The Influence Of High Salt Diet On Liver Fibrosis

Backgrounds:Sodium is an indispensable nutrient,which is necessary to human health.However,today's eating habits,especially western eating habits,cause salt intake significantly exceed daily physiological needs.Excessive salt intake may increase the risk of many diseases,such as cardiovascular-related diseases,autoimmune diseases,metabolic diseases.And high-salt diet has been implicated in breakdown of our immune system balance.Increasing studies demonstrate that excessive salt intake can promote pro-inflammatory macrophage polarization thereby aggravates autoimmune diseases.In addition,high-salt diet can cause disturbances in the intestinal flora,which can aggravate EAE by affecting the Th17 type immune response.For a long time,liver fibrosis has seriously affected human health,and it may eventually develop into cirrhosis and even death.Poor eating habits are one of the common causes of liver fibrosis,and immune dysfunction is also considered to be an important factor in aggravating liver fibrosis.Studies have shown macrophages play a key role in liver fibrosis,which can secrete transforming growth factor(TGF-?),platelet derived growth factor(PDGF)and other profibrotic factors to promote the activation of hepatic stellate cells(HSC).In addition,the gut-liver axis determines that intestinal microbiota disorders and impaired intestinal barriers can cause bacteria and toxins translocate to the liver,and then aggravate liver fibrosis.However,the effects of high-salt diet on liver fibrosis have rarely been reported.Therefore,the purpose of this study is to investigate the influence of high-salt diet on liver fibrosis and its immune mechanisms..vMethods:ND and HSD mice model were established by given different salt content in food and drinking water.Combined special diet with carbon tetrachloride(CC14)treatment,we established four groups:normal diet(ND),high salt diet(HSD),liver fibrosis with normal diet group(ND CC14)and liver fibrosis with high salt diet group(HSD CC14).Differences in severity of liver fibrosis were determined by pathological staining and the levels of profibrogenic cytokines.We analyzed the macrophages in liver and small intestine by flow cytometry.The expression level of profibrogenic cytokines in intestinal tissue and portal vein blood was compared by enzyme-linked immunosorbent assay.In vitro experiments confirmed the effects of high salt on macrophages.The difference of intestinal flora between ND CC14 group and HSD CC14 group was detected by 16S sequencing,and the changes of intestinal permeability and liver bacteria translocation were confirmed by quantitative PCR.The role of the translocated bacteria in liver fibrosis was evaluated by in vitro experiments.Results:To investigate the effect of high-salt diet on liver injury and fibrosis,we compared the pathological changes of four groups of mice.Compared with the ND group,the liver pathological staining of HSD group mice showed no obvious liver injury or liver fibrosis.However,in the CC14 group,compared with the ND CC14 group of mice,H&E staining,masson staining and alpha-SMA immunohistochemistry showed that the degree of liver injury and fibrosis in the HSD CC14 group was increased,and the expression of TGF-(3,TNF-a and MCP-1 in the liver was also significantly increased.We further compared the effect of high-salt diet on the distribution and activation of macrophages in the liver,and found that compared with the ND CC14 group,the proportion and activation level of macrophages in the liver of HSD CC14 group mice were increased.At the same time,the proportion of macrophages in the laminar propria of the small intestine of mice in the HSD CC14 group was increased,and the expression of fibrosis promoting factors and monocyte chemokines in the small intestine tissues were increased.In addition,HSD increased the levels of pro-fibrosis related factors and monocyte chemotactic factors in portal vein.In vitro experiments showed that high-salt environment can directly increase the expression of fibrosis related factors in macrophages,thereby enhancing the activation of hepatic stellate cells.The proportion of macrophages in the liver and small intestine increased simultaneously in the HSD CC14 group,while increase of fibrosis-promoting factors and monocyte chemokine in the liver,portal vein and intestinal tract were consistent.All of these results suggested that the increased number and activation of macrophages in the liver may due to the change in intestine.As the high-salt diet may affect the liver immune status by changing the intestinal flora,we performed 16srDNA sequencing analysis.The results revealed that HSD have changed the intestinal flora structure in liver fibrosis mice,destroyed the intestinal barrier function,and promoted intestinal bacteria,especially the Enterococcus translocate to the liver.In vitro experiments have shown that Enterococcus can promote macrophage activate and enhance the ability of macrophages to promote hepatic stellate cells activation.Conclusion:This study found that high-salt diet can significantly aggravate liver fibrosis in mice,possibly by increasing the proportion and activation of macrophages in liver,and then promote hepatic stellate cell activation.On the one hand,HSD can cause disorder in the intestinal flora,increase the permeability of the intestinal,and promote the translocation of intestinal bacteria to the liver.On the other hand,the high-salt diet increases the number of macrophages in the lamina propria of small intestine,accompanied by increasing pro-fibrotic factors and monocyte chemotactic factor in the small intestine,and these factors can reach the liver through the portal vein.These two aspects may be the reason for the increase in the number and activation of macrophages in liver,which ultimately aggravates liver fibrosis.In summary,high-salt diet can aggravate liver fibrosis,and reasonable salt intake is important for the prevention and treatment of liver fibrosis.