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Objective To investigate the PI3K/Akt/GSK-3β pathway in human renal tubular epithelial cells (HK-2) ischemia-reperfusion injury in the regulation of apoptosis and recombinant human erythropoietin (rHuEPO) pre-protective effect. Normal culture HK-2 cells were divided into seven groups, the normal control group, ischemia-reperfusion (I / R) group, the LY294002 intervention group (PI3K/Akt blockers, 10 umol / L), Licl intervention group ( GSK-3β blockers, 20 umol / L) of rHuEPO intervention group (20 U / L), LY294002 dual intervention group of rHuEPO, HuEPO Licl dual intervention group. RT-PCR and Western blotting of protein kinase B (Akt), glycogen synthase kinase 3β (GSK-3β) and cysteine ??aspartyl protease expression of Caspase-3 gene level and protein activity levels; MTT assay cells vitality; AnnexinV / PI staining and flow cytometry to detect apoptosis. Results ischemia-reperfusion injury induced HK-2 cells apoptosis rate increase (15.2 ± 1.43%) of Akt activity levels, raised the level of activity of GSK-3β, Caspase-3 mRNA and protein activity levels increase, compared with the normal control group, The difference was statistically significant (P lt; 0.05), Akt, GSK-3βmRNA levels basically unchanged (P gt; 0.05). Compared with I / R group, LY294002 intervention apoptosis rate further increase (18.2 ± 2.06%), the level of activity of Akt down GSK-3β activity increases, Caspase-3 mRNA and protein activity raised, Licl intervention apoptosis rate reduced (12.3 ± 0.85%) of Akt activity level raised to lower the activity of GSK-3β, Caspase-3 mRNA and protein activity down the differences were statistically significant (P lt; 0.05). rHuEPO intervention compared with the I / R group, the apoptosis rate decreased (11.1 ± 1.62%), elevated levels of Akt activity and GSK-3β activity down, Caspase-3 mRNA and protein activity down, the difference was statistically significant (P lt ; 0.05). Compared with rHuEPO intervention of rHuEPO LY294002 dual intervention increased apoptosis rate (13.4 ± 1.94%), the levels of Akt activity and GSK-3β activity raised Caspase-3 mRNA and protein activity rates, rHuEPO Licl double intervention apoptosis rate down (7.5 ± 1.31%) of Akt activity levels increased GSK-3β activity decreased Caspase-3 mRNA and protein decreased activity, and the differences were statistically significant (P lt; 0.05). Conclusion 1. Ischemia-reperfusion injury can cause apoptosis of renal tubular epithelial cells, Akt activity decreased GSK-3β activity increased impact of exogenous Caspase-3-dependent apoptosis pathway may be one of the mechanisms of apoptosis. 2.rHuEPO through enhanced Akt activity, reduce GSK-3β and Caspase-3 enzyme activity, thereby reducing apoptosis, a protective effect of HK-2 ischemia-reperfusion injury.
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