n-caproylsphingosine and Hyperplasia

We have previously demonstrated that hexanoyl-D-erythro-sphingosine (C(6)-ceramide), an anti-mitogenic cell-permeable lipid metabolite, limited vascular smooth muscle growth by abrogating trauma-induced Akt activity in a stretch injury model of neointimal hyperplasia. Furthermore, ceramide selectively and directly activated protein kinase C zeta (PKC zeta) to suppress Akt-dependent mitogenesis. To further analyze the interaction between ceramide and PKC zeta, the ability of ceramide to localize within highly structured lipid microdomains (rafts) and activate PKC zeta was investigated. Using rat aorta vascular smooth muscle cells (A7r5), we now demonstrate that C(6)-ceramide treatment results in an increased localization and phosphorylation of PKC zeta within caveolin-enriched lipid microdomians to inactivate Akt. In addition, ceramide specifically reduced the association of PKC zeta with 14-3-3, a scaffold protein localized to less structured regions within membranes. Pharmacological disruption of highly structured lipid microdomains resulted in abrogation of ceramide-activated, PKC zeta-dependent Akt inactivation, whereas molecular strategies suggest that ceramide-dependent PKC zeta phosphorylation of Akt3 at Ser(34) was necessary for ceramide-induced vascular smooth muscle cell growth arrest. Taken together, these data demonstrate that structured membrane microdomains are necessary for ceramide-induced activation of PKC zeta and resultant diminished Akt activity, leading to vascular smooth muscle cell growth arrest.

Topics: 14-3-3 Proteins; Animals; Caveolins; Cell Line; Ceramides; Enzyme Activation; Hyperplasia; Membrane Microdomains; Mitosis; Muscle, Smooth, Vascular; Protein Kinase C; Proto-Oncogene Proteins c-akt; Rats; Tunica Intima

Neointimal hyperplasia at the site of surgical intervention is a common and deleterious complication of surgery for cardiovascular diseases. We hypothesized that direct delivery of a cell-permeable growth-arresting lipid via the balloon tip of an embolectomy catheter would limit neointimal hyperplasia after stretch injury. We have previously demonstrated that sphingolipid-derived ceramide arrested the growth of smooth muscle cell pericytes in vitro. Here, we show that ceramide-coated balloon catheters significantly reduced neointimal hyperplasia induced by balloon angioplasty in rabbit carotid arteries in vivo. This ceramide treatment decreased the number of vascular smooth muscle cells entering the cell cycle without inducing apoptosis. In situ autoradiographic studies demonstrated that inflating the balloon catheter forced cell-permeable ceramide into the intimal and medial layers of the artery. Intercalation of ceramide into vascular smooth muscle cells correlated with rapid inhibition of trauma-associated phosphorylation of extracellular signal-regulated kinase and protein kinase B. These studies demonstrate the utility of cell-permeable ceramide as a novel therapy for reducing neointimal hyperplasia after balloon angioplasty.

Topics: Angioplasty, Balloon, Coronary; Animals; Apoptosis; Carotid Artery Injuries; Carotid Stenosis; Ceramides; Disease Models, Animal; Hyperplasia; Mitogen-Activated Protein Kinases; Muscle, Smooth, Vascular; Postoperative Complications; Rabbits; Tunica Intima