Hall M C, Li Y, Pong R C, Ely B, Sagalowsky A I, Hsieh J T
Department of Urology, University of Texas Southwestern Medical Center, Dallas 75235-9110, USA.
J Urol. 2000 Mar;163(3):1033-8.
To evaluate whether p21 (WAF-1/CIP1) should be considered a potential candidate for human bladder cancer gene therapy, we determined: (1) the basal level of p21 expression in bladder cancer cell lines, (2) the response of bladder cancer cells to increased p21 expression following p21 adenovirus infection, and (3) the mechanism of growth inhibition produced by p21 overexpression.
Five established human bladder cancer cell lines and one primary culture derived from an invasive transitional cell carcinoma were used in this study. To examine the effect of p21 protein on the growth of human bladder cancer cells, a recombinant adenovirus vector system containing p21 cDNA, under the control of cytomegalovirus promoter, was constructed. A control virus containing p21 in an antisense orientation was used to eliminate potential artifacts caused by viral toxicity.
Human bladder cancer cell lines exhibit variable endogenous p21 levels which correlate with the in vitro growth status. Significant, but highly variable increases in the steady-state level of p21 were detected in p21 adenovirus infected cells. Human bladder cancer cell lines responded heterogeneously to p21 adenovirus infection. Growth of the WH cell line was substantially inhibited in a dose and time-course dependent fashion. The mechanism of p21 growth inhibition was found to be due to G0/G1 arrest and not the induction of apoptosis. In contrast, p21 adenovirus failed to inhibit the growth of T24 bladder cancer cells because T24 cells were resistant to viral infection. The 253J bladder cancer cells exhibited marked sensitivity to adenovirus; substantial growth inhibition was seen with both sense and antisense p21 very early in the time course of infection.
We found significant variation in the basal level of p21 protein expression in several human bladder cancer cell lines. Increased p21 expression as a result of adenoviral infection may be a potent growth suppressor in some human bladder cancer because it elicits cell cycle arrest in G0/G1 stage, but not the induction of apoptosis. Bladder cancer cells exhibit a wide spectrum of sensitivity to adenoviral infection that may be caused by the presence of viral receptor heterogeneity. This wide spectrum of sensitivity has significant basic scientific and clinical implications and warrants further study.
为了评估p21（WAF-1/CIP1）是否应被视为人类膀胱癌基因治疗的潜在候选者，我们进行了以下研究：（1）测定膀胱癌细胞系中p21的基础表达水平；（2）检测p21腺病毒感染后膀胱癌细胞对p21表达增加的反应；（3）研究p21过表达产生生长抑制的机制。
本研究使用了五种已建立的人类膀胱癌细胞系和一种源自浸润性移行细胞癌的原代培养细胞。为了检测p21蛋白对人类膀胱癌细胞生长的影响，构建了一种重组腺病毒载体系统，该系统包含在巨细胞病毒启动子控制下的p21 cDNA。使用含有反义p21的对照病毒来消除由病毒毒性引起的潜在假象。
人类膀胱癌细胞系呈现出可变的内源性p21水平，这与体外生长状态相关。在p21腺病毒感染的细胞中检测到p21稳态水平有显著但高度可变的增加。人类膀胱癌细胞系对p21腺病毒感染的反应存在异质性。WH细胞系的生长以剂量和时间依赖性方式受到显著抑制。发现p21生长抑制的机制是由于G0/G1期停滞，而非诱导细胞凋亡。相反，p21腺病毒未能抑制T24膀胱癌细胞的生长，因为T24细胞对病毒感染具有抗性。253J膀胱癌细胞对腺病毒表现出明显的敏感性；在感染过程的早期，正义和反义p21均能显著抑制其生长。
我们发现几种人类膀胱癌细胞系中p21蛋白的基础表达水平存在显著差异。腺病毒感染导致的p21表达增加在某些人类膀胱癌中可能是一种有效的生长抑制因子，因为它能引发细胞周期在G0/G1期停滞，而非诱导细胞凋亡。膀胱癌细胞对腺病毒感染表现出广泛的敏感性，这可能是由病毒受体异质性导致的。这种广泛的敏感性具有重要的基础科学和临床意义，值得进一步研究。
