Effect of recombinant human erythropoietin pretreatment on PI3K-Akt-GSK-3β signaling pathway in human renal tubular epithelial cells apoptosis induced by ischemia-reperfusion injury / 中华肾脏病杂志

ABSTRACT

Objective To study the role of PI3K-Akt-GSK-3β signaling in the apoptosis of renal tubular cells after ischemia-reperfusion injury and the protective mechanism of recombinant human erythropoietin(rHuEPO). Methods The human kidney tubular epithelial cells(HK-2)were cultured in vitro in different conditions as control group with serum, ischemia-reperfusion(IR)group, LY294002 group with LY294002(AKT inhibitor)10 μmol/L 30 minutes before IR treatment, LiCl group with LiCl(GSK-33 inhibitor)20 μtmol/L 30 minutes before IR treatment, rHuKPO group with EPO 20 U/ml 30 minutes before IR treatment, rHuEPO + LY294002 group with EPO 20 U/ml and in the presence of LY294002(10 μmol/L)30 minutes before IR treatment, rHuEPO +LiCl group with EPO 20 U/ml and in the presence of LiCl(20 μmol/L)30 minutes before IR treatment. Akt, GSK-33 and caspase-3 activation were measured by Western blotting. The apoptotic ratio of HK-2 cells was measured by flow cytometry. Cell viability was detected by MTT. Results In comparison with the control group, the apoptotic ratio raised up to 15.20%±1.43%, the expression of Akt activity decreased, GSK-33 activity and caspase-3 activity markedly elevated in IR group(P<0.05). LY294002 group up-regulated the apoptotic ratio(18.20%±2.06%), decreased the expression of Akt activity, increased GSK-33 activity and caspase-3 activity, however, LiCl group down-regulated the apoptotic ratio(12.30%±0.85%), increased the expression of Akt activity, decreased GSK-33 activity and caspase-3 activity compared with IR group(P<0.05). rHuEPO group remarkably decreased the apoptotic ratio(11.10%±1.62%), increased the expression of Akt activity, decreased GSK-33 activity and caspase-3 activity compared with IR group(P<0.05). rHuEPO+LY294002 group elevated the apoptotic ratio(13.40%±1.94%), decreased the expression of Akt activity, increased GSK-33 activity and caspase-3 activity, meanwhile, rHuEPO +LiCl group down-regulated the apoptotic ratio(7.50%±1.31%), increased the expression of Akt activity, decreased GSK-33 activity and caspase-3 activity compared with rHuEPO group(P<0.05). Conclusions PI3K-Akt-GSK-3β signaling pathway is involved in HK-2 cells apoptosis induced by ischemia-reperfusion injury and rHuEPO may be used as a new therapy.