomidine and sevoflurane are anesthetics widely used in clinical practice, but the effect of their combined application on artificial hip replacement is not clear. This study explored and analyzed the effects of this program on the immune function and stress response of patients, in order to provide more directions for the formulation and selection of anesthetic plans.

    Methods: The study selected the medical records of 160 patients undergoing artificial hip replacement in our hospital from November 2020 to November 2022 for retrospective analysis. According to the different anesthetic plans, the patients were divided into the study group (SG, 83 patients receiving dexmedetomidine combined with sevoflurane in anesthesia) and the reference group (RG, 77 patients receiving sevoflurane anesthesia) to compare the immune function indexes, anesthesia monitoring indexes, postoperative recovery indexes, stress response indexes, and postoperative complications between the two groups.

    Results: The SG had lower heart rate (HR) and mean arterial pressure at extubation (T1) and 30 min after extubation (T2) than the RG (p < 0.001), but there was no significant difference in respiratory rate and pulse blood oxygen saturation (p > 0.05). The SG had distinctly lower postoperative awakening time and hospitalization time than the RG (p < 0.05). There was no significant difference in the activities of daily living scores between the two groups (p > 0.05). The SG had remarkably higher levels of serum immunoglobulin, CD3+, CD4+, CD8+, and CD4+/CD8+ than the RG after surgery (p < 0.05). In contrast, the SG had notably lower levels of HR, norepinephrine, epinephrine, and cortisol than the RG (p < 0.001). There was no significant difference in the incidence of adverse reactions between the two groups (p > 0.05).

    Conclusion: Dexmedetomidine combined with sevoflurane is an efficient and safe anesthetic scheme for artificial hip replacement. It not only has little effect on the immune function of patients but also can reduce the stress response of the human body, providing strong support for a smooth surgical procedure.

  • Article
    Shaohua Zeng, Zhenyu Feng, Jinpeng Gu, Hexin Xu, Jianwen Zeng, Tiejun Pan
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6397-6409. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.607
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    Objectives: Prostate cancer is becoming increasingly common, and aggressive and metastatic growth continues to be the main factor in patient fatalities. Zeylenone (Zey) is a natural plant extract with anti-cancer effects, but its mechanism is unclear. This study aims to decipher the effect and mechanism of Zey in prostate cancer.

    Methods: After using Zey to inhibit androgen non-dependent prostate cancer cells PC3 and DU145, the survival rate of cells was detected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) to select the best dose. The capacity of cells to proliferate, migrate, and invade was then determined through colony formation test and Transwell® migration/invasion assay, respectively. Next, nuclear morphology of apoptotic cells and cell cycle were detected. In addition, the signaling pathways of Wnt/β-catenin (Wnt5a, β-catenin, cylin), phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) [PI3K, AKT, mechanistic target of rapamycin (mTOR)], glycogen synthase kinase-3beta (GSK-3β), and epithelial-mesenchymal transition (EMT) signaling [Vimentin, N-cadherin, E-cadherin, alpha-smooth muscle actin (α-SMA)] were explored using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR). We also conducted a tumor-forming model of nude mice and the tumor volume and weight were measured. Then, hematoxylin and eosin (HE) staining of the tumor tissues was performed and mRNA levels of the above signaling protein genes were also detected in the tumor tissues.

    Results: Cellular experiments revealed that Zey inhibited both PC3 cells and DU145 cells growth, colonization, invasion, and migration in a dose-dependent manner (p < 0.01). Medium and high doses of Zey significantly promoted PC3 cell apoptosis after 12, 24, and 48 h (p < 0.01), while low doses of Zey significantly promoted PC3 cell apoptosis only after 48 h (p < 0.01). All Zey treatment groups promoted DU145 cell apoptosis after 12, 24, and 48 h (p < 0.05 and p < 0.01), with a stronger effect observed with increasing time and dose. Besides, all Zey treatment groups boosted the ratio of G0/G1 stage cells (p < 0.01), reduced G2/M stage (p < 0.05 and p < 0.01) and S stage cells (p < 0.05 and p < 0.01) in both PC3 and DU145 cells. Mechanism research showed that all Zey treatment groups decreased the mRNA expression of key genes related to the PI3K/AKT signaling pathway (p < 0.05 and p < 0.01), Wnt/β-catenin signaling pathway (p < 0.05 and p < 0.01), EMT-related pathway (p < 0.05 and p < 0.01), and GSK-3β mRNA expression (p < 0.05 and p < 0.01) in both PC3 and DU145 cells. Animal experiments revealed that Zey could reduce the tumor dimensions of prostate cancer mouse tumor-forming model (p < 0.01), promote tumor cell necrosis (p < 0.01), and reduce the key genes mRNA expression of EMT (p < 0.01) and AKT/GSK-3β/β-catenin signaling pathway (p < 0.01).

    Conclusions: Zey can inhibit the proliferation, migration, and invasion of prostate cancer cells. Its mechanism of action may be linked to the synergistic inhibition of the AKT/GSK-3β and Wnt5/GSK-3β signaling pathways, which further inhibits EMT in prostate cancer cells.

  • Article
    Omkulthom Al kamaly, Ashwag S. Alanazi, Omar Noman, Hamada Imtara
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6411-6418. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.608
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    Background: Medicinal plants and natural products have proven effective in inhibiting the growth of cancer cells due to their rich content of active compounds, with polyphenols being among the most prominent. This study aims to assess the antiproliferative activity of Heliotropium bacciferum extracts on breast (MCF-7) and lung (A549) cancer cell lines and identify the extract's key active compounds.

    Methods: The antiproliferative activity of Heliotropium bacciferum extract and fractions on breast (MCF-7) and lung (A549) cancer cell lines was evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. Subsequently, the hexane extract was selected as the most effective extract for investigating the mechanism of its antiproliferative activity on A549 cell lines. The investigation involved the evaluation of cell cycle alterations using propidium iodide staining and the induction of apoptosis using the Annexin V-FITC/PI kit. Finally, gas chromatography/mass spectrometry (GC/MS) analysis was conducted to identify the chemical compounds in the hexane extract of Heliotropium bacciferum.

    Results: According to the MTT assay, the hexane fraction of Heliotropium bacciferum effectively halted the growth of A549 and MCF-7 cancer cells with IC50 values of 104.14 and 83.84 μg/mL, respectively. Additionally, methyl lineoleate (36.5%), linoleic acid (19.1%), and 3′-Acetyllycopsamine (10.7%) were the major components in the Heliotropium bacciferum hexane fraction, according to GC/MS analysis.

    Conclusions: In order to produce pharmaceutical anticancer medicines for cancer therapies, Heliotropium bacciferum hexane fraction can be employed.

  • Article
    Md. Showkoth Akbor, Mehedi Hasan Bappi, Abdullah Al Shamsh Prottay, Mst. Farjanamul Haque, Md. Sakib Al Hasan, Shoyaeb Ahammed, Rokibul Islam Chowdhury, Md. Sabbir Hosain, Clara Mariana Gonçalves Lima, Sheikh F. Ahmad, Sabry M. Attia, Talha Bin Emran, Henrique Douglas Melo Coutinho, Micheline Azevedo Lima, Muhammad Torequl Islam
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6419-6435. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.609
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    Background: Sleep is essential to human homeostasis and affects the immune system. Sleep disorders are a group of conditions that disturb normal sleep patterns and can affect overall health, safety, and quality of life. Poor or insufficient sleep has been associated with various dysfunctions in most body systems, such as endocrine, metabolic, higher cortical function, and neurological disorders. This study aims to evaluate the effects of the natural plant derivatives Sesamol (SES) and Thymol (THY) on sodium pentobarbital-induced sleeping mice.

    Methodology: The animals were given Sesamol (SES) (25, 50 mg/kg), Thymol (THY) (30 mg/kg), Diazepam (DZP) (3 mg/kg), and Caffeine (CAF) (10 mg/kg) orally (p.o.) in the respective groups individually and in combination. After 30 minutes, the treated mice were given sodium thiopental (10 mg/kg) intraperitoneally (i.p.) to induce sleep, and latency of sleeping time and duration were observed. Additionally, an in silico study was undertaken to predict the involvement of gamma-aminobutyric acid (GABA) receptors in the sleep mechanism.

    Results: In the current study, we observed that SES and THY increased the duration of sleeping time and decreased the latency of sleep induction. When SES, DZP, and THY were administered together, they demonstrated the greatest hypnotic activity. SES and THY exhibited strong binding affinity with different GABA receptor subtypes in in silico studies. The pharmacokinetic analysis of SES and THY using SwissADME indicated good absorption, distribution, metabolism, and excretion properties.

    Conclusions: SES and THY produced a hypnotic-like effect in the mice model, possibly through the GABAA and GABAB receptor interaction pathways.

  • Letter
    Chien Ing Yeo, Yee Seng Tan, Bey Hing Goh, Long Chiau Ming, Sibrandes Poppema
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(11): 6437-6442. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233711.610
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    Nuclear magnetic resonance (NMR) metabolomics is a powerful tool in cancer research that functions to unveil disease-specific metabolic signatures and potential biomarkers while aiding in the development of personalized therapies and the understanding of drug mechanisms. Recent innovations, such as high-resolution magic angle spinning (HR-MAS) and 2D NMR spectroscopy, offer comprehensive insights into metabolite structures and identification. Its potential application in Hodgkin lymphoma (HL) research shows promise in uncovering unique metabolic pathways specific to this malignancy. Furthermore, the integration of NMR data with genetics facilitates the identification of cancer biomarkers for diagnostic, prognostic, and therapeutic purposes. Nonetheless, persistent challenges encompass variability in sample preparation, data acquisition, and analysis methods, necessitating standardization and validation efforts. Despite collaborative data-sharing initiatives enhancing transparency and fostering cooperation, centralization and standardization of data remain ongoing objectives. Overall, NMR metabolomics presents the potential to transform the understanding of cancer metabolism and leverage the development of personalized anticancer therapies, although it is necessary to address challenges and enhance data integration.