Ahn Woong Shick, Bae Su Mi, Lee Keun Ho, Lee Joon Mo, Namkoong Sung Eun, Chun Heung Jae, Kim Chong Kook, Kim Yong-Wan
Department of Obstetrics and Gynecology, The Catholic University of Korea, Seoul, South Korea.
Gynecol Oncol. 2004 Feb;92(2):611-21. doi: 10.1016/j.ygyno.2003.10.033.
We investigated the time-course expression patterns of p53 and E6 on cervical cancer cells to obtain a molecular level understanding of cell-dependent tumor growth suppression effects of recombinant adenovirus expressing p53 in vitro and in vivo.
Four human papillomavirus (HPV)-infected human cervical cancer cell lines (HPV 16-positive cells, CaSki and SiHa cells; and HPV 18-positive cells, HeLa and HeLaS3 cells) were used. Also, HPV negative C33A and HT3 cell line that has a mutation on p53 gene were used. After infection with AdCMVp53, the cell growth inhibition was studied via cell count assay, MTT assay, and Neutral red assay. After transfecting AdCMVp53 and AdCMVLacZ into the cancer cells-xenografted nude mice, antitumor effects were investigated for 1 month, respectively.
For each cervical cancer cell, IC50 was as follows; CaSki (68.5 multiplicity of infection, or MOI), SiHa (43.5 MOI), HeLa (31 MOI), HeLaS3 (42 MOI), C33A (21 MOI), and HT3 (62 MOI). In particular, complete inhibition of cell growth was observed at 125 MOI in both CaSki and SiHa cells. However, the complete inhibition was detected at 62.5 MOI in HeLa and HeLaS3. In contrast, at these MOI, no suppression of cell growth was observed when cells were infected with recombinant adenovirus expressing beta-gal as a negative control. The levels of p53 protein were notably expressed in CaSki and HeLa more than in SiHa and HeLaS3 on days 2 and 4. However, the p53 was only detected in HeLaS3 on day 6. In contrast, p53 expression was continually maintained in C33A and HT3 during the same periods. After transfection AdCMVp53 into CaSki- and SiHa-xenografted nude mice, the size of tumor was remarkably decreased in SiHa cells as compared to AdCMVLacZ transfection.
The adenovirus-mediated p53 gene transfection was done effectively in vitro and in vivo. Also, the antitumor effects were accomplished via differential role of p53-specific apoptotic cell death, which is dependent upon the cervical cancer cell line.
我们研究了p53和E6在宫颈癌细胞上的时间进程表达模式，以从分子水平了解表达p53的重组腺病毒在体外和体内对细胞依赖性肿瘤生长的抑制作用。
使用了四种感染人乳头瘤病毒（HPV）的人宫颈癌细胞系（HPV 16阳性细胞，CaSki和SiHa细胞；以及HPV 18阳性细胞，HeLa和HeLaS3细胞）。此外，还使用了p53基因发生突变的HPV阴性C33A和HT3细胞系。用AdCMVp53感染后，通过细胞计数法、MTT法和中性红法研究细胞生长抑制情况。将AdCMVp53和AdCMVLacZ转染到接种了癌细胞的裸鼠体内后，分别观察1个月的抗肿瘤效果。
对于每种宫颈癌细胞，半数抑制浓度（IC50）如下：CaSki（68.5感染复数，即MOI），SiHa（43.5 MOI），HeLa（31 MOI），HeLaS3（42 MOI），C33A（21 MOI）和HT3（62 MOI）。特别地，在CaSki和SiHa细胞中，在125 MOI时观察到细胞生长完全抑制。然而，在HeLa和HeLaS3细胞中，在62.5 MOI时检测到完全抑制。相比之下，当用表达β-半乳糖苷酶的重组腺病毒作为阴性对照感染细胞时，在这些MOI下未观察到细胞生长抑制。在第2天和第4天，CaSki和HeLa中p53蛋白的表达水平明显高于SiHa和HeLaS3。然而，在第6天仅在HeLaS3中检测到p53。相比之下，在同一时期C33A和HT3中p53表达持续维持。将AdCMVp53转染到接种CaSki和SiHa细胞的裸鼠体内后，与转染AdCMVLacZ相比，SiHa细胞中的肿瘤大小明显减小。
腺病毒介导的p53基因转染在体外和体内均有效进行。此外，抗肿瘤作用是通过p53特异性凋亡性细胞死亡的不同作用实现的，这取决于宫颈癌细胞系。
