DLPC decreases TGF-β1-induced collagen mRNA by inhibiting p38 MAPK in hepatic stellate cells

Abstract

Dilinoleoylphosphatidylcholine (DLPC), the active component of polyenylphosphatidylcholine extracted from soybeans, decreases collagen accumulation induced by TGF-β1 in cultured hepatic stellate cells (HSCs). Because DLPC exerts antioxidant effects and TGF-β1 generates oxidative stress, we evaluated whether the antifibrogenic effect of DLPC is linked to its antioxidant action. In passage 1 culture of rat HSCs, TGF-β1 induced a concentration-dependent increase in procollagen-α1(I) mRNA levels and enhanced intracellular H2O2 and superoxide anion formation and lipid peroxidation but decreased GSH levels. These changes were prevented by DLPC. Upregulation of collagen mRNA by TGF-β1 was likewise inhibited by catalase and p38 MAPK inhibitor SB-203580, suggesting involvement of H2O2 and p38 MAPK signaling in this process. TGF-β1 or addition of H2O2 to HSCs activated p38 MAPK with a rise in procollagen mRNA level; these changes were blocked by catalase and SB-203580 and likewise by DLPC. α-Smooth muscle actin abundance in HSCs was not altered by TGF-β1 treatment (with or without DLPC), indicating that downregulation of procollagen mRNA by DLPC was not due to alteration in HSC activation. These results demonstrate that DLPC prevents TGF-β1-induced increase in collagen mRNA by inhibiting generation of oxidative stress and associated H2O2-dependent p38 MAPK activation, which explains its antifibrogenic effect. DLPC, an innocuous phospholipid, may be considered for prevention and treatment of liver fibrosis.

Apoptotic Body Engulfment by a Human Stellate Cell Line Is Profibrogenic

“…It has been shown that the PI-3K inhibitor, LY294002, and the p38 MAPK inhibitor, SB203580, both reduced collagen mRNA levels in primary rodent stellate cells (Cao et al, 2002;Reif et al, 2003;VarelaRey et al, 2002). Similarly, we demonstrated that inhibiting PI-3K or p38 MAPK reduced collagen and TGF-␤1 mRNA, respectively, suggesting that both PI-3K and p38 MAPK pathways are activated in human stellate cells after apoptotic body engulfment.…”

“…Three major classes of MAP kinases have been described including p42/p44 MAPK, p38 MAPK, and SAPK/JNK (stress-activated protein kinase/c-Jun N-terminal kinase). Of these, the p38 MAPK signaling pathway is involved in the induction of procollagen ␣1(I) mRNA levels by TGF-␤1 (Cao et al, 2002;Varela-Rey et al, 2002). An additional kinase, phosphatidylinositol-3-kinase (PI3-K), also contributes to both type I collagen mRNA and protein expression in stellate cells (Reif et al, 2003).…”

Leptin Stimulates Tissue Inhibitor of Metalloproteinase-1 in Human Hepatic Stellate Cells

“…Leptin was used at concentrations of 25-100 ng/ml. The inhibitors and their concentrations were: 50 M JAK inhibitor AG 490 (45)(46)(47); 1000 units/ml catalase (49); 20 M p38 inhibitor SB203580 (39,48); 20 M SB202474 (an inactive analog of SB203580) (52); and 30 M ERK1/2 inhibitor PDO98059 (39,50). Me 2 SO was used as vehicle control for the inhibitors.…”

“…Rat Hepatic Stellate Cells-HSCs were isolated from male SpragueDawley rats and cultured in DMEM supplemented with 10% FCS and antibiotics until subconfluence as previously described (39). Cells were trypsinized, subcultured, and used 3 days later (as passage 1).…”

“…Furthermore, leptin was found to generate increased amounts of H 2 O 2 in isolated vascular endothelial cells in association with atherogenic processes (37,38). H 2 O 2 was shown to stimulate p38 in cultureactivated rat HSC, resulting in ␣1(I) procollagen mRNA up-regulation (39), and to phosphorylate ERK1/2 in human fibrosarcoma cells, leading to increased matrix metalloproteinase-1 transcription (40). In addition, Sohara et al (41) reported that ERK1/2 mediates TIMP-1 production in liver myofibroblasts in response to oncostatin M, which is, like leptin, a member of the interleukin-6 cytokine family that signals through class I cytokine receptors.…”