蛋白激酶CK2α对喉癌细胞凋亡和超微结构的影响

Effects of protein kinase CK2α on apoptosis and ultrastructure of laryngeal carcinoma cells

目的 探讨蛋白激酶CK2α对人喉癌细胞凋亡和超微结构的影响及其可能机制.方法 用脂质体转染法分别将蛋白激酶CK2α特异性siRNA表达质粒psiRNA-hH1neo-CK2α及非特异性siRNA表达质粒psiRNA-hH1neo-cont转染人喉癌Hep-2细胞.Western印迹法检测转染细胞蛋白激酶CK2α蛋白表达,膜联蛋白V-异硫氰酸荧光素（Annexin V-FITC）和碘化丙啶（PI）双染色法检测转染细胞凋亡率的变化,透射电镜观察转染细胞形态学变化,Western印迹法检测转染细胞bcl-2和Bax蛋白的表达.结果 转染psiRNA-hH1neo-CK2α质粒后,Hep-2细胞蛋白激酶CK2α蛋白表达明显下降（P＜0.01）.和未转染细胞组和psiRNA-hH1neo-cont转染细胞组比较,psiRNA-hH1neo-CK2α转染组细胞出现典型的凋亡征象,如核固缩、染色质凝集靠近核膜和凋亡小体形成.psiRNA-hH1neo-CK2α转染细胞凋亡率明显高于未转染细胞组和psiRNA-hH1neo-cont转染细胞组（25.66%±0.83%比3.66%±0.43%、5.18%±0.22%,均P＜0.05）;与其他2组比较,psiRNA-hH1neo-CK2α转染组细胞bcl-2蛋白表达较低（相对吸光度比值为0.20±0.09 vs 0.72±0.16、0.56±0.11,均P＜0.01）,Bax蛋白表达较高（相对吸光度比值为0.81±0.17 vs 0.26±0.12、0.33±0.17,均P＜0.01）,bcl-2/Bax较低（0.25±0.05 vs 2.76±0.21、1.70±0.22,均P＜0.01）.结论 蛋白激酶CK2α与喉癌细胞凋亡密切相关,该作用可能与bcl-2/Bax降低有关,蛋白激酶CK2α可能是一个有潜力的喉癌治疗靶点.

Objective To investigate the effect of protein kinase CK2α on apoptosis and ultrastructure of human laryngeal carcinoma cells and its possible mechanism. Methods The siRNA expression plasmid psiRNA-hH1neo-CK2αt specific to protein kinase CK2α and non-specific siRNA expression plasmid psiRNA-hH1neo-cont were transfected into Hep-2 cells respectively by lipofectamine method. Western blot was used to detect the expression of kinase CK2α protein. The apoptotic rate was measured by Annexin V-FITC/PI double-staining. The morphological changes of Hep-2 cells were observed under transmission electron microscope （ TEM ）. The expressions of bcl-2 and Bax protein were measured by Western blot. Results The expression of protein kinase CK2α protein significantly decreased in the Hep-2 cells transfected with psiRNA-hH1 neo-CK2α（ P 〈 0.01 ）. Compared with the untransfected cells and siRNA-hH1neo-cont transfected group, psiRNA-hH1neo-CK2α transfected group presented with classical ultrastructural features of apoptosis, such as karyopyknosis, chromatic agglutination adjacent to nuclear membrane and apoptotic body. The apoptotic rate of psiRNA-hH1 neo-CK2α transfected group was obviously higher than that in untransfected cells and siRNA-hH1neo-cont transfected group （25.66% ± 0.83% vs 3.66% ±0.43%, 5.18% ±0.22%, both P 〈 0.05）. Compared with two other groups, the bcl-2 protein expression of psiRNA-hH1 neo-CK2α transfected group decreased （0.20 ± 0.09 vs 0.72 ±± 0. 16, 0. 56 ±0.11, both P 〈 0.01 ）, the Bax protein expression increased （0.81 ± 0.17 vs 0.26 ± 0. 12, 0.33 ± 0.17,both P 〈 0.01 ） while the ratio of bcl-2 to Bax decreased （0.25 ±0.05 vs 2.76 ±0.21, 1.70 ±0. 22, both P 〈 0.01 ）. Conclusion Protein kinase CK2a plays an important role in the apoptosis of human laryngeal carcinoma cells possibly by decreasing bcl-2/Bax. Protein kinase CK2a may provide a potential therapeutic target against human laryngeal carcinoma