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(Circulation Research. 2002;90:609.)
© 2002 American Heart Association, Inc.

Integrative Physiology

Inhibition of Protein Kinase C Prevents Endothelial Cell Migration and Vascular Tube Formation In Vitro and Myocardial Neovascularization In Vivo

Aihong Wang, Motohiro Nomura, Sybill Patan, J. Anthony Ware

From the Cardiovascular Division, Department of Medicine, Department of Molecular Pharmacology, Albert Einstein College of Medicine, Bronx, NY.

Correspondence to Dr J. Anthony Ware, Vice President, Cardiovascular Research and Clinical Investigation, Eli Lilly and Co, Lilly Corporate Center, Indianapolis, IN 46285. E-mail ware_j_anthony{at}lilly.com

Although protein kinase C (PKC) activation is required for endothelial cell (EC) growth, migration, adhesion, and vessel formation, the role of individual PKC isoenzymes in these events is not defined. Because PKC has been previously linked with enhanced EC migration and response to angiogenic growth factors, we characterized a specific phosphorothioate-modified 21-mer antisense PKC (AS-PKC). AS-PKC (500 nmol/L) prevented the expression of PKC protein by 90% in human ECs and did not reduce the expression of any other PKC isoenzyme. AS-PKC reduced human EC migration by 64% compared with its control oligonucleotide in a "scratch" wounding assay, and AS-PKC reduced human EC adhesion to the extracellular matrix protein vitronectin by 18%. Phosphorylation of mitogen-activated protein kinase (extracellular signal–regulated kinase 1/2) induced by vascular endothelial growth factor was inhibited by 30% in human ECs transfected with AS-PKC. Compared with control, AS-PKC also reduced the number of EC tubes formed in a 3D type I collagen gel assay by 37.5%. Finally, using an osmotic minipump, we infused AS-PKC into mice in which myocardial infarction was induced by coronary ligation and found that the oligonucleotide was primarily taken up by intramyocardial blood vessels. Compared with the results with control oligonucleotide, AS-PKC oligonucleotide inhibited the number of anti-PKC–stained blood vessels by 48% and reduced the total vessel number by 72% as well. In conclusion, the expression of PKC is required for full EC migration, adhesion to vitronectin, vascular endothelial growth factor–induced extracellular signal–regulated kinase activation, and tube formation and is likely to be of importance in myocardial angiogenesis in vivo after ischemia.

Key Words: angiogenesis • protein kinase C • endothelial cells • myocardial function

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