Gut Dysbiosis and Increased Intestinal Permeability Drive microRNAs, NLRP-3 Inflammasome and Liver Fibrosis in a Nutritional Model of Non-Alcoholic Steatohepatitis in Adult Male Sprague Dawley Rats

Abstract

Larisse Longo,1,2 Jéssica Tonin Ferrari,2 Pabulo Henrique Rampelotto,2,3 Gustavo Hirata Dellavia,4 Amanda Pasqualotto,2 Claudia P Oliveira,5 Carlos Thadeu Schmidt Cerski,1,4,6 Themis Reverbel da Silveira,2 Carolina Uribe-Cruz,1,2 Mário Reis Álvares-da-Silva1,2,4,7 1Graduate Program in Gastroenterology and Hepatology, Universidade Federal do Rio Grande do Sul, Porto Alegre, Rio Grande do Sul, Brazil; 2Experimental Laboratory of Hepatology and Gastroenterology, Center for Experimental Research, Hospital de Clínicas de Porto Alegre, Porto Alegre, Rio Grande do Sul, Brazil; 3Graduate Program in Pharmaceutical Sciences, Universidade Federal do Rio Grande do Sul, Porto Alegre, Rio Grande do Sul, Brazil; 4School of Medicine, Universidade Federal do Rio Grande do Sul, Porto Alegre, Rio Grande do Sul, Brazil; 5Department of Gastroenterology (LIM07), Faculdade de Medicina da Universidade de São Paulo, São Paulo, Brazil; 6Unit of Surgical Pathology, Hospital de Clínicas de Porto Alegre, Porto Alegre, Rio Grande do Sul, Brazil; 7Division of Gastroenterology, Hospital de Clínicas de Porto Alegre, Porto Alegre, Rio Grande do Sul, BrazilCorrespondence: Larisse Longo Email [email protected]/Aim: The interactions between the gut and liver have been described in the progression of non-alcoholic steatohepatitis (NASH). The aim of this study was to develop an experimental nutritional model of NASH simulating metabolic changes occurring in humans.Materials and Methods: Adult male Sprague Dawley rats were randomized into two groups: controls (standard diet) and intervention (high-fat and choline-deficient diet) for 16 weeks, each experimental group with 10 animals. Biochemical analysis, hepatic lipid content, microRNAs, inflammatory, gut permeability markers and gut microbiota were measured.Results: Animals in the intervention group showed significantly higher delta Lee index (p=0.017), abdominal circumference (p< 0.001), abdominal adipose tissue (p< 0.001) and fresh liver weight (p< 0.001), as well as higher serum levels of alanine aminotransferase (p=0.010), glucose (p=0.013), total cholesterol (p=0.033), LDL cholesterol (p=0.011), and triglycerides (p=0.011), and lower HDL cholesterol (p=0.006) compared to the control group. Higher TLR4 (p=0.041), TLR9 (p=0.033), MyD88 (p=0.001), Casp1 (p< 0.001), NLPR3 (p=0.019), liver inflammation index interleukin (IL)-1β/IL10 (p< 0.001), IL6/IL10 (p=0.002) and TNFα/IL10 (p=0.001) were observed in the intervention group, and also lower permeability markers Ocln (p=0.003) and F11r (p=0.041). Gene expression of miR-122 increased (p=0.041) and miR-145 (p=0.010) decreased in the intervention group. Liver steatosis, inflammation and fibrosis, along with collagen fiber deposition increment (p< 0.001), were seen in the intervention group. Regarding gut microbiota, Bray-Curtis dissimilarity index and number of operational taxonomic units were significantly different (p< 0.001) between the groups. Composition of the gut microbiota showed a significant correlation with histopathological score of NAFLD (r=0.694) and index IL-1β/IL-10 (r=0.522).Conclusion: This experimental model mimicking human NASH demonstrated gut and liver interaction, with gut microbiota and intestinal permeability changes occurring in parallel with systemic and liver inflammation, miRNAs regulation and liver tissue damage.Keywords: fatty liver disease models, fibrosis, gut microbiota, non-alcoholic fatty liver disease, non-alcoholic steatohepatitis

Keywords

• fatty liver disease models
• fibrosis
• gut microbiota
• non-alcoholic fatty liver disease
• non-alcoholic steatohepatitis