Mouse Hepatic Oval Cells Require Met-Dependent PI3K to Impair TGF-β-Induced Oxidative Stress and Apoptosis

We have previously shown that oval cells harboring a genetically inactivated Met tyrosine kinase (Met(-/-) oval cells) are more sensitive to TGF-beta-induced apoptosis than cells expressing a functional Met (Met(flx/flx)), demonstrating that the HGF/Met axis plays a pivotal role in oval cell survival. Here, we have examined the mechanism behind this effect and have found that TGF-beta-induced a mitochondria-dependent apoptotic cell death in Met(flx/flx) and Met(-/-) oval cells, associated with a marked increase in levels of the BH3-only proteins Bim and Bmf. Bmf plays a key role during TGF-beta-mediated apoptosis since knocking down of BMF significantly diminished the apoptotic response in Met(-/-) oval cells. TGF-beta also induced oxidative stress accompanied by NADPH oxidase 4 (Nox4) mRNA up-regulation and decreased protein levels of antioxidant enzymes. Antioxidants inhibit both TGF-beta-induced caspase 3 activity and Bmf up-regulation, revealing an oxidative stress-dependent Bmf regulation by TGF-beta. Notably, oxidative stress-related events were strongly amplified in Met(-/-) oval cells, emphasizing the critical role of Met in promoting survival. Pharmacological inhibition of PI3K did impair HGF- driven protection from TGF-beta-induced apoptosis and increased sensitivity of Met(flx/flx) oval cells to TGF-beta by enhancing oxidative stress, reaching apoptotic indices similar to those obtained in Met(-/-) oval cells. Interestingly, both PI3K inhibition and/ or knockdown itself resulted in caspase-3 activation and loss of viability in Met(flx/flx) oval cells, whereas no effect was observed in Met(-/-) oval cells. Altogether, results presented here provide solid evidences that both paracrine and autocrine HGF/Met signaling requires PI3K to promote mouse hepatic oval cell survival against TGF-beta-induced oxidative stress and apoptosis.