Targeting Endothelial Dysfunction in Vascular Complications Associated with Diabetes

Cardiovascular complications associated with diabetes remain a significant health issue in westernized societies. Overwhelming evidence from clinical and laboratory investigations have demonstrated that these cardiovascular complications are initiated by a dysfunctional vascular endothelium. Indeed, endothelial dysfunction is one of the key events that occur during diabetes, leading to the acceleration of cardiovascular mortality and morbidity. In a diabetic milieu, endothelial dysfunction occurs as a result of attenuated production of endothelial derived nitric oxide (EDNO) and augmented levels of reactive oxygen species (ROS). Thus, in this review, we discuss novel therapeutic targets that either upregulate EDNO production or increase antioxidant enzyme capacity in an effort to limit oxidative stress and restore endothelial function. In particular, endogenous signaling molecules that positively modulate EDNO synthesis and mimetics of endogenous antioxidant enzymes will be highlighted. Consequently, manipulation of these unique targets, either alone or in combination, may represent a novel strategy to confer vascular protection, with the ultimate goal of improved outcomes for diabetes-associated vascular complications

Identification of Golgi-localized acyl transferases that palmitoylate and regulate endothelial nitric oxide synthase

Lipid modifications mediate the subcellular localization and biological activity of many proteins, including endothelial nitric oxide synthase (eNOS). This enzyme resides on the cytoplasmic aspect of the Golgi apparatus and in caveolae and is dually acylated by both N-myristoylation and S-palmitoylation. Palmitoylation-deficient mutants of eNOS release less nitric oxide (NO). We identify enzymes that palmitoylate eNOS in vivo. Transfection of human embryonic kidney 293 cells with the complementary DNA (cDNA) for eNOS and 23 cDNA clones encoding the Asp-His-His-Cys motif (DHHC) palmitoyl transferase family members showed that five clones (2, 3, 7, 8, and 21) enhanced incorporation of [3H]-palmitate into eNOS. Human endothelial cells express all five of these enzymes, which colocalize with eNOS in the Golgi and plasma membrane and interact with eNOS. Importantly, inhibition of DHHC-21 palmitoyl transferase, but not DHHC-3, in human endothelial cells reduces eNOS palmitoylation, eNOS targeting, and stimulated NO production. Collectively, our data describe five new Golgi-targeted DHHC enzymes in human endothelial cells and suggest a regulatory role of DHHC-21 in governing eNOS localization and function

Caveolin as a Potential Drug Target for Cardiovascular Protection

Caveolae and caveolin are key players in a number of disease processes. Current research indicates that caveolins play a significant role in cardiovascular disease and dysfunction. The far-reaching roles of caveolins in disease and dysfunction make them particularly notable therapeutic targets. In particular, caveolin-1 and caveolin-3 have been identified as potential regulators of vascular dysfunction and heart disease, and might even confer cardiac protection in certain settings. Such a central role in vascular health therefore makes manipulation of caveolin-1/3 function or expression levels clear therapeutic targets in a variety of cardiovascular related disease states. Here, we highlight the role of caveolin-1 and -3 in cardiovascular health and explore the potential of caveolin-1 and caveolin-3 derived experimental therapeutics

Myoferlin is critical for endocytosis in endothelial cells

Myoferlin is a member of the ferlin family of proteins that promotes endomembrane fusion with the plasma membrane in muscle cells and endothelial cells. In addition, myoferlin is necessary for the surface expression of vascular endothelial growth factor receptor 2 through the formation of a protein complex with dynamin-2 (Dyn-2). Since Dyn-2 is necessary for the fission of endocytic vesicles from the plasma membrane, we tested the hypothesis that myoferlin may regulates aspects of receptor-dependent endocytosis. Here we show that myoferlin gene silencing decreases both clathrin and caveolae/raft-dependent endocytosis, whereas ectopic myoferlin expression in COS-7 cells increases endocytosis by up to 125%. Interestingly, we have observed that inhibition of Dyn-2 activity or caveolin-1 (Cav-1) expression impairs endocytosis as well as membrane resealing after injury, indicating that Dyn-2 and Cav-1 also participate in both membrane fission and fusion processes. Mechanistically, myoferlin partially colocalizes with Dyn-2 and Cav-1 and forms a protein complex with Cav-1 solubilized from tissue extracts. Together, these data describe a new role for myoferlin in receptor-dependent endocytosis and an overlapping role for myoferlin-Dyn-2-Cav-1 protein complexes in membrane fusion and fission events