Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology最新文献

Objective To explore the effects of microRNA-155-5p (miR-155-5p) on lipopolysaccharide (LPS)-induced neuroinflammatory damage of human SH-SY5Y neuroblastoma cells. Methods SH-SY5Y cells line was overexpressed miR-155-5p or transfected with negative control (miR-155-5p mimic group, mimic-NC group), down-expressed miR-155-5p or transfected with its negative control (miR-155-5p inhibitor group, inhibitor-NC group). The cells with successful transfection in the above groups were treated with LPS for 24 hours. The cells without SH-SY5Y cells transfection and those with LPS treatment were included into control group and LPS group, respectively. The activity of SH-SY5Y cells was detected by MTT assay. The apoptosis rate was detected by flow cytometry. The mRNA levels of tumor necrosis factor α (TNF-α), interleukin 1β (IL-1β), interleukin 6 (IL-6) and interleukin 10 (IL-10), and expression of miR-155-5p were detected by reverse transcription PCR. The levels of cleaved caspase-3 (c-caspase-3), B-cell lymphoma/leukemia-2 (Bcl2), Bcl2-associated X protein (BAX), phosphorylated nuclear factor κB p65/nuclear factor κB p65 (p-NF-κB p65/NF-κB p65) and phosphorylated p38 mitogen-activated protein kinase/p38 mitogen-activated protein kinase (p-p38 MAPK/p38 MAPK) were detected by Western blot analysis. The expression of miR-155-5p in SH-SY5Y cells was regulated by miR-155-5p mimic and miR-155-5p inhibitor. The target relationship between miR-155-5p and suppressor of cytokine signaling 1(SOCS1) was predicted by bioinformatics, which was verified by luciferase assay. SH-SY5Y cells with down-regulation of both miR-155-5p and SOCS1 were constructed (miR-155-5p inhibitor/si-SOCS1 group). The cells activity, apoptosis, mRNA expressions of inflammatory cytokines, expression of SOCS1 protein, ratios of p-NF-κB p65/NF-κB p65 and p-p38 MAPK/p38 MAPK were detected by the above methods. Results Compared with control group, activity of SH-SY5Y cells, expression of Bcl2 protein, level of IL-10 mRNA and expression of SOCS1 protein decreased in LPS group while apoptosis rate, expressions of c-caspase-3 and BAX proteins, and levels of TNF-α, IL-1β and IL-6 mRNA were increased, along with the increased miR-155-5p level and ratios of p-NF-κB p65/NF-κB p65 and p-p38 MAPK/p38 MAPK. The activity of SH-SY5Y cells, expression of Bcl2 protein, level of IL-10 mRNA and expression of SOCS1 protein increased in miR-155-5p inhibitor group, compared with LPS group, whereas decreased miR-155-5p level, apoptosis rate, expressions of c-caspase-3 and BAX proteins, levels of TNF-α, IL-1β and IL-6 mRNA, and expressions of p-NF-κB p65/NF-κB p65 and p-p38 MAPK/p38 MAPK proteins were observed. Compared with LPS group, activity of SH-SY5Y cells, expression of Bcl2 protein, level of IL-10 mRNA and expression of SOCS1 protein were decreased in miR-155-5p mimic group, while miR-155-5p level, apoptosis rate, expressions of c-caspase-3 and BAX proteins, levels of TNF-α, IL-1β and IL-6 mRNA, and ratios o

Objective To investigate the immunoregulatory effects of CD226 on the chronic restraint stress (CRS)-induced depression-like behavior and its underlying mechanism in mice. Methods Male C57/BL6J mice and CD226 gene knockout (KO) mice with the same strain (4-6 week old) were adopted to establish CRS models. The stress-induced depression scores of mice were evaluated through behavioral testing such as forced swimming test and sucrose preference test. Flow cytometry was used to analyze the differences of the intraepithelial lymphocytes in the spleens, peyer's patches, and intestines between the two groups. Results Compared with WT CRS group, mice in CD226KO CRS group showed significantly decreased immobility time in forced swimming test and increased sucrose preference rate. The ratio of CD4₊ T cells to CD8₊ T cells in spleen was significantly reduced, combined with the remarkably elevated proportion of TCRαβ and TCRαβCD8αβ cells in the small intestinal IELs of CD226 KO mice with CRS. Conclusion Knockout of CD226 alleviates CRS-induced depression-like behavior in mice, alters the proportion of immune cells in murine spleen and intestine, and improves the overall immune status of mice under stress.

Objective To analyze the clinical significance of calneuron 1 (CALN1) expression in glioma and its role in tumor immune cell infiltration by bioinformatics. Methods The expression of CALN1 gene in glioma in the Cancer Genome Atlas (TCGA) database was analyzed by Xiantao Academic Online. Kaplan-Meier survival analysis was used to evaluate its prognostic value, and receiver operating characteristic (ROC) curve was employed to evaluate its clinical diagnostic efficiency. Gene Set Enrichment Analysis (GSEA) was adopted to identify the potential mechanism of CALN1 in glioma. The relationship between CALN1 mRNA and glioma immune cell infiltration was discussed. Results The expression of CALN1 decreased significantly in glioma, and its expression level was negatively correlated with tumor grade. Compared with the control group, the expression level of CALN1 in isocitrate dehydrogenase mutant and 1p/19q co-deletion gliomas increased significantly. Glioma patients with low expression of CALN1 had poor prognosis and significantly reduced overall survival, disease specific survival and progression-free interval. ROC curve analysis showed that CALN1 expression level had good clinical diagnostic value. The results of GSEA gene enrichment suggested that the expression level of CALN1 was negatively correlated with mitosis and neutrophil degranulation. Immunoinfiltration analysis showed that T helper type 2 (Th2) cells, macrophages and neutrophils were significantly increased in the group with low expression of CALN1. Conclusion The expression level of CALN1 in glioma is positively correlated with the prognosis. The abnormal decrease of CALN1 expression may lead to the invasion of tumor-promoting immune cells. CALN1 can be used as a potential prognostic marker and therapeutic target for glioma.

Objective To investigate the effects of neutrophil extracellular traps (NETs) on proliferation, migration, and invasion of human prostate cancer and its related mechanisms. Methods The expressions of interleukin-8 (IL-8), lymphocyte antigen 6G (LY6G) and citrullinated histone H3 (H3CIT) in 28 cases of tumor tissues and adjacent tissues were detected by immunohistochemical staining. Neutrophils were extracted from the patients and stimulated with PMA to form NETs in vitro. The up-regulated genes of DU145 cells stimulated by NETs were detected by RNA-seq, and verified by real time quantitative PCR and Western blot analysis. KEGG and GO analyses of upregulated genes were performed using the DAVID database. The proliferation, invasion and migration ability of DU145 cells was assessed by CCK-8 assay, Transwell^TM and scratch assay. After knockdown of IL-8 expression in DU145 cells, the changes of proliferation, invasion and migration ability of DU145 cells were tested over again. Results The expression levels of IL-8, LY6G, and H3CIT in tumor sites were significantly higher than those in adjacent tissues. After co-incubated with NETs, the expression of 638 genes including IL-8 were up-regulated in DU145 cells, which related to phosphatidylinositol 3 kinase/protein kinase B (PI3K/AKT) signaling pathway, cell proliferation and invasion. NETs can promote the proliferation, invasion and migration of DU145. After silencing IL-8, the ability of NETs to promote the proliferation, invasion and migration of DU145 was decreased. Conclusion NETs promote proliferation, migration, and invasion of DU145 by upregulating the expression of IL-8 in DU145 cells.

Objective To construct the prokaryotic expression plasmid of human mitoferrin 2 (SLC25A28), and to express and purify the protein for preparing its rabbit polyclonal antibody. Methods The prokaryotic expression plasmid pET28a(+)-SLC25A28-His was constructed and transferred into E. coli BL21 (DE3), and induced with Isopropyl-β-D-thiogalactopyranoside (IPTG). The SLC25A28 protein was extracted in form of inclusion bodies, and was further purified by His-NTA column after dissolved in 8 mol/L urea. The anti-SLC25A28 polyclonal antibody was prepared by immunizing rabbits, and its specificity was determined by Western blot analysis. Results pET28a(+)-SLC25A28-His was constructed and SLC25A28 protein was successfully expressed in E. coli BL21 (DE3) with the purity up to 90%. The Western blot results indicated that anti-SLC25A28 polyclonal antibody was capable to recognize specifically the SLC25A28 protein in testis. Conclusion The human SLC25A28 is successfully expressed in E. Coli, and the rabbit polyclonal antibody specific to SLC25A28 is prepared.

Objective To observe the correlation of stromal cell-derived factor 1 (SDF-1) with bone marrow mesenchymal stem cell (BMSCs) migration and airway inflammation in asthmatic rats. Methods Twenty-four clean SD rats were randomly divided into normal control (NC) group, model control (MC) group, and BMSCs group. Asthma model was established by OVA. In the BMSCs group, 1×10⁶ BMSCs (1 mL) were transplanted into the tail vein on the day the model was completed. Pathological changes in lung tissues were evaluated by HE staining. The count of inflammatory cells in bronchoalveolar lavage fluid(BALF) was evaluated by Wright-Giemsa staining. The concentrations of IL-4, IL-5, IL-13, IgE, IgG1 and IgG2a in BALF were tested by ELISA. The expression of SDF-1 and STAT6 mRNA in lung tissue was measured by real time quantitative PCR. The expression of SDF-1 protein in bronchial epithelial cells were evaluated by Immunofluorescence staining. The expression of SDF-1 and STAT6 protein in lung tissue were measured by Western blot analysis. Results Compared with the normal group, the number of relative inflammatory cell counts and the concentrations of IL-4, IL-5, IL-13, IgE, IgG1, and IgG2a in BALF of the MC group increased significantly. The mRNA and protein expression of SDF-1 and STAT6 in lung tissue increased significantly. Compared with the MC group, inflammatory cells and inflammatory cytokines of BALF of BMSCs group were decreased in numbers, as was the expression of SDF-1 and STAT6 in lung tissues. Compared with the MC group, the expression of SDF-1 gene in lung tissues was increased, as was the expression of SDF-1 protein in bronchial epithelial cells. Conclusion In the process of asthmatic inflammation, the expression of chemokine SDF-1 in the damaged site increases, and promotes the migration of exogenous BMSCs to the lung tissue of asthmatic rats. BMSCs can regulate immune imbalance of Th1/Th2 cells by homing to damaged lung tissue, thus inhibiting asthmatic airway inflammation.

Objective To identify the expression of Toll-like receptor 2 (TLR2) on peripheral blood monocytes and B cells of patients with allergic rhinitis (AR), allergic rhinitis combined with allergic asthma (ARA) before and after allergen challenge. Methods The peripheral venous blood from patients with AR and ARA were recruited and stimulated with Artemisia sieversiana wild allergen extract (ASWE), house dust mite allergen extract (HDME), and Platanus pollen allergen extract (PPE). Flow cytometry was then used to detect the expression of TLR2 on peripheral monocytes and B cells. Results Compared with healthy control (HC) group, the percentage of TLR2₊ monocytes and decreased mean fluorescence intensity (MFI) of TLR2 on monocytes in AR and ARA patients decreased. After being challenged with the above mentioned three allergens, the portion of TLR2₊ monocytes in HC group and MFI of TLR2 on monocytes in AR patients also decreased. Meanwhile, MFI of TLR2 on B cells also showed a decrease after challenged with ASWE and HDME. Conclusion The expression of TLR2 on monocytes and B cells decreases in AR and ARA patients.

Fatty liver disease is one of the most prevalent chronic liver disease worldwide and the gut-liver axis is recognized as increasingly prominent in fatty liver disease. Intestinal dysfunction can affect the occurrence or progression of liver disease, therein, validating the critical role of the intestinal immune cells. Enormous literature reported that macrophages, lymphocyte, dendritic cells (DCs) and other immune cells in the gut as well as their subsets may regulate the fatty liver disease progression via different mechanisms, including disruption of intestinal barrier, dysregulation of intestinal lipid transporters and mediating immune cell migration to liver.

Objective To investigate the anti-inflammatory effects of sinoline on adjuvant arthritis (AA) rats and the changes of NOD like receptor pyrin-domain containing 3 (NLRP3). Methods Wistar rats were randomly divided into control group, AA model group, (100, 200, 400) mg/kg sinomenine group and 100 mg/kg Tripterygium wilfordii group, with 10 rats in each group. Except for the control group, AA model was established by Freund complete adjuvant. 12 days after modeling, control group and model group were given the same amount of normal saline, and other groups were given drugs by intragastric administration, once a day, for consecutive 16 days. Joint conditions of AA rats were evaluated by multiple arthritis and joint swelling. The level of NLRP3 protein in synovial tissues was detected by Western blot, and the expression and distribution of NLRP3 in synovial tissues were detected by immunohistochemical staining. Serum levels of interleukin-1β (IL-1β), IL-6 and tumor necrosis factor α (TNF-α) were detected by ELISA. Results Compared with the control group, multiple arthritis and joint swelling significantly increased in model group, while those significantly decreased in sinomenine treatment groups and Tripterygium wilfordii group. Decreased expression of NLRP3 protein in synovial tissue was observed, along with the significantly reduced levels of IL-1β, IL-6 and TNF-α in serum in sinoline treatment groups. Conclusion Sinolin can improve joint inflammation in AA rats by inhibiting NLRP3 and downstream inflammatory factors.

Objective To investigate the effect and mechanism of compound 21(C21), an agonist of angiotensin II-2 receptor (AT2R) on the cytokine levels of NRK-52E cells stimulated by advanced glycation end products bovine serum albumin (AGE-BSA). Methods NRK-52E cells were divided into control and (25, 50, 100, 200)mg/L AGE-BSA groups and cultured for 48 hours. The mRNA and protein expression levels of leukin-6 (IL-6) and tumor necrosis factor α (TNF-α) were detected by real-time quantitative PCR and ELISA. The NRK-52E cells stimulated by AGE-BSA(25 mg/L) for 48 hours were then treated with (0.01, 0.05, 0.1)mmol/L C21 for 24 hours. The mRNA and protein expression levels of protein kinase C (PKC), nuclear factor κB p65 (NF-κB p65) and transforming growth factor β1 (TGF-β1) were detected by qRT-PCR and Western blot analysis. Results The mRNA expression levels of IL-6 and TNF-α significantly increased in NRK-52E cells stimulated by AGE-BSA at different doses, with the greatest increase in the 25 mg/L AGE-BSA group. The mRNA and protein expression levels of PKC, NF-κB p65 and TGF-β1 in AGE-BSA-induced NRK-52E cells significantly decreased by (0.01, 0.05, 0.1)mmol/L C21. Conclusion AGE-BSA promotes the expression of IL-6, TNF-α, PKC, NF-κB p65 and TGF-β1 in NRK-52E cells, while C21 inhibits the effect of AGE-BSA on NRK-52E cells.