Metformin and Probiotics Interplay in Amelioration of Ethanol-Induced Oxidative Stress and Inflammatory Response in an In Vitro and In Vivo Model of Hepatic Injury

Abstract

Alcohol-induced liver injury implicates inflammation and oxidative stress as important mediators. Despite rigorous research, there is still no Food and Drug Administration (FDA) approved therapies for any stage of alcoholic liver disease (ALD). Interestingly, metformin (Met) and several probiotic strains possess the potential of inhibiting alcoholic liver injury. Therefore, we investigated the effectiveness of combination therapy using a mixture of eight strains of lactic acid-producing bacteria, commercialized as Visbiome® (V) and Met in preventing the ethanol-induced hepatic injury using in vitro and in vivo models. Human HepG2 cells and male Wistar rats were exposed to ethanol and simultaneously treated with probiotic V or Met alone as well as in combination. Endoplasmic reticulum (ER) stress markers, inflammatory markers, lipid metabolism, reactive oxygen species (ROS) production, and oxidative stress were evaluated, using qRT-PCR, Oil red O staining, fluorimetry, and HPLC. In vitro, probiotic V and Met in combination prevented ethanol-induced cellular injury, ER stress, oxidative stress, and regulated lipid metabolism as well as inflammatory response in HepG2 cells. Probiotic V and Met also promoted macrophage polarization towards the M2 phenotype in ethanol-exposed RAW 264.7 macrophage cells. In vivo, combined administration of probiotic V and Met ameliorated the histopathological changes, inflammatory response, hepatic markers (liver enzymes), and lipid metabolism induced by ethanol. It also improved the antioxidant markers (HO-1 and Nrf-2), as seen by their protein levels in both HepG2 cells as well as liver tissue using ELISA. Hence, probiotic V may act, in addition to the Met, as an effective preventive treatment against ethanol-induced hepatic injury.

Lactic Acid Bacteria Exert a Hepatoprotective Effect against Ethanol-Induced Liver Injury in HepG2 Cells

“…Treatment with five LAB led to the inhibition of SREBP1C and FAS mRNA expression. Our results are consistent with a report by Farhin et al who revealed that probiotics downregulated srebp1c and FAS in HepG2 cells exposed to ethanol [28]. Chronic alcohol consumption impedes lipid oxidation due to erroneous lysosomal biosynthesis, thereby delaying lipid degradation [30].…”

“…By investigating LAB, L. brevis MG5280 and MG5311, L. reuteri MG5458, L. fermentum MG4237, and MG4294 were found to show excellent efficacy at relieving oxidative stress by elevating GSH and antioxidant enzymes. Our results were similar to those obtained with Probiotic V, a product that includes various LAB, which was found to reduce oxidative stress by inhibiting lipid peroxidation and the expression of CYP2E1 in HepG2 cells exposed to ethanol [28]. Moreover, the activity of enzymes, such as AST and ALT, has been reported as one of the most sensitive markers of hepatotoxicity [29].…”

Macrophage Polarization and Its Role in Liver Disease

“…Furthermore, the study by Patel et al. ( 139 ) has shown that probiotics and metformin, as well as their combination, can promote M2 polarization and inhibit M1 polarization, contributing to the alleviation of alcoholic liver injury. The role and mechanisms of macrophage polarization in ALD are shown in Table 4 .…”

“…For example, inulin can inhibit short chain fatty acid-induced M1 polarization, and promote M2 polarization, attenuating the inflammation of ALD mice (138). Furthermore, the study by Patel et al (139) has shown that probiotics and metformin, as well as their combination, can promote M2 polarization and inhibit M1 polarization, contributing to the alleviation of alcoholic liver injury. The role and mechanisms of macrophage polarization in ALD are shown in Table 4.…”

Probiotics: Evolving as a Potential Therapeutic Option against Acetaminophen-Induced Hepatotoxicity

“…Probiotics and postbiotics have emerged as potential nutraceuticals with promises to improve the quality of life and prevent diseases [ 13 ]. Probiotics and postbiotics have also shown promise to rescue the liver from diseases and xenobiotic-induced hepatotoxicity, and restore normal liver functions [ 14 , 15 , 16 , 17 , 18 , 19 , 20 , 21 , 22 ].…”