594 Delivery of Recombinant Adeno Associated Virus Mediated Human Tissue Kallikrein Gene Attenuates Insulin Resistance and Prevents Renal Complications in Diabetic Rats Molecular Therapy �������� ���[.]
CARDIOVASCULAR 593 Delivery of Pleiotrophin Plasmid Induces Neovasculature Formation in Ischemic Myocardium Karen L Christman,1 Qizhi Fang,2 Richard E Sievers,3 Anne J Kim,1 Hubert H Fok,3 Albert F Candia,4 Kenneth J Colley,4 Randall J Lee.2,3 Joint Bioengineering Graduate Group, University of California Berkeley and San Francisco, San Francisco, CA, United States; Cardiovascular Research Institute, University of California San Francisco, San Francisco, CA, United States; 3Medicine, University of California San Francisco, San Francisco, CA, United States; 4Angiogenix, Inc., Burlingame, CA, United States Pleiotrophin (PTN) is a secreted heparin-binding cytokine with diverse functions In vitro, PTN has been shown to induce angiogenesis and stimulate endothelial cell proliferation We have recently demonstrated that the PTN gene is upregulated during ischemia in the heart, thus indicating a role in angiogenesis in the myocardium In this study, we hypothesized that delivery of PTN to ischemic myocardium will induce neovasculature formation We first developed a pCMV plasmid encoding the PTN gene and demonstrated via an ELISA that mammalian cells transfected with this plasmid are capable of producing and secreting PTN The produced PTN also induced proliferation of serum-starved SW13 cells, indicating that it is biologically active 250 μg of the pCMVPTN plasmid was injected directly into the infarcted myocardium of female Sprague-Dawley rats (n=3) following occlusion of their left anterior descending portion of the left coronary artery for 17 minutes Another set of rats (n=2) was injected with 250 μg of a pCMV-β-gal plasmid to serve as a control The rats were sacrificed after a minimum of five weeks The hearts were rapidly excised, fresh frozen, and sectioned into 10 μm slices sections from each heart were stained for capillaries using a Griffonia simplicifolia lectin which binds to a carbohydrate domain on endothelial cells The average number of capillaries within the infarct scar was quantified for each group using Fovea 2.0 plug-in for Adobe Photoshop Another five sections were stained for arterioles using an anti-smooth muscle actin antibody The average number and diameter of arterioles within each infarct were also quantified for each group using SPOT 3.5.5 software Injection of PTN plasmid induced formation of neovasculature compared to the β-gal plasmid control The capillary density following delivery of PTN plasmid significantly increased to 1258±157 capillaries/mm² compared to 782±166 capillaries/mm² with β-gal plasmid (P