Study On The Effect And Mechanism Of Chronic Psychological Stress On Intestinal Flora And Liver Pathology In Rats With Liver Fibrosis

BACKGROUND:With the change of the medical model from the "biomedical model" to the "bio-psycho-social medical model",chronic psychological stress as an influencing factor to promote the development of liver fibrosis has gradually been valued,but the mechanism is unclear for now.Progressive liver disease is often accompanied by intestinal endotoxemia(IETM)and intestinal bacterial translocation.Lipopolysaccharide(LPS),the main component of endotoxin,can initiate intracellular signal transduction via liver Toll-like receptor 4(TLR4),mediate the inflammatory cascade,activate Kupffer cells and hepatic stellate cells,promote the formation of extracellular matrix,and eventually leads to the aggravation of liver fibrosis.The above process is called "gut-liver" axis.There is also an interactive effect between psychological stress and the intestine,called "brain-gut" axis,whose existence has become a consensus,and psychological stress can cause intestinal flora disturbance and promote intestinal bacterial translocation through the "brain-gut" axis.Therefore,we imagine that chronic psychological stress may indirectly affect the liver through the gut,that is,chronic psychological stress causes intestinal flora disturbances and bacterial translocations to increase blood LPS levels,and then aggravate liver damage and liver fibrosis through the "gut-liver" axis.PERPOSE:To observe the effects of chronic psychological stress on intestinal flora and liver pathology in rats with liver fibrosis,detect the changes in signal transduction pathways in hepatocytes of the "gut-liver" axis during this process,and explore the possible mechanism of chronic psychological stress affecting the progression of fibrosis.METHODS:Sixty normal SD rats were selected and randomly divided into 6 groups:(1)Normal group: normal rats,no modeling;(2)Olive oil group: liver fibrosis negative control model,subcutaneous injection of olive oil,3ml / kg,2 times / week,double the first dose,and completed in 8 weeks;(3)Liver fibrosis group: liver fibrosis model,40% carbon tetrachloride is injected subcutaneously in the same dose and time as the oliveoil group;(4)Psychological stress group: normal rats were subjected to chronic psychological stress for 4weeks from the 5th week;(5)olive oil + psychological stress group: chronic psychological stress was performed for 4 weeks from the 5th week of the establishment of the liver fibrosis negative control model;(6)Liver fibrosis + psychological stress group: chronic psychological stress was performed for 4 weeks from the 5th week of the establishment of the liver fibrosis model.All rats were sacrificed after 8 weeks of rearing.The cecum contents(intestinal stool)was taken to detect the diversity of intestinal flora in the intestinal stool by 16 S r RNA sequencing analysis;The portal vein blood was taken to detect the blood LPS level by azochromogenic limulus test;The liver tissue was taken to detect collagen deposition by Masson staining and to assess the degree of liver fibrosis by semi-quantitative staging.Method of Immunohistochemisyry and IOD measuring were used to detect and quantify the expression of liver intracellular protein,which including TLR4,My D88,NF-κβ,TNF-α,IL-1,IL-6,TRIF,TRAM,IRF-3 and IFN-β.RESULTS:1、The beta diversity of rat intestinal flora was shifted to varying degrees under the effects of olive oil,psychological stress and liver fibrosis.Compared with the normal group,chronic psychological stress significantly increased the abundance of WPS-2 bacteria in the gut of rats(P <0.01),liver fibrosis caused a decrease in the abundance of thick-walled mycoplasma in the intestinal tract,and an increase in the abundance of actinomycota,spirochaeta,and soft-walled mycoplasma.The liver fibrosis group also showed an increase in the abundance of Bacteroides,while the liver fibrosis + psychological stress group showed an extra increase in the abundance of Patescibacteria(P = 0.047)and WPS-2(P = 0.000).2、Low level LPS remained in portal vein blood of normal group and olive oil group rats.In contrast,the LPS level of portal vein blood of rats in psychological stress group,liver fibrosis group and liver fibrosis + psychological stress group significantly and gradually increased(P <0.05).3、Compared with the normal group and the olive oil group rats,the liver fibrosis group had obvious collagen fiber deposition and lobular structure disorder,and the liver fibrosis stage score increased significantly(P <0.05).Under the influence of chronic psychological stress,pathological damage occurred in the liver tissue of each group of rats.liver fibrosis rats had a large amount of collagen fibers deposited and pseudolobules formed after the chronic psychological stress was applied.The staging score of liverfibrosis in liver-fibrosis rats with chronic psychological stress was further increased,compared with that in liver-fibrosis rats without stess although the statistical difference was not significant(P > 0.05).4、Compared with the normal group and the olive oil group,the Liver intracellular protein expression levels of TLR4,My D88,NF-κβ,AP-1,TNF-α,IL-6,TRIF,TRAM,IRF-3 and IFN-βin liver fibrosis rats was significantly increased(P <0.05).Under chronic psychological stress,The expression levels of the above-mentioned proteins were all significantly higher than before(P <0.01).CONCLUSION:1、Chronic psychological stress can mediate intestinal flora disturbance through the "brain-gut" axis,causing bacterial translocation,leading to an increase in portal blood LPS levels.2、Chronic psychological stress can be used as an independent factor to cause and aggravate liver pathological damage and promote the progress of liver fibrosis in rats with liver fibrosis.3、The "gut-liver" axis hepatocyte transduction pathway mediated by the LPS-TLR4 signal pathway plays an important role in the development of liver fibrosis.4、Chronic psychological stress can promote the activation of the "brain-gut" axis and the "gut-liver" axis,which may be an in vivo mechanism that indirectly aggravates liver pathological damage and promotes the development of liver fibrosis.