The grape-derived polyphenol resveratrol differentially affects epidermal and platelet-derived growth factor signaling in human liver myofibroblasts

https://doi.org/10.1016/j.biocel.2005.11.001Get rights and content

Abstract

The grape-derived polyphenol resveratrol is anti-proliferative for human liver myofibroblasts, which may be beneficial for the treatment of liver fibrosis. However, its mechanism of action is ill understood. Here, we have studied how resveratrol interfered with signaling pathways used by epidermal or platelet-derived growth factors to induce the proliferation of these cells.

We found that resveratrol inhibited epidermal growth factor or platelet-derived growth factor-induced DNA synthesis. Resveratrol did not, however, decrease epidermal growth factor receptor autophosphorylation or activation of extracellular regulated kinases, but strongly inhibited the phosphorylation of Akt and of its substrate forkhead related transcription factor. This suggested that resveratrol inhibited epidermal growth factor-induced mitogenic signaling through inhibition of the phosphatidylinositol 3-kinase /Akt pathway. The phosphatidylinositol 3-kinase inhibitor LY 294002, also, inhibited epidermal growth factor-dependent DNA synthesis and Akt phosphorylation but did not decrease extracellular regulated kinases phosphorylation. In contrast, resveratrol inhibited platelet-derived growth factor-stimulated receptor autophosphorylation and every subsequent signaling step. Resveratrol did not directly inhibit phosphatidylinositol 3-kinase activity measured on immunoprecipitates from epidermal growth factor-stimulated myofibroblasts, but it strongly reduced the autophosphorylation of the phosphatidylinositol 3-kinase downstream target phospho-inositide-dependent kinase-1 that phosphorylates Akt.

We, thus, show that resveratrol has growth factor-specific effects: it inhibits platelet-derived growth factor signaling via reduced receptor activation, whereas it reduces epidermal growth factor-dependent DNA synthesis via inhibition of the phosphatidylinositol 3-kinase/Akt pathway, possibly through inhibition of phospho-inositide-dependent kinase-1 activity.

Section snippets

Materials

Culture medium, additives and recombinant human EGF and PDGF-BB were from Invitrogen (Life Technologies, Cergy-Pontoise, France). Human AB serum was from Etablissement Français du Sang (Bordeaux, France). Polyvinylidene difluoride membranes were from Millipore (Saint-Quentin en Yvelines, France). Chemicals were from Sigma–Aldrich (Saint-Quentin Fallavier, France). Hyperfilm and the ECL kit were from Amersham (Les Ulis, France). [3H] thymidine was from ICN Biomedicals (Orsay, France).

Resveratrol

Effect of resveratrol on EGF- and PDGF-BB-dependent DNA synthesis

As expected, stimulation with EGF caused a two-fold increase in [3H]thymidine incorporation. Treatment with resveratrol resulted in a dose-dependent inhibition of EGF-induced [3H]thymidine incorporation with an IC50 value of 26.0 ± 1.4 μM (Fig. 1A). Results were normalized with those of a MTT assay conducted in replicate wells; it should be stressed that the MTT results did not differ significantly in any condition (not shown), in agreement with our extensive previous results showing that

Discussion

In a previous study, we have reported that resveratrol was anti-proliferative for human liver myofibroblasts grown in the presence of serum (Godichaud et al., 2000). Here, we show that resveratrol inhibits DNA synthesis when stimulated by EGF or PDGF-BB. As previously described, this was not accompanied by any cytotoxicity (Godichaud et al., 2000). In order to find the target of the anti-proliferative effect of resveratrol, we first examined the effect of resveratrol on growth factor receptor

Acknowledgments

The study was supported in part by grants from Institut de Recherches sur les Boissons and Institut Vin et Santé. SG was the recipient of a fellowship from Berkem Chimie Fine, Gardonne, France.

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