Mammalian target of rapamycin mediates the angiogenic effects of leptin in human hepatic stellate cells.
Leptin modulates the angiogenic properties of hepatic stellate cells (HSC), but the molecular mechanisms involved are poorly understood. We investigated the pathways regulating hypoxia-inducible factor 1α (HIF-1α) and vascular endothelial growth factor (VEGF) in leptin-stimulated myofibroblastic HSC. Exposure to leptin enhanced the phosphorylation of TSC2 on T1462 residues and of p70 S6 kinase and the translational inhibitor 4E-binding protein-1, indicating the ability of leptin to activate the mammalian target of rapamycin (mTOR) pathway. Similar findings were observed when HSC were exposed to PDGF. Both leptin and PDGF increased the expression of HIF-1α and VEGF in HSC. In the presence of rapamycin, a specific mTOR inhibitor, leptin and PDGF were no longer able to activate mTOR, and expression of VEGF was reduced, whereas HIF-1α abundance was not affected. Moreover, knockdown of Raptor, a component of the mTORC1 complex, reduced the ability of leptin to increase VEGF. mTOR was also necessary for leptin- and PDGF-dependent increase in HSC migration. Leptin increased the generation of reactive oxygen species in HSC, which was reduced by NADP(H) oxidase inhibitors. Both N-acetyl cysteine and diphenylene iodonium, a NADP(H) inhibitor, inhibited the expression of HIF-1α and VEGF stimulated by leptin or PDGF. Finally, conditioned media from HSC treated with leptin or PDGF induced tube formation in cultured human umbilical vein endothelial cells. In conclusion, in HSC exposed to leptin or PDGF, increased expression of VEGF requires both activation of mTOR and generation of reactive oxygen species via NADPH-oxidase. Induction of HIF-1α requires NADP(H) oxidase but not mTOR activation.
Dipartimento di Medicina Interna, University of Florence, Italy.
Sara
S
Aleffi
Nadia
N
Navari
Wanda
W
Delogu
Sara
S
Galastri
Erica
E
Novo
Krista
K
Rombouts
Massimo
M
Pinzani
Maurizio
M
Parola
Fabio
F
Marra
D002460:Cell Line; D002465:Cell Movement / Q000502:physiology; D056945:Hep G2 Cells; D055166:Hepatic Stellate Cells / Q000378:metabolism / Q000473:pathology / Q000502:physiology*; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism* / Q000502:physiology; D020738:Leptin / Q000378:metabolism / Q000502:physiology*; D008099:Liver / Q000098:blood supply*; D019255:NADPH Oxidases / Q000502:physiology; D009389:Neovascularization, Pathologic; D018919:Neovascularization, Physiologic; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000502:physiology; D017382:Reactive Oxygen Species / Q000378:metabolism; D038762:Ribosomal Protein S6 Kinases, 70-kDa / Q000737:chemistry; D015398:Signal Transduction / Q000502:physiology*; D058570:TOR Serine-Threonine Kinases / Q000502:physiology*; D000077005:Tuberous Sclerosis Complex 2 Protein; D025521:Tumor Suppressor Proteins / Q000737:chemistry; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism* / Q000502:physiology
D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D020738:Leptin; D010982:Platelet-Derived Growth Factor; D017382:Reactive Oxygen Species; C000624653:TSC2 protein, human; D000077005:Tuberous Sclerosis Complex 2 Protein; D025521:Tumor Suppressor Proteins; D042461:Vascular Endothelial Growth Factor A; D019255:NADPH Oxidases; C546842:MTOR protein, human; D038762:Ribosomal Protein S6 Kinases, 70-kDa; D058570:TOR Serine-Threonine Kinases