Red wine-derived resveratrol exacerbates the effects of free fatty acids on hepatic stellate cells in vitro whereby promoting fibrogenesis

Aims: At moderate consumption, the red wine-derived resveratrol (i.e., trans-3,5,4′-trihydroxystilbene) has been shown to improve the health and survival of obese mice. This nonflavonoid polyphenol also acts antioxidative, antineoplastic and antiinflammatory. In contrast, free fatty acids (FFAs) contribute to liver fibrogenesis by acting profibrotically. FFAs also induce apoptosis as a prominent feature of non-alcoholic fatty liver disease (NAFLD).

Aim: To assess potential downregulation by resveratrol of FFA-induced pro-fibrogenic genes (TGFβ1, α-SMA, TIMP1) and/or the expression of death receptors (Fas/CD95, TNFR1) and apoptosis-related molecules in the LX-2 stellate cell line.

Methods: LX-2 cells were first (48h) incubated with 0.5 mM FFAs (2:1 oleate:palmitate) and subsequently (24h) treated with or without resveratrol at 5, 10, or 15µM. The expression of fibrosis- and apoptosis-related genes was quantified by real-time PCR. The LD50 of resveratrol in the additional presence of 0.5 mM FFAs was determined by the MTT cytotoxicity assay.

Results: The LD50 of resveratrol plus FFAs was calculated as 23.2µM. Incubation of LX-2 cells with 0.5 mM FFAs only slightly upregulated the activation- and fibrosis-related genes α-SMA (x2.9), TGFβ1 (x1.6), and TIMP1 (x1.6x). We also found upregulation of the expression of select death receptors (Fas/CD95: x3.5; TNF-R1: x1.4), as well as Bcl-2 (x2.3) and Mcl1 (x1.3). Further addition of 15µM resveratrol further upregulated these genes (α-SMA: x3.7; TGFβ1: x1.9; TIMP1: x2.3; Fas/CD95: x7.5; TNF-R1: x2.1; Bcl2×3.4; and Mcl1: x1.8), while resveratrol had no significant effect in the absence of FFAs.

Conclusion: FFAs activate LX-2 hepatic stellate cells. Unexpectedly, resveratrol further upregulated these FFA-mediated effects on the molecular level. The additional finding that resveratrol has virtually no effect in the absence of FFAs suggests that both act synergistically and that, therefore, and despite current recommendation, obese patients should be discouraged from consuming red wine even at moderate amounts.