| The human gut is the natural habitat for a large and dynamic bacterial community. The intestinal microflora is a positive health asset that crucially influences the normal structural and functional development of the mucosal immune system. Gut microflora disequilibrium would be happened in many liver diseases, resulting in endotoxemia or bacterial translocation, and these changes are mainly due to the destruction of gut immune barrier. Gut immune system can distinguish commensal from pathogenic bacteria because it can response to pathogens but be tolerant to resident intestinal microflora. Recent studies have defined a previously unknown arm of the CD4+ T cell effector response, the Th17 lineage, which promises to change our understanding of immune regulation, immune pathogenesis and host defense. Intestinal Thl7 cells differentiation and development rely on the presence of gut commensal microflora. The appropriate presence of Th17 cells can maintain the function of gut immune system, but redundant development of Th17 cells will result in the breakage of the gut immune barrier. Remarkably, Th17 development depends on the pleiotropic cytokine TGF-β, which is also linked to regulatory T cell (Treg cell) development and function, providing a key role in the tolerance of gut immune to commensal bacteria. Therefore, Th17 and Treg cells are key cell subsets of lymphocytes in connecting the intestinal flora and intestinal immune function. It is proposed that the fine balance between Th17 and Treg cells is crucial for maintenance of immune homeostasis.In addition, emerging evidence suggests that inflammatory responses mediated by T cells play a key role in the development and progression of many forms of acute and chronic liver disorders such as chronic hepatitis B, acute liver failure, and so on. So, it is valuable to investigate the effect of commensal bacteria on the intestinal immunity focusing especially on the phenotypic composition of the T cells such as Thl7 and Treg cells and the altered cytokine production within it before and after interfering by probiotics in a mouse model of acute liver injury induced by D-galactosamine (D-GalN). We can analyze the relationship between commensal bacteria and gut immune system, and possible mechanism of commensal bacteria disturbance affecting on the gut immune barrier and the prognosis of acute liver injury.Regarding Salmonella infections as the control of gut flora disturbance, a mouse model of acute liver injury induced by D-GalN intraperitoneal injection was established. We analyzed the change of gut microflora and bacterial structural features using real-time fluorescence quantitative PCR and PCR-DGGE before and after the pretreatment by the application of Bifidobacterium and Lactobacillus as a probiotic in this acute liver injury model. The results showed that obvious disequilibrium of gut microflora was occurred in mice of acute liver injury model group compared with normal control group. The number of gene copy of potential pathogens such as Enterobacter, Enterococcus, and Bacteroides was significantly increased, while which in Bifidobacterium and Lactobacillus was significantly decreased. Furthermore, DGGE fingering patterns reflected the difference of structure feature of gut microflora disturbance between the mice of acute liver injury and Salmonella infection. The number of potential pathogens was significantly decreased and the number of Bifidobacterium and Lactobacillus was significantly increased after the probiotic pretreatment. The gut flora imbalance obviously had been corrected and the probiotic pretreatment also showed protective effect on liver injury and liver function to some extent.To further examine the changes of body and gut immune function after disturbance of gut flora under conditions of acute liver injury, we still established a mouse model of acute liver injury induced by D-GalN intraperitoneal injection regarding Salmonella infections as the control of gut flora disturbance. PBMC and intestinal lamina propria lymphocytes were isolated 48 hours after the animal model was established. Peripheral CD4/CD8 ratio, peripheral and intestinal Th17 and Treg cells were analysed by flow cytometry. Serum concentrations of the corresponding cytokines were measured by enzyme-linked immunosorbent assay (ELISA). Results showed that peripheral CD4/CD8 ratio was increased significantly, serum cytokines IL-6, IL-17, IL-23 and TGF-p were increased significantly in mice with acute liver injury. Consistent with the cytokine profile, the peripheral and intestinal Thl7 cell population was increased markedly. In contrast, the peripheral and intestinal Treg cell population was decreased dramatically in mice of acute liver injury group. The balance between Thl7 and Treg cells has been destroyed. Probiotic pretreatment can significantly reduce the proportion of Th17 cells in the peripheral blood and intestinal lamina propria, increase the proportion of Treg cells and to some extent restored Th17/Treg cells balance. Moreover, a positive correlation between Th17 cells in the peripheral blood and intestinal lamina propria and serum ALT and AST level was found in this study, which suggests that Th17 cells may be a new parameter indirectly reflexing the disease activity.Combination of above two parts of studies about the relationship between the changes of gut microflora and Thl7/Treg cells balance of body and gut immune system before and after probiotics pretreatment under conditions of acute liver injury, we draw the following conclusions:(1) Obvious disequilibrium of gut microflora was occurred when acute liver injury model was established. The number of gene copy of potential pathogens was significantly increased, while which of probiotics was decreased markedly. The structure feature of gut microflora disturbance of mice with acute liver injury was significantly different from which of mice with Salmonella infection.(2) Thl7/Treg cells balance in body and intestinal and related cytokine profile may play a key role in the pathogenesis of acute liver injury and gut microflora disturbance, and Th17 cells may be a new parameter indirectly reflexing the severity of acute liver injury. Future research on the Th17/Treg cells balance may provide an opportunity to illustrate the pathogenesis of liver disease and gut microflora disturbance, prevention and regulation of Th1 7/Treg cells balance may improve the development of new therapeutic targets for these diseases.(3) Probiotic application can significantly regulate gut microflora disturbance, and then improve the body and intestinal immune barrier and immune function and protect the liver injury and liver function to a certain extent through regulating the Th17/Treg cells balance. |
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