斑蝥素对胰腺癌细胞系PANC1、CFPAC-1的凋亡诱导作用及机制研究

Cantharidin induces apoptosis in pancreatic cancer cell lines PANC1 and CFPAC-1

目的研究斑蝥素对胰腺癌细胞系PANCl、CFPAC-1的凋亡诱导作用及机制。方法用斑蝥素处理PANCl、CFPAC-1细胞。四甲基偶氮唑蓝（MTF）法检测细胞增殖；流式细胞术检测细胞凋亡；酶化学法检测caspase活性；RT—PCR及蛋白质印迹法检测凋亡相关基因的表达。结果斑蝥素呈剂量依赖性明显抑制胰腺癌细胞系PANCl、CFPAC-1增殖及诱导细胞凋亡。10μmol／L斑蝥素处理细胞72h，PANCl、CFPAC-1细胞的增殖抑制率最高，分别达（52．95±6．34）％和（71．21±6．30）％。处理24h，PANCI细胞的早期和晚期凋亡细胞分别从7．35％增加到24．89％、6．36％增加到17．73％；CFPAC-1细胞从6．39％增加到24．70％、9．21％增加到12．58％（P值均〈0．01）。PANCl细胞的caspase8和caspase9活性分别为对照组的（155．84-11．5）％和（194．64-14．7）％；CFPAC-1细胞分别为对照组的（182．54±24．3）％和（215．84±12．2）％（P值均〈0．01）。促凋亡基因TNF-α、TRAILRI、TRAILR2、Bad、Bak和Bid表达增加，抗凋亡基因Bcl-2表达减少。结论斑蝥素通过激活Caspase、上调促凋亡基因及下调抗凋亡基因的表达从而诱导胰腺癌细胞凋亡。

Objective To investigate the apoptosis induction effect of Cantharidin on pancreatic cancer cell line PANC1 and CFPAC-1 and possible mechanism. Methods PANC1 and CFPAC-1 was treated with Cantharidin. Cell growth was determined by MTI＇. Apoptosis was measured by flow cytometry. Caspase activity was measured by using enzyme chemical method. Apoptosis-related gene expressions were determined by using RT-PCR and Western blotting. Results Cantharidin significantly inhibited the growth of pancreatic cancer cells PANC1, CFPAC-1 and induced apoptosis in a dose-dependent manner. Seventy-two hours after 10 μmol/L Cantharidin treatment, the inhibitory rates of PANC1, CFPAC-1 were （52.95 ± 6.34）% and （71.21± 6.30）%. Twenty-four hours after treatment, the early and later period apoptotic cell of PANC1 was increased from 7.35% to 24.89%, from 6.36% to 17.73%. The early and later period apoptotic cell of CFPAC was increased from 6.39% to 24.70%, from 9.21% to 12.58% （P 〈0.01）. Activity of caspase 8 and caspase 9 in PANC1 cells was （ 155.8 ± 11.5 ） % and （ 194.6 ± 14.7 ） % when compared with that of control group. Activity of caspase 8 and easpase 9 in CFPAC-1 was （ 182.5 ± 24.3 ） % and （215.8 ± 12.2） % when compared with that of control group （ P 〈 0.01 ）. The expression of pro-apoptotic genes, TNF-α, TRAILR1, TRAILR2, Bad, Bak and Bid was elevated, the expression of anti-apoptotic Bcl-2 gene was decreased. Conclusions Cantharidin can induce apoptosis in pancreatic cancer cell lines by activating caspase,up-regulating the expression of pro-apoptotic genes and down-regulating the expression of anti-apoptotic genes.