Receptor Interacting Protein 3 Suppresses Vascular Smooth Muscle Cell Growth by Inhibition of the Phosphoinositide 3-Kinase-Akt Axis

Abstract

Proliferation of vascular smooth muscle cells (VSMCs) is a primary mechanism underlying cardiovascular proliferative disorders. Phosphoinositide 3-kinase (PI3K)-Akt (or protein kinase B) axis has been assigned at the center of pathways that regulate cell proliferation. Here we demonstrate that enhanced PI3K-Akt signaling by mitogenic stimulation or arterial injury profoundly elevates expression of receptor interacting protein 3 (RIP3) in primary cultured rat VSMCs and in vivo and that the up-regulation of RIP3 leads to VSMC growth arrest and apoptosis via inhibiting the PI3K-Akt signaling pathway, thereby alleviating balloon injury-induced neointimal formation. Specifically, mitogenic stimulation with platelet-derived growth factor-BB or angiotensin II leads to a profound increase in RIP3 expression, which is abolished by inhibition of PI3K or Akt, and increased PI3K-Akt signaling by expression of a constitutively active PI3K mutant also elevates RIP3 expression. Importantly, adenoviral overexpression of RIP3 not only triggers apoptosis but also causes cell cycle arrest at G 1 /G 0 phases that is associated with suppressed Akt activation. In sharp contrast, RIP3 gene silencing enhances serum-and platelet-derived growth factorinduced cell proliferation and Akt activation. In vivo adenoviral gene delivery of rat RIP3 (rRIP3) increased apoptosis and reduced VSMC proliferation, thus, effectively alleviating balloon injury-induced neointimal formation. The growth-suppressive and pro-apoptotic effects are independent of rRIP3 Ser/Thr kinase activity, because overexpression of a kinase-inactive mutant of rRIP3, similar to its wild type, is sufficient to induce growth arrest and apoptosis. These findings reveal a novel growth-suppressive action of RIP3, marking RIP3 as an important factor to prevent excessive mitogenic stimulation-or injury-induced vascular smooth muscle cells hyperplasia.To maintain tissue homeostasis, eukaryotic cells must keep a balance of cell proliferation and cell death in response to various sources of injury or stress stimuli. Cell hyper-proliferation has long been considered as an important etiological factor of cardiovascular diseases and cancer. Vascular smooth muscle cells (VSMCs) 4 are normally maintained in a non-proliferative state in the arterial tunica media. But arterial injury, inflammation, or excessive mitogenic stimulation triggers transmigration of VSMCs from the media into the intima layer of the arterial wall, where the VSMCs proliferate and synthesize extracellular matrix proteins, resulting in expansion of the arterial intima, i.e. neointimal formation (1-3). Proliferation of neointimal VSMCs is the most common causes of severe cardiovascular diseases such as hypertension, ischemic heart disease, and subsequent myocardial infarction, strokes, and congestive heart failure (4).Multiple factors, including growth factors, neurohormones, inflammatory cytokines, and reactive oxygen species, have been implicated in vascular proliferative disorders. For instance, platelet-derived gr...

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