20 March 2023, Volume 37 Issue 3

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  • Article
    Xianghe Chen, Peng Sun, Aiguo Chen, Haixia Peng, Xianliang Zhang, Kang Yang, Huilin Yu, Renqing Zhao
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1163-1171. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.119
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    Background: Substantial evidence has demonstrated that impact exercise can help adolescents to accumulate more bone mass. However, the effect of OB (osteoblast) on promoting bone formation during impact exercise remains to be explored. This study aimed to examine the mechanism for downhill running promoting bone formation ability and bone mineralization ability by activating the TGF-β (transforming growth factor-β)/Smad (drosophila mothers against decapentaplegic protein) pathway in growing mice.

    Methods: Four-week-old C57BL/6 female mice (n = 28) were randomly divided into exercise and control groups. Mice in the exercise group received a forty-minute downhill training on a descending treadmill (descent 9°) every day for eight weeks. After eight weeks, BMSCs (bone marrow mesenchymal stem cells) were collected and cultured for assessment of osteoblastic differentiation and the capacity of osteoblastic mineralization. We determined the protein and mRNA expression of TGF-β, Smad2/3/4, and Runx2 (runt-related transcription factor 2) in bone tissues. The BMD (bone mineral density) and histomorphological changes were also examined.

    Results: The study showed that the eight-week downhill training significantly increased BMD and ALP (alkaline phosphatase-positive) activity in the tibial epiphysis. Also, downhill running promoted BV/TV (bone volume fraction), Tb.N (trabecular number), and Tb.Th (trabecular thickness), while it decreased Tb.Sp (trabecular separation). Additionally, the numbers of ALP+CFU-f (colony forming units-fibroblastic) cells and the area of mineralized nodule formation were significantly higher in the exercise group compared with that of the control group. Lastly, the mRNA expression of TGF-β, Smad2/3/4, and Runx2 was significantly elevated in the exercise group, and the protein expression of p-Smad2/3/4, and Runx2 was also increased.

    Conclusions: The findings indicated that downhill running enhanced bone mass accumulation and bone tissue morphology in growing mice by activating the TGF-β/Smad pathway.

  • Article
    Mingxuan Feng, Jingsheng Zhang, Xiao Teng, Zhaobo Zhang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1173-1186. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.120
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    Background: Although studies have highlighted the potential role of monocytes in osteoporosis, the detailed mechanism of action of monocytes in the microenvironment and their diagnostic value in osteoporosis have not been fully elucidated. This study identified immune-related genes in osteoporosis using transcriptional data from monocyte samples obtained from public datasets.

    Methods: Microarray datasets associated with osteoporosis were downloaded from the Gene Expression Omnibus database. Differentially expressed genes (DEGs) between the low bone mineral density (BMD) and high BMD groups were screened, with a false-discovery rate (FDR) <0.05 and |log2fold change (FC)| > 0.263 as thresholds. The type of immune infiltration was assessed in monocyte samples using the single-sample gene set enrichment (ssGSEA) algorithm, and immune-related DEGs were identified. Enrichment analysis was performed on immune-related DEGs. Subsequently, optimal immune-related DEGs were screened using least absolute shrinkage and selection operator (LASSO) and recursive feature elimination (RFE) algorithms. Finally, a risk diagnostic model was constructed based on optimal immune-related DEGs.

    Results: A total of 428 DEGs were identified in patients with low BMD compared with those in the control group. Four types of immune cells with significant differences and 310 immune-related DEGs were identified. These immune-related DEGs were mainly enriched in the peroxisome, metabolic pathways, and alanine, aspartate, and glutamate metabolism. In addition, seven optimal immune-related genes were identified as immune-related markers for osteoporosis, and a diagnostic model was constructed. The risk diagnostic model of the seven signature genes showed high area under the curve (AUC) values, suggesting a reliable predictive ability for osteoporosis.

    Conclusions: Our results suggest a potential role for these seven optimal genes in osteoporosis progression and immune cell infiltration. Risk models based on the seven optimal genes are useful tools for osteoporosis diagnosis and osteoporotic fracture risk prediction.

  • Article
    Hong Peng, Lizhu Nie, Zheng Jiang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1187-1200. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.121
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    Background: Colorectal cancer (CRC) is the main cause of cancer-related mortality globally. Thus, it is critical to find new therapeutic targets for CRC. This study aimed to evaluate the role and potential mechanism of procollagen-lysine, 2-oxoglutarate 5-dioxygenase 3 (PLOD3) on CRC cells.

    Methods: PLOD3 expression was identified by immunohistochemistry (IHC) and Western blot (WB) assay in CRC tissue and cells. PLOD3 roles in HCT116 (PLOD3 knockdown) and RKO (PLOD3 overexpression) cells were evaluated. In vitro by Cell Counting Kit-8 (CCK-8) assay, colony formation, transwell, and wound healing assays, and in vivo by xenograft and hepatic metastasis model. RNAseq was conducted between PLOD3 knockdown and control in HCT116 cells. The key genes β-catenin, c-myc and Sex-determining region Y-box 4 (SOX4) in CRC were determined by WB assay in vitro. Immunofluorescence assay was used to assess PLOD3 effect on the expression and location of β-catenin.

    Results: PLOD3 was screened as one of the main proliferative factors in CRC. In addition, CRC cell lines and clinical CRC tissues overexpressed PLOD3. In individuals with CRC, overexpressed PLOD3 was positively related with poor prognosis and metastasis. It was found that PLOD3 increased CRC cell proliferation, colony formation, migration, and invasion in vitro by gain- and loss-of-function tests. Additionally, a xenograft mouse model and a hepatic metastasis mouse model were used to show that PLOD3 ablation decreased CRC outgrowth and metastasis in vivo. In contrast, PLOD3 overexpression had the opposite effect. PLOD3 upregulated SOX4 in a mechanism that enhanced c-myc and β-catenin expression. Immunofluorescence assay indicated that PLOD3 expression was positively related to β-catenin in CRC cells.

    Conclusions: PLOD3 promotes tumor progression and poor prognosis through SOX4/Wnt/β-catenin signaling pathway in CRC.

  • Article
    Changhui He, Ting Wang, Wei Zhang, Chenyang Zhao, Na Li, Lixiu Peng
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1201-1214. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.122
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    Background: Endometrial cancer (EC) is the most common malignancy in the female reproductive system in developed countries and exhibits high heterogeneity. Our aim was to construct a ferroptosis-related lncRNA prognostic model to guide tailored treatment for EC patients.

    Methods: RNA-sequencing data and clinical information about EC were obtained from the cancer genome atlas (TCGA). A risk model was established by least absolute shrinkage and selection operator (LASSO) analysis. Subsequently, gene set enrichment analyses (GSEA) was applied to explore the differences in biological characteristics. Immune cells abundance profile was evaluated using different methods, including XCELL, MCPcounter, EPIC, TIMER, and CIBERSORT. The R package “oncopredict” was used to estimate drug sensitivity.

    Results: In this study, 35 normal samples and 500 tumor samples were included. Five lncRNAs (AC092969.1, AL356489.2, AP000757.1, AP001189.3, and LINC01936) were selected to compose the ferroptosis-related lncRNA prognostic model, which possessed better predictive ability than individual indicators to predict 5-year survival (AUC (area under the ROC curve) = 0.75). A nomogram for overall survival was established, which integrated risk score, stage, grade, and age. There was a higher proportion of TP53-mut subtype serous papillary carcinoma in high-risk group, poorly differentiated. Ferroptosis-related pathways are significantly enriched in the GSEA of the low-risk group. We observed differences between low-risk and high-risk groups in their immune landscapes, immune checkpoint expression levels, drug sensitivity and mutation landscapes.

    Conclusions: The prognostic risk model is a reliable and robust predictor for survival outcomes in EC patients. It can identify patients that likely will derive benefit from immunotherapy and other drug therapies.

  • Article
    Bowen Hu, Lingyu Du, Hongya Xie, Guangda Yuan, Yong Yang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1215-1226. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.123
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    Background: Nucleotide-binding domain leucine-rich family caspase-activation and recruitment domain (CARD) domain-containing protein 4 (NLRC4) has been linked to several human diseases. However, its function in lung adenocarcinoma is uncertain.

    Methods: NLRC4 expression was investigated using The Cancer Genome Atlas (TCGA) and pan-cancer Genotype-Tissue Expression data. The researchers examined the link between NLRC4 expression and patient outcome by using TCGA clinical survival data. NLRC4 enrichment was analyzed using the clusterProfiler R package. The TCGA database was also used to investigate the association between NLRC4 expression and immune cell infiltration. Western blot was used to confirm NLRC4 expression and its effect on apoptosis-related proteins in lung adenocarcinoma. Immunohistochemistry was used to investigate the effect of NLRC4 expression on immune cell infiltration.

    Results: NLRC4 expression is low in lung adenocarcinoma. Low NLRC4 expression was associated with related genes involved in apoptosis and immune cell activation. Further investigation showed that low NLRC4 expression was associated with high infiltration scores of macrophages, dendritic cells, T cells, B cells, and natural killer (NK) cells. NLRC4 knockdown inhibited apoptosis-related protein production in lung adenocarcinoma cells. The number of peripheral macrophages, dendritic cells, T cells, B cells, and NK cells was significantly reduced in pathological tissues with low NLRC4 expression.

    Conclusions: Our findings suggest that NLRC4 may be a risk gene for lung adenocarcinoma and affects the infiltration of the tumor immune cells.

  • Article
    Shiqin Xi, Junsong Zhu, Qunwei Qu, Lei Fang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1227-1236. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.124
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    Background: An increasing number of microRNAs (miRNAs) are involved in the progression of knee osteoarthritis (OA). In this study, we identified a previously unknown role of miR-1285-5p in inflammation during knee OA and its underlying mechanism.

    Methods: RT-qPCR (real time flurocent qualitative PCR) was used to detect the expression of miR-1285-5p and PAK1 (p21 activated kinase 1) in cartilage from subjects with knee OA, normal tissues, and chondrocytes. After establishing in vitro chondrocyte inflammation model, we detected cell proliferation and apoptosis using CCK-8 (cell counting kit-8), EdU proliferation, flow cytometry, and caspase-3 activity assays. ELISA (enzyme-linked immunosorbent assay) was also used to test IL-1β (interleukin-1β), TNF-α (tumor necrosis factor alpha), and IL-6 levels in the chondrocyte culture. The targeted binding of miR-1285-5p to the PAK1 3′UTR (untranslational region) was verified using luciferase reporter assays.

    Results: MiR-1285-5p was downregulated in cartilage from subjects with knee OA and lipopolysaccharide (LPS)-treated chondrocytes. MiR-1285-5p overexpression inhibited the increased secretion of IL-1β, TNF-α, and IL-6 in chondrocytes after LPS treatment. Similarly, inhibition overexpression was observed on LPS-induced chondrocyte apoptosis after miR-1285-5p overexpression. In contrast, miR-1285-5p overexpression restored the proliferative impairment in chondrocytes exposed to LPS. MiR-1285-5p targeted and suppressed PAK1 expression. PAK1 overexpression abrogated the protective effect of the miR-1285-5p mimic on LPS-stimulated chondrocytes. The relieving effect of miR-1285-5p on LPS-increased p-IKKβ (phospho-IKK beta), IκB-α (IkappaB-alpha), and NF-κBp65 (nuclear factor NF-kappa-B p65) levels was offset by PAK1 overexpression.

    Conclusions: By downregulating PAK1 expression via NF-κB signaling, miR-1285-5p protection against knee OA was demonstrated, indicating that miR-1285-5p/PAK1 may be a suitable therapeutic target for the management of human knee OA.

  • Article
    Chao Qiu, Shengcai Li, Gang Dai
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1237-1243. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.125
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    Objective: To analyze the impact of glargine and glimepiride combined with appropriate exercise on improving lower extremity function and blood glucose level in patients with lower extremity arterial occlusive disease (LEAOD).

    Methods: One hundred thirty patients with diabetic LEAOD were randomly divided on a 1:1 basis (n = 65) in control and research groups according to a random number table. The research group was prescribed glargine and glimepiride combined with appropriate exercise.

    Results: There were no statistically significant differences in fasting plasma glucose (FPG), 2h postprandial blood glucose (2h PG), glycosylated hemoglobin (HbA1c), insulin function index (HOMA-islet), and insulin resistance index (HOMA-IR) pre-treatment between groups. FPG, 2h PG, HbA1c, HOMA-islet, and HOMA-IR levels were significantly lower in the research group than in the control group post-treatment. Moreover, the rate of improvement was greater in the research group than the control group. The incidence of adverse events during treatment was lower in the research group than in the control group.

    Conclusions: Glargine and glimepiride combined with appropriate exercise in patients with diabetic LEAOD effectively controlled blood glucose level, improved insulin activity, and promoted recovery of lower extremity function.

  • Article
    Kun-heng Fan, Zhi-yu Li, Tian-cheng Liu, Qian Xi, Jian-hua Zhang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1245-1252. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.126
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    Objective: To study the correlation between gadolinium ethoxybenzyl diethylenetriamine pentaacetic acid (Gd-EOB-DTPA)-enhanced magnetic resonance imaging (MRI) liver parenchyma enhancement and liver function and the choice of hepatobiliary phase (HBP) in patients with different liver functions.

    Data and Methods: A total of 88 patients who underwent Gd-EOB-DTPA-enhanced MRI were analyzed retrospectively. Ac-cording to Child–Pugh classifications, they were divided into Groups A, B, and C, and Group D was established for patients with normal liver function. The liver parenchyma signal intensity (SI) at 5, 10, 15, and 20 min (SI5min, SI10min, SI15min, and SI20min) and biliary excretion times of a c ontrast medium were obtained.

    Results: After contrast medium injection, the SI of Group D rose significantly within 10 min and stabilized within 10–20 min. The SI in Group A increased significantly within 15 min and stabilized after 15 min, with no significant difference identified between SI15min and SI20min (t = 0.071, p = 0.944). In Groups B and C, the degree of hepatic parenchyma enhancement exhibited no significant difference after a 5 min delay. There were significant differences in SI5min and SI10min between Groups D and A; In SI5min, SI10min, SI15min, and SI20min between Group D and Groups B and C; In SI20min between Group A and Groups B and C; And in SI15min between Groups A and C (p < 0.05). SI10min and SI20min had a negative correlation with the total bilirubin and prothrombin time and a positive correlation with serum albumin.

    Conclusions: Gd-EOB-DTPA-enhanced liver parenchyma is correlated with liver function and the corresponding biochemical indexes. A delay time of 15 min for the hepatobiliary phase was sufficient for most patients who get Gd-EOB-DTPA-enhanced MRI.

  • Article
    Yujing Guo, Sheng Yong, Qifu Long, Ying Hu, Yuzhen Xu, Jidong Li, Zengqiang Miao, Huiqian Li
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1253-1265. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.127
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    Background: Hypoxia induces lymphocyte apoptosis and affects their function. However, there is limited information about the mechanisms underlying the hypoxia-induced apoptosis of lymphocytes.

    Objective: Apoptosis is mainly mediated by Fas (factor associated suicide) receptor-dependent apoptosis pathway and mitochondria pathway. Therefore, this study aimed to investigate whether hypoxia-induced lymphocyte apoptosis occurs via the Fas receptor-dependent pathway and/or the mitochondrial pathway.

    Methods: Mouse spleen lymphocytes were exposed to hypoxia (1%) and normoxia (21%) for 6, 12, and 24 h. Cell viability, cell morphological changes and apoptosis were studied. Reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) production were also measured. The expression of apoptosis-related proteins was analyzed by quantitative real-time PCR (qRT-PCR) and western blotting.

    Results: Hypoxia-treated reduced lymphocyte survival and increased lymphocyte apoptosis, inducing morphological changes characteristic to early apoptosis. Hypoxia exposure enhanced the ROS formation. The MMP of lymphocytes decreased after 12 and 24 h of hypoxia exposure. Factor associated suicide (Fas), factor associated suicide ligand (FasL), Bcl-2 homologous antagonist-killer protein (Bak), Cytochrome C, apoptotic peptidase activating factor 1 (Apaf-1), and cysteinyl aspartate specific proteinase (caspase)-3, -6, -7, -8, and -9 transcript expression was upregulated and B-cell lymphoma 2 (Bcl-2) transcript expression was downregulated in lymphocytes exposed to hypoxia for different time points. Then, the expression of Fas, FasL Bak, Cytochrome C, and caspase-3 were upregulated and that of Bcl-2 was downregulated at the protein level after hypoxia exposure.

    Conclusions: Hypoxia may induce lymphocyte apoptosis through both the Fas death receptor-dependent and mitochondria-dependent apoptosis pathways, thus mediating lymphocyte injury and causing immune dysfunction. Our findings provide the experimental basis for the mechanism of action of hypoxia in immune function impairment.

  • Article
    Peizhi Zhang, Jiechuan Qiu, Qingliang Wang, Yingkun Xu, Zicheng Wang, Fan Peng, Yuhu Hao, Guangzhen Wu, Qinghua Xia
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1267-1283. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.128
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    Background: Cuproptosis is a novel discovered cell death mechanism in cancers. This study aimed to investigate the expression and prognostic value of DLAT (dihydrolipidamide s-acetyltransferase), one of the Cuproptosis-related genes, in kidney renal clear cell carcinoma (KIRC) using a comprehensive analysis integrating bioinformatics and experimental assays.

    Methods: High-throughput data and clinical information of KIRC patients were downloaded from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases. We used various R packages and bioinformatics analysis websites to analyze DLAT expression and function. Clinical tissue samples were used to verify DLAT expression in KIRC based on real-time quantitative reverse transcription (qRT-PCR) and immunohistochemistry. Subsequently, we validated DLAT differential expression in normal renal cells versus different KIRC cell lines using qRT-PCR. CCK-8 (cell counting kit-8) assay was used to assess the effect of DLAT overexpression on KIRC cells proliferation. Wound healing and Transwell assays were used to assess the effect of DLAT overexpression on KIRC cells invasion and migration.

    Results: DLAT mRNA expression in KIRC tumor tissue was lower than in normal kidney tissue. DLAT high expression suggested a good prognosis in KIRC. Gene ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that DLAT was involved in a variety of biological functions and pathways. Infiltration analysis indicated that DLAT might affect the prognosis of KIRC by affecting immune infiltration in the KIRC tumor microenvironment. qRT-PCR and immunohistochemistry further DLAT low expression in KIRC compared to normal kidney tissue. Besides, we found that DLAT overexpression could not only inhibit KIRC cells growth, but also effectively inhibit the migration and invasion of KIRC cell lines.

    Conclusions: This study explored the expression of DLAT in KIRC and the effect of DLAT on the proliferation, migration, and invasion of renal cancer cell lines, which has a positive role in guiding the diagnosis and treatment of KIRC.

  • Article
    Xiao-dong Zhu, Qi Zhang, Yang Wang, Ming-rui Jiang, Muhammad Ali, Qian-nan Sun, Dao-rong Wang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1285-1291. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.129
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    Background: Gastric cancer (GC) is one of the most common malignant tumors worldwide, but due to its complex pathogenesis, current treatments are still very limited. Studies have shown that galectin-8 expression is significantly increased in GC and is associated with poor prognosis in patients, but the mechanism of galectin-8 in GC remains unclear. Therefore, this study aimed to investigate the expression, prognostic value, and molecular mechanism of galectin-8 in GC.

    Methods: UALCAN was used to analyze galectin-8 expression levels in GC and their correlation with clinical stage, histological subtype, and lymph node metastasis. Western blotting and immunohistochemical staining were used to detect the expression levels of galectin-8 in GC tissues. siRNA was transfected using lentivirus to knock down galectin-8. Galectin-8 expression levels and transfection efficacy in GC cells were detected using Western blot. The Cell Counting Kit-8 (CCK-8) assay and colony formation assay were used to detect the level of cell proliferation. Western blotting was used to detect the rat sarcoma viral oncogene homolog (Ras) signaling pathway-related protein expression levels. Immunofluorescence staining was performed for co-localization analysis of galectin-8 and Kirsten Ras (K-Ras) proteins.

    Results: The expression levels of galectin-8 were significantly increased in GC tissues and cells, and that was positively correlated with clinical stage, histological subtype and lymph node metastasis, and negatively correlated with overall survival. Knockdown of galectin-8 significantly inhibited Human Gastric Adenocarcinoma (AGS) cell proliferation and decreased RAF proto-oncogene serine/threonine-protein kinase (Raf), mitogen-activated protein kinase (Mek), and K-Ras protein expression levels. Immunofluorescence staining results indicated that galectin-8 and K-Ras protein co-localized in the cytoplasm.

    Conclusions: Galectin-8 promotes GC proliferation by activating the Ras signaling pathway and is a potential therapeutic target and prognostic biomarker for GC.

  • Article
    Luying Huang, Ziyi Zhao, Chenglong Li
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1293-1302. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.130
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    Objective: Colon cancer stem-like cells (CSCs) are self-renewing and tumor-initiating. They are resistant to chemotherapy drugs, such as doxorubicin. The presence of this subpopulation leads to poor treatment outcomes, recurrence and metastasis in colon cancer. Therefore, current therapeutic strategies targeting stem cell self-renewal pathways are a reasonable approach for cancer prevention and treatment. In this study, we aimed to evaluate the potential antitumor effects of Lup-20(29)-en-3b-ol (lupeol) by targeting this subpopulation.

    Methods: CSCs were enriched by being cultured in serum-free medium, and the effects of lupeol on CSC malignancies were measured. These effects included proliferation, migration, invasion and tumor formation. The stemness hallmarks were measured by western blotting. The effects of lupeol on the chemosensitivity of CSCs were measured by detecting apoptosis.

    Results: We found that lupeol did not affect cell proliferation and viability but remarkably inhibited the invasion and tumor formation abilities of CSCs. When colon cancer cells were cultured in serum-free medium, lupeol inhibited sphere formation and decreased stemness hallmarks. Moreover, when lupeol was added to the formed spheres for 48 h, the spheres dissociated and tended to grow adherently. Additionally, only Nestin and β-catenin, but not Sox2, Oct4 (Octamer-binding transcription factor) or Nanog, were significantly decreased. These results demonstrate that lupeol disturbed stemness in colon cancer, potentially by regulating Nestin or β-catenin. Additionally, lupeol treatment or β-catenin inhibitor treatment significantly increased DOX (Doxorubicin)-induced apoptosis, demonstrating that lupeol increased chemosensitivity potentially by downregulating β-catenin in CSCs.

    Conclusions: Taken together, our results demonstrate that lupeol is a potential chemoagent that specifically targets CSCs in colon cancer.

  • Article
    Ying Li, Wei He
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1303-1311. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.131
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    Background: This study aimed to explore Smad7 function and possible mechanism in human epidermoid cancer cells A431.

    Methods: A431 cells were divided into Control group, NC group (transfected with empty vetor) and miR-Smad7 group (transfect with Smad7 recombinant plasmids). Transwell and WST-1 assay were performed to determine A431 cells’ cell migration, invasion and growth. ELISA (enzyme-linked immunosorbent assay) was used to assess secreted TGF-β1 (transforming growth factor beta1) level, and immunofluorescence assay and confocal microscopy was used to assess Smad4 subcellular distribution in A431 cells. TGF-βRI (transforming growth factor beta receptors), Smad, PCNA (proliferating cell nuclear antigen), C-myc, Bcl-2 (B-cell lymphoma-2), Bax and EGFR (epidermal growth factor receptor) protein levels of A431 cells were determined by Western blot. A431 cell cycle and apoptosis was determined using flow cytometry. A431 cells morphology was determined by transmission electron microscope (TEM) micrographs and scanning electron microscope (SEM).

    Results: The results showed that three Smad7-targeting miR-Smad7s were successfully constructed. A431 cells Smad7 silencing reduced cell invasion, migration and proliferation (p < 0.01) and contributed to enhance PCNA, C-myc, and EGFR protein expression levels (p < 0.01). In addition, miR-Smad7 induced G1 A431 cells arrest and apoptosis (p < 0.01). Specifically, morphology apoptosis phenotype was observed in miR-Smad7 group. Smad4 subcellular distribution showed the presence of Smad4 in the nucleus in miR-Smad7 group. Western blot demonstrated higher TGF-βRI and Smad4 expression levels in the miR-Smad7-2 group compared to the Control group (p < 0.01).

    Conclusions: In summary, Smad7 affected A431 cells proliferation, migration, invasion and apoptosis, most probably by interfering with the TGF-β signaling pathway.

  • Article
    Jianke Li, Yangyang Lei, Guoping Li, Ningzi Tian, Shanshan Gao, Junhao Li, Tianzhu Yu, Xiaolin Wang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1313-1323. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.132
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    Purpose: Cystatin C (Cyst C) is involved in various human cancers. However, Cyst C has not yet been studied in pancreatic cancer (PC). This study aimed to examine Cyst C expression and its prognostic and biological role in PC.

    Methods: Cyst C expression in PC patients was assessed searching on the Clinical Proteomic Tumor Analysis Consortium (CPTAC) database. It was further validated in microarray tissue by immunohistochemistry (IHC) analysis. The biological role of Cyst C and its mechanism were explored by several molecular biology methods.

    Results: Cyst C was downregulated in PC tissues compared to the pericarcinoma tissue. Cyst C low expression was associated with high tumor grade, high N stage and poor prognosis of PC patients. Furthermore, Cyst C overexpression inhibited PANC-1 (human pancreatic epithelioid carcinoma cell line) and MIA PaCa2 cells migration, while Cyst C expression inhibition promoted the cellular migration. In addition, Cyst C knockdown leaded to autophagic deficiency and SQSTM1 accumulation in pancreatic cancer cells, followed by increased CDH2 and ZEB1 expression and decreased CDH1 expression. However, the expression patterns of these genes were reversed after SQSTM1 or RELA (V-Rel Reticuloendotheliosis Viral Oncogene Homolog A) down-regulation.

    Conclusions: This study revealed that Cyst C downregulation induces epithelial-to-mesenchymal transition (EMT) and promoted pancreatic cancer metastasis, at least partially due to SQSTM1/RELA signaling pathway activation. Cyst C might act as a promising biomarker for PC and provide a novel treatment strategy.

  • Article
    Yuewen Gao, Yanqin Wang, Lingxin Meng, Kai Wang, Xiaoyong Liu, Lei Chen
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1325-1334. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.133
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    Introduction: MicroRNAs (miRNAs) play vital roles in tumorigenesis and metastasis. The role of miR-155-5p in various cancers has been widely reported; However, its role in gastric cancer (GC) remains unknown. The aim of the present study was to investigate the role of miR-155-5p in GC pathogenesis and drug resistance.

    Methods: The miRNA profiles of GC were downloaded from the University of California Santa Cruz Xena database, and miR-155-5p levels in normal and GC tissues were analyzed. Adjacent and cancer tissues were collected from patients with GC. Real-time quantitative polymerase chain reaction (RT-qPCR) was used to analyze miR-155-5p levels in GC tissues and cell lines. Then, the NCI-N87 (human GC cells NCI-N87) cells were divided into four groups: Control, miR-155-5p-OE (overexpression), LP (lobaplatin), and miR-155-5p-OE+LP groups. The cells in the miR-155-5p-OE, and miR-155-5p-OE+LP groups were both transfected with miR-155-5p mimics, and then the cells in the miR-155-5p-OE+LP group were treated with 5 µg/mL lobaplatin. The cells in the LP group were only treated with lobaplatin, while the cells in the control group were without treatments. After culture, Transwell and cell counting kit-8 assays were used to assess the migratory and invasive capacities and viability, respectively, of lobaplatin-treated GC cells.

    Results: Bioinformatic analysis and RT-qPCR results showed that miR-155-5p was highly abundant in GC tissues and cell lines. Enrichment of miR-155-5p facilitated the growth of NCI-N87 cells. Additionally, lobaplatin treatment significantly decreased miR-155-5p levels in NCI-N87 cells and inhibited the viability and migratory and invasive capacities, whereas miR-155-5p enrichment rescued the GC cells from the inhibitory effects of lobaplatin treatment.

    Conclusions: Our results revealed that miR-155-5p promotes the growth of NCI-N87 cells, and its enrichment rescues NCI-N87 cells from the inhibitory effects of lobaplatin treatment. Thus, miR-155-5p may be a potential molecular target for the treatment of GC.

  • Article
    Hui Deng, Ting Xiong, Hanxiang Yu, Mingzhang Li, Zhijun Chen, Jun Liu, Shenliang Chen, Jun Tao, Lifeng Xie
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1335-1351. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.134
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    Background: Knee osteoarthritis (OA) is the most common osteoarthritic condition and a major contributor to disability. Increasing evidence indicates the pathological changes in the synovial membrane occur earlier than OA. Cuproptosis is a novel kind of copper-induced cell death. However, the role of cuproptosis-related genes (CRGs) in OA is still unknown. Our aim was to explore the role of CRGs in the expression and immunomodulation of osteoarthritis disease.

    Methods: The gene expression profile-related datasets including 27 normal samples and 27 OA samples were retrieved from the Gene Expression Omnibus (GEO) database. Cluster specific differentially expressed genes were identified using the weighted gene co-expression network analysis (WGCNA) algorithm. Based on least absolute shrinkage (LASSO), support vector machine–recursive feature elimination (SVM-RFE) and RandomForest (RF) algorithm, we constructed a CRG-based predictive model of osteoarthritis. Next, we used the nomogram, calibration curves to test the prediction power of our predictive model, used quantitative real-time PCR (polymerase chain reaction) (qRT-PCR) to verify differences of key genes expression in human synovial tissues. The functional enrichment of CRGs in OA was then evaluated using the gene set enrichment analysis (GSEA) approach. Finally, we constructed targeting drugs and ceRNA network based on online database.

    Results: We constructed a predictive model consisted of six CRGs (FDX1, CDKN2A, GSS, ATP7B, NDUFA1 and NDUFB1) by the LASSO, SVM-RFE and RF algorithm. The nomogram, calibration curves and qRT-PCR manifested that the predictive model had satisfactory performance. Analysis of immune infiltration revealed immune heterogeneity between OA and healthy individuals. Further immune correlation analysis showed that four genes (FDX1, CDKN2A, GSS and ATP7B) was closely associated with the immune status of OA. Single-cell RNA sequencing analysis showed that the cells type of the six CRGs aggregation was mainly related to immunity. Finally, we identified CDKN2A, ATP7B and five miRNAs (has-miR-30b-3p, has-miR-3163, has-miR-570-3p, has-miR-548x-3p, and has-miR-576-5p) as potential therapeutic targets.

    Conclusions: We systematically evaluated the immunological traits between individuals with normal tissues and OA tissues and the CRGs that were closely associated with immunity in OA. In addition, we constructed a CRG-based predictive model to disclose the patients of OA with higher immune scores. These findings may offer additional evidence for the function of cuproptosis in the immune regulation of OA and indicate that CRGs are potential diagnostic biomarkers of OA.

  • Article
    Xiaoliang Wang, Jin Zhang, Zhixin Sun, Sainan Zhao, Wenjuan Deng
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1353-1360. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.135
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    Objectives: The purpose of this study is to unveil the modulatory effect of microRNA (miRNA)-15b-5p on the differentiation of dental pulp stem cells (hDPSCs).

    Methods: Following the differentiation of hDPSCs into odontoblasts, qRT-PCR (quantitative reverse transcription-polymerase chain reaction) was used to determine miR-15b-5p expression in the cells. Dual-luciferase reporter (DLR) assay was adopted to confirm the downstream target gene of miR-15b-5p. Post transfection of miR-15b-5p mimic, inhibitor or si-insulin-like growth factor 1 (IGF1) into the differentiated cells, alkaline phosphatase (ALP) activity and mineralized nodule formation were evaluated by a commercial assay kit and alizarin red staining. Differentiation-related protein levels were evaluated using Western blotting. Transwell and extracellular matrix adhesion cell assays were performed to observe the invasion and adhesion of differentiated cells.

    Results: Downregulation of miR-15b-5p was observed in odontoblasts. MiR-15b-5p targeted IGF1, and miR-15b-5p mimic repressed IGF1 expression. Additionally, miR-15b-5p inhibitor strengthened the ALP activity and mineralized nodule formation, enhanced the invasion and adhesion of hDPSCs, and promoted the expression levels of dentin sialophosphoprotein (DSPP), dentin matrix protein 1 (DMP1), osteopontin (OPN), bone sialoprotein (BSP) and osteocalcin (OCN) during the differentiation of hDPSCs. However, the miR-15b-5p mimic exerted the effects opposite to the miR-15b-5p inhibitor. Besides, silencing of IGF1 reversed the effects of miR-15b-5p inhibitor on the above indexes.

    Conclusions: MiR-15b-5p regulates the differentiation of hDPSCs into odontoblasts by targeting IGF1.

  • Article
    Jin Tong, Ting Ye, Xiaofang Yang, Hong Peng, Xiao Xiao, Bingjue Xie, Xiaoqiang Wan, Wen Li, Li Zhou, Jia Chen, Jinjun Guo
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1361-1370. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.136
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    Background: Hepatitis B e antigen (HBeAg) precursor proteins, which share an identical encoding PreC/C open reading frame (PreC/C-ORF) of hepatitis B virus (HBV) core, can inhibit viral replication in vitro. The apolipoprotein B mRNA (messenger ribonucleic acid) editing enzyme catalytic polypeptide 3A (APOBEC3A) can inhibit HBV replication by interacting with the HBV core protein. The aim of the study was to explore whether the HBeAg precursor proteins can regulate HBV replication by interacting with APOBEC3A.

    Methods: Plasmids harboring the coding sequence of HBeAg precursor proteins (P25 and P22) and APOBEC3A were individually or co-transfected into human hepatoma and embryonic kidney cells. P25, P22, and APOBEC3A expressions were quantified by Western blotting. The co-localization and interaction of P25, P22, and APOBEC3A were determined by confocal microscopy and co-immunoprecipitation (Co-IP). We constructed serial precursor proteins of HBeAg deletion mutants for further Co-IP analysis. Enzyme-linked immunosorbent assay (ELISA) was employed to evaluate the levels of HBeAg and HBsAg. HBV replication intermediates from the transfected cells were detected by Southern blotting.

    Results: P25, P22, and APOBEC3A proteins were successfully expressed in transfected cells, and both P25 and P22 protein co-localization with APOBEC3A protein in the cytoplass. Co-IP with APOBEC3A was mediated by the central region of the HBeAg precursor (amino acids 107–178). The levels of HBsAg were significantly lower in cells co-transfected with HBV e-, P22/P25, and APOBEC3A, than in cells co-transfected with HBV e-, APOBEC3A, or in cells co-transfected with HBV e-, P22/P25. The levels of HBeAg were lower in cells co-transfected with HBV e-, P22/P25, and APOBEC3A, than in cells co-transfected with HBV e-, P22/P25. Compare with cells transfected with HBV e-, APOBEC3A, or in cells co-transfected with HBV e-, P25/P22, HBV replication intermediates were lower in the cells co-transfected with HBV e-, P25, APOBEC3A, and significantly lower in the cells with HBV e-, P22, APOBEC3A.

    Conclusions: HBV replication can be inhibited by the interaction of P25/P22 with APOBEC3A protein in the cytoplasm.

  • Article
    Xiaoying Hao, Caihong Gao, Yuanyuan Xiao, Huifeng Jia, Jingjing Yu, Xiaoqing Tian, Yongli Hou
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1371-1380. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.137
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    Background: C-X-C motif chemokine ligand 12 (CXCL12)/C-X-C motif chemokine receptor 4 (CXCR4) and T helper (Th)17/regulatory T (Treg) cells are reportedly involved in the immune response during pregnancy. The aim of this study was to investigate the effect of CXCL12/CXCR4 on Th17/Treg balance in a mouse model of lipopolysaccharide (LPS)-induced abortion.

    Methods: Abortion was induced by injection of LPS and AMD3100 was used to block CXCL12/CXCR4. The fetus absorption rates and spleen index were calculated, and hematoxylin and eosin staining was carried out. Quantitative reverse transcriptase-polymerase chain reaction was employed to determine the expression levels of chemokine ligand 12 (CXCL12), CXC receptor 4 (CXCR4), interleukin 17 (IL-17), IL-6, IL-21, IL-22, IL-2, IL-10, retinoid-related orphan nuclear receptor-γt (RORγt), interferon-γ (IFN-γ), forkhead box P3 (Foxp3), and transforming growth factor-β (TGF-β). Protein levels of CXCL12 and CXCR4 were determined, using western blot analysis. Flow cytometry was applied to quantify IL-17 and Foxp3 expression levels.

    Results: AMD3100 significantly reversed the higher levels of CXCL12 and CXCR4, fetus absorption rates, and spleen indices induced by LPS (p < 0.05). Histopathological examination showed that decidual integrity was damaged and the trophoblasts were thinner after LPS treatment, which were blocked by AMD3100. Immunohistochemistry (IHC) analysis revealed that AMD3100 significantly attenuated Th17 infiltration but significantly promoted Foxp3 expression. AMD3100 significantly downregulated the proportion of Th17 cells and significantly upregulated that of Treg cells in the peripheral blood and spleen. Besides, the expression levels of Th17-related cytokines (IL-17, RORγt, IFN-γ, IL-6, IL-21 and IL-22) were significantly downregulated, while Treg-related cytokines (Foxp3, TGF-β, IL-2 and IL-10) were significantly upregulated by AMD3100.

    Conclusions: The inhibition of C-X-C motif chemokine ligand 12 (CXCL12)/C-X-C motif chemokine receptor 4 (CXCR4) could alleviate the miscarriage in LPS-induced abortion by regulating the balance of Th17/Treg cells in mice. These findings may provide a promising strategy for the clinical management of recurrent miscarriage.

  • Article
    Zelin Xu, Xu Deng, Yuntao Luo, Huzhi Cai, Xinyu Chen
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1381-1391. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.138
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    Objectives: To explore how the mechanism of miR-18a-5p overexpression through the Notch signaling pathway improves cardiac function and inhibits cardiomyocyte apoptosis in chronic heart failure rats.

    Methods: Forty rats were randomly divided into contrast group, negative control (NC) group, high-expression group, and low-expression group by random number table, with ten rats in each group. NC group rats, high-expression group rats, and low-expression group rats were injected intraperitoneally with 2 mg/mL adriamycin solution to establish the rat model of chronic heart failure. The miR-18a-5p overexpression and inhibition lentivirus vector was constructed, and the miR-18a-5p overexpression and inhibition lentivirus suspension were injected into the myocardium of rats in the high-expression group and low-expression group respectively. No treatment was made in the contrast group and NC group. The expression levels of miR-18a-5p were detected in the myocardial tissue of rats and the cardiac function of rats. In addition, the apoptosis of rat cardiomyocytes was observed, and the expression levels of apoptosis-related proteins (Cleaved-caspase-3, Bax, and Bcl-2) were detected in the myocardial tissue of rats. The possible binding sites of miR-18a-5p and Notch2 3′untranslated region (UTR) region were predicted, and the interaction between miR-18a-5p and Notch2 was verified. Finally, the expression levels of Notch signal pathway-related genes and proteins (Notch2, Hes1, and Hes5) were detected in the myocardial tissue of rats.

    Results: In comparison with the NC group, miR-18a-5p expression levels in myocardial tissue of high-expression group rats increased (p < 0.05), and that of low-expression group rats decreased (p < 0.05). In contrast with the NC group and low-expression group, the values of left ventricular end-diastolic dimension (LVEDD) and left ventricular end-systolic dimension (LVESD) in the high-expression group decreased (p < 0.05), and the values of left ventricular ejection fraction (LVEF) and fraction shortening (FS) of the left ventricle in the high-expression group increased (p < 0.05). In comparison with the NC group and low-expression group, myocardial cell apoptosis in the high-expression group decreased (p < 0.05), and the protein expression levels of Cleaved-caspase-3 and Bax in myocardial tissue of high-expression group rats decreased (p < 0.05). Moreover, the Bcl-2 protein expression levels in the myocardial tissue of high-expression group rats increased (p < 0.05). Through prediction and verification, miR-18a-5p might bind to the “GCACCUUA” site of the Notch2 3′UTR region, which could inhibit the activity of wild-type Notch2 3′UTR luciferase reporter plasmid but could not inhibit the activity of mutant Notch2 3′UTR luciferase reporter plasmid. In comparison with the NC group and low-expression group, the messenger ribonucleic acid (mRNA) and protein expression levels of Notch signal pathway-related factors Notch2, Hes1, and Hes5 in myocardial tissue of high-expression group rats were decreased (p < 0.05).

    Conclusions: miR-18a-5p overexpression could improve cardiac function and inhibit cardiomyocyte apoptosis in chronic heart failure rats, which might be related to the regulation of the Notch signal pathway.

  • Article
    Pingping Liu, Chunmiao Chen, Guozhe Zhang, Qi Zhang, Hengyue Yang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1393-1400. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.139
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    Objectives: Rotavirus (RV) is the main pathogen of infantile viral gastroenteritis in the world. The purpose of this study was to explore the mechanism of shikonin inhibiting the replication and activity of RV.

    Methods: Caco-2 cells were treated with shikonin at different concentrations. The inhibitory effect of shikonin on RV proliferation in vitro was estimated by CytoPathic Effect (CPE) inhibition assay and virus production reduction assay. An autophagy flow detection kit was used to detect the autophagy of different subgroups of cells. The levels of oxidative stress indexes such as reactive oxygen species (ROS), malondialdehyde (MDA), superoxide dismutase (SOD) and glutathione (GSH) were estimated. The levels of the Sirt1/FoxO1/Rab7 pathway proteins and autophagy proteins Beclin-1 and LC3II/LC3I were estimated by western blot. Caco-2 cells were treated with Nicotinamide (100 µg/mL), and the role of Shikonin in inhibiting RV was confirmed by salvage experiment.

    Results: When the concentration of Shikonin was less than 100 µM, there was no obvious cytotoxicity. Shikonin therapy induced dose-dependent inhibition of SA11 and WA virus replication in Caco-2 cells. When Caco-2 cells were infected with RV with different concentrations of the multiplicity of infection (MOI), shikonin produced a similar inhibitory effect on RV. The number of autophagy bodies and autophagy proteins Beclin-1 and LC3II/LC3I decreased notably in shikonin-treated cells. Shikonin can alleviate oxidative stress induced by RV and inhibit RV replication through the Sirt1/FoxO1/Rab7 pathway.

    Conclusions: Shikonin can inhibit the replication and amplification of RV, which may be related to the inhibition of the Sirt1/FoxO1/Rab7 pathway and the regulation of autophagy and oxidative stress.

  • Article
    Kaining Jia, Hongyan Zhang, Jiawei Wang, Qiang Lin
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1401-1413. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.140
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    Background: The pathogenesis of neuropathic pain (NP) remains a mystery, and no ideal treatment methods or prevention measures exist. Our goal was to identify potential therapeutic targets for NP using the RNA sequencing (RNA-Seq) dataset GSE126611 from the Gene Expression Omnibus (GEO) database and experimentally validate the potential target.

    Methods: Dataset GSE126611 was used for screening differentially expressed genes (DEGs) and weighted gene co-expression network analysis (WGCNA). KEGG (Kyoto Encyclopedia of Genes and Genomes) and GO (Gene Ontology) enrichment analyses were performed on the DEGs and key modules. Tryptase delta 1 (TPSD1) was screened out for experimental validation. A chronic constriction injury (CCI) model of a rat was used to induce NP. The levels of mechanical nociceptive threshold (MNT) and thermal pain threshold (TPT) of rats were measured on days 1, 7 and 14 after modeling. On day 14, levels of interleukin (IL)-1β, tumor necrosis factor (TNF)-α, IL-6, IL-10, and transforming growth factor (TGF)-β in serum were measured by enzyme-linked immunosorbent assay (ELISA). Expression levels of TPSD1 in the meningeal tissue, p65 and NF-κB (nuclear factor kappa-B) inhibitor alpha (IκBα) in the spinal dorsal horn tissue were measured by quantitative PCR (qPCR) and Western blot. Immunofluorescence measured the expression of ionized calcium-binding adapter protein 1 (Iba1) in the spinal dorsal horn tissue. TPSD1 and NF-κB inhibitor BAY 11-7082 were used to induce microglia. Cell supernatant was collected for IL-1β, TNF-α, IL-6, IL-10, and TGF-β detection. Cells were collected for p65, phosphorylated p65 (p-p65), IκBα, phosphorylated IκBα (p-IκBα), Iba1, CD86, and CD206 expression levels detection.

    Results: In vivo results showed that compared with the sham group, the levels of MNT, TPT, TGF-β, IL-10, IκBα, and TPSD1 in the CCI group were notably down-regulated, whereas the levels of IL-1β, TNF-α, IL-6, p-IκBα, and Iba1 were obviously up-regulated. Besides, we cultured microglia with TPSD1, and results showed that compared to the conventional culture microglia, the levels of TGF-β, IL-10, p-p65, p-IκBα, and M1 marker CD86 were significantly decreased, while the levels of IL-1β, TNF-α, IL-6, IκBα, and M2 marker CD206 were significantly increased.

    Conclusions: The TPSD1 gene induces the activation of the NF-κB signaling pathway, which activates microglia and polarizes them toward the M1 phenotype, leading to the secretion of pro-inflammatory factors and causing pain. The findings suggest that the management of neuropathic pain could be improved by targeting TPSD1.

  • Article
    Feng Wang, Yajun Gu, Xiaohui Shen, Peng Wang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1415-1425. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.141
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    Objective: To characterize oral squamous cell carcinoma (OSCC) subtypes and prognostic models with early region 2 binding factor (E2F) families.

    Methods: Expression and clinical information of OSCC and normal samples were downloaded from the Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases as testing and validation sets, respectively. Based on E2Fs, unsupervised cluster analysis was used to classify the different subtypes of OSCC. We then screened for genes that were differentially expressed (DEGs) between tumor and normal tissues and used weighted gene co-expression network analysis (WGCNA) to screen for genes in the modules most significantly associated with prognosis. DEGs that were significantly associated with prognosis were used to construct a risk model. Finally, a nomogram combining clinically independent factors was constructed.

    Results: OSCC samples were divided into two subtypes. There was a significant difference in prognosis between the two subtypes of patients (p < 0.05). Twenty-five DEGs that were significantly associated with prognosis were screened from the 153 subtype-related genes. Subsequently, five genes were used to construct a risk model using the least absolute shrinkage and selection operator (LASSO) method. In both the TCGA and GEO datasets, it was found that the prognosis of patients in the high-risk group was significantly worse than that in the low-risk group (p < 0.05). Age, N stage, and risk group were independent prognostic factors for OSCC and were used to construct the nomogram, which had a good prognostic performance.

    Conclusions: The risk model based on E2F-related subtypes and DEGs showed good predictive performance for the prognosis of patients with OSCC. Nomograms constructed using independent prognostic factors, including risk score, may play an important role in personalizing prognosis and treatment in the future.

  • Article
    Zemin Ding, Xia Hong, Xiaohui Li, Yueyue Wang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1427-1441. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.142
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    Background: This study aims to explore the active components of Panax Ginseng (PG)-Schisandrae Chinensis Fructus (SC) and reveal the mechanism of improving neurodegenerative diseases (NDD) based on network pharmacology and molecular docking.

    Methods: Detailed information about the PG-SC herbal pair was obtained from traditional Chinese medicine systems pharmacology (TCMSP) and screened using OB (oral bioavailability) ≥30% and DL (drug-likeness) ≥0.18 as criteria. Genes were collected using OMIM, Genecards, and PharmGKB (Pharmacogenetics and Pharmacogenomics Knowledge Base) methods. The string database was utilized to obtain protein-protein interaction (PPI) data. The core targets were entered into the “Data Source” of the eFP (electronic Fluorescent Pictograph) browser (http://bar.utoronto.ca/efp_human/) for organ localization and created a target-organ localization map. We constructed and analyzed the network using Cytoscape 3.9.1 (National Institute of General Medical Sciences, Bethesda, MD, USA). Encyclopedia of genomes (KEGG) pathway-enriched targets were used for analysis. After identifying the core network, a protein-protein interaction (PPI) network of PG-SC targets was constructed. Autodock Vina and PyMOL software were used to validate the results for its active ingredients and key targets.

    Results: There were 14 active ingredients in Ginseng and 8 in Schisandra, with 8320 corresponding protein targets and 80 common targets with neurodegenerative diseases (NDD), of which 21 highly abundant targets were highly expressed in the brain. Six core targets were identified, including AKT1 (RAC-alpha serine/threonine-protein kinase), CASP3 (Caspase 3), MAPK8 (Mitogen-Activated Protein Kinase 8), 1L1B (Interleukin 1 Beta), JUN (Jun Proto-Oncogene, AP-1 Transcription Factor Subunit), and PTGS2 (Prostaglandin-Endoperoxide Synthase 2). The 142 KEGG signaling pathways were enriched, including the pathways of neurodegeneration-multiple diseases and neuroactive ligand-receptor interaction. Molecular docking confirmed that the core targets (AKT1, CASP3, MAPK8, 1L1B, JUN, PTGS2) could bind to the active compounds (Kaempferol, Beta_sitosterol, Ginsenoside Rh2, Stigmasterol, Inermin, Dianthramine, Arachidonate, Frutinone A, Girinimbin, Suchilactone, Fumarine).

    Conclusions: Based on network pharmacology and molecular docking, the PG-SC herbal pair could act on NDD through neuronal programmed cell death targets and related signaling pathways. Our findings provide the basis for further research using herbs to treat NDD and develop anti-NDD drugs.

  • Article
    Chunyuan Hao, Tuowei Zhang, Tonghua Li, Weihua Gao, Yang Ren, Junzhe Wang, Yuyang Zhang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1443-1451. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.143
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    Purpose: Leptin was reported to be associated with and is considered a risk factor for acute myocardial infarction (AMI). The aim of this study was to investigate the role and related regulation mechanism of Leptin in the myocardial infarction induced dysfunctions.

    Methods: MI (myocardial infarction) rat model was established to explore the role of Leptin on myocardial functions. Eighteen rats are divided into three group, including control rat, Leptin rat and MI rat. Echocardiography was performed to check left ventricular function. The hypoxic cell model was established, using H9C2 cardiomyocytes cell, to investigate the related controlling mechanism of Leptin. The miR-27a/b-3p expression in MI-rats was measured via quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). The interaction between miR-27a/b-3p and mitochondrial fission factor (MFF) was checked for using double luciferase reporter gene. The glucose consumption and the lactic acid production in hypoxic cell model were utilized to estimate the cellular energy metabolism.

    Results: Leptin expression was significantly increased and miR-27a/b-3p expression was significantly decreased in MI-rats (p < 0.01). Leptin knockdown significantly alleviated the myocardial dysfunction of MI-rats (p < 0.01). miR-27a/b-3p expression was significantly downregulated in the hypoxic cell model and the cardiomyocytes of MI-rats (p < 0.01). MFF was proved to be a downstream target of miR-27a/b-3p, and its overexpression reversed the influence of miR-27a/b-3p on the lactic acid production and the cellular energy metabolism of the hypoxic cell model. Moreover, miR-27a/b-3p significantly elevated the abundances of mitochondrial proteins optic atrophy (OPA1), mitofusin-1 (MFN1) and mitofusin-2 (MFN2) (p < 0.01), and decreased the levels of proteins hexokinase 2 (HK2), lactate dehydrogenase A (LDHA) and phosphorylated Akt (p-AKT), and MFF upregulation could significantly reverse these phenomena.

    Conclusions: Leptin participates in ventricular remodeling and the mitophagy of cardiomyocytes, by regulating miR-27a/b-3p/mitochondrial fission factor (MFF)/optic atrophy 1 protein (OPA1) axis.

  • Article
    Yue Zhang, Hongliang Sun, Hailing Chong, Qingfeng Li, Mingming Tian, Jianxi Jiang, Chunjiao Liu, Qingchun Li
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1453-1462. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.144
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    Background: Aurora B is one of the spindle assembly checkpoint proteins. It plays a crucial role in the regulation of mammalian female mitosis and meiosis. In this study, we examined the mechanism of small ubiquitin-related modifier modification (SUMOylation) of Aurora B on the chromosome arrangement of mice oocytes.

    Methods: Oocytes were randomly divided into three groups: Control group, AuroraB_K207R group, and the small ubiquitin-related modifier (SUMO) inhibitor group. The number of oocytes that developed at 2.5, 8, and 14 h were counted. Metaphase II (MII) oocytes in each group were selected for observing the arrangement of chromosomes and spindles. Meanwhile, MII oocytes in the control and AuroraB_K207R groups were collected for use in the Smart-seq2 protocol.

    Results: Messenger ribonucleic acid (mRNA) was obtained from the AuroraB_K207R group via the transcription of a linearized pMD 18T-T7-AuroraB_K207R plasmid. The proportion of metaphase I (MI) oocytes after 8 h in the SUMO inhibitor group was significantly lower than in the AuroraB_K207R and control groups (p < 0.05). The MII oocytes in the AuroraB_K207R and SUMO inhibitor groups decreased significantly compared with the control group after 14 h (p < 0.05). Furthermore, the rate of abnormal chromosome arrangement in the AuroraB_K207R and SUMO inhibitor groups increased significantly compared with the control group (p < 0.05). Using Smart-seq2, a total of 3288 differentially expressed genes were identified; 2874 of these genes involving Aurora B were upregulated and 414 genes were downregulated in the AuroraB_K207R group.

    Conclusions: Small ubiquitin-related modifier modifications may directly affect the regulatory function of Aurora B on chromosome arrangement in mice oocyte meiosis, leading to oocyte maturation disorder. Therefore, the SUMOylation of Aurora B plays an important role in chromosome arrangement in mice oocyte meiosis.

  • Article
    Bo Liu, Zheng-ya Yu, Xiao Chen, Meng Lian, Jia-ming Chen, Xuan Li, Fang-Jian Shang, Ju-gao Fang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1463-1472. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.145
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    Objective: Tumor microenvironment (TME) is complex, with crosstalk between cancer cells and other cells contributing to cancer development. FGD5-AS1 is an oncogenic long noncoding ribonucleic acid (lncRNA) derived from exosome in cancer cells. It was shown that cancer cells can regulate macrophage polarization to a state favorable to cancer development. The aim of this study was to explore the role of exosome-derived FGD5-AS1 in the interaction between thyroid cancer (TC) cells and macrophages, and provide new insights into the mechanisms underlying TC development.

    Methods: Macrophage M2 polarization in the collected clinical TC cell tissues was observed through flow cytometry. To examine the effect of TC cells on macrophage polarization, macrophages were co-incubated with SW1736 cells and then exosome secretion inhibitor (GW4869) was added. To observe the role of FGD5-AS1 in SW1736-derived exosomes, SW1736 cells were transfected with FGD5-AS1 overexpression vector, and SW1736-derived exosomes (SW1736FGD5-AS1-Exos) were later isolated, followed by a treatment of macrophages. The number of macrophages expressing cluster of differentiation 163 (CD163) under different treatment was detected with flow cytometry, and the expression of Arginase 1 (Arg-1), cluster of differentiation 206 (CD206), interleukin 10 (IL-10) and FGD5-AS1 was measured using quantitative reverse transcription-polymerase chain reaction (qRT-PCR). After DiO labeling of exosomes (SW1736-Exo), the endocytosis of exosomes in macrophages were observed. In order to explore the effect of macrophages on SW1736 cells, the medium of macrophages treated by different exosomes was incubated with SW1736 cells. Finally, the migration and invasion of SW1736 cells were observed by scratch assay and Transwell.

    Results: TC cell tissues appeared to be infiltrated by significantly larger number of M2 macrophages, compared with paracancerous tissues (p < 0.05). When macrophages were co-incubated with SW1736 cells, a significant increase in macrophage M2 polarization was observed, but the increase was significantly inhibited by GW4869. SW1736-Exo could be endocytosed by macrophages and significantly promote macrophage M2 polarization, while SW1736FGD5-AS1-Exos was able to significantly enhance the promotion (p < 0.05). Macrophages treated with SW1736FGD5-AS1-Exos significantly increased the migration and invasion ability of SW1736 cells (p < 0.05).

    Conclusions: FGD5-AS1 carried by TC cell-derived exosomes induces macrophage M2 polarization, while M2 macrophages promote migration and invasion of TC cells.

  • Article
    Siyuan Cheng, Hongfan Chen, Qin Fang, Yuhao Wang, Shanxin Liu
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1473-1484. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.146
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    Background: Our preliminary research has suggested that hydroxysafflor yellow A (HSYA) has an alleviative effect on myocardial ischemia/reperfusion (I/R) injury. Here, we investigate the possible mechanism implicated.

    Methods: A hypoxia/reoxygenation (H/R) model, which mimicked the I/R model in vitro, was constructed in rat cardiomyocytes H9c2. Following the intervention with HSYA and transfection, the viability and apoptosis of cardiomyocytes were evaluated by the cell counting kit-8 (CCK-8) and flow cytometry assays. The mitochondrial membrane potential (∆Ψm) was determined with JC-1 analysis with both a confocal microscope and flow cytometry. The content of adenosine triphosphate (ATP) was quantified with a commercial kit. Reverse-transcription quantitative PCR (RT-qPCR) and western blot were employed to quantify the levels of hypoxia inducible factor-1α/BCL2 Interacting Protein 3 (HIF-1α/BNIP3) pathway-, mitochondrial fission- and mitophagy-related factors.

    Results: HSYA enhanced the viability, prevented apoptosis, restored the ∆Ψm loss, and promoted the ATP content in H/R-treated cardiomyocytes. HSYA inhibited the expressions of mitochondrial fission factor (Mff), Fission, Mitochondrial 1 (Fis1), and P62 yet promoted those of HIF-1α, BNIP3, and Beclin1 and increased LC3II/LC3I ratio in H/R-treated cardiomyocytes. BNIP3 silencing was associated with the suppression of viability and ATP levels, and the promotion of apoptosis and ∆Ψm loss in H/R-modeled cardiomyocytes, in addition to the observation that the modulatory effects of HSYA on both factors related to mitochondrial fission- and HIF-1α/BNIP3 pathway-mediated mitophagy were reversed after BNIP3 silencing.

    Conclusions: HSYA exerts a preventive effect on H/R-modeled cardiomyocytes by enhancing the HIF-1α/BNIP3 signaling pathway-mediated mitophagy.

  • Article
    Jia Liu, Zhengkun Wang, Yifei Chen, Lan Chen, Xiu Liu, Lingyun Li, Ning Bao
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1485-1492. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.147
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    Background: Hysteroscopic surgical dilatation fluid overload is a serious complication that can lead to circulatory overload, increased extravascular lung water (EVLW) and even acute heart failure pulmonary edema. Because bipolar hysteroscopy uses electrolyte fluid as dilatation fluid, we cannot determine dilatation fluid overload by the traditional method of blood sodium ion concentration, and currently we urgently need a method that can quickly diagnose distended fluid overload during hysteroscopy early to prevent operative hysteroscopy intravascular absorption (OHIA).

    Objective: To construct a predictive model of fluid overload caused by bipolar hysteroscopic procedures monitored by bedside cardiac ultrasound and evaluate its predictive ability. Screening bedside cardiac ultrasound indicators to advance the diagnosis of fluid overload-induced increased EVLW.

    Methods: 110 patients undergoing bipolar electric knife hysteroscopic release of uterine adhesions were selected. Cardiac ultrasound indices included inferior vena cava collapse rate (IVC-CI), cardiac output (CO), left ventricular ejection fraction (LVEF), E-peak to A-peak ratio (E/A), the ratio of peak mitral valve early diastolic flow velocity (E) on pulsed Doppler ultrasound to peak mitral annular early diastolic motion velocity (e’) on tissue Doppler ultrasound (E/e’), mitral annular systolic displacement (MAPSE), and tricuspid annular systolic displacement (TAPSE). Mann-Whitney U non-parametric test was used to screen the significant single-factor indicators mentioned above for multi-factor logistic regression analysis, identify the independent influencing factors indicators, establish a prediction model, draw the ROC (receiver operating characteristic) curves, and evaluate predictive ability of the prediction model.

    Results: Multifactor logistic regression analysis showed that IVC-CI and E/A were independent risk factors for increased lung water in bipolar hysteroscopic surgery. The expression of the joint detection factor model was obtained as Logit (p) (y = lung water increase) = 3.893 – 0.081 × ICV-CI – 5.839 × E/A. The area under the ROC curve of the constructed joint prediction model was 0.938 (95% CI (confidence interval): 0.896–0.980), with a sensitivity of 84.6% and a specificity of 94.8%.

    Conclusions: The combined predictive model of cardiac ultrasound indicators constructed based on logistic regression analysis is of high diagnostic value for early detection of complications of fluid overload leading to increased EVLW during bipolar hysteroscopy.

  • Article
    Chen Chen, Hong-ying Wang, Cheng-yu Chu, Wei Zhang, Yi-ting Jin, Qiang Zou
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1493-1500. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.148
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    Background: Hypoxia induces microenvironmental changes in breast cancer and other solid tumors. Tumor cells that survive hypoxia are claimed to acquire more aggressive properties through a cascade of activated signaling pathways. Here in this study, we focus on Rab11a protein function during hypoxia and its role in breast cancer migration.

    Methods: MDA-MB-468 and T47D cell lines were used in this study. Rab11a was overexpressed or suppressed by lentivirus transfection. Cobalt chloride (CoCl2) was used to mimic a hypoxic environment. Transwell assays were performed to assess the role of Rab11a in tumor cell migration during hypoxic stimulation. The effect of hypoxia on Rab11a and epithelial-mesenchymal transition (EMT) related proteins were determined by western blot assay. Immunohistochemistry was used to determine Rab11a expression levels in breast cancer patients with axillary lymph node metastases.

    Results: Hypoxia-inducible factor-1α (HIF-1α) was upregulated in T47D and MDA-MB-468 cells during CoCl2-hypoxia stimulation while Rab11a downregulation was only detected in MDA-MB-468 cells with an increased migration ability. Rab11a knockdown promoted MDA-MB-468 cells migration with a decreased E-cadherin and increased β-catenin, Vimentin and N-cadherin in hypoxic environments. In breast cancer patients, Rab11a levels were down-regulated in the metastatic axillary lymph nodes compared to the primary tumor site.

    Conclusions: Our study is the first to show Rab11a downregulation in triple negative breast cancer in the presence of hypoxia and raises the question of a paradoxical role of Rab11a in tumor progression.

  • Article
    Ying Zhang, Ning Zhang, Jitian Li, Qiushi Wei, Qiang Yuan, Wei He, Yiping Dong, Zhenhao Jing, Xiangyang Cao, Youwen Liu, Leilei Zhang, Haibin Wang, Wuyin Li
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1501-1509. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.149
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    Background: We aimed to explore the molecular mechanisms underlying the effect of Gugutouhuaisiyu Capsule (GGTHSYC) on traumatic osteonecrosis of the femoral head (ONFH) using rabbit models.

    Methods: New Zealand rabbits (n = 24) were randomly placed into the control, model, and 30 and 60 mg/kg GGTHSYC treatment groups. Animals in the model and GGTHSYC treatment groups were subjected to surgically induced ONFH. Rabbits in the 30 and 60 mg/kg GGTHSYC treatment groups were administered GGTHSYC via the gastrointestinal route. MRI (magnetic resonance imaging) was employed to assess osteonecrosis. The mRNA and protein expression levels of BMP-2 (bone morphogenetic protein 2), RUNX2 (runt related transcription factor 2), VEGF (vascular endothelial growth factor), HIF-1α (hypoxic-induced factor-1α), and OPG (osteoprotegerin) in femoral head tissues were measured using reverse transcription-quantitative polymerase chain reaction and western blotting and immunochemistry, respectively.

    Results: GGTHSYC significantly enhanced osteocalcin activities in rabbits with traumatic ONFH by increasing ALP (alkaline phosphatase) expression (p = 0.008). Administration of GGTHSYC also significantly increased the mRNA and protein levels of HIF-1, VEGF, RUNX2, BMP-2, OPG, and BSP (bone sialoprotein) (p < 0.05).

    Conclusions: Administration of GGTHSYC improved bone regeneration following traumatic ONFH, which was accompanied by increased expression of the transcription factors HIF-1α and RUNX2 as well as other related proteins, suggesting that the transcription factors HIF-1α and RUNX2 may be a potential target for the treatment of traumatic ONFH.

  • Article
    Fan Zhang, Long Li, Wen-tao Huang, Jun-hua Liao, Ye-jun Huang, Jing-xin Huang, Yu-hui Zeng
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1511-1519. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.150
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    Background: Septic shock, the most severe form of sepsis syndrome, is life-threatening and requires immediate medical attention. However, due to its poorly understood mechanism, there is no effective treatment for septic shock. Serum exosomes are important mediators of systemic inflammation due to septic shock, and we sought to uncover key signaling molecules in mediation.

    Methods: Serum exosomes from patients with septic shock (SS-exo) and healthy controls (Normal-exo) were isolated and characterized by western blot, nanoparticle tracking assay (NTA) and transmission electron microscopy (TEM). The long noncoding RNA (lncRNA) actin filament associated protein 1-antisense RNA 1 (AFAP1-AS1) expression levels in exosomes and THP-1 (the human leukemic cell line) cells co-incubated with the exosomes were measured by real-time quantitative reverse transcription PCR (polymerase chain reaction) (qRT-PCR). THP-1 cells were divided into lipopolysaccharide (LPS, 100 ng/mL) group, SS-exo group, SS-exo-shRNA group, and SS-exo-shRNA-AFAP1-AS1 group. The enzyme-linked immunosorbent assay (ELISA) was employed to examine the inflammatory cytokine interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α) expression in each group. Flow cytometry was applied to examine cell surface antigens CD86 (cluster of differentiation 86) and CD206 expression in each group. In addition, qRT-PCR was conducted to determine IL-6, inducible nitric oxide synthase (iNOS), Arginase I (Arg1), and interleukin-10 (IL-10) expression levels in each group, and western blotting was employed to examine p-ERK1 (p-extracellular regulated protein kinases 1), ERK1/2, p38 MAPK (mitogen-activated protein kinase), and p-p38 MAPK protein expression levels in each group.

    Results: We successfully isolated and characterized SS-exo. The AFAP1-AS1 content was significantly increased in SS-exo. AFAP1-AS1 could be transmitted to THP-1 cells and stimulate THP-1 cell polarization to pro-inflammatory macrophages (M1 macrophages), thereby promoting the production of inflammatory factors IL-6 and TNF-α. Western blot revealed that AFAP1-AS1 significantly activated the MAPK signaling pathway.

    Conclusions: AFAP1-AS1, carried by SS-exo, excites THP-1 cell polarization in M1 macrophages and promotes an inflammatory response by activating the MAPK signaling pathway.

  • Article
    Bo Hou, Bing Shen, Wei-Yong Zhang, Shi-bin Zhang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1521-1527. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.151
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    Objective: This study aimed to assess the diagnostic value of combined detection of computed tomography (CT), serum homocysteine (Hcy), cystatin C (Cyst-C), and uric acid (UA) concentrations for hypertensive left ventricular hypertrophy.

    Methods: The present study matched 50 patients with primary hypertension without left ventricular hypertrophy (control group) with 50 patients with primary hypertension with left ventricular hypertrophy (HLVH group) from March 2019 to May 2020. The serum Hcy, Cyst-C, UA levels, IVST (interventricular septum thickness), LVDD (left ventricular end-diastolic diameter), LVPWT (left ventricular posterior wall thickness), LVEF (left ventricular ejection fraction), LVFS (left ventricular fraction shortening), and blood lipid levels were compared between the two groups. The specificity, sensitivity, and accuracy of CT combined with serum Hcy, Cyst-C, and UA levels were compared between the two groups, and the correlation of serum Hcy, Cyst-C, and UA levels with HLVH was analyzed.

    Results: Patients with left ventricular hypertrophy showed poorer outcomes of serum Hcy, Cyst-C, UA levels, IVST, LVDD, LVPWT, LVEF, LVFS, and blood lipid levels versus those without (p < 0.05). Combined detection was associated with higher specificity, sensitivity, and accuracy versus single detection (p < 0.05). Serum Hcy, Cyst-C, and UA levels were positively correlated with hypertensive left ventricular hypertrophy (p < 0.05).

    Conclusions: Hybrid detection of CT and serum Hcy, Cyst-C, and UA levels substantially improve the diagnostic accuracy for hypertensive left ventricular hypertrophy.

  • Article
    Andrea Mazzatenta, Susi Zara, Sara Invitto, Andrea Porzionato, Amelia Cataldi, Camillo Di Giulio, Mieczyslaw Pokorski
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1529-1535. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.152
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    Background: Transduction mechanisms of the hypoxic chemoreflex elicited by carotid body (CB) chemoreceptor cells remain unclear. Recent studies direct attention to the plausible link between CB and olfactory chemoreceptor functions.

    Methods: Here we used immunohistochemistry to investigate the distribution and localization of olfactory marker protein (OMP) in human CB. Carotid bodies were collected post-mortem from hospital patients aged 27–76 years who died from reasons unrelated to chronic pulmonary or cardiovascular disorders. We used specific antibodies to selectively identify CB cells and OMP in tissue sections. The binding of antibodies to target antigens was visualized with the Ultra Vision detection system.

    Results: We show that OMP is abundantly present in the cytoplasm of CB chemoreceptor cells. The presence of OMP in these cells indicates that the olfactory system may participate in shaping the chemosensory CB function.

    Conclusions: The findings support the notion that the transduction mechanisms of chemoreceptive systems contain a degree of homology, irrespective of the anatomical localization and the functional role these systems fulfill. The ectopic presence of OMP in CB broadens the current understanding of the mechanisms underlying chemosensory responses.

  • Article
    Wei Lv, Fei Jing, Chong Geng, Na Chen
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1537-1546. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.153
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    Background: MicroRNAs (miRNA) are biomarkers and potential therapeutic targets for autoimmune diseases, including Hashimoto’s thyroiditis (HT). However, there are limited studies to evaluate the mechanism of microRNAs associated with HT. This study explores the mechanism of action and anti-inflammation activity of miR-216a-5p in thyroid follicular epithelial cells.

    Methods: In this research, 100 ng/mL LPS (lipopolysaccharide) was used to induce Nthy-ori 3-1 cells to simulate HT in vitro, and then the cell activity and cytotoxicity were measured using CCK-8 (Cell Counting Kit-8) and flow cytometry. The KIAA0101 and miR-216a-5p expressions in the thyroid follicular epithelial cells and HT patients were measured by quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR) and western blot assay. Luciferase activity was used to characterize the interaction between miR-216a-5p and KIAA010. In addition, the ability of thyroid follicular epithelial cells to secrete inflammatory factors was measured using an enzyme-linked immunosorbent assay (ELISA) kit.

    Results: The expression of miR-216a-5p was abnormally down-regulated in the lipopolysaccharide (LPS)-induced thyroid follicular epithelial cells and the thyroid gland of HT patients. Down-regulation of miR-216a-5p severely inhibited the proliferation activity of cells, induced apoptosis, and enhanced the inflammatory response. In addition, LPS significantly up-regulated KIAA0101 expression (p < 0.05) in HT patients. miR-216a-5p overexpression substantially inhibited KIAA0101 expression due to the existence of binding sites between miR-216a-5p and KIAA0101, enhanced cell proliferation, inhibited apoptosis and exerted anti-inflammatory activity. Furthermore, its overexpression significantly reduced the level of inflammatory factors such as tumor necrosis factor-alpha (TNF-α) and interleukin 1 beta (IL-1β) (p < 0.05).

    Conclusions: miR-216a-5p could bind with KIAA0101 and regulate KIAA0101 to achieve effective therapy for Hashimoto’s thyroiditis.

  • Article
    Haibo Wang, Guohua Zeng, Xianghong Yang, Bangchuan Hu, Yan Shen, Jinsong Shen
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1547-1559. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.154
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    Background: The involvement of long non-coding RNA (lncRNA) metastasis associated lung adenocarcinoma transcript 1 (MALAT1) in inflammation-related kidney damage has been widely demonstrated. However, the mechanisms by which MALAT1 regulate glomerular endothelial cells in sepsis remain undefined, which is the subject in this study.

    Methods: MALAT1-targeted gene was predicted, and verified through Starbase and dual-luciferase reporter experiment. ShMALAT1 and miR-126-5p inhibitors were transfected into lipopolysaccharide (LPS)-induced human renal glomerular endothelial cells (HRGECs) and murine glomerular endothelial cells (MGECs). The viability, reactive oxygen species (ROS) content, apoptosis and expressions of inflammatory cytokines of HRGECs and MGECs were examined by cell counting kit-8, immunofluorescence assay, flow cytometry and enzyme-linked immunosorbent assay (ELISA), respectively. Expressions of vascular cellular adhesion molecule 1 (VCAM1), inter-cellular adhesion molecule-1 (ICAM-1), monocyte-chemoattractant protein-1 (MCP-1), Nuclear P65 and Cytoplasmic P65 were analyzed by Western blotting.

    Results: MALAT1 silencing promoted viability and Cytoplasmic P65 expression, but inhibited ROS content, apoptosis, inflammatory cytokine (TNF-α (tumor necrosis factor-alpha) and IL-6 (interleukin 6)) levels, and VCAM1, ICAM-1, MCP-1 and Nuclear P65 expressions in LPS-induced HRGECs and MGECs. MiR-126-5p had a targeted binding relationship with MALAT1 in HRGECs, and MALAT1 silencing increased miR-126-5p expression. MiR-126-5p inhibitor had the opposite effect to MALAT1 silencing, and reversed the effect of MALAT1 silencing on viability, apoptosis and inflammatory cytokines in LPS-induced HRGECs and MGECs. Moreover, miR-126-5p inhibitor increased VCAM1, ICAM-1, MCP-1 and Nuclear P65 expressions, but reduced that of Cytoplasmic P65 in LPS-induced HRGECs and MGECs, and partially offset the effect of shMALAT1 on the above protein expressions.

    Conclusions: MALAT1 silencing reduces LPS-induced glomerular endothelial cell damage by promoting miR-126-5p expression.

  • Article
    Xuemei Liu, Ze Zhang, Huanayi Yekeben, Zulikaier Tuerxun, Yumaierjiang Balati, Pengfei Gong, Zhen Bao, Yuchun Yang, Pengyi He, Muhuyati
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1561-1571. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.155
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    Background: This study aimed to evaluate the correlation between the level of periostin (POSTN) in plasma and acute myocardial infarction (AMI).

    Methods: Patients with a definite diagnosis of AMI were included in the present study, and normal controls were established. The levels of POSTN in the peripheral blood leukocytes (WBCs) of patients with AMI within 24 h of infarction were measured by polymerase chain reaction. The plasma levels of POSTN within 24 h, on day 7, and on day 30 after AMI were measured by enzyme-linked immunosorbent assay, and the data were statistically analyzed.

    Results: The expressions of POSTN in plasma and WBCs were significantly lower in patients with AMI within 24 h. The plasma levels of POSTN in patients with AMI were the lowest at 24 h after infarction; They increased significantly on day 7 and approached normal levels on day 30. The most rapid elevation of POSTN occurred within 7 d. When the POSTN level was 302.3 ng/mL, the sensitivity and specificity for the diagnosis of AMI were 73% and 85%, respectively.

    Conclusions: The plasma level of POSTN was significantly lower during the early stage of AMI, and it changed with the development of the infarction. The plasma level of POSTN within 24 h might be of diagnostic value for AMI.

  • Article
    Dong Luo, Yunfei Liu, Zhiqiang Li, Liandong Ji, Xuejun Gong, Xiao Yu
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1573-1587. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.156
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    Background: Pancreatic ductal adenocarcinoma (PDAC) is a solid malignancy with adverse outcomes. Our aim was to explore promising critical genes related to the development of PDAC.

    Methods: Microarray datasets were downloaded, and differentially expressed genes (DEGs) were identified. Gene Ontology (GO) function and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were conducted by the Database for Annotation, Visualization and Integrated Discovery (DAVID, version 6.8, national cancer institute, Frederick, MD, USA). A protein-protein interaction (PPI) network of DEGs was built with the Search Tool for the Retrieval of Interacting Genes (STRING) (version 11.0, Global Biodata Coalition and ELIXIR, Zurich, Switzerland, http://string-db.org/), and hub genes were determined using Cytoscape (Version 3.7.2, UC San Francisco & Gladstone Institute, San Diego, CA, USA, http://www.cytoscape.org/). Then, expression of hub genes was confirmed by querying Gene Expression Profiling Interactive Analysis (GEPIA) and Human Protein Atlas (HPA) (version 22.0, the Knut & Alice Wallenberg foundation, Wallenberg, Sweden, https://www.proteinatlas.org/) databases. These results were further confirmed by quantitative real time PCR (qRT-PCR) of PDAC cell lines. The prognostic significance of hub genes was evaluated through Kaplan–Meier plotter, and the diagnostic value was assessed using UCSC (University of California Santa Cruz) Xena.

    Results: Two downregulated DEGs and 196 upregulated DEGs were identified. These overexpressed genes were primarily enriched in extracellular exosomes, calcium ion binding, cell adhesion, the Phosphatidylinositol 3 kinase (PI3K)/protein kinase B (PKB, also known as AKT) signaling pathway and cancer. 10 highly expressed hub genes, namely Cyclin-Dependent Kinase 1(CDK1), Ribonucleotide Reductase Regulatory Subunit M2 (RRM2), DNA Topoisomerase II Alpha (TOP2A), Abnormal Spindle-Like Microcephaly-Associated Protein (ASPM), Protein Regulator of Cytokinesis 1 (PRC1), ZW10 Interacting Kinetochore Protein (ZWINT), Maternal Embryonic Leucine Zipper Kinas (MELK), Centrosomal Protein 55 (CEP55), Denticleless E3 Ubiquitin Protein Ligase Homolog (DTL), and NIMA Related Kinase 2 (NEK2), were determined, and the increased expression of these screened genes was further confirmed in PDAC cells compared with immortalized human pancreatic ductal epithelial cells (HPDE6). Increased expression of the selected genes had shorter overall survival (OS) and relapse-free survival (RFS) in PDAC. The analyses of receiver operating characteristic (ROC) predicted the area under curve (AUC) of hub genes in PDAC ranged from 0.970 to 0.990.

    Conclusions: In conclusion, our study suggested that upregulation of CDK1, RRM2, TOP2A, ASPM, PRC1, ZWINT, MELK, CEP55, DTL, and NEK2 is associated with poor OS and RFS, and these genes exhibit high diagnostic value in PDAC. These genes may be potential independent diagnostic markers. Further studies are needed to validate their potential diagnostic and therapeutic values in PDAC.

  • Article
    Jun Fu, Fang Wang, Siyuan Ha
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1589-1595. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.157
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    Background: A large proportion of cancer patients suffer from devastating neuropathic pain. Vincristine (Vin), a first-line anticancer therapy, usually induces this pain, which negatively affects the life quality of patients.

    Purpose: To explore propentofylline’s effect on Vin-induced neuropathic pain and its potential mechanism of action.

    Methods: A Vin-induced rat model was used in this study. Vin (125 µg/kg) was administered intraperitoneally every other day for 8 days (days 1~8), to establish neuropathic pain model in rats. Then, the rats were randomized into the model group and three treatment groups (low-, mid- and high-dosage groups). On days 8~14, the three treatment groups received daily intraperitoneal injections of propentofylline (PPF) at 6.25, 12.5, and 25 mg/kg/day, while all groups received equal volumes of physiological saline. Behavioral tests were conducted, to check pain hypersensitivity on days 0, 8, 10, 14 and 21. Glial activation was assessed using immunohistochemical method on day 14. Spinal cord pro-inflammatory cytokines and chemokine levels were estimated using enzyme-linked immunosorbent assay (ELISA) on days 8, 14 and 21.

    Results: Vin-induced pain hypersensitivity was significantly suppressed by mid-dosage and high-dosage PPF treatment, starting from day 10 (p < 0.05 or 0.01). On day 14, activated glial numbers decreased significantly on mid- and high-dosage PPF treatments (p < 0.05 or 0.01). High-dosage PPF treatment significantly reduced MCP-1 (monocyte chemoattractant protein-1) up-regulation (p < 0.01). The up-regulation of pro-inflammatory cytokines were significantly reduced by mid-dosage and high-dosage PPF (p < 0.01 or 0.05).

    Conclusions: Neuroinflammation plays a crucial role in the progression of Vin-induced neuropathic pain. PPF could attenuate this pain, through weakening the recruitment of neuroinflammation, thereby relieving pain hypersensitivity in rat.

  • Article
    Tengteng Chen, Di Chen
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1597-1605. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.158
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    Objectives: As the worst and most aggressive prognostic subtype of clinical breast cancer, triple-negative breast cancer (TNBC) accounts for 25% of total breast cancer (BC) deaths. This study aims to explore the relationship between Adenosine 5′-monophosphate (AMP)-activated protein kinase (AMPK) pathway activity and adriamycin (ADM) resistance in TNBC.

    Methods: MDA-MB-231 cells (a TNBC cell line) were treated with ADM to obtain ADM-resistant MDA-MB-231 (MDA-MB-231/ADM) cells. MDA-MB-231/ADM cells treated with A-769662 (an activator of AMPK) and Compound C (an inhibitor of AMPK) were set as the A-769662 and Compound C groups, respectively. Western blot was utilized to detect the AMPK pathway-related protein expression in TNBC tissues and MDA-MB-231 cells. Cell function assays [3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), flow cytometry and colony formation assay] were employed for investigating the roles of A-769662 and Compound C in ADM resistance, apoptosis, and colony formation of MDA-MB-231/ADM cells and MDA-MB-231 cells.

    Results: In this study, AMPK pathway activity was inhibited in TNBC tissues. In addition, AMPK pathway activity exhibited a significant decrease in MDA-MB-231/ADM cells and ADM-resistance TNBC tissues compared with the corresponding ADM-sensitive TNBC tissues and MDA-MB-231 cells, respectively. After ADM treatment, the AMPK pathway activation in MDA-MB-231/ADM cells induced by A-769662 significantly reduced the half-maximal inhibitory concentration (IC50) and colony formation ability of ADM while increasing the apoptosis rate. Nevertheless, inhibiting the AMPK signaling pathway using Compound C notably increased the IC50 of ADM and colony formation ability and reduced the apoptosis of cells.

    Conclusions: After the treatment with ADM, the activation of the AMPK pathway greatly enhanced the MDA-MB-231/ADM cells’ sensitivity to ADM, promoted apoptosis, and inhibited cell growth. Furthermore, the activation of AMPK signaling may be an important strategy to inhibit ADM resistance in TNBC.

  • Article
    Jinjin Li, Guangjie Liu, Qiumei Zhang, Rongna Dong, Jingyun Zhang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1607-1616. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.159
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    Background: Diabetic kidney disease (DKD) is a systemic microvascular complication of poorly controlled diabetes, caused by chronic hyperglycemia. Metformin, as a first-line drug in oral treatment of patients with type 2 diabetes mellitus (T2DM), has attracted increasing attention because in addition to its significant hypoglycemic effect, it has protective effect on DKD. The purpose of this study was to explore the regulatory role of metformin in DKD.

    Methods: Twelve 8-week-old male Specific-Pathogen-Free (SPF) C57BL/KsJ-db/db mice were used as animal models of T2DM. Six 8-week-old male C57BL/6 mice were used as control subgroup (NC subgroup); db/db mice were randomly divided into two subgroups: DKD subgroup (n = 6) and metformin (MET) intervention subgroup (DKD + MET subgroup, n = 6). The amount of 24-hour urinary protein and microalbumin, and levels of serum creatinine, and blood urea nitrogen were measured. Hematoxylin-Eosin (HE) staining and Periodic Acid-Schiff (PAS) staining were used to evaluate the morphological and pathological changes of the glomeruli. The expression of glomerular podocyte labeled protein was observed by immunofluorescence. The expressions of podocyte marker protein and autophagy related marker protein were detected by western blot.

    Results: In contrast with the normal control subgroup, metformin markedly decreased the body weight of DKD mice. Compared with the DKD subgroup, in the DKD + MET subgroup the pathological damage of the renal structure was alleviated, serum creatinine (Scr), blood urea nitrogen (BUN), 24-hour urinary protein and urinary mindin were decreased, while the expressions of microtubule-associated protein light chain 3 (LC3), nephrin, and podocin were increased. Light microscopic evaluation using PAS staining confirmed that the above injuries were alleviated by metformin. The protein of PTEN (phosphatqase and tensin homologue)/Akt (protein kinase B)/mTOR (mechanistic target of rapamycin) pathway in drug group tended to be increased, but did not reach statistical significance.

    Conclusions: Metformin can improve the pathological damage of DKD, improve the renal functional indicators Scr and BUN, and decrease urinary microalbumin (UMA) and urinary mindin, increase the levels of autophagy proteins LC3 and Beclin-1, and inhibit the podocyte damage in DKD, and then delay the development of DKD.

  • Article
    Yu Chen, Wenwu Luo, Guangyao Ning, Menglong Jiang, Renquan Zhang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1617-1627. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.160
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    Background: The development of tumors is accompanied by abnormal cell proliferation. Centromere-Associated Protein E (CENPE) is involved in controlling cell proliferation. The purpose of this study is to explore the function of CENPE in non-small cell lung cancer (NSCLC).

    Methods: In this study, we extracted the datasets from The Cancer Genome Atlas (TCGA) and the Gene Expression Omnibus (GEO) to analyze the CENPE mRNA expression in tumor and normal tissues of NSCLC. And using the Kaplan–Meier Plotter to explore the correlation between the expression of CENPE and the prognosis of NSCLC patients. Furthermore, we use TIMER2.0 (Shirley Liu Lab, Dana Farber Cancer Institute, Boston, MA, USA) for immune cell infiltration analysis to elucidate the underlying mechanism of CENPE in promoting the progression of NSCLC. To validate the results from the bioinformatic analysis, we collected 86 patient samples of NSCLC and did immunohistochemical staining of CENPE, the cluster of differentiation molecule 11b (CD11b), and CD33.

    Results: We found that CENPE was highly expressed in lung adenocarcinomas and it was associated with poor prognosis. Further studies revealed that smoking and mutations in the TP53 gene were involved in the regulation of CENPE expression in cancer cells. The high expression of CENPE in tumor cells is involved in the regulation of the composition of tumor microenvironment cells and mainly promotes the infiltration of immunosuppressive cells, myeloid-derived suppressor cells (MDSC), which may boost the progression of lung adenocarcinoma by inhibiting immune function.

    Conclusions: Therefore, CENPE can be used as a potential therapeutic target for lung adenocarcinoma.

  • Article
    Wei Zheng, Bin He, Min Qiu, Qiong Zhang, Na Liu, Xiaoqing Li, Hua Xu, Yan Zhang, Feng Li
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1629-1639. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.161
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    Background: The present study aimed to systemically evaluate the safety and clinical efficacy of Shenqi Fuzheng injection (SFI) combined with conventional treatment for cervical cancer.

    Methods: Studies investigating the effect of SFI plus conventional treatment versus conventional treatment until 19 April 2021, were systematically reviewed. RevMan 5.3 software (The Nordic Cochrane Centre, Copenhagen, Denmark) was utilized for the present data analysis.

    Results: Eight articles were included in this study. Patients in the SFI group had better complete remission (CR) [risk ratio (RR) 95% confidence interval (95% CI) = 1.69 (1.02, 2.82), p = 0.04], partial remission (PR) [RR (95% CI) = 1.78 (1.21, 2.64), p = 0.004], objective response rate (ORR) [RR (95% CI) = 1.76 (1.36, 2.30), p < 0.0001], improved quality of life (RR (95% CI) = 1.80 (1.30, 2.49), p = 0.0004), improved plus stable quality of life (RR (95% CI) = 1.27 (1.10, 1.46), p = 0.001), more CD3+ T cell [weighted mean difference (WMD) (95% CI) = 9.76 (5.79, 13.73), p < 0.00001], CD4+ T cell (WMD (95% CI) = 8.16 (6.36, 9.95), p < 0.00001), NK (natural killer) cell (WMD (95% CI) = 3.16 (0.02, 6.31), p = 0.05), higher CD4/CD8 (WMD (95% CI) = 0.64 (0.23, 1.05), p = 0.002), less CD8+ T cell (WMD (95% CI) = –5.66 (–8.18, –3.13), p < 0.0001), and less risk of leukopenia (RR (95% CI) = 0.52 (0.38, 0.69), p < 0.00001) and nausea or vomiting (RR (95% CI) = 0.59 (0.45, 0.77), p = 0.0001).

    Conclusions: SFI combined with conventional treatment exhibited better therapeutic efficacy and safety than those of conventional chemotherapy.

  • Article
    Guofei Zhu, Yeying Wang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1641-1647. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.162
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    Background and objective: Early diagnosis and timely management of refractory Mycoplasma pneumoniae pneumonia (MPP) helps improve the prognosis of patients. The aim of this study was to explore the relationship between the peripheral granulocyte-macrophage colony-stimulating factor (GM-CSF), high mobility group box 1 (HMGB1) and the clinical features as well as the prognosis of children with refractory MPP.

    Methods: A total of 130 MPP children were enrolled. They were divided into two groups based on the clinical diagnosis: Refractory MPP group (n = 57) included children whose clinical and radiological features deteriorated despite receiving appropriate antibiotic treatment for more than 7 days; And Non-refractory group (n = 73) which included the other children. Clinical data, including physical examination, treatment and outcome, were collected. The peripheral GM-CSF and HMGB1 level before and after treatment were determined, through enzyme-linked immunosorbent sorbent assay (ELISA) kits.

    Results: Compared with the non-refractory MPP group, the level of GM-CSF and HMGB1 were significant higher in the refractory MPP group (p < 0.05). The receiver operator characteristic (ROC) curve analysis results indicated that GM-CSF and HMGB1 levels before treatment can predict refractory MPP attack (area under the curve, AUC = 0.706, 0.665, p < 0.05). The combined predictive value of the GM-CSF and HMGB1 levels for refractory MPP attack was higher than their independent predictive value (AUC = 0.776). Moreover, the levels of GM-CSF and HMGB1 at 1 week after treatment could predict long hospital stay (>28 d) (AUC = 0.738, 0.776, p < 0.05) and poor prognosis of refractory MPP (AUC = 0.817, 0.804, p < 0.05).

    Conclusions: GM-CSF and HMGB1 can predict the occurrence and development of refractory MPP, thus, they can be used as the auxiliary indicators for clinic diagnoses and prognosis in patients, providing specific guidance for an appropriate clinical treatment.

  • Article
    Gao Chen, Shu-Shu Liu, Meng-Zheng Wu, Chun-Jun Qin, Fei-Fei Liu, Jian-Ying Liu, Li Liu
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1649-1654. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.163
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    Background: To investigate the role of tuberculosis infection interferon-γ release assay (TB-IGRA) combined with pleural effusion TB-RNA (tuberculosis RNA detection) detection in the diagnosis of tuberculous pleural effusion.

    Method: In this observational study, serum TB-IGRA and pleural effusion with TB-RNA were detected in patients with tuberculous and malignant pleural effusion, respectively, and the diagnostic effects of the two groups of patients were compared.

    Result: The content of pleural globulin in the tuberculous pleural effusion group was higher than that in the malignant pleural effusion group; The difference was significant (t = 2.03, p = 0.04). The serum IFN-γ (interferon-gamma) content in patients who were in the tuberculous pleural effusion group was significantly higher than that in the malignant pleural effusion group; The positive rate of TB-IGRA combined with TB-RNA was 97.5% in the tuberculous pleural effusion group and 14.5% in the malignant pleural effusion group; The diagnostic sensitivity of TB-IGRA was significantly better than that of TB-RNA, and the specificity was significantly lower than that of TB-RNA; The diagnostic sensitivity and specificity of TB-IGRA combined with TB-RNA was considerably higher than that of TB-RNA alone, and these differences were statistically significant (p < 0.05).

    Conclusions: The sensitivity, specificity, and accuracy of TB-IGRA combined with TB-RNA in diagnosing tuberculous pleural effusion are high, and its application may prevent some missed diagnoses of complicated pleural effusion.

  • Article
    Jinying Hao, Lei Zhou, Zhengyu Zhang, Yaqin Geng, Yu Zhang, Yan Wang, Bo Gao
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1655-1667. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.164
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    Background: Overexpression of long non-coding RNA (lncRNA) Maternally Expressed 3 (MEG3) significantly alleviates the inflammatory response caused by rheumatoid arthritis (RA). However, the regulatory effect of lncRNA MEG3 combined with transcription factors remains unknown in RA.

    Methods: Clinical synovial tissues were collected from The Second People’s Hospital of Changzhou. Histological changes of synovial tissues and cellular morphology were examined by hematoxylin-eosin and immunofluorescent staining. A real-time quantitative polymerase chain reaction as well as a western blot test were used to analyze gene transcript levels. Cell mobility and proliferation of fibroblast-like synoviocytes (FLSs) were assessed according to colony formation, wound healing and transwell assays. Cytokine contents were quantified using enzyme-linked immuno sorbent assay kits. Interactions were analyzed using RNA pull-down and chromatin immunoprecipitation assays.

    Results: lncRNA MEG3 was significantly downregulated in RA participants’ synovial tissues (p < 0.05) and rats (p < 0.001). Short interference RNA of lncRNA (si-lncRNA) MEG3 significantly reduced cell growth, migration, invasion and inflammatory level of FLSs, while MEG3 overexpression caused in the reverse results. BTG Anti-Proliferation Factor 2 (BTG2) was shown to be significantly inhibited in RA, and the regulatory effects of BTG2 overexpression on cell activity and inflammation were significantly inhibited by si-lncRNA MEG3. Enhancer of Zeste 2 Polycomb Repressive Complex 2 Subunit (EZH2) was confirmed to be regulated by lncRNA MEG3, further to control the transcription of BTG2. Si-EZH2 significantly inhibited cell activity and inflammatory level of FLSs, which was similar to the effects of si-lncRNA MEG3.

    Conclusions: lncRNA MEG3 regulated the expression of BTG2 to participate in RA progression via targeting EZH2, which provided a novel molecular mechanism of RA.

  • Article
    Wen-Yu Liu, Shi-Yu Wang, Jia-Rong Zhang, Cong Lu, Xian-Ming Xu, Hao Wu
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1669-1678. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.165
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    Aim: Preeclampsia (PE) and intrauterine growth restriction (IUGR) are two significant obstetrical diseases that cause serious harm to maternal and infant health. Worldwide, PE is the most frequent cause of IUGR; The latter is regarded as a serious complication of PE but its underlying mechanism and molecular biological changes are poorly understood. Thus, few effective medical therapies for its treatment are available. PE and IUGR share the same etiological background but their connections at the molecular level were rarely known. Consequently, it is of urgency to create an effective method to evaluate their molecular signature. The objective of this study was to identify the hub genes related to PE with IUGR (PE-IUGR) by conducting a weighted gene co-expression network analysis (WGCNA).

    Methods: The GSE147776 data set containing 28 samples of placental tissue (n = 6 with PE-IUGR) was downloaded from the Gene Expression Omnibus database. The gene expression profile was correlated with phenotypic data and analyzed using a WGCNA. Additionally, a WGCNA was used to construct a gene co-expression network, and hub genes were further identified by identifying modules related to the clinical traits of PE-IUGR.

    Results: Nine genes, i.e., TDRKH, XPOT, AMACR, NBN, ALS2, CLYBL, CENPQ, PCGF6 and COQ3 were obtained by the WGCNA. These were considered the key genes that were likely involved in IUGR in PE.

    Conclusions: We constructed a co-expression network of PE-IUGR and identified 9 hub genes related to the condition.

  • Article
    Zhenli Tang, Demei Xin, Wei Liu, Bing Yan, Rui Liu
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1679-1684. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.166
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    Objective: To study and compare the clinical effects of targeted treatments with Nilaparib and Olaparib on recurrent ovarian cancer.

    Methods: A total of 98 patients with recurrent ovarian cancer who were diagnosed and treated in our hospital from February 2019 to July 2021 were enrolled and randomly assigned to group A and group B, with 49 patients in each group. Patients in both groups received combined chemotherapy regimen of paclitaxel and cisplatin (TP). In addition, group A received Nilaparib while group B was treated with Olaparib. The two groups were compared with respect to clinical therapeutic effects and safety of medication.

    Results: The objective response rate (ORR) and disease control rate (DCR) were significantly higher in group A than in group B (p < 0.05). After treatment, the serum level of cancer antigen 125 (CA125) in group A was markedly lower than that in group B (p < 0.05). However, there were no significant differences in total incidence of adverse reactions between the two groups.

    Conclusions: Nilaparib produced a relatively better therapeutic effect in the treatment of recurrent ovarian cancer, when compared with Olaparib, and it decreased serum level of CA125. Therefore, the overall clinical effect of targeted treatment with Niraparib on recurrent ovarian cancer was better than that of Olaparib.

  • Article
    Zhenyu Liao, Erqiang Wang, Huijiao Jiang, Jun Hou, Xiangwei Wu, Xueling Chen
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1685-1698. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.167
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    Background: The primary pathological features of cystic echinococcosis (CE) are the formation of fibrotic pericytes and space-occupying lesions in the liver, but fibrogenesis has received little attention in CE. This study aimed to investigate the possible mechanisms underlying CE-induced fibrogenesis from the perspective of competing endogenous RNAs (ceRNAs).

    Methods: Hydatid fluid was collected from sheep livers infected with Echinococcus granulosus and was used to treat rat hepatic stellate cells (HSC-T6) to establish a fibrotic cell model. Next, RNA sequencing and bioinformatic analyses were conducted. Finally, two important ceRNA axes were chosen for quantitative real-time polymerase chain reaction (qRT-PCR) analysis.

    Results: After hydatid fluid treatment, the both mRNA and protein expression levels of fibrosis-related factors (anti-alpha-smooth muscle actin, type III collagen, and transforming growth factor-β receptor II) were significantly higher than those in control cells. After sequencing, 569 differentially expressed (DE) mRNAs, 151 DE long non-coding RNAs (lncRNAs), and 14 DE miRNAs were identified in infected cells. Next, ceRNA regulatory networks consisting of 207 mRNAs, 13 miRNAs, and 54 lncRNAs were constructed. Based on the comparative toxicogenomics database, 8 fibrosis-related pathways were identified, including focal adhesion, extracellular matrix-receptor interaction, the hypoxia-inducible factor 1 signaling pathway, and pathways in cancer, as well as disease pathway-related regulatory networks associated with 38 DE lncRNAs, 43 DE mRNAs, 11 DE miRNAs, and 8 pathways. The qRT-PCR results of LNC_003335-novel_559-TNC and LNC_002273-miR-125-2-3p-PDGFD were consistent with the RNA sequencing results.

    Conclusions: Our study showed that the two ceRNA axes LNC_003335-novel_559-TNC and LNC_002273-miR-125-2-3p-PDGFD may be potential critical targets for CE-induced fibrogenesis.

  • Article
    Zi-Yi Wang, Zhe Guo, Xue-Song Wang, Hai-Yan Liao, Feng Chen, Yu-Xin Liu, Zhong Wang
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1699-1707. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.168
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    Background: Alveolar macrophages accumulation is the leading cause of sepsis acute lung injury (SALI). AKT (protein kinase B, PKB) signaling pathway is closely related to macrophages apoptosis and polarization. This study aimed to assess AKT inhibitor MK-2206 effect on lipopolysaccharide (LPS)-induced macrophages and on acute lung injury in a septic mice model.

    Methods: Cell Counting Kit-8 (CCK-8) was used to evaluate the impact of MK-2206 on macrophages proliferation in vitro. AKT-mTOR (mammalian target of rapamycin) pathway proteins expression level and apoptosis-related proteins were measured by western blot (WB). Macrophages phenotypic modulation and cell apoptosis level were assessed by flow cytometry. In vivo, survival rate and lung tissue wet: Dry weight ratio were evaluated to assess MK-2206 influence on SALI in septic mice. Immunohistochemistry (IHC) was used to assess alveolar macrophages infiltration. The level of inflammation-related factors were determined by ELISA in vivo and ex vivo.

    Results: Ex vivo, MK-2206 critically restrained Raw264.7 cell proliferation in a dose-dependent way. MK-2206 at 100 ng/mL could up-regulate macrophages apoptosis and skew macrophages from M1 towards M2, followed by an increased in IL (interleukin)-10 level and decreased TNF-α (tumor necrosis factor-α) level. In vivo, MK-2206 could significantly mitigate alveolar macrophages aggregation, reduce lung tissue edema, down-regulate IL-6 and TNF-α levels in peripheral blood, and improve survival in sepsis mice.

    Conclusions: This research revealed that MK-2206 may serve as a promising drug in SALI by regulating macrophage apoptosis and macrophage polarization, which consequently inhibits macrophage pro-inflammatory response by down-regulating AKT signaling pathway.

  • Article
    Xuan ZHOU, Zhen-Zhen XU, He-Hua MA, Zu-Yi WENG, Juan LI
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1709-1718. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.169
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    Background: Acute myeloid leukemia (AML) is the most common type of leukemia that has a poor prognosis. Biomarkers and effective treatments for AML are still under research. Cuproptosis is a type of copper-dependent programmed cell death that has been linked to cancer progression. However, the clinical impacts of cuproptosis-related genes (CRGs) remain unclear.

    Purpose: This study aimed to evaluate the potential role of CRGs to prognose AML.

    Methods: It was examined the expression of 14 CRGs, and a 3-gene risk model using RNA-sequencing data from The Cancer Genome Atlas (TCGA) cohort was constructed. The least absolute shrinkage and selection operator (LASSO)-penalized Cox regression analyses were used to generate a risk score and discriminate the cohort into low- and high-risk AML groups. Then they were analyzed for chemotherapeutic and immune responses. Findings were further validated with AML clinical samples.

    Results: Thirteen CRGs were differentially expressed between AML patients and healthy controls. Patients with high-risk exhibited shorter overall survival (p = 0.003 in TCGA, p = 0.0216 in GSE37642), worse therapeutic response and increased inflammation. Thus, risk-score generated from the 3-gene risk model (risk score = (–0.01161) × GCSH expression + (–0.40387) × LIPT1 expression + (0.248985) × PDHA1 expression) could act as an independent prognostic factor. At last, GCSH expression decreased was validated with the clinical samples (p = 0.0312).

    Conclusions: Cuproptosis has a strong relationship with AML. The 3 CRG-related risk model could well predict AML prognosis.

  • Article
    Yang-kun Wang, Wei Zhang, Tian Yun, Junling Zhou, Jinli Gao, Ping Li, Su-nan Wang, Xiaodong Chen
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1719-1727. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.170
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    Objective: This study aims to investigate the occurrence, development, and histopathological staging of gastric adenocarcinoma of the fundic glands (GA-FG).

    Methods: Detailed histological observations, immunohistochemistry staining as well as follow-ups were conducted on specimens from 215 patients with abnormal proliferative lesions of the gastric fundic glands, who had undergone endoscopic biopsies and endoscopic submucosal dissection.

    Results: Four stages were observed in GA-FG. The first stage was the benign proliferative stage, where the lesions were located in the local proliferative region of the fundic glands, and the lesions could be divided into two types: Ductal proliferation and cystic dilatation. The second stage was the gastric fundic glandular adenoma stage, where the lesions were mainly located in the gastric mucosa layer and could be divided into two types: Gastric fundic microadenoma and gastric fundic adenoma. The third stage was the atypical proliferative stage of the gastric fundic glands, in which the lesion invaded the mucosal muscle, and ≤50%of the mucosal muscle was interspersed with abnormal proliferative glandular ducts. The lesions could be divided into two types: Atypical ductal proliferation of the glands and atypical cystic dilatation of the glands. The fourth stage was the GA-FG stage, in which the lesion directly invaded the mucosal muscle layer as far as the submucosa, and the lesions could be divided into three types: GA-FG with low proliferative activity, GA-FG with high proliferative activity, and cystic GA-FG.

    Conclusions: These lesions can be divided into the following four types based on the stage of occurrence: Benign hyperplasia of the gastric fundic glands, adenoma of the gastric fundic glands, atypical hyperplasia of the gastric fundic glands, and GA-FG. This staging should enable clinicians to administer targeted treatment and track cellular transformation, which is important for monitoring the development of GA-FG.

  • Article
    Chen Du, Ting Liu, Xiujuan Chen
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1729-1736. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.171
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    Objective: The present study aims to provide new ideas for research on the molecular mechanism of polycystic ovary syndrome (PCOS) by (1) investigating the expression characteristics of long non-coding RNAs (lncRNAs) in cumulus granulosa cells (GCs) in patients with PCOS and (2) conducting in vitro validation.

    Methods: The differential expression profiles of lncRNAs in GCs from patients with PCOS and women with normal ovulation were obtained by sequencing using the HiSeq platform. The key lncRNAs were screened, and the target genes were verified by real-time polymerase chain reaction (RT-PCR) and Western blot in GCs of different quality (high-quality embryos and non-high-quality embryos).

    Results: A total of 204 significantly differentially expressed lncRNAs were identified using the results of transcriptome gene sequencing; The expression level was up-regulated in 83 and down-regulated in 121. The RT-PCR validation showed that the expressions of target lncRNA ENST00000433673 and messenger RNA (mRNA) NPY1R (Neuropeptide Y Receptor Y1) were significantly higher in the GCsof non-high-quality embryos from patients with PCOS than in the GCs of non-high-quality embryos from healthy subjects; Meanwhile, the expression of the NPY1R protein was significantly higher in the GCs of non-high-quality embryos from healthy subjects than in the GCs of non-high-quality embryos from patients with PCOS. For high-quality embryos, the expressions of lncRNA ENST00000433673 and mRNA NPY1R did not change significantly in GCs from both healthy subjects and patients with PCOS. The expression of NPY1R protein have similar pattern with NPY1R mRNA in different groups, the expression level of NPY1R protein was the highest in control group, and the expression of lncRNA ENST00000433673 increased while the protein expression of NPY1R decreased.

    Conclusions: lncRNA ENST00000433673 might have important regulatory roles during processes such as embryonic development; This suggests that lncRNAs might be involved in the pathogenesis of PCOS.

  • Article
    Ping Zhao, Xin Li, Yumeng Gu, Wenjun Feng, Xiaolin Tian, Xiaoshuang Xia, Yanfen Du
    Journal of Biological Regulators and Homeostatic Agents. 2023, 37(3): 1737-1743. https://doi.org/10.23812/j.biol.regul.homeost.agents.20233703.172
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    Objective: In this study, we aimed to explore the features of motor symptoms, non-motor symptoms, and magnetic resonance imaging (MRI) of vascular parkinsonism (VP) in patients with cerebral small vessel disease (CSVD).

    Methods: The clinical data of consecutive patients who visited the Neurological Department of the Second Hospital of Tianjin Medical University were collected—We collected 40 cases of CSVD patients who met the diagnostic criteria of VP as the CSVD-VP group, at the same time, 46 patients without VP symptoms were collected as the control group (CSVD group). Several scales were used to evaluate the patients’ motor and non-motor functions, and all patients underwent 3.0T MRI.

    Results: The rate of diabetes, cerebral infarction, and hypercholesterolemia in the CSVD-VP group was significantly higher than that in the CSVD group (p < 0.05). The mean Barthel index of the CSVD-VP group was 72.00 (10.05), which was significantly lower than that of the CSVD group 82.75 (8.39) (p < 0.05). Among the least scoring items were urinary incontinence and difficulty walking up and down stairs. The total score of Unified Parkinson’s Disease Rating Scale (UPDRS) III in the CSVD-VP group was 16.53 (5.89), which was significantly higher than that in the CSVD group 3.51 (2.05) (p < 0.05). The scores of Tinetti balance 10.15 (2.15) and gait 9.03 (1.45) in the CSVD-VP group were significantly lower than those in the CSVD group (p < 0.05). The scores of Mini-Mental State Examination 23.85 (2.41) and Montreal Cognitive Assessment 21.35 (2.50) in the CSVD-VP group were significantly lower than those in the CSVD group 25.00 (2.00) and 23.00 (1.99), respectively (p < 0.05). Lacunes and white matter lesion (Fazekas ≥2) in the CSVD-VP group were more significant than those in the CSVD group (p < 0.05).

    Conclusions: Patients with CSVD-VP had a lower daily living ability score and higher UPDRS III score than those with CSVD alone, the symptoms of parkinsonism in the CSVD-VP patients were accompanied by cognitive dysfunction, urinary incontinence, anxiety, depression, and other non-motor symptoms. The MRI scans of CSVD-VP patients had more obvious lacunar infarction and white matter hyperintensities around the basal ganglia.