乌司他丁对缺血诱导的大鼠骨髓间充质干细胞凋亡的影响

Effects of Ulinastatin on apoptosis induced by ischemia in rat bone marrow mesenchymal stem cells

目的研究乌司他丁是否能够抑制缺血诱导的大鼠骨髓间充质干细胞(BMSC)凋亡。方法采用25只SPF级C57BL/6J雄性大鼠为研究对象,将大鼠随机平均分为正常对照组、缺血模型组、乌司他丁组、乌司他丁+AKT通路抑制剂组、乌司他丁+ERK通路抑制剂组,其中正常对照组不进行任何前处理;缺血模型组仅进行缺血处理;乌司他丁组、乌司他丁+AKT通路抑制剂(LY294002)组、乌司他丁+ERK通路抑制剂(PD98059)组,每组大鼠在缺血处理BMSC前,分别加入乌司他丁、乌司他丁+LY294002、乌司他丁+PD98059预处理1h,然后在药物作用下再进行缺血处理。乌司他丁浓度为500U/ml。结果培养6h后,细胞凋亡比例显著高于培养3h时,其余相邻时间点之间无显著差异。正常对照组凋亡率与缺血模型组凋亡率之间差异具有统计学意义,缺血模型组大鼠caspase-3显著高于正常对照组。乌司他丁+AKT通路抑制剂组与乌司他丁+ERK通路抑制剂组大鼠caspase-3水平均显著高于乌司他丁组,且乌司他丁+AKT通路抑制剂组大鼠caspase-3水平显著低于乌司他丁+ERK通路抑制剂组大鼠(t=3.289,P=0.046)。结论培养6h效率最高,较适宜作为最佳缺血时间。乌司他丁能够有效抑制大鼠BMSC凋亡。乌司他丁通过ERK抑制通路发挥作用更优于AKT通路。

Objective To investigate the effect of Ulinastatin on inhibiting the apoptosis of rat bone marrow-derived mesenchymal stem cells( BMSC) induced by ischemia. Methods Twenty-five SPF grade C57BL/6J male rats were randomly divided into normal control group,ischemic model group,Ulinastatin group,Ulinastatin + AKT pathway inhibitor group,Ulinastatin + ERK pathway inhibitor group,and normal control group without any pretreatment. Ischemic model group only treated with ischemia; Ulinastatin group,Ulinastatin + AKT pathway inhibitor ( LY294002) group and Ulinastatin + ERK pathway inhibitor ( PD98059) group were pretreated with Ulinastatin,Ulinastatin + LY294002 and ulinastatin + PD98059 respectively for 1 hour before ischemic treatment of BMSC. Ulinastatin concentration was 500 U/ml. Results After 6 hours of culture,the percentage of apoptosis was significantly higher than that at 3 hours of culture,and there was no significant difference between the other adjacent time points. There was significant difference between the apoptosis rate of normal control group and ischemic apoptosis model group. Caspase-3 of ischemic group was significantly higher than that of normal control group. Caspase-3 levels in rats treated with Ulinastatin + AKT pathway inhibitor and ulinastatin + ERK pathway inhibitor were significantly higher than those in rats treated with Ulinastatin,and caspase-3 levels in rats treated with Ulinastatin + AKT pathway inhibitor were significantly lower than those in rats treated with Ulinastatin + ERK pathway inhibitor ( t = 3. 289,P = 0. 046) . Conclusion The culture of 6 h is the most efficient and suitable for the best ischemia time. Ulinastatin can effectively inhibit the apoptosis of rat BMSC. Ulinastatin is more effective by ERK pathway than the AKT inhibitory pathway.