Regulation of TGFβ1-mediated collagen formation by LOX-1 -: Studies based on forced overexpression of TGFβ1 in wild-type and LOX-1 knock-out mouse cardiac fibroblasts

Transforming growth factor beta(1) (TGF beta(1)) activation leads to tissue fibrosis. Here, we report on the role of LOX-1, a lectin-like 52-kDa receptor for oxidized low density lipoprotein, inTGF beta(1)-mediated collagen expression and underlying signaling in mouse cardiac fibroblasts. TGF beta(1) was overexpressed in wildtype (WT) and LOX-1 knock-out mouse cardiac fibroblasts by transfection with adeno-associated virus type 2 vector carrying the active TGF beta(1) moiety (AAV/TGF beta(ACT)(1)). Transfection of WT mouse cardiac fibroblasts with AAV/TGF beta(ACT)(1) markedly enhanced the expression of NADPH oxidases (p22(phox), p47(phox), and gp91(phox) subunits) and LOX-1, formation of reactive oxygen species, and collagen synthesis, concomitant with an increase in the activation of p38 and p44/42 mitogen-activated protein kinases (MAPK). The TGF beta(1)-mediated increase in collagen synthesis was markedly attenuated in the LOX-1 knock-out mouse cardiac fibroblasts as well as in WT mouse cardiac fibroblasts treated with a specific anti-LOX-1 antibody. Treatment with anti-LOX-1 antibody also reduced NADPH oxidase expression and MAPK activation. The NADPH oxidase inhibitors and gp91phox small interfering RNA reduced LOX-1 expression, MAPK activation, and collagen formation. The p38 MAPK inhibitors as well as the p44/42 MAPK inhibitors reduced collagen formation without affecting LOX-1 expression in cardiac fibroblasts. These observations suggest that collagen synthesis in cardiac fibroblasts involves a facilitative interaction between TGF beta(1)-NADPH oxidase and LOX-1. Further, the activation of MAPK pathway appears to be downstream of TGF beta(1)-reactive oxygen species-LOX-1 cascade.