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

Uveitis is a common ocular disease and may cause serious damage of visual function. The pathogenesis of uveitis is closely related to its autoimmune response. The Th1 cells and Th17 cells were identified to play a key role in the occurrence of autoimmune uveitis and experimental autoimmune uveitis, whilst the Th17 cells were found to be closely associated with disease recurrence. The regulatory T cells (Tregs) were found to be involved in the regression of inflammation. The dysfunction and ratio of Tregs were the major causes for persistence and recurrence of uveitis. Th17 cells and Tregs can also interconvert with each other under certain conditions. Regulating the Th17/Tregs balance might be a potential novel approach to alleviating inflammation and hence providing the treatment for uveitis.

Objective To explore how alveolar macrophages from chronic obstructive pulmonary disease (COPD)-model rats affect proliferation and secretion of 16HBE human bronchial epithelial cells and investigate the associated mechanism. Methods Alveolar macrophages were extracted from COPD rats induced by cigarette smoke exposure and LPS instillation through bronchoalveolar lavage, then co-cultured with 16HBE cells in vitro. Exosomes were extracted from alveolar macrophages of rats with exosome isolation kit. The differentially expressed miRNA in exosomes derived from macrophages of rats in COPD group and control group was detected by PCR. miR-380 was overexpressed with miR-380 mimic while the expression of cystic fibrosis transmembrane transduction regulator (CFTR) was knocked down with siRNA in 16HBE cells. The proliferation of 16HBE cells was detected with CCK-8 assay. The migration ability of 16HBE cells was evaluated with Transwell^TM migration assay. The levels of mucins (MUC5AC, MUC5B, MUC2) and CFTR expressed by 16HBE cells were detected with Western blot analysis. The expression of TNF-α and IL-6 in the supernatant of 16HBE cells was detected with ELISA. Results The alveolar macrophages from COPD rats enhanced the proliferation and migration of 16HBE cells. The production of mucins and TNF-α as well as IL-6 in 16HBE cells were increased by COPD macrophages. The expression of miR-380 was significantly elevated in exosomes derived from COPD alveolar macrophages. Both overexpression of miR-380 and inhibition of CFTR decreased the expression of CFTR, resulting in the significantly enhanced proliferation and migration of 16HBE cells as well as increased expression of MUC5AC, MUC5B, MUC2 and TNF-α, IL-6. Conclusion The alveolar macrophages from COPD rats can enhance the proliferation and mucin expression as well as inflammatory cytokine secretion of 16HBE cells. This process may be involved with abnormal expression of miR-380 in exosomes of COPD alveolar macrophages and down-regulation of CFTR in bronchial epithelial cells.

Objective To identify the characteristic genes and immune infiltration in dilated cardiomyopathy (DCM) by bioinformatic analysis. Methods We identified differentially expressed genes (DEG) on two DCM gene expression data sets, and performed gene ontology (GO), disease ontology (DO), and gene set enrichment analysis (GSEA) functional enrichment to obtain potential pathways. Two machine learning algorithms including support vector machine recursive feature elimination (SVM-RFE) algorithm and Least Absolute Shrinkage and Selection Operator (LASSO) algorithm were used to determine diagnostic markers. Finally, we used the cell type analysis tool CIBERSORT for immune cell infiltration analysis. Results A total of 51 DEGs were finally identified. Thioredoxin interacting protein (TXNIP), crystallin Mu (CRYM), heat shock 70kDa protein 1-like (HSPA1L), and eukaryotic elongation factor 1A-1 (EEF1A1) were considered candidate diagnostic markers. Enrichment analysis focused on features including cardiac processes, outer membranes of mitochondria and organelles, ubiquitin-like protein ligase, natural killer cell-mediated cytotoxicity, Th1, and Th2 cell differentiation, T cell receptor signaling pathways, and Th17 cell differentiation. Immune cell infiltration found naive B cells, neutrophils, and γT cells may be involved in the pathogenesis of DCM. Besides, neutrophils, T follicular helper cells, and M1 macrophages were highly correlated with four characteristic genes. Conclusion The four characteristic genes identified by machine learning, TXNIP, CRYM, HSPA1L, and EEF1A1, show potentially close relation to DCM. At the same time, immune cell infiltration analysis can better showcase the pathophysiological process of DCM.

Objective To investigate how vitamin D (VD) affects fine particulate matter (PM_2.5)-induced autophagy and cytokines production in A549 human alveolar epithelial cells. Methods PM_2.5 samples were prepared at the upper part of an acetylene diffusion flame burner. A549 cells were treated with PM_2.5 in vitro, and/or were treated with VD or autophagy inhibitor 3-methyladenine (3-MA). Western blot analysis was employed to analyze the level of LC3-II/I and Beclin-1 in A549 cells with different groups. Real time quantitative PCR was used to detect the mRNA expression of interleukin-25 (IL-25), IL-33 and thymic stromal lymphopoietin (TSLP). The formation of autophagosomes was observed by transmission electron microscopy. Results LC3-II/I ratio and beclin-1 protein expression were found increased in A549 cells after PM_2.5 treatment, and autophagosome were increased too. There was a marked decrease of PM_2.5-induced autophagy with VD treatment. After 3-MA treatment, the autophagy was inhibited. Then, PM_2.5 continued to induce autophagy, while VD could also reverse it. PM_2.5 promoted the secretion of IL-25, IL-33 and TSLP by inducing autophagy in A549 cells while this process was inhibited by 3-MA and VD. Conclusion VD can inhibit PM_2.5-induced autophagy and cytokine release in A549 cells, thus playing a protective role.

Objective To investigate the effect of PDS5B on the biological function of A549 human lung cancer cells and possible molecular mechanism. Methods The proliferation of lung cancer cells was detected by MTT assay and colony formation assay after silencing or overexpressing PDS5B of A549 cells. The cell migration was detected by scratch assay and Transwell^TM assay. The protein expression of PDS5B and Wnt5a in A549 cells was detected by Western blot analysis. Cell migration was detected by Transwell^TM after PDS5B small interference RNA(siRNA) and Wnt5a siRNA were co-transfected. Results Compared with the negative control group, the protein expression of PDS5B decreased significantly after transfected with PDS5B siRNA. The proliferation ability , colony formation rate and migration ability of A549 cells significantly improved, and the expression of Wnt5a was increased. The opposite results were observed after PDS5B over-expression. The co-transfer experiment showed that Wnt5a could resist the inhibition of A549 cells by PDS5B. Conclusion PDS5B inhibits lung cancer cell proliferation by down-regulating Wnt5a expression.

Objective To express and purify the dormancy survival regulator Rv2628 of Mycobacterium tuberculosis, so as to prepare and identify its rabbit polyclonal antibody. Methods Using the genome of Mycobacterium tuberculosis H37Rv strain as a template, the Rv2628 gene was amplified to construct a recombinant expression plasmid which was transformed into an Escherichia coli protein expression strain and induced expression by IPTG. The target protein was purified using Ni-NTA chromatography column, and the rabbit polyclonal antibody was obtained by immunizing New Zealand white rabbits with purified Rv2628 protein. The specificity of polyclonal antibodies was verified by Western blot analysis and indirect ELISA, respectively. Results The PET-30A-RV2628 recombinant carrier was successfully constructed. After induction by IPTG, the RV2628 protein was mainly expressed in the form of inclusion. The high-purity Rv2628 protein was obtained by Ni-NTA column purification. Rabbit anti-Rv2628 polyclonal antibody was obtained after immunizing the rabbit. The antibody had good antigen binding properties and the antibody titer reached 1:1 093 500. Conclusion The high-purity Rv2628 protein and high-titer rabbit anti-Rv2628 polyclonal antibody were successfully prepared.

Objective To examine the effects of Coxsackie virus B3 (CVB3) on the NLR family, pyrin domain containing protein 3 (NLPR3) of mouse macrophages and its mechanisms. Methods RAW264.7 cells, primary mouse macrophages (bone marrow-derived macrophages or peritoneal macrophages), and short hairpin RNA (shRNA)-NLRP3 lentivirus infected RAW264.7 cells were stimulated by different dosages of CVB3. The transcript levels of NLRP3 and IL-1β were measured by quantitative real-time PCR. IL-1β in the supernatants of cell cultures was determined by ELISA. The protein level of NLRP3 was tested by Western blot analysis and the interacting proteins of NLRP3 were detected by co-immunoprecipitation (Co-IP). Results The transcript levels of NLRP3 and IL-1β were significantly up-regulated in the CVB3 stimulated RAW264.7 cells and primary mouse macrophages (bone marrow-derived macrophages or peritoneal macrophages). The expression level of NLRP3 presented CVB3-dose dependence and demonstrated the highest expression level at 6 hours after CVB3 treatment. The transcript level of IL-1β significantly increased the most at 6 hours after CVB3 treatment, while the protein level of IL-1β peaked at 24 hours after CVB3 treatment. In the GFP-shRNA-NLRP3 lentivirus infected RAW264.7 cells, NLRP3 was obviously inhibited, and with CVB3 stimulation, IL-1β in the supernatants of cell cultures decreased significantly. Moreover, NLRP3 antibody was used for Co-IP experiment, in which the resultant protein complex was then stained with silver nitrate. The differential protein band between different groups was identified as nicotinamide adenine dinucleotide kinase 2 (NADK2) by mass spectrometry. This result demonstrated that CVB3 induced the interaction between NADK2 and NLRP3. Conclusion CVB3 stimulation promotes the activation of NLRP3 in macrophages, thereby enhancing the expression and secretion of pro-inflammatory cytokine IL-1β by activating NADK2.

Age-related thymus involution that comes with age, can lead to degenerative diseases, autoimmune diseases, tumors and opportunistic infections. Transcription factors such as forkhead box N1 (FoxN1), p63 and autoimmune regulator (AIRE) that can affect the process of age-related thymus involution and the differentiation, development and output of T cells by regulating the proliferation, development and differentiation of thymic epithelial cells(TECs). Here, we reviewed the mechanism of regulating age-related thymus involution and the research into the new treatment schemes progresses in the upstream and downstream targets of FoxN1, p63 and AIRE, miRNAs and signal pathways that affect their functions.

Objective To investigate whether miR-141-3p promote the migration of CNE-2 human nasopharyngeal carcinoma (NPC) cells by regulating vimentin. Methods Real-time quantitative PCR was used to detect the expression of miR-141-3p in NPC tissues and adjacent tissues and the expression level of vimentin was detected by immunohistochemical staining and Western blot analysis. Real-time quantitative PCR and Western blot were used to screen 5-8F, CNE-2, HNE1 human NPC cell lines with the highest relative expression of miR-141-3p. Real-time quantitative PCR and Western blot analysis were used together to verify the relationship between miR-141-3p and vimentin expression. Small interfering RNA (si-miR-141-3p) was used to down-regulate miR-141-3p of CNE-2 cells. MTT assay tested the proliferating inhibition rate of CNE-2 cells. Transwell^TM chamber assay was performed to detect cell invasion and migration and Western blot analysis to detect the expression of vimentin. Results Compared with the paracancerous tissues, the expression of miR-141-3p and vimentin in NPC tissues increased significantly. Compared with NP69 cells, the expressions of miR-141-3p and vimentin increased significantly in CNE-2 cells. The down-regulation of miR-141-3p in CNE-2 cells has induced significant decrease of cell invasion and migration capabilities, cell proliferation capabilities, as well as vimentin protein expression. Conclusion miR-141-3p can enhance the proliferation and migration of CNE-2 cells by promoting vimentin expression.

Objective To identify hub genes and key pathways in ulcerative colitis (UC) by bioinformatics. Methods The mRNA expression microarray GSE134025 related to ulcerative colitis was retrieved from Gene Expression Omnibus (GEO) database. The differentially expressed genes (DEGs) between ulcerative colitis samples and normal intestinal mucosal cells were screened by the R limma package, and then GO and KEGG pathway analysis was carried out. The protein-protein interaction (PPI) network was constructed by STRING database, and Cytoscape software was used to screen the hub genes in the PPI network. KEGG mapper was employed to mark the location of the hub genes in signal pathway. Results A total of 190 DEGs were screened, of which 147 were up-regulated and 43 were down-regulated. GO function enrichment showed hub genes were mainly involved in biological processes including inflammatory response and positive regulation of proliferation. Cellular component mainly enriched in membrane and plasma membrane, maintaining the molecular functions of mediating heparin binding and cytokines. KEGG analysis showed that the enriched pathways are mainly related to cytokine-cytokine receptor interaction, chemokine signaling pathway and other signal transduction pathway. Ten hub genes such as IL-6, CXCL8, CXCL10, CXCL1, CXCL9, ANXA1, IL-1β, CCL20, CXCL2, and CXCL11 were screened out of the PPI network, which were located downstream of the signaling pathway regulated by IL-17. Conclusion The 10 hub genes related to the risk of UC were likely to be regulated by IL-17 during the occurrence and development of UC.