Harada Hideki, Nakagawa Hiroshi, Oyama Kenji, Takaoka Munenori, Andl Claudia D, Jacobmeier Birgit, von Werder Alexander, Enders Gregory H, Opitz Oliver G, Rustgi Anil K
Gastroenterology Division, University of Pennsylvania, Philadelphia, PA 19104, USA.
Mol Cancer Res. 2003 Aug;1(10):729-38.
Normal human somatic cells have a finite life span and undergo replicative senescence after a limited number of cell divisions. Erosion of telomeric DNA has emerged as a key factor in senescence, which is antagonized during cell immortalization and transformation. To clarify the involvement of telomerase in the immortalization of keratinocytes, catalytic subunit of telomerase (hTERT) expression was restored in normal human esophageal epithelial cells (EPC2). EPC2-hTERT cells overcame senescence and were immortalized without p16INK4a genetic or epigenetic alterations. p16INK4a was expressed at moderate levels and remained functional as evidenced by induction with UV treatment and binding to cyclin-dependent kinase 4 and 6. There were no mutations in the p53 gene, and p53 was functionally intact. Importantly, senescence could be activated in the immortalized EPC2-hTERT cells by overexpression of oncogenic H-ras or p16INK4a. Furthermore, the EPC2-hTERT cells yielded basal cell hyperplasia in an innovative organotypic culture system in contrast to a normal epithelium from parental cells. These comprehensive results indicate that the expression of telomerase induces immortalization of normal human esophageal keratinocytes without inactivation of p16INK4a/pRb pathway or abrogation of the p53 pathway.
正常人类体细胞具有有限的寿命，并在有限次数的细胞分裂后经历复制性衰老。端粒DNA的侵蚀已成为衰老的关键因素，在细胞永生化和转化过程中受到拮抗。为了阐明端粒酶在角质形成细胞永生化中的作用，在正常人食管上皮细胞（EPC2）中恢复了端粒酶催化亚基（hTERT）的表达。EPC2-hTERT细胞克服了衰老并实现了永生化，且没有p16INK4a基因或表观遗传改变。p16INK4a以中等水平表达并保持功能，紫外线处理诱导以及与细胞周期蛋白依赖性激酶4和6结合证明了这一点。p53基因没有突变，且p53功能完整。重要的是，通过致癌性H-ras或p16INK4a的过表达，可以在永生化的EPC2-hTERT细胞中激活衰老。此外，与来自亲代细胞的正常上皮相比，EPC2-hTERT细胞在创新的器官型培养系统中产生了基底细胞增生。这些综合结果表明，端粒酶的表达诱导正常人食管角质形成细胞永生化，而不会使p16INK4a/pRb途径失活或废除p53途径。
