过氧化物酶增殖物活化受体γ活化对胰腺癌细胞凋亡的诱导作用

Activation of PPAR-γ induces apoptosis in human pancreatic carcinoma cell line

目的探讨过氧化物酶增殖物活化受体γ(PPARγ)在诱导胰腺癌细胞凋亡中的调节作用。方法胰腺癌细胞系SW1990经10μmol/L PPARγ配体15-脱氧-前列腺素J2(15d-PGJ2)、20μmol/L维甲酸受体α(RXRα)配体9-顺式-维甲酸(9-cis-RA)及其两者联合作用培养48 h后,应用电子显微镜观察细胞形态学变化;流式细胞仪研究细胞凋亡比例的变化。30只雌性裸鼠皮下接种1×107个SW1990细胞,2周后待肿瘤可触及时,将其随机分为对照组和治疗组(予罗格列酮)。11周后处死裸鼠并取下移植瘤,采用脱氧核糖核酸末端转移酶介导的生物素缺口末端标记法(TUNEL)评估移植瘤中凋亡细胞的情况,并用逆转录聚合酶链反应(RT-PCR)检测bcl-2凋亡相关基因家族成员bcl-2、bcl-xl、bak、bax以及凋亡抑制基因Survivin的表达。结果电子显微镜结果显示15d-PGJ2、9-cis-RA及其两者联合作用均可导致SW1990细胞发生形态学变化。可见细胞核和胞质内容物密集皱缩,核内染色质凝集成块并边集,形成1个或数个沿核膜的月芽小体。在联合应用组中还可观察到凋亡小体的存在。流式细胞术检测提示对照组凋亡比率为0.95%,15d-PGJ2、9-cis-RA及其联合应用可引起SW1990细胞凋亡比率增加,15d-PGJ2组为8.05%,9-cis-RA组为8.36%,联合作用组为6.86%。TUNEL染色显示治疗组移植瘤组织中凋亡指数(AI)为15.25±4.57,明显高于对照组的6.25±2.62(P<0.05)。RT-PCR揭示在SW1990移植瘤中,促凋亡基因bak、bax表达上调,抗凋亡基因bcl-xl、Suvivin下调,抗凋亡基因bcl-2几乎不表达。结论PPARγ的活化可诱导SW1990胰腺癌细胞凋亡。其机制可能是通过上调促凋亡基因bak、bax及下调抗凋亡基因bcl-xl、Suvivin表达而实现的。

Objective To appraise the effects of PPARγ activation on induction of apoptosis in human pancreatic carcinoma cell line. Methods SW1990 pancreatic carcinoma cell line, treated with ligand of PPARγ, 15dPGJ2, ligand of RXRa, 9-cis-RA, and their combination, was used to study the effects of PPARγ activation on cell apoptosis by flow cytometry, and on morphology by electron-microscopy. Thirty female nude mice were inoculated subcutaneously with 1 million SW1990 cells until tumors were palpable 2 weeks later and the mice were randomized to receive either vehicle （control） or Rosiglitazone via drinking water for 9 weeks. The tumors were measured at regular intervals. After a 11-week experimental period,the mice were euthanized and the tumors were immuno-histochemically assayed for apoptosis （TUNEL）. The expression of bcl-2, bcl-x1, bak and bax, members of the bcl-2 family of apoptosis-associated genes and Survivin were also assayed by using RT-PCR. Results Flow eytometry demonstrated a significant increase of AnnexinV-positive-PI negative apoptotic cells after treatment with 15d- PGJ2, 9-cis-RA, and the combination. The apoptotic ratios were 0.95% （control）,8.05% （15d- PGJ2 ）,8.36%（9- cis- RA） .6.86 %（combination）,respectively. SW1990 cells were cultured with 15d-PGJ%, 9-cis- RA inducing dramatic morphological changes by electron-microscopy and showing nuclear and cytoplasmic shrinkage, condensation and margination of chromatin against the nuclear membrane, which were the earliest changes of apoptosis with furthermore formation of apoptotic bodies. Apoptotic cells were detected in both groups, but TUNEL assay showed a marked up-regulation trend in experimental group FAIt （15.25±4.57 ）] in comparing with the control group[AI： （6.25 ± 2.62）]. RT PCR demonstrated that the expressions of bak, bax, the proapoptotic members of bcl-2 family were increased; and bcl-x1, Suvivin, the anti-apoptotic gene, were slightly decreased. The expression of bcl-2 could be hardly detected in SW1990 cell xenograft tumor. Conclusions Activation of PPARγ induces apoptosis in SW1990 cells. The mechanism is probably associated with up-regulating the expression of hak, bax and down regulating the expression of bcl-x1 and Survivin. （Shangliat Med J, 2006, 29： 81 84）