Leptin as a novel profibrogenic cytokine in hepatic stellate cells: mitogenesis and inhibition of apoptosis mediated by extracellular regulated kinase (Erk) and Akt phosphorylation

Abstract

A key feature in the molecular pathogenesis of liver fibrosis requires maintenance of the activated hepatic stellate cell (HSC) phenotype by both proliferation and inhibition of apoptosis. We provide evidence that leptin is a potent HSC mitogen and dramatically inhibits stellate cell apoptosis. Leptin proved to be as potent an HSC mitogen as platelet-derived growth factor (PDGF) as assessed by bromodeoxyuridine (BrdU) incorporation in isolated primary HSCs; data using fluorescent propidium iodide (PI) uptake revealed that leptin, like PDGF, increased HSC populations in the S-and G 2 /M-phases of the cell cycle. Leptin resulted in a robust increase in cyclin D1 expression. Using the chemical inhibitor of Janus kinase 2 (Jak2) activity, AG 490, and overexpression of the suppressor of cytokine signaling 3 (SOCS-3), we show that blockade of leptin receptor (Ob-Rb) phosphorylation blocks leptin-induced HSC proliferation. Leptinassociated phosphorylation of both extracellular regulated kinase (p44/p42, Erk) and Akt is also prohibited. Further, the PI-3 kinase inhibitor LY294002 and MAPK inhibitor PD98059 were found to significantly reduce leptin-induced HSC proliferation, thereby indicating that leptin induced HSC proliferation is Akt-and Erk-dependent. Akt was also protective against HSC apoptosis. Leptin abolished both cycloheximide-induced and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis, demonstrated by reduced caspase-3 activity, HSC-TUNEL staining, and DNA fragmentation. We conclude that leptin acts as a direct hepatic stellate cell survival agonist. Importantly, we have demonstrated that leptin-induced HSC proliferation and survival by Ob-Rb phosphorylation are both Erk-and Akt-dependent. Keywords nonalcoholic steohepatitis (NASH); liver fibrosis; proliferation; MAP kinase; PI-3 kinaseThe role of leptin, a 16 kDa protein hormone, in liver disease may be distinctly important as human obesity and diabetes mellitus type II become more prevalent (1). Nonalcoholic steatohepatitis (NASH) is characterized by fatty change of the liver with various degrees of inflammation and fibrosis (2,3). The exact mechanism by which leptin promotes liver fibrosis is unknown. Recently, a role for leptin in the molecular pathogenesis of mammalian fibrosis has been substantiated in glomerulosclerosis (4) as well as in liver fibrosis (5)(6)(7)(8) NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript peripheral tissues, leptin exerts control on body weight homeostasis via inhibitory actions on glucose metabolism and insulin secretion (9). Ob-Rb is the functional form of the membrane-associated leptin receptor. It is related to the class II cytokine group receptors binding interleukin-2, interferon, and growth hormone and is closely related to the gp130 signal-transducing component of the interleukin-6 and the G-CSF receptor (10,11). The long form of the leptin receptor, Ob-Rb, is present in various peripheral organs, including inflammatory blood cells, lung, kidney, liver,...

Sonic hedgehog is an autocrine viability factor for myofibroblastic hepatic stellate cells

“…These findings extend and confirm data derived from earlier studies of clonal HSC lines, in which Shh antibodies increased cellular apoptosis and recombinant Shh stimulated growth [29]. The present study also demonstrates that proliferative actions of PDGF-BB, a major HSC mitogen [45,46], are mediated via the Hh pathway because three distinct approaches that abrogate Hh signaling each significantly inhibited PDGF-related increases in HSC incorporation of BrdU. Both agents that specifically antagonize Hh signaling by preventing activation of discrete Hh pathway signaling components (Anti-Shh antibodies prevent Shh from activating Ptc, and cyclopamine prevents Ptc from activating Smo) [60], exerted comparable effects on HSC DNA synthesis, reducing the stimulatory actions of PDGF-BB by about 50%.…”

“…As expected [32,45,46], PDGF-BB elicited rapid phosphorylation of AKT on Thr308 and Ser473, with peak AKT activation occurring within the initial hour after PDGF-BB addition (Fig 4A). The ensuing accumulation of Shh protein was inhibited by addition of either the PI3 kinase inhibitor, Ly294002, which inhibits PDGF-BB activation of AKT [45] (Fig 4B) or adenovirally-mediated delivery of dominant negative AKT (Ad5dnAkt) (Fig 4C). The effects of Ad5dnAkt were specific because treatment with a control adenoviral vector (Ad5GFP) did not inhibit PDGF induction of Shh.…”

“…PDGF-BB-activation of AKT mediates PDGF-BB's trophic actions [32,45,46]. Therefore, subsequent studies addressed the effect of AKT activation on HSC expression of Shh.…”

Cellular and molecular mechanisms in liver fibrogenesis

“…Moreover, leptin has been shown to up-regulate type I procollagen expression, to potentiate TGFβ1-mediated effects [53], to induce tissue inhibitor of metalloproteinase-1 (TIMP1) expression [54] as well as to stimulate HSC proliferation and survival [55], and up-regulate expression of chemokines (like CCL2) in a NF-kB-dependent way [52]. Although leptin may stimulate multiple signaling pathways, literature data suggest that its pro-fibrogenic effects are mainly NADPH oxidase and ROS dependent [56] and may also be mediated by inhibition of the expression and activity of peroxisome proliferator-activated receptor- (PPAR-), which maintains HSC quiescence and reverses HSC trans-differentiation to MFs.…”

JAK-STAT pathway in carcinogenesis: Is it relevant to cholangiocarcinoma progression?

“…Leptin acts as a direct hepatic stellate cell survival agent [81][82][83] . Serum leptin is also elevated after common bile duct ligation and CCl4 induced liver fibrosis that are both risk factors for cholangiocarcinoma development [84] .…”