Expression of hTERT immortalises normal human urothelial cells without inactivation of the p16/Rb pathway
Expression of hTERT immortalises normal human urothelial cells without inactivation of the p16/Rb pathway
复制标题
DOI：
10.1038/sj.onc.1209513
复制
发表时间：
2006-08-17
期刊：
影响因子：
8
通讯作者：
Knowles, M. A.
中科院分区：
文献类型：
--
作者：
Chapman, E. J.;Hurst, C. D.;Knowles, M. A.
CDKN 2A基因座在尿路上皮细胞癌（UCC）中经常失活，但这种改变如何导致膀胱肿瘤发生尚不清楚。虽然大多数UCC表达端粒酶，但p16/Rb通路的失活通常是体外永生化所必需的。这和p16参与正常人尿路上皮细胞（NHUC）的衰老表明，CDKN 2A缺失可能有助于绕过衰老，并允许永生化。CDKN 2A编码p16和p14（ARF），因此该基因座的失活可以破坏Rb和p53肿瘤抑制途径。逆转录病毒介导的转导被用于特异性地调节NHUC中的p16/R带/或p53肿瘤抑制通路并表达人端粒酶逆转录酶（hTERT）。hTERT的表达绕过Rb和p53通路依赖的增殖和永生化NHUC的障碍。TERT-NHUC具有正常核型，非致瘤性，并且意外地保留了CDKN 2A。因此，UCC中CDKN 2A失活的表型意义可能不仅仅与衰老的旁路有关。人尿路上皮细胞的表型测定依赖于来源于浸润性肿瘤或通过表达SV 40-大T细胞而永生化的NHUC的细胞株。遗传正常但永生的NHUC系的生产现在为实验提供了一个有价值的平台，以检查UCC肿瘤发生所需的事件的时间和组合。
The CDKN2A locus is frequently inactivated in urothelial cell carcinoma (UCC), yet how this alteration contributes to bladder tumorigenesis is not known. Although most UCC express telomerase, inactivation of the p16/Rb pathway is generally required for in vitro immortalisation. This and the involvement of p16 in senescence of normal human urothelial cells (NHUC) suggest that CDKN2A deletion may aid bypass of senescence and allow immortalisation. CDKN2A encodes p16 and p14(ARF) and therefore inactivation of this locus can disrupt both the Rb and p53 tumour suppressor pathways. Retrovirus-mediated transduction was used to specifically modulate the p16/Rband/or p53 tumour suppressor pathways in NHUC and to express human telomerase reverse transcriptase (hTERT). Expression of hTERT bypassed Rb and p53 pathway-dependent barriers to proliferation and immortalised NHUC. TERT-NHUC had normal karyotypes, were non-tumorigenic and unexpectedly retained CDKN2A. Thus, the phenotypic significance of inactivation of CDKN2A in UCC may not be solely related to bypass of senescence. Phenotypic assays in human urothelial cells have relied on cell strains derived from invasive tumours or NHUC immortalised by expression of SV40-large T. The production of genetically normal but immortal NHUC lines now provides a valuable platform for experiments to examine the timing and combination of events necessary for UCC tumorigenesis.