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目的:观察不同浓度威灵仙水提物(Clematis)对多氯联苯(PCB118)诱导的胰岛β细胞凋亡的保护作用。方法:将大鼠胰岛素瘤(INS-1)细胞株分为对照组(control)、PCB118(25μmol/L)组、Clematis低剂量组(4mg/L)以及Clematis高剂量组(40mg/L),分别于药物干预9、24、48h后于倒置显微镜下观察INS-1细胞形态,通过CCK-8法检测各组INS-1细胞活性,并采用流式细胞术检测细胞的凋亡率,Western Blot检测Cleaved Caspase-3、Caspase-3蛋白的表达。结果:倒置显微镜下观察结果显示:PCB118刺激9、24、48h后INS-1细胞凋亡,Clematis高浓度及低浓度组PCB118诱导的INS-1细胞凋亡减少;CCK-8结果显示PCB118处理后细胞活性显著下降(P<0.01),而Clematis高、低浓度组均改善PCB118的抑制作用,其中高浓度组与PCB118组比较,差异有统计学意义(P<0.05);流式细胞术检测结果显示PCB118诱导INS-1细胞早期凋亡明显(P<0.05),而Clematis干预后细胞早期凋亡改善,高浓度组更为显著(P<0.05);同时,PCB118处理后凋亡相关蛋白Cleaved Caspase-3表达上调,Clematis可逆转Cleaved Caspase-3过表达(P<0.05)。结论:威灵仙水提物可以抑制PCB118诱导的INS-1细胞凋亡,增强PCB118毒性环境下的INS-1细胞活性,为中药保护胰岛β细胞提供了实验依据。
OBJECTIVE: To observe the protective effect of Clematis with different concentrations on the islet β-cell apoptosis induced by PCB118. Methods: INS-1 cell lines were divided into control group, PCB118 (25μmol / L), Clematis low dose group (4mg / L) and Clematis high dose group (40mg / L) The morphological changes of INS-1 cells were observed under inverted microscope 9, 24 and 48 hours after drug intervention. The activity of INS-1 cells in each group was detected by CCK-8 assay. The apoptosis rate of INS-1 cells was detected by flow cytometry. Western Blot The expression of Cleaved Caspase-3, Caspase-3 protein was detected. Results: The results of inverted microscopy showed that INS-1 cells apoptosis was induced by PCB118 stimulation for 9, 24 and 48 hours, and PCB118 induced by Clematis reduced the apoptosis of INS-1 cells. The results of CCK-8 showed that after PCB118 treatment (P <0.01), while Clematis high and low concentration groups all improved the inhibition effect of PCB118, the difference was statistically significant (P <0.05) between high concentration group and PCB118 group; the results of flow cytometry The results showed that PCB118 induced early apoptosis in INS-1 cells significantly (P <0.05), while the cleaved cells treated with Clematis improved significantly (P <0.05), while the apoptosis-related protein Cleaved Caspase -3 expression, Clematis can reverse Cleaved Caspase-3 overexpression (P <0.05). Conclusion: The extract of Clematis can inhibit INS-1 cells apoptosis induced by PCB118 and enhance the activity of INS-1 cells under PCB118 toxic environment, which provides the experimental evidence for the protection of islet β cells by Chinese herbs.