Insulin Stabilizes Microvascular Endothelial Barrier Function via Phosphatidylinositol 3-Kinase/Akt-Mediated Rac1 Activation

Abstract

Objective-Insulin is a key regulator of metabolism, but it also confers protective effects on the cardiovascular system.Here, we analyze the mechanism by which insulin stabilizes endothelial barrier function. Methods and Results-Insulin reduced basal and antagonized tumor necrosis factor-␣-induced macromolecule permeability of rat coronary microvascular endothelial monolayers. It also abolished reperfusion-induced vascular leakage in isolated-perfused rat hearts. Insulin induced dephosphorylation of the regulatory myosin light chains, as well as translocation of actin and vascular endothelial (VE)-cadherin to cell borders, indicating a reduction in contractile activation and stabilization of cell adhesion structures. These protective effects were blocked by genistein or Hydroxy-2-naphthalenylmethylphosphonic acid tris acetoxymethyl ester (HNMPA-[AM] 3 ), a pan-tyrosine-kinase or specific insulin-receptor-kinase inhibitor, respectively. Insulin stimulated the phosphatidylinositol 3-kinase (PI3K)/Akt pathway and NO production, and it activated Rac1. Inhibition of PI3K/Akt abrogated Rac1 activation and insulin-induced barrier protection, whereas inhibition of the endothelial nitric oxide synthase/soluble guanylyl cyclase pathway partially inhibited them. Inhibition of Rac1 abrogated the assembly of actin at cell borders. Accordingly, it abolished the protective effect of insulin on barrier function of the cultured endothelial monolayer, as well as the intact coronary system of ischemic-reperfused hearts. Key Words: capillary permeability Ⅲ coronary circulation Ⅲ endothelium Ⅲ nitric oxide Ⅲ vascular biology I nsulin is an essential hormone of metabolic homeostasis. Recent clinical findings show that intensive insulin therapy has vasoprotective effects under inflammatory conditions 1,2 and reduces major cardiovascular events in diabetics. 3 These effects of insulin seem to be independent of its metabolic effects on endothelial cells. Conclusion-InsulinVascular endothelial cells forming the inner lining of all vessels play an important role in the regulation of vascular homeostasis. They provide a semiselective barrier for water, solutes, macromolecules, and blood-borne components and are also involved in regulating the trafficking of blood cells across the vessel wall. This function can be altered by a variety of diverse circulating vasoactive inflammatory mediators and hormones such as insulin. It has recently been demonstrated that endothelial cells of macrovascular origin express insulin receptors. 4 Binding of insulin to the receptors induces nitric oxide production via phosphatidylinositol 3-kinase (PI3K)-mediated activation of endothelial nitric oxide synthase (eNOS) 5 and causes relaxation of the smooth muscles. Recently, it has been shown that insulin reduces mesenteric venular albumin leakage on systemic insulin administration in rats. 6 However, the underlying signaling mechanism of this effect is largely unknown.The integrity of the endothelial barrier is highly dependent on the endothelial actomyosi...

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