Inhibitory Effect of Adenoviral Vector-Mediated Delivery of p21WAF1/CIP1 on Retinal Vascular Endothelial Cell Proliferation and Tube Formation in Cultured Rhesus Monkey Cells (RF/6A)

Objective: To investigate the inhibitory effect(s) of adenovirus (Ad) vector-mediated delivery of p21(WAF1/CIP1) (Ad-p21) on proliferation and tube formation in Rhesus monkey choroid-retina vascular endothelial cells (RF/6A). Methods: In vitro-cultured RF/6A cells were divided into three groups: phosphate-buffered saline (PBS), Ad-p21-transfected, and negative control. Plasmid vectors were transfected via Ad-p21. The mRNA and protein expressions of p21 and cyclin-dependent kinase (CDK)2 in RF/6A cells were measured by reverse transcription-PCR (RT-PCR) and western blot analyses. Cell-cycle distributions were analyzed by flow cytometry. Matrigel was used as a matrix for endothelial cell tube formation. Results: Expressions of p21 mRNA and protein were greater, and expressions of CDK2 mRNA and protein lower, in the Ad-p21-transfected group than in either the PBS or negative control groups. Cell-cycle distribution analysis indicated that the proportion of G0/G1 cells was significantly higher in the Ad-p21 transfected group than in either the PBS or negative control groups (p = 0.000). There were significantly fewer endothelial cell tubes in the Ad-p21-transfected group than in either the PBS or negative control groups (p = 0.004). Conclusions: Ad-p21 inhibits RF/6A cell proliferation and tube formation. The underlying mechanism to account for this may be that overexpression of p21 arrests the cell-cycle transition from the G1- to the S-phase via inhibition of CDK2 activity.