ObjectiveNon-alcoholic fatty liver disease (NAFLD)-associated hepatocellular carcinoma (HCC) is an increasing healthcare burden worldwide. We examined the role of dietary cholesterol in driving NAFLD–HCC through modulating gut microbiota and its metabolites.DesignHigh-fat/high-cholesterol (HFHC), high-fat/low-cholesterol or normal chow diet was fed to C57BL/6 male littermates for 14 months. Cholesterol-lowering drug atorvastatin was administered to HFHC-fed mice. Germ-free mice were transplanted with stools from mice fed different diets to determine the direct role of cholesterol modulated-microbiota in NAFLD–HCC. Gut microbiota was analysed by 16S rRNA sequencing and serum metabolites by liquid chromatography–mass spectrometry (LC–MS) metabolomic analysis. Faecal microbial compositions were examined in 59 hypercholesterolemia patients and 39 healthy controls.ResultsHigh dietary cholesterol led to the sequential progression of steatosis, steatohepatitis, fibrosis and eventually HCC in mice, concomitant with insulin resistance. Cholesterol-induced NAFLD–HCC formation was associated with gut microbiota dysbiosis. The microbiota composition clustered distinctly along stages of steatosis, steatohepatitis and HCC. Mucispirillum, Desulfovibrio, Anaerotruncus and Desulfovibrionaceae increased sequentially; while Bifidobacterium and Bacteroides were depleted in HFHC-fed mice, which was corroborated in human hypercholesteremia patients. Dietary cholesterol induced gut bacterial metabolites alteration including increased taurocholic acid and decreased 3-indolepropionic acid. Germ-free mice gavaged with stools from mice fed HFHC manifested hepatic lipid accumulation, inflammation and cell proliferation. Moreover, atorvastatin restored cholesterol-induced gut microbiota dysbiosis and completely prevented NAFLD–HCC development.ConclusionsDietary cholesterol drives NAFLD–HCC formation by inducing alteration of gut microbiota and metabolites in mice. Cholesterol inhibitory therapy and gut microbiota manipulation may be effective strategies for NAFLD–HCC prevention.

ObjectiveHelicobacter pylori is associated with gastric inflammation, precancerous gastric atrophy (GA) and intestinal metaplasia (IM). We aimed to identify microbes that are associated with progressive inflammation, GA and IM 1 year after H. pylori eradication.DesignA total of 587 H.pylori–positive patients were randomised to receive H. pylori eradication therapy (295 patients) or placebo (292 patients). Bacterial taxonomy was analysed on 404 gastric biopsy samples comprising 102 pairs before and after 1 year H. pylori eradication and 100 pairs before and after 1 year placebo by 16S rRNA sequencing.ResultsAnalysis of microbial sequences confirmed the eradication of H. pylori in treated group after 1 year. Principal component analysis revealed distinct microbial clusters reflected by increase in bacterial diversity (p<0.00001) after H. pylori eradication. While microbial interactions remained largely unchanged after placebo treatment, microbial co-occurrence was less in treated group. Acinetobacter lwoffii, Streptococcus anginosus and Ralstonia were enriched while Roseburia and Sphingomonas were depleted in patients with persistent inflammation 1 year after H. pylori eradication. A distinct cluster of oral bacteria comprising Peptostreptococcus, Streptococcus, Parvimonas, Prevotella, Rothia and Granulicatella were associated with emergence and persistence of GA and IM. Probiotic Faecalibacterium praustznii was depleted in subjects who developed GA following H. pylori eradication. Functional pathways including amino acid metabolism and inositol phosphate metabolism were enriched while folate biosynthesis and NOD-like receptor signalling decreased in atrophy/IM-associated gastric microbiota.ConclusionThis study identified that gastric microbes contribute to the progression of gastric carcinogenesis after H. pylori eradication.

See Covering the Cover synopsis on page 2; See editorial on page 38.BACKGROUND AND AIMS: Dietary fat intake is associated with increased risk of colorectal cancer (CRC). We examined the role of high-fat diet (HFD) in driving CRC through modulating gut microbiota and metabolites. METHODS: HFD or control diet was fed to mice littermates in CRC mouse models of an azoxymethane (AOM) model and Apc min/þ model, with or without antibiotics cocktail treatment. Germ-free mice for fecal microbiota transplantation were used for validation. Gut microbiota and metabolites were detected using metagenomic sequencing and high-performance liquid chromatography-mass spectrometry, respectively. Gut barrier function was determined using lipopolysaccharides level and transmission electron microscopy. RESULTS: HFD promoted colorectal tumorigenesis in both AOMtreated mice and Apc min/þ mice compared with control diet-fed mice. Gut microbiota depletion using antibiotics attenuated colon tumor formation in HFD-fed mice. A significant shift of gut microbiota composition with increased pathogenic bacteria Alistipes sp. Marseille-P5997 and Alistipes sp. 5CPEGH6, and depleted probiotic Parabacteroides distasonis, along with impaired gut barrier function was exhibited in HFD-fed mice. Moreover, HFDmodulated gut microbiota promotes colorectal tumorigenesis in AOM-treated germ-free mice, indicating gut microbiota was essential in HFD-associated colorectal tumorigenesis. Gut metabolites alteration, including elevated lysophosphatidic acid, which was confirmed to promote CRC cell proliferation and impair cell junction, was also observed in HFD-fed mice. Moreover, transfer of stools from HFD-fed mice to germ-free mice without interference increased colonic cell proliferation, impaired gut barrier function, and induced oncogenic genes expression. CONCLUSIONS: HFD drives colorectal tumorigenesis through inducing gut microbial dysbiosis, metabolomic dysregulation with elevated lysophosphatidic acid, and gut barrier dysfunction in mice.