目的 探讨多聚左旋精氨酸(PLA)诱导NCI-H292细胞凋亡的信号通路及分子机制.方法 将加入PLA 的浓度梯度分5组:0、10、20、40、60 mg/L,作用NCI-H292细胞24 h后Western blot法检测每组c-Jun氨基末端激酶( JNK)的磷酸化水平;设对照组、PLA 组(40 mg /L)、SP组(加入特异性JNK通路抑制剂SP600125)、PLA+SP组,加药24 h后流式细胞仪检测每组NCI-H292细胞凋亡率,Western blot法检测各组NCI-H292细胞内凋亡相关蛋白 Bcl-2/Bax、Caspase-3、P-JNK/JNK和β-actin蛋白的表达水平.结果 对照组、PLA组、SP组、 PLA+ SP组 NCI-H292细胞凋亡率分别为(5.13 ±1.07)%、(22.62 ± 1.66)%、(5.69 ± 0.14)%、(8.99 ± 3.73)% ;Western blot 法检测 PLA 诱导 NCI-H292 细胞内BCL-2/Bax比值降低( P＜0. 01),Caspase 3 表达升高( P＜0. 001),PLA诱导NCI-H292 细胞JNK 磷酸化水平增加(P＜0.001 ), SP600125 抑制 PLA 诱导 JNK 磷酸化( P ＜0.001).结论 PLA可能介导JNK信号通路引起NCI-H292细胞凋亡水平增加.%Objective To investigate the signaling pathway and molecular mechanism of PLA-induced NCI-H292 apoptosis. Methods The concentration gradients with PLA were divided into five groups: 0, 10, 20, 40 and 60 mg/L. After NCI-H292 cell treatment for 24 hours, Western blot was employed to detect the phosphorylation level of JNK in each group. The set groups included the control group, PLA group (40 mg/L), SP (JNK inhibitor) group, and PLA+SP group. After 24 hours’ treatment, flow cytometer was used to detect NCI-H292 apoptotic rate in each group, while Western blot was adopted to detect their Bcl-2/Bax, Caspase-3, P-JNK/JNK and β-actin pro-tein expression levels in NCI-H292 cells. Results The NCI-H292 apoptotic rates in the black control group, PLA group, SP group, and PLA+SP group were (5.13 ± 1.07 ) % , (22.62 ± 1.66 ) % , (5.69 ± 0.14 ) % ,(8.99 ± 3.73)% , respectively. According to the detection results by Western blot, the BCL-2/Bax ratio in PLA-induced NCI-H292 cells was reduced ( P＜0.01) , the expression of Caspase-3 was increased ( P＜0.001) . In PLA-in-duced NCI-H292 cells, the phosphorylation level of JNK was enhanced (P＜0.001), and SP600125 inhibited the JNK phosphorylation induced by PLA ( P＜0.001) . Conclusion PLA may lead to the increase of NCI-H292 ap-optosis level by inducing JNK signaling pathway.
展开▼
