【摘要】 目的 探讨高糖损伤人腹膜间皮细胞(HPMC)的机制。方法 (1)分离培养HPMC,以细胞形态、免疫细胞化学染色作细胞鉴定;(2)第2代HPMC分别经含1.5%、2.5%、4.25%葡萄糖的M199培养基培养48h后(以正常M199培养基和含4.25%甘露醇的M199培养基为对照),检测HPMC caspase-3活性和HPMC凋亡情况;(3)分别用25μmol/L、50μmol/L、和100μmol/LZ-VAD.fmk和4.25%葡萄糖共同刺激体外培养的HPMC(以DMSO和4.25%葡萄糖为对照),流式细胞仪观察HPMC凋亡情况,同时检测50μmol/L Z-VAD.fmk组和葡萄糖对照组caspase-3活性。结果 (1)HPMC caspase-3活性和凋亡率与葡萄糖浓度呈正相关(r分别为0.9506和0.9868,P<0.01),且HPMC caspase-3活性与凋亡率呈正相关(r=0.9860,P<0.01)。与M199对照组相比,2.5%葡萄糖组和4.25%葡萄糖组HPMC caspase-3活性和凋亡率显著增加(P<0.05),而1.5%葡萄糖组及4.25%甘露醇组HPMC caspase-3活性和凋亡率无显著差异(P>0.05)。(2)4.25%葡萄糖组caspase-3活性显著高于4.25%甘露醇组(P<0.05),但4.25%甘露醇组caspase-3活性与对照组相比无显著差异(P>0.05)。(3)与对照组相比,Z-VAD,fmk组HPMC凋亡率(P<0.05)和caspase-3活性(P<0.05)显著下降,且HPMC凋亡率与Z-VAD.fmk呈负相关(r=-0.8836,P<0.01)。结论 (1)高糖能以更多还原

【Abstract】 Objective To investigate the mechanism of high-glucose-induced injury to human peritoneal mesothelial cells(HPMC). Methods (1)The cultured HPMCs were exposed to culture medium containing different concentrations of glucose(1. 5% , 2. 5% , 4. 25% )for 48 hours and 4. 25% mannitol and normal culture medium were as control. Then apoptosis was observed by flow cytometry and caspase-3 activity was measured by ApoAlert?CPP33/Caspase-3 Assay kits. (2) The cultured HPMCs were exposed to 4.25% glucose culture medium containing different concentrations of caspases inhibitor, Z-VAD. fmk (25, 50, 100 μmol/L) for 48 hours and 4. 25% glucose culture medium containing DMSO was as control. Then apoptosis was observed by flow cytometry and caspase-3 activity was measured by ApoAlert?CPP33/ Caspase-3 Assay kits as well. Results (1) Glucose increased caspase-3 activity in a concentration-dependent manner. Compared to control, caspase-3 activity was significantly higher in 4. 25% glucose group and 2. 5% glucose group, but not significantly different in 1. 5% glucose group and 4. 25% mannitol group. (2) Apoptotic rate of HPMC was significantly lower in Z-VAD. fmk group than that in control. Z-VAD. fmk decreased the number of apoptotic cells in a concentration-dependent manner. Also, caspase-3 activity of HPMC was significantly lower in Z-VAD. fmk group than that in control. Conclutions (1) High-glucose can induce apoptosis and caspase-3 activation of HPMC in a dose-dependent manner. (2) Z-VAD. fmk inhibits high glucose-induced apoptosis of HPMC in a dose-dependent manner. (3)High glucose induces apoptosis of human peritoneal mesothelial cells by caspase-3 activation.更多还原