异丙酚预处理对HK-2细胞缺氧/复氧损伤的保护作用及机制

Protective effects and possible mechanism of propofol preconditioning on HK-2 cell with anoxia-reoxygenation injury

目的探讨P38MAPK对异丙酚预处理的HK-2细胞缺氧/复氧损伤的保护作用的影响。方法取离体培养的HK-2细胞,随机分成5组:对照组(A组)。CoCl2组(B组):加入300μMCoCl2处理1h,然后更换正常的培养基培养24h,之后更换无血清的培养基培养。异丙酚组(C组)培养孔中加入25μmol/L异丙酚预处理1h后加入300μM的CoCl2处理1h,然后更换正常的培养基培养24h,之后更换无血清的培养基培养。脂肪乳剂组(D组):加入10%脂肪乳剂90μL预处理1h后加入300μMCocl2。SB组(E组),培养孔中加入10mol/LSB203580预孵育10min后处理同C组。MTT法测定细胞增殖,流式细胞技术测定细胞的凋亡。RT-PCR技术检测Bcl-2,Caspase-3和P38mRNA表达。结果脂肪乳剂组与CoCl2组比较,差异无统计学意义,25μmol/L异丙酚预处理可以明显增加HK-2细胞的增殖能力,减少凋亡(P<0.05或P<0.01)。预处理上调P38mRNA的表达,同时可以上调凋亡抑制基因Bcl-2的表达,下调促凋亡基因caspase-3的表达,而这些调节作用可被SB203580抑制(P<0.05或P<0.01)。结论25μmol/L异丙酚预处理对缺氧/复氧后的HK-2细胞有保护作用,P38MAPK和凋亡相关基因在预处理中起到重要的作用。

[Objective] To investigate the effect of propofol preconditioning on human proximal renal tubular epithelial （HK-2） cells with anoxia-reoxygenation injury and the role of P38 protein kinase. [Methods] HK-2 cell were randomly assigned to 5 groups： Control group （group A）, CoCl2 group （group B）, propofol preconditioning group （group C） pretreated theHK-2 cell with 25 μmol/L propofol lhour, then added 300 μM CoCl2, intralipid group （group D）： pretreated the HK-2 cell with 10% intralipid 90 μL 1 hour, then added 300 μM COC12, SB203580 （inhibitor of P38MAPK） group （group E）, pretreated with 10 umol/L SB203580 10min, then 25 umol/L propofol were added, after 1 hour, added 300 μM COCl2, stimulated with no serum media 24 hours later. We used MTT method and FACS to detect the proliferation and apoptosis of HK-2 cell, and RT-PCR method was used to show the regulation of P38 and apoptosis related gene. [Results] After pretreated with 25 μmol/L propofol, HK-2 cell proliferation increased and apoptosis decreased when cultured with no serum media 24 hours later （P 〈0.05 or 0.01）, Intralipid did not affect the HK-2 cell, P38 was upregulated after preconditioning, meanwhile Bcl-2 gene was upreguahed and caspase-3 gene was down-regulated. These regulation can be reversed by SB203580 （P 〈0.05 or 0.01）. [Conclusion] Precondit/oning with 25 μmol/L propofol can protect HK-2 cell against anoxia-reoxygenation injury by attenuating HK-2 cell apoptosis, P38 protein kinase may be useful to control apoptosis related gene.