| Background: Type 1 diabetes is a chronic autoimmune disease that usually starts in children.There are more than 1.25 million type 1 diabetes patients in the United States,and 13,000 new diabetes in China each year,The annual incidence of type 1 diabetes in China is 1.01 per 100,000 people,among which 1.93 per 100,000 people are aged 0-14.The incidence of type 1 diabetes is increasing year by year,and the overall incidence is increasing by about 2% to 3% each year.The incidence of type 1 diabetes in adolescents is also increasing.Type 1 diabetes accounts for 90% of juvenile diabetes patients,and the pathogenesis of type 1 diabetes is still unclear.Previously,type 1 diabetes was thought to be a single autoimmune disease caused by the attack of T lymphocytes on pancreatic islet beta cells.Currently,it is believed to be caused by the interaction of environmental factors,microbial factors,genes,metabolism and immune system.Many environmental exposures have been associated with type 1 diabetes,including diet in infancy,vitamin D supplementation,early exposure to viruses associated with islet inflammation,such as enteroviruses,and decreased intestinal microbial diversity.In terms of genes,people used to pay more attention to protein-coding genes,but high-throughput sequencing shows that protein-coding genes only account for about 2%.Most of the genes are non-coding genes,and most of the non-coding genes are non-coding RNAs,while lncRNAs are a kind of non-coding RNAs with a length of more than 200 nucleotides.In the early stage,lncRNA53106 expression was significantly increased in the type 1 diabetes model through gene chip screening.The purpose of this study was to investigate the regulatory mechanism of lncRNA53106 in the apoptosis model of MIN6 cells stimulated by inflammatory factors,and to reveal the role of lncRNA53106 in the pathogenesis of type 1 diabetes.It provides new ideas for further discovering the pathogenesis of type 1 diabetes and treating type 1 diabetes.Methods: 1.Mouse islet cell lines(MIN6 cells)were cultured,and inflammatory factors(IL-1 beta 10ng/ml,TNF-alpha 50ng/ml,IFN-gamma 50ng/ml)were combined to stimulate MIN6 cells to establish a MIN6 cell apoptosis model.The apoptosis rate of MIN6 cells in the normal culture group and the inflammatory factor stimulation group was measured by flow cytometry.The effects of lncRNA53106,CXCL10 and apoptosis-related factors on the apoptosis of MIN6 cells were detected by q PCR and Western blot.2.Lnc RNA53106 smart silencer was constructed and transfected into MIN6 cells using lipo2000 as vector.After q PCR confirmed the inefficiency of knockdown,inflammatory cytokines were combined for stimulation and the apoptosis rate of MIN6 cells was determined by flow cytometry.The effects of lncRNA53106,CXCL10 and apoptosis-related factors on the apoptosis of MIN6 cells were detected by q PCR and Western blot.3.CXCL10 si RNA was constructed and transfected into MIN6 cells with lipo2000 as the vector.After q PCR verification of low knockdown efficiency,inflammatory cytokines were used for combined stimulation.The apoptosis rate of MIN6 cells was measured by flow cytometry,and the m RNA expression level of lncRNA53106 was detected by q PCR.Results: 1.In the combined stimulation of inflammatory factors on the apoptosis of MIN6 cells,the apoptosis rate in the inflammatory factor group was significantly increased and statistically significant(P<0.01),and the m RNA expression levels of lncRNA53106 and CXCL10 were significantly increased.2.When lncRNA53106 was knocked down and stimulated with inflammatory factors,the apoptosis rate in the knockdown group was significantly reduced compared with that in the control group(P<0.05).The m RNA and protein expressions of CXCL10 in the knockdown group were also reduced by q PCR and Western blot,and the m RNA expressions of apoptotic factors Bax and Caspase3 were also reduced.3.CXCL10 was knocked down and then combined with cytokine stimulation,The apoptosis rate in the knockout group was significantly lower than that in the control group(P<0.05),and lncRNA53106 expression was slightly increased but no significant difference(P=0.6087).Conclusion: Lnc RNA53106 may promote the expression of apoptosis factor by upregulation of CXCL10,and promote the apoptosis of beta cells of the pancreas,thus may leading to the occurrence of type 1 diabetes. |
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