To investigate the protective effect of lithocholic acid (LCA), originating from the gut microbiota, on liver damage in septic mice.

A total of fifteen C57BL/6J mice were randomly divided into a sham surgery group (Sham group), a sepsis group (Sep group) and a fecal microbiota transplantation group (Sep-fmt group), five in each group. The sepsis model was induced using cecal ligation and puncture (CLP), while the Sham group underwent the abdominal wall incision and closure only. The Sep-fmt group received fecal microbiota solution for 3 d on the 2nd day after CLP, and the Sham group and Sep group were given an equal dose of sterile phosphate buffer containing 10% glycerol for 3 d according to body mass. Faecal samples were collected for 16S ribosomal RNA (rRNA) sequencing to analyze differences in bile acid metabolism. Then thirty C57BL/6J mice were assigned to a sham surgery group (Sham group, n = 6), a sepsis group (Sep group, n = 12) and a LCA group (LCA-Sep group, n = 12). Six mice were guaranteed in each group for subsequent experimental analysis. The LCA-Sep group received intragastric administration of 100 mg/kg LCA for five consecutive days before sepsis modeling. The Sham group and Sep group were administered an equal dose of solvent according to body weight for five consecutive days. Faecal samples were analyzed by 16S rRNA sequencing to assess changes in gut microbiota. The mortality, clinical severity score (CSS), serum tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6) and IL-10 were measured, and the liver tissue pathology was evaluated.

Bile acid metabolism analysis showed that there were significant differences in secondary bile acids among the Sham group, Sep group and Sep-fmt group (all P < 0.001). The increase of LCA was the largest in the Sep-fmt group, so LCA was selected for subsequent experiments. At 24 h after CLP, no mice died in the Sham group, four mice died in the Sep group, and two mice died in the LCA-Sep group. The CSS of surviving mice among these three groups showed statistically significant difference [(1.0 ± 0.0), (3.5 ± 0.5), (2.5 ± 0.5); F = 47.500, P < 0.001]. The α diversity analysis showed that the intestinal microflora richness [Chao1 and abundance-based coverage estimator (ACE) indexes] and intestinal microflora diversity (Shannon and Simpson indexes) of the three groups were statistically significantly different (H = 8.766, 8.766, 9.704, 10.994; all P < 0.05), and the intestinal microflora diversity was higher in the LCA-Sep group than in the Sep group (P < 0.05). The β diversity analysis showed that there was a significant difference in the bacterial community structure among the three groups (H = 18.322, P = 0.001). The average relative abundance of Bacteroidetes in the Sham group, Sep group and LCA-Sep group was 62.17%, 11.10% and 34.47% respectively, and the average relative abundance of Proteobacteria was 10.99%, 62.81% and 40.58% respectively. Comparing the serum TNF-α, IL-6, IL-10 and the number of TUNEL positive cells in liver tissue of mice in these three groups, the differences were statistically significant (H = 14.749, 14.363, 15.158; F = 131.688; all P < 0.001), with higher TNF-α, IL-6 and TUNEL positive cells in the Sep group and higher IL-10 in the LCA-Sep group (all P < 0.05). Immunofluorescence showed that M2 macrophages in the Sham group increased and M2 macrophages in the Sep group decreased; after LCA pretreatment, the expression of M2 macrophages in the LCA-Sep group increased compared with the Sep group.

Sepsis leads to gut microbiota imbalance, increased inflammatory factors and liver cell damage; LCA, a gut microbiota metabolite, can downregulate the abundance of potential pathogenic bacteria in the gut microbiota, promote M2 macrophage polarization, and reduce systemic inflammation and hepatocyte apoptosis of septic mice.