Constitutively active glycogen synthase kinase-3beta gene transfer sustains apoptosis, inhibits proliferation of vascular smooth muscle cells, and reduces neointima formation after balloon injury in rats.

Abstract

Glycogen synthase kinase (GSK)-3beta is a crucial factor in many cellular signaling pathways and may play an important role in smooth muscle proliferation and apoptosis after angioplasty. To investigate the effect of GSK-3beta modulation on neointima formation, smooth muscle proliferation, and apoptosis after balloon injury in vivo, we delivered adenoviral vectors expressing the constitutively active form of GSK-3beta (GSK-S9A: 9th serine switched to alanine) or a control gene into rat carotid arterial segments after balloon injury with a 2F Fogarty catheter. Viral infusion mixtures (5x108 pfu) were incubated in the arterial lumen for 20 minutes, and the effects of gene delivery were evaluated 3 days and 2 weeks after gene delivery with morphometry and immunohistochemical staining for proliferating and apoptotic cells. There were no significant differences in intimal, medial, and lumen areas at 3 days after the procedure. However, 2 weeks after gene delivery, the active GSK-3beta gene transfer resulted in a significantly lower intima to media ratio (0.29+/-0.06 versus 0.86+/-0.09, P<0.01) and a greater lumen area (0.41+/-0.02 versus 0.31+/-0.01 mm2, P<0.01) compared with the control gene transfected group. This was attributable to a significant reduction in intimal area (0.05+/-0.01 versus 0.15+/-0.02 mm2, P<0.01), whereas the medial area was similar (0.17+/-0.01 versus 0.18+/-0.01 mm2, P=0.21). Proliferation index was significantly reduced both at 3 days and 2 weeks in the active GSK-3beta gene transferred group (2.97+/-0.29% versus 5.71+/-0.50%, P<0.01). In addition, apoptotic index, which was not significantly different between the 2 groups at 3 days, was significantly higher in the active GSK-3beta gene transferred group at 2 weeks (3.14+/-0.68% versus 22.7+/-1.63%, n=10, P<0.01, for control versus active GSK-3beta gene transfer). In vivo delivery of the active GSK-3beta gene inhibits smooth muscle proliferation, sustains apoptosis, and reduces neointima formation after balloon injury in rats and may be a future therapeutic target to limit neointima hyperplasia after angioplasty.