Huang Chen, Liu Li-ying, Song Tu-sheng, Ni Lei, Song Li-ping, Si Lü-sheng
Key Laboratory of Environment and Genes Related to Diseases, Xi'an Jiaotong University, Xi'an 710061, China.
Nan Fang Yi Ke Da Xue Xue Bao. 2006 Jan;26(1):11-5.
To observe the effect of small interfering RNA (siRNA)-induced MAPK p42 silencing on the survival of HeLa cells.
Two siRNAs targeting at the MAPK p42 gene and one random siRNA were synthesized respectively by Silencer siRNA Construction Kit and transfected into HeLa cells by Lipofectamin 2000. The expression of p42(MAPK) in the transfected HeLa cells was analyzed by Western blotting and immunohistochemistry, and the morphology of cells were observed with electron microscope. TUNEL assay and Annexin V/PI staining were employed for detecting the cell apoptosis.
The expression of p42(MAPK) in the HeLa cells was remarkably suppressed after transfection with the two siRNAs, reduced by about 2.5 and 3.2 folds respectively in comparison with the negative control. Chromatin margination in the cell nuclei were observed in the transfected cells, and TUNEL assay and Annexin V/PI staining further confirmed the occurrence of cell apoptosis.
In vitro MAPK p42 siRNA-1 and siRNA-2 transfection can specifically silence the gene expression and induce apoptosis of HeLa cells.
观察小干扰RNA（siRNA）诱导的丝裂原活化蛋白激酶（MAPK）p42沉默对人宫颈癌HeLa细胞存活的影响。
用Silencer siRNA构建试剂盒分别合成两条靶向MAPK p42基因的siRNA和一条随机siRNA，并用Lipofectamin 2000转染HeLa细胞。采用蛋白质免疫印迹法和免疫组织化学法分析转染后HeLa细胞中p42（MAPK）的表达情况，并用电子显微镜观察细胞形态。采用脱氧核糖核苷酸末端转移酶介导的缺口末端标记法（TUNEL）和膜联蛋白V/碘化丙啶（Annexin V/PI）染色检测细胞凋亡情况。
转染两条siRNA后，HeLa细胞中p42（MAPK）的表达明显受到抑制，与阴性对照相比，分别降低了约2.5倍和3.2倍。在转染细胞的细胞核中观察到染色质边集现象，TUNEL法和Annexin V/PI染色进一步证实了细胞凋亡的发生。
体外转染MAPK p42 siRNA-1和siRNA-2可特异性沉默基因表达并诱导HeLa细胞凋亡。
