Sirolimus Inhibits Platelet-Derived Growth Factor–Induced Collagen Synthesis in Rat Vascular Smooth Muscle Cells

Abstract

Vascular smooth muscle cell (VSMC) proliferation and extracellular matrix (ECM) accumulation play key roles in the development and the progression of vascular remodeling such as transplant arteriosclerosis and restenosis. The present study examined the effects of sirolimus (SRL) on platelet-derived growth factor (PDGF)-induced fibronectin secretion, collagen synthesis, and the related signaling pathways including reactive oxygen species (ROS) and mitogen-activated protein kinases (MAPK) in rat VSMCs. Primary rat VSMCs were isolated from male Sprague-Dawley rats. Growth arrested, synchronized cells were treated with various concentrations of SRL before the addition of PDGF at 10 ng/mL. Proliferating cell nuclear antigen expression, fibronectin secretion, and the activation of extracellular signal-regulated protein kinase (ERK) and p38 MAPK were assessed by Western blot analysis, collagen synthesis by [(3)H]-proline incorporation, and cellular ROS by flow cytometry. PDGF (10 ng/mL) increased VSMC proliferation by 1.7-fold, fibronectin secretion by 1.5-fold, collagen synthesis by 2.1-fold, cellular ROS by 1.6-fold, and activation of ERK and p38 MAPK by 3.3- and 3.9-fold compared to controls. SRL above 1 nmol/L inhibited PDGF-induced VSMC proliferation and collagen synthesis but not PDGF-induced fibronectin secretion, cellular ROS, and activation of ERK and p38 MAPK. These data demonstrated that PDGF increased ECM synthesis as well as proliferation through cellular ROS and subsequent MAPK activation and that SRL inhibited PDGF-induced VSMC proliferation and collagen synthesis in a cellular ROS- and MAPK activation-independent way.