Zhang Ming-sheng, Zhou Yun-feng, Xie Cong-Hua, Zhang Wen-jie, Zhou Fu-xiang, Qu Xue-ju
Depmartment of Cancer Radio-Chemotherapy, Zhongnan Hospital of Wuhan University, Wuhan 430071, China.
Zhonghua Yi Xue Za Zhi. 2006 Jan 10;86(2):76-81.
To investigate the relationship between the signal transducer and activator of transcription 6 (STAT6) and human colon cancer.
Four STAT6-specific recombinant plasmid vectors, pshRNA-STAT6-1, 2, 3, and 4 were constructed and transfected into the cultured human colon cancer cells of the line HT-29. Seventy-two hours later RT-PCR was used to detect the mRNA expression of STAT6 and the apoptosis-related genes Bcl-2 and Bax, flow cytometry (FCM) was used to detect the protein expression of phopho-STAT6 (pSTAT6). HT-29 cells were inoculated into a plate and transfected with pshRNA-STAT6-1 or pshRNA-STAT6-4, and HT-29 cells without transfection were used as controls. Seventy-two hours later FCM was used to observe the cell apoptosis. Another HT-29 cells were inoculated into a plate and transfected with pshRNA-STAT6-1 or pshRNA-STAT6-4, or blank liposome as controls. Seventy-two hours later. Western blotting was used to detect the protein expression of Bcl-2 and Bax genes.
The p-STAT6 protein expression rate was 3.2% +/- 0.6% in the pshRNA-STAT6-1 group, significantly lower than that of the blank control group (18.2% +/- 0.9%, P < 0.01) with an inhibition rate of 82.4%, and was 7.9% +/- 0.4% in the pshRNA-STAT6-4 group, significantly lower than that in the blank control group too (P < 0.01) with an inhibition rate of 56.6%. And the p-STAT6 protein expression rates of the pshRNA-STAT6-2 and pshRNA-STAT6-3 groups were 16.6% +/- 0.5% and 17.1% +/- 0.7% respectively, both not significant different from that of the blank control group (both P > 0.05). The early cell apoptosis rates of the pshRNA-STAT6-1 and pshRNA-STAT6-4 groups were 13.0% and 8.8% respectively, both significantly higher than that of the blank control group (0.4%, both P < 0.05). The mRNA expression of Bcl-2 was significantly lower and the mRNA expression of Bax was significantly higher in the pshRNA-STAT6-1 and pshRNA-STAT6-4 groups than in the blank control and blank liposome groups (all P < 0.01). The protein expression patterns of Bcl-2 and Bax was consistent with that of their protein expression.
STAT6 signaling pathway inhibits the apoptosis of colon cancer cells by regulation of the Bcl-2 and Bax genes.
探讨信号转导及转录激活因子6（STAT6）与人类结肠癌的关系。
构建4种STAT6特异性重组质粒载体，即pshRNA-STAT6-1、2、3和4，并将其转染至培养的人结肠癌HT-29细胞系。72小时后，采用逆转录聚合酶链反应（RT-PCR）检测STAT6及凋亡相关基因Bcl-2和Bax的mRNA表达，采用流式细胞术（FCM）检测磷酸化STAT6（pSTAT6）的蛋白表达。将HT-29细胞接种于培养板，用pshRNA-STAT6-1或pshRNA-STAT6-4转染，未转染的HT-29细胞作为对照。72小时后，用FCM观察细胞凋亡情况。另将HT-29细胞接种于培养板，用pshRNA-STAT6-1或pshRNA-STAT6-4转染，或以空白脂质体作为对照。72小时后，采用蛋白质免疫印迹法检测Bcl-2和Bax基因的蛋白表达。
pshRNA-STAT6-1组p-STAT6蛋白表达率为3.2%±0.6%，显著低于空白对照组（18.2%±0.9%，P<0.01），抑制率为82.4%；pshRNA-STAT6-4组p-STAT6蛋白表达率为7.9%±0.4%，也显著低于空白对照组（P<0.01），抑制率为56.6%。pshRNA-STAT6-2组和pshRNA-STAT6-3组p-STAT6蛋白表达率分别为16.6%±0.5%和17.1%±0.7%，与空白对照组相比均无显著差异（均P>0.05）。pshRNA-STAT6-1组和pshRNA-STAT6-4组早期细胞凋亡率分别为1上0%和8.8%，均显著高于空白对照组（0.4%，均P<0.05）。pshRNA-STAT6-1组和pshRNA-STAT6-4组中Bcl-2的mRNA表达显著低于空白对照组和空白脂质体组，Bax的mRNA表达显著高于空白对照组和空白脂质体组（均P<0.01）。Bcl-2和Bax的蛋白表达模式与其mRNA表达一致。
STAT6信号通路通过调控Bcl-2和Bax基因抑制结肠癌细胞凋亡。
