Regulation of Retinal Endothelial Cells

Human retinal endothelial cells (Cell system, Kirkland, WA) from 7–8^th passage were incubated in NG or HG media containing 1% fetal calf serum, 9% Nu-serum and 1 µg/mL endothelial growth supplement for 4 days^{13 (link),14 (link)}. Cell incubated in high glucose were divided into two groups, cells in group 1 remained in high glucose for eight days, in the absence or presence of 1 µM 5-aza-2′-deoxycytidine (Aza; Sigma-Aldrich Corp, St. Louis, MO, USA), respectively. In group 2, after four days of high glucose exposure, the cells were incubated in normal glucose for four additional days, in the absence or presence of Aza (HG-NG and HG-NG/Aza). For osmotic/ metabolic control, 20 mM D-glucose was replaced by 20mM L-glucose (L-Gl). As reported previously, under these incubation conditions, morphology of HRECs was maintained^{22 (link)}.