Microbiota-Gut-Metabolomics Analysis Reveals the Hepatotoxicity Induced by Longdan Xiegan Decoction on Chronic Liver-Injured Rat and Potential Mechanisms

Ethnopharmacological relevance: Longdan Xiegan Decoction (LXD) is a traditional Chinese prescription, which is composed of 10 kinds of herbs. LXD has been used to purge dampness-heat of liver and gallbladder for thousands of years in China and first recorded in "Lan Shi Mi Zang", then included in "Yi Fang Ji Jie". Although the utilization of LXD remains prevalent in the treatment of chronic liver injury in China, there still might exist toxicity problems induced by long-term and/or excessive use of LXD.Aim of the study: This study aimed to explore hepatotoxicity of LXD by excessive use during the treatment of chronic liver injury and revealed the potential mechanisms based on microbiota-gut-metabolomics analysis.Materials and methods: LC-MS was used to identify the main active ingredients of LXD. D-galactose (D-gal) was used to induce chronic liver injury model on rat. LXD was intragastrically administrated at doses of 15, 30 and 60mg/mL for 6 weeks, respectively. Histopathological changes, liver index and liver function indexes were used to assess the degree of hepatotoxicity. Western blot was used to detect the protein expression of inflammatory factors including IL-1 beta, IL-6, TNF-alpha and COX-2. Metabolomics methods basing on GC-MS combined with 16S rRNA sequencing technology was used to carry out microbiota-gut-metabolomics analysis in order to reveal the potential hepatotoxicity mechanisms of LXD during the treatment of liver injury.Results: 60mg/mL of LXD could aggravate D-gal induced liver dysfunction, histopathological injury, overexpression of hepatocyte inflammation, and could cause disturbance of gut microbiota and and metabolites. Changes of the relative abundance of unclassified_Lachnospiraceae, unclassified_Peptostreptococcaceae, Desulfovibrio and disturbed tryptophan metabolites especially for the indoles compounds might participate in the hepatotoxicity occurrence. Conclusion: Excessive use of LXD could cause hepatotoxicity on rats with liver injury during long-term administration. The mechanisms of hepatotoxicity occurrence was inextricably linked to LXD interference with the balance of gut microbiota and metabolites.