The effect of high glucose on the expression of erythropoietin receptor in human umbilical vein endothelial cells / 眼科研究

ABSTRACT

Objective Although vascular endothelial growth factor(VEGF)is a primary mediaor in diabetic retinopathy(DR).VEGF inhibition alone is insufficient for preventing retinal neovascularization.Some studies showed that erythropoietin(EPO)is a potent retinal angiogenic factor of independent of VEGF in DR.The present study is to investigate the effect of high glucose on the expression of mRNA and protein of the erythropoietin receptor(EPOR)in cultured human umbilical vein endothelial cells(UVECs)in vitro.MethodsHuman UVECs from the cell center of the hospital were cultured in vitro and passaged in DMEM containing 10% neonatal bovine serum with 22mmol/L of glucose for 12,24,48 and 72 hours in the experimental group.Cells cultured in 5.5mmol/L glucose were used as control group Ⅰ and mannitol + 22mmol/L of glucose(isotope)as control group Ⅱ.The expression of EPOR mRNA in Human UVECs were detected by semi-quantitative reverse transcriptase(RT)-polymerase chain reaction(PCR)and detected at A_(260mm)/A_(280mm).The PCR product was calculated as the A value of EPOR mRNA amplification/the A value of GAPDH mRNA amplification.The expression of EPOR protein in Human UVECs was detected by immunocytochemistry.ResultsThe A_(260mm)/A_(280mm) value of EPOR mRNA receptor expression was 0.32±0.02 in the 5.5mmol/L glucose group,and 0.34±0.02 in the mannitol+22mmol/L glucose group(P>0.05).In 12 hours,24 hours and 72 hours after the experiment,the A260mm/A280mm value of EPOR mRNA was 0.82±0.01,0.96±0.02 and 1.02±0.01,respectively,indicating a significant increase in comparison with the 5.5mmol/L glucose group.The expression of Human UVECs protein was gradually increased with passage in the experimental group.Expression of Human UVECs protein was stronger in various time points in the 22mmol/L glucose group than in the 5.5mmol/L glucose group.ConclusionHigh glucose elevates the expression of EPOR mRNA and protein in Human UVECs in a time-dependent manner.The effect of high glucose(22mmol/L glucose)on the expression of EPOR mRNA and protein in Human UVECs is not related to osmotic pressure.