Re-Endothelialization Study on Endovascular Stents Seeded by Endothelial Cells through Up- or Downregulation of VEGF.

Abstract

We studied the effects of gene transfection of endothelial cells with vascular endothelial growth factor (VEGF) on re-endothelialization and inhibition of in-stent restenosis. Transfected endothelial cells (ECs) exposed to different VEGF levels were seeded on a stent surface for evaluation in vitro. VEGF121(++) ECs and VEGF121(--) ECs were established using lentiviral-mediated HUVECs transfection. VEGF RNA transcription level and VEGF protein expression were detected by qPCR, Western blot, and ELISA. Methyl thiazolyl tetrazolium (MTT) assay, wound healing assay, and in vitro HUVEC tube formation assay showed that VEGF overexpression promoted cell proliferation, migration, and endothelial capillary-like tube formation. Downregulation of VEGF expression inhibited these activities. Using a rotational culturing system, cells tightly adhered on the stent surface. Stents seeded with transfected ECs at different VEGF levels were implanted in abdominal aortas of New Zealand white rabbits to study re-endothelialization and inhibition of in-stent restenosis. Stents with cells exposed to excess VEGF expression were almost completely covered with cells after stent implantation for 1 week (w). In the VEGF interference group this process was delayed over 4 w due to RNAi-mediated silencing of VEGF. Cryosectioning after 12 w showed that stents seeded with HUVECs exposed to excess VEGF expression significantly reduced the neointima area and stenosis when compared with bare metal stents and stents from the VEGF interference group. Transgenic HUVECs were not found in tissues of experimental animals. Furthermore, cells from these tissues were similar to those from normal tissue. In conclusion, VEGF-mediated endothelialization was found. Furthermore, ECs exposed to VEGF overexpression reduced neointimal hyperplasia, promoted endothelialization, and reduced in-stent restenosis.