Compatibility with Semen Sojae Praeparatum attenuates hepatotoxicity of Gardeniae Fructus by regulating the microbiota, promoting butyrate production and activating antioxidant response

Herb-induced liver injury is a leading cause of drug-induced liver injury in China and its incidence is also increasing worldwide. Gardeniae Fructus (ZZ) has aroused wide concern for hepatotoxicity in recent decades. But when ZZ is administered in combination with Semen Sojae Praeparatum (DDC) to compose a herbal pair Zhizichi Decoction (ZZCD), lower hepatotoxicity is observed. The mechanism involved in the attenuated effect remains to be investigated. Our previous studies showed that DDC benefited host metabolism by regulating the gut microbiota and it reduced the exposure of major toxic components of ZZ. The present study was aimed to investigate how DDC attenuated hepatotoxicity of ZZ from the perspective of gut microbiota. Rats received ZZ and ZZCD treatment of different dosages and antibiotic treatment was applied to explore the involvement of gut microbiota. Biochemical assays and histopathological analysis were conducted to evaluate liver injury. Gut microbiota in caecal contents was profiled by 16S rRNA sequencing. Short-chain fatty acids (SCFAs) in caecal contents were measured by gas chromatography mass spectrometry (GCMS). To verify the protective effect of butyrate, it was administered with genipin, the major hepatotoxic metabolite of ZZ, to rats and HepG2 cells. Plasma lipopolysaccharide (LPS) level and colon tissue section were used to evaluate gut permeability. Expression level of Nuclear factor erythroid-derived 2-like 2 (Nrf2) was detected by immunohistochemistry in vitro and by western blot in vivo. Our study showed that ZZCD displayed lower hepatotoxicity than ZZ at the same dosage. ZZ induced gut dysbiosis, significantly reducing Lactobacillus and Enterococcus levels and increasing the Parasutterella level. In combination with DDC, these alterations were reversed and beneficial genus including Akkermansia and Prevotella were significantly increased. Besides, butyrate production was diminished by ZZ but was restored when in combination with DDC. Butyrate showed detoxification on genipin-induced liver injury by promoting colon integrity and promoting Nrf2 activation. Besides, it protected genipin-induced hepatocyte damage by promoting Nrf2 activation. DDC attenuates ZZ-induced liver injury by regulating the microbiota, promoting butyrate production and activating antioxidant response.