The Study On The Mechanism Of Lactobacillus Plantarum And Its Metabolite Acetate In Regulating Type 1 Diabetes Based On Inhibition Of NLRP3 Inflammasome

The interaction between gut microbiota and gut cells contributes to the health of the body.Microorganisms and their metabolites can promote the differentiation and development of immune cells,reduce intestinal inflammation and permeability,and achieve intestinal homeostasis.The imbalance of intestinal flora can lead to a variety of autoimmune diseases such as type 1 diabetes(T1D).Most probiotics,such as Lactobacillus casei,Lactobacillus reuteri,Bifidobacterium and Streptococcus thermophilus can improve the intestinal flora structure of the host,reduce intestinal permeability and alleviate the symptoms of T1D patients when taken in sufficient quantities.The mechanism by which Lactobacillus Plantarum NC8regulates T1D is still unclear.NLRP3 inflammasome,as a member of the inflammatory family,can enhance the inflammatory response by promoting the production and secretion of pro-inflammatory cytokines.Many studies have shown that NLRP3 inflammasomes play an important role in the development of T1D.However,it remains to be elucidated whether Lactobacillus Plantarum NC8 can alleviate T1D by regulating NLRP3 inflammasomes.In view of this,this study used streptozotocin(STZ)to construct a type 1 diabetes mouse model,to explore the effect of Lactobacillus Plantarum NC8 and its metabolite acetate on T1D,and further study its mechanism of alleviating T1D by regulating NLRP3 inflammasome.In order to reveal whether Lactobacillus Plantarum NC8 can alleviate T1D,mice were divided into four groups:control,T1D,Lactobacillus Plantarum NC8 prophylaxis,and Lactobacillus Plantarum NC8 treatment.In addition to the control group,other groups used STZ to construct mice T1D models for subsequent experiments,and mice in the prophylaxis and treatment groups used Lactobacillus Plantarum NC8(5×10~9 CFU/m L)for 3 weeks,and PBS was used for the other groups.Through weekly testing of mice weight and blood glucose,the results found that after suffering from T1D,the weight of mice decreased,blood glucose significantly increased.The results of tissue paraffin section and tissue immunofluorescence showed that the pancreatic islets of T1D mice were severely damaged and the number of macrophages in the pancreas increased.The results of flow cytometry showed that Th1/Th17cells increased in spleen and PLN of T1D mice.The body weight of mice in both the prophylaxis group and the treatment group of Lactobacillus Plantarum NC8 was protected to some extent.The damage of pancreatic islets in both groups was reduced,and the number of macrophages in the pancreas was reduced.The serum insulin level of Lactobacillus Plantarum NC8 prophylaxis group was also significantly higher than that of T1D group.Compared with T1D group,Th1/Th17 cells in spleen and PLN of mice in Lactobacillus Plantarum NC8 prophylaxis group and treatment group were significantly decreased.The above results reveal the mechanism by which Lactobacillus Plantarum NC8 reduces pancreatic islet damage in mice,reduces macrophages in the pancreas and Th1/Th17 cells in the spleen and pancreatic lymph nodes of T1D mice,thereby reducing inflammatory damage and weight loss caused by T1D and alleviating T1D.Through further detection of short chain fatty acids in mouse feces,it was found that the content of acetate in the feces of T1D mice was significantly lower than that of the control group,while the content of acetate in the feces of mice in the prevention and treatment groups of Lactobacillus Plantarum NC8 increased.To determine whether the metabolite acetate of Lactobacillus Plantarum NC8 can alleviate T1D,mice were also divided into four groups:Control,T1D,Acetate prophylaxis group and Acetate treatment group.The results showed that both the Acetate prophylaxis group and the Acetate treatment group decreased blood glucose,increased body weight,and reduced islet damage.Compared with T1D mice,the serum insulin level of mice in the Acetate prophylaxis group and treatment group was higher,the number of Th1/Th17 cells in spleen and PLN was significantly decreased,and the number of macrophages in pancreas was also decreased.These results were similar to those of T1D mice treated with Lactobacillus Plantarum NC8,indicating that acetate,the metabolite of Lactobacillus Plantarum NC8,had a certain effect on relieving T1D.In order to clarify the mechanism of Lactobacillus Plantarum NC8 and its metabolite acetate alleviating T1D by regulating NLRP3 inflammatory bodies,this study used J774A.1mouse macrophage system to establish an inflammatory model for in vitro experiments and C57BL/6J mice were used for animal experiments.The animal experimental groups were the same as in the above experiments.The results of q-PCR and Western blotting showed that acetate could inhibit the expression of NLRP3 in macrophages.The results of cell supernatant ELISA showed that the level of IL-1 in the supernatant of macrophages treated with acetate was lower than that in the untreated group.The q-PCR and Western Blot test using mice pancreas also found that the NLRP3 in the pancreas of T1D mice increased significantly,while the expression of NLRP3 in the pancreas of mice in the Lactobacillus Plantarum NC8 prophylaxis group,treatment group of Lactobacillus Plantarum NC8,acetate treatment group and acetate prophylaxis group decreased.At the same time,the results of serum ELISA showed that the level of IL-1βin T1D group was significantly higher than that in the control group,while the level of IL-1βof mice in the prophylaxis group and treatment group of Lactobacillus Plantarum NC8,acetate treatment group and acetate prophylaxis group was lower than that in T1D group.The comprehensive results showed that Lactobacillus Plantarum NC8 and its metabolite acetate could alleviate the damage caused by T1D in mice,reduce the number of macrophages in the pancreas of T1D mice,and reduce the Th1/Th17 cells in the spleen and pancreatic lymph nodes,thereby alleviating T1D in mice.At the same time,in vivo and in vitro experiments proved that Lactobacillus Plantarum NC8 and its metabolite acetate probably regulated T1D by inhibiting NLRP3 inflammatory bodies.Lactic acid bacteria can alleviate T1D,providing data for the application of probiotics in T1D and new ideas for the prevention and treatment of autoimmune diseases with probiotics.