Perlecan deficiency causes endothelial dysfunction by reducing the expression of endothelial nitric oxide synthase

Perlecan is a major heparan sulfate proteoglycan found in the subendothelial extracellular matrix of the vascular wall. The aim of this study was to investigate the role of perlecan in the regulation of vascular tone. A previously developed conditional perlecan-deficient mouse model was used to measure changes in the isometric force of isolated aortic rings. The vessels were first precontracted with phenylephrine, and then treated with increasing concentrations of vasorelaxants. Endothelium-dependent relaxation, elicited by acetylcholine, was significantly reduced in the perlecan-deficient aortas, whereas endothelium-independent relaxation caused by the exogenous nitric oxide donor sodium nitroprusside remained well preserved. The expression of the endothelial nitric oxide synthase (eNOS) gene, detected by real-time polymerase chain reaction, was significantly decreased in the perlecan-deficient aortas. The expression of eNOS protein detected using Western blotting was also significantly decreased in the perlecan-deficient aortas. We examined the role of perlecan in eNOS gene expression by creating perlecan knockdown human aortic endothelial cells using small interfering RNA (siRNA) for perlecan. Perlecan gene expression was significantly reduced in the perlecan siRNA-treated cells, resulting in a significant decrease in eNOS gene expression. Perlecan deficiency induced endothelial dysfunction, as indicated by a reduction in endothelium-dependent relaxation due, at least partly, to a reduction in eNOS expression. These findings suggest that perlecan plays a role in the activation of eNOS gene expression during normal growth processes.