Mouse hepatic oval cells require Met-dependent PI3K to impair TGF-β-induced oxidative stress and apoptosis

PLoS One. 2013;8(1):e53108. doi: 10.1371/journal.pone.0053108. Epub 2013 Jan 2.

Abstract

We have previously shown that oval cells harboring a genetically inactivated Met tyrosine kinase (Met(-/-) oval cells) are more sensitive to TGF-β-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-β 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-β-mediated apoptosis since knocking down of BMF significantly diminished the apoptotic response in Met(-/-) oval cells. TGF-β also induced oxidative stress accompanied by NADPH oxidase 4 (Nox4) mRNA up-regulation and decreased protein levels of antioxidant enzymes. Antioxidants inhibit both TGF-β-induced caspase 3 activity and Bmf up-regulation, revealing an oxidative stress-dependent Bmf regulation by TGF-β. 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-β-induced apoptosis and increased sensitivity of Met(flx/flx) oval cells to TGF-ß 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-β-induced oxidative stress and apoptosis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antioxidants / metabolism
  • Apoptosis*
  • Caspase 3 / metabolism
  • Cell Line
  • Glutathione / metabolism
  • Hepatocyte Growth Factor / metabolism
  • Hepatocytes / cytology*
  • Membrane Potentials
  • Mice
  • Mitochondria / metabolism
  • Oxidative Stress*
  • Phosphatidylinositol 3-Kinases / metabolism*
  • Phosphatidylserines / metabolism
  • Proto-Oncogene Proteins c-met / metabolism*
  • RNA, Small Interfering / metabolism
  • Reactive Oxygen Species / metabolism
  • Signal Transduction
  • Transforming Growth Factor beta / pharmacology*

Substances

  • Antioxidants
  • Phosphatidylserines
  • RNA, Small Interfering
  • Reactive Oxygen Species
  • Transforming Growth Factor beta
  • Hepatocyte Growth Factor
  • Phosphatidylinositol 3-Kinases
  • Proto-Oncogene Proteins c-met
  • Casp3 protein, mouse
  • Caspase 3
  • Glutathione

Grants and funding

AMP was recipient of a research-training contract (grant SAF2006-12025) from the Ministry of Education and Science. GC was recipient of a research-training contract from the Ministry of Education and Science. ASC was recipient of an Alban scholarship program and then a research assistant contract (grant SAF2009-12477). MGA is recipient of a research assistant contract (grant S2010/BMD-2402). This work has been supported by grants SAF2006-12025 from Ministry of Education and Science (Spain), SAF2009-12477 from Ministry of Science and Innovation (Spain), and 920359 (CAM-UCM, BSCH-UCM). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.