Investigation of Microbiome Metabolites and Mitochondrial Function in a model of Non-Alcoholic Fatty Liver Disease

Abstract

Several dietary metabolites produced by gut bacteria have been linked to disease including non-alcoholic fatty liver disease (NAFLD). Trimethylamine N-oxide (TMAO) and phenylacetic acid (PAA) are microbiome-derived metabolites that have been associated with early onset of NAFLD. Hypothesising that these metabolites contribute to lipid deposition in the liver by altering hepatic mitochondrial function, we assessed how TMAO and PAA affect hepatocyte bioenergetics in cell models of liver steatosis. Proliferative and differentiated HepaRG cells were cultured under standard conditions, and steatosis was established by 48h exposure to oleate and palmitate (2:1 molar ratio). Lipid accumulation was assessed by BODIPY™ staining and quantified by CellProfiler software. To gain insight into HepaRG mitochondrial respiration, we measured a number of bioenergetic parameters with the Seahorse extracellular flux analyser in control cells and cells exposed to PAA (100 µM and 200 µM) or TMAO (20 µM, 50 µM, 100 µM and 200 µM). PAA and TMAO led to an increase in lipid deposition in HepaRG cells. TMAO caused a 42% increase in lipid accumulation in proliferative cells and 1.6% in differentiated HepaRG cells while PAA exacerbated intracellular lipid droplets by 54% in proliferative HepaRG and by 10% in fully differentiated cells. The same effects were seen in mitochondrial function. PAA and TMAO lowered spare respiratory capacity dose-dependently, maximal respiration and 7 basal oxygen consumption in HepaRG cells. In addition, the decline in mitochondrial function preceded lipid accumulation. Our data indicate that microbiome-derived compounds decrease mitochondrial capacity significantly and exacerbate lipid deposition suggesting a potential link between microbiome metabolite driven mitochondrial dysfunction and NAFLD onset.

Date of Award	2023
Original language	English
Awarding Institution	University of Plymouth
Supervisor	David Sheridan (Other Supervisor)