Study On The Pathogenesis Of Follicular Helper T Cell-associated Intestinal Immunity In Non-obese Type 2 Diabetes

Objective:The pathogenesis of type 2 diabetes(T2D)is complex and diverse.Most current studies on type 2 diabetes focus on the pathogenesis based on obesity levels.However,many individuals develop T2D at the non-obese body mass index(BMI)level.It is unclear whether the etiological mechanisms found in these obesity models are all applicable to non-obese T2D patients.Recent studies have shown that intestinal flora,intestinal immunity and inflammatory responses may play an important role in the pathogenesis of T2D.We have tried to find out the potential therapeutic targets by studying the role of follicular helper T cells(Tfh;CD4~+CXCR5~+T cells)in intestinal immunity.Point to provide new ideas for the treatment of type 2 diabetes and the regulation of intestinal immunity and intestinal inflammation.Methods:First,we performed an alignment analysis of the composition of immunoglobulin(Ig)in stool samples from 24 non-obese T2D patients,14 BMI-matched healthy controls,and 11 active Crohn's disease patients.A total of 4 healthy controls and10 non-obese T2D patients in these subjects were willing to donate an intestinal mucosal biopsy sample.Four of the 10 non-obese T2D patients provided 2,3 provided 3,and 3provided 4 intestinal mucosal biopsy samples.We performed Ig subtype composition and Tfh cell cytokine secretion analysis on B cells of intestinal mucosa samples of all subjects who were willing to provide samples.We then used 6 non-obese T2D patient samples to remove CD4~+CXCR5~+T cells and neutralize IFN-?.Finally,we used 4 healthy control mucosal samples and 7 non-obese T2D mucosal samples for E.coli stimulation experiments and their IFN-?expression was measured by flow cytometry.The statistical method is as shown in the legend,and the t test,paired t test or Mann-Whitney U test,Wilcoxon signed rank test is applied accordingly.The comparison of multiple groups of samples was based on the analysis of variance and Kruskal-Wallis H test according to the normality and homogeneity of variance.If the comparison between groups was statistically significant,the method of multiple comparison was used for post hoc test(post-Hoc test).Statistical analysis was performed using Graphpad Prism 5 software,and P<0.05 was considered to be statistically significant.Results:1.Fecal IgG levels in non-obese T2D patients are higher than those in BMI-matched healthy controls,but lower than in active Crohn's disease patients.2.The percentage of cells expressing IFN-?in CD4~+CXCR5~+T cells in the intestinal mucosa of non-obese T2D patients was significantly increased,while the percentage of cells expressing IL-4 and IL-17 was significantly decreased.3.Significant reduction in IgG production after removal of CD4~+CXCR5~+T cells and addition of polyclonal rabbit anti-human IFN-?neutralizing antibody.4.In non-obese T2D patients and healthy controls,intestinal mucosal CD4~+CXCR5~+T cells increased IFN-?secretion after E.coli stimulation,but IFN-?secretion was more significant in non-obese T2D patients.Conclusion:1.The level of intestinal inflammatory response in non-obese T2D patients is higher than that of healthy controls and lower than that of active Crohn's disease patients.2.CD4~+CXCR5~+T cells in the intestinal mucosa of non-obese T2D patients express IFN-?and low expression of IL-4 and IL-17.3.CD4~+CXCR5~+T cells and IFN-?may play important roles in intestinal IgG inflammation in non-obese T2D patients.4.Intestinal CD4~+CXCR5~+T cells in non-obese T2D patients had a stronger inflammatory response to intestinal bacteria than the BMI-matched healthy controls.