摘要
目的检测蛋白激酶B（PKB）抑制剂SH-6联合抗肿瘤药物奥沙利铂对诱导人大肠癌细胞凋亡过程的影响。方法采用非放射性PKB活性免疫沉淀检测试剂盒检测SH-6干预前后细胞中PKB活性。将大肠癌细胞系lovo细胞分为4组：空白对照组、SH-6组、奥沙利铂组、SH-6＋奥沙利铂组。按照上述分组用药物干预大肠癌lovo细胞24小时，分别采用DNA ladder法和Hochest33258法检测各组细胞的凋亡情况，Western blot检测caspase-9活性变化。结果（1）SH-6可有效抑制PKB的活性；（2）DNA ladder检测法显示，空白对照组未见梯形DNA裂解片段，而其他3组均出现明显的DNA梯形裂解带。Hochest法显示，空白对照组细胞的细胞核均质蓝染，而药物处理的3组均出现明显的细胞核致密浓染及核碎裂等细胞凋亡形态学改变；（3）Western blot结果提示奥沙利铂干预后，lovo细胞中Caspase-9的活性与未处理组比较，明显增强，加用SH-6处理后，大肠癌lovo细胞中Caspase-9的活性与未处理组比较明显增强，与单用奥沙利铂组比较有所增强。结论PKB抑制剂能促进大肠癌细胞发生凋亡，增强抗肿瘤化疗药物的疗效。
Objective To investigate the effect of the inhibitor of protein kinase B （PKB）on inducing apoptosis of human colon cancer cells interfered by chemotherapeutics. Methods PKB activity was measured by immunoprecipitation. According to the different treatment,the colon cancer cell line lovo were divided into four groups as control group, SH-6 group, Oxaliplatin group, and SH-6 ＋ Oxaliplatin group. After 24 hours＇ treatment, cell apoptosis were detected by DNA ladder and hochest33258 staining. The changes of caspase-9 activity were determined by Western blot analysis. Results （1）SH-6 can effectively inhibit the activity of PKB. （2）DNA agarose gel electrophoresis showed that there were no apoptotic oligonucleosomal DNA fragmentation in the control group, while DNA fragmentation were found in the other groups;Hochest33258 staining showed that both chromatin condensationnor and nuclear fragmentation was observed in drug-treatment group,but these morphological hallmarks of apoptosis can not be found in the control group. （3）Western blot showed that the activity of caspase-9 in the groups treated by Oxaliplatin or SH-6 were higher than in the control group,and the activity of the groups treated by Oxaliplatin plus SH-6 was higher than those treated by only one drug. ConclusionThe inhibitor of PKB can induce the apoptosis of colon ceils and enhance the effect of ehemotherapeutics.
出处
《临床内科杂志》
CAS
2009年第10期694-696,共3页
Journal of Clinical Internal Medicine
