Primary Culture Of Vascular Endothelial Cells And Study On The Effect Of CXCL16 On VECs

Objective: To investigate and establish a easy accessible utility and reproducibility approach of isolation and primary culture of rat aortic endothelial cells (RAECs) in vitro, to identify ECs and analyze bionomics of ECs ,and to provide a valuable technical resource. We will investigate the effect of CXCL16 on rat aortic endothelial cells proliferation in vitro culture, too.Methods: 1. Isolated and cultured rat aorta endothelial cells by explant technique. Then, adopt syn-magneto glass beads coupled with platelet endothelial cell adhesion molecule (PECAM or CD31) antibody to screen ECs , cell-depuration , determine the cells growth rate and , draw VECs Growth -curve. The cells were identified by morphocytology and by immunofluorescence to detect von Willebrand's factor (vWF) or Factor VIII. 2. The cells obtained on passages 2 and 3 were used for the experiment. 6X103 cells grown in a 96-well plate were partially cultivated in M199 culture solution supplemented with 1% FBS for 24h and then stimulated by either various concentrations of CXCL16 (OnM, O.OlnM, 0.1nM, 1nM,10nM) and concentration of basic fibroblast growth factor (bFGF,10ng/mL) for another 48h (n = 10) .After incubation 48h, cell proliferation was examined using WST-1 assay. Result: 1. Endothelial cells of rat aorta exhibited polygonal, when the cells reach confluence, they appear as cobblestone-like monolayers . Another characteristic of ECs is their capacity to organize in tubules or capillary-like microtubes when plated on three-dimensional Matrigel and when stimulated with serum or growth factors. After immunostaining, ECs showed green positive reaction in cytoplasm. 2. After RAECs were cultured with the indicated concentrations (0nM, 0.01nM, 0.1nM, lnM,...