PTEN can inhibit in vitro organotypic and in vivo orthotopic invasion of human bladder cancer cells even in the absence of its lipid phosphatase activity
PTEN can inhibit in vitro organotypic and in vivo orthotopic invasion of human bladder cancer cells even in the absence of its lipid phosphatase activity
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DOI：
10.1038/sj.onc.1207599
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发表时间：
2004-09-02
期刊：
影响因子：
8
通讯作者：
Theodorescu, D
中科院分区：
文献类型：
--
作者：
Gildea, JJ;Herlevsen, M;Theodorescu, D
最近的研究发现，在浸润性膀胱癌中PTEN抑癌基因的改变频率高于浅表性疾病，表明PTEN在这一过程中是重要的。PTEN在膀胱癌侵袭中的作用进一步由以下事实表明：PTEN是细胞运动的调节剂，细胞运动是肿瘤侵袭的必要组分。然而，它是未知的，是否PTEN是机械地参与“体内”肿瘤侵袭或仅仅是一种附带现象，如果前者是真的，这个过程是否依赖于其蛋白或脂质磷酸酶活性。为了解决这些问题，我们用野生型PTEN构建体或脂质磷酸酶（G129 E）缺陷的构建体或蛋白和脂质磷酸酶（G129 R）活性两者稳定转染了几种常用的具有已知侵袭性表型的人膀胱癌细胞系。在这里，我们表明，趋化性抑制的野生型和G129 E突变体的PTEN，但不是由G129 R转染的细胞。使用一种新的器官型体外侵袭试验，我们评估了野生型和突变型PTEN转基因表达对T24 T（一种具有功能受损的PTEN的人膀胱癌细胞系）侵袭能力的影响。结果表明G129 E突变体与野生型PTEN转染一样有效地阻断侵袭。与野生型基因相反，该突变体在琼脂中对细胞克隆形成没有影响。为了进一步确定PTEN在肿瘤侵袭中的作用，我们在原位体内试验中评估了载体和PTEN转染的T24 T细胞，该试验忠实地再现了人类疾病。在本实验完成时对小鼠膀胱进行的显微镜检查与器官型试验获得的结果相似。我们的研究结果首次证明：（1）PTEN对细胞运动的抑制作用转化为对体内侵袭的抑制;（2）即使在缺乏其脂质磷酸酶活性的情况下，PTEN也能抑制肿瘤的侵袭;（三）体外器官型方法如何用作体内侵袭的替代物，允许快速解剖导致这种表型的分子过程，同时减少研究中使用的动物数量。
Recent studies have found a higher frequency of the PTEN tumor-suppressor gene alterations in invasive bladder carcinoma than in superficial disease, suggesting that PTEN is important in this process. A role of PTEN in bladder cancer invasion is further suggested by the fact that PTEN is a regulator of cell motility, a necessary component of tumor invasion. However, it is unknown whether PTEN is mechanistically involved in 'in vivo' tumor invasion or merely an epiphenomenon and, if the former is true, whether this process is dependent on its protein or lipid phosphatase activities. To address these issues, we stably transfected several commonly used human bladder cancer cell lines with known invasive phenotypes with either wild-type PTEN constructs or those deficient in the lipid phosphatase (G129E) or both protein and lipid phosphatase (G129R) activities. Here we show that chemotaxis was inhibited by both the wild-type and G129E mutant of PTEN but not by G129R-transfected cells. Using a novel organotypic in vitro invasion assay, we evaluated the impact of wild-type and mutant PTEN transgene expression on the invasive ability of T24T, a human bladder cancer cell line with a functionally impaired PTEN. Results indicate that the G129E mutant blocks invasion as efficiently as wild-type PTEN transfection. In contrast to the wild-type gene, this mutant has no effect on cell clonogenicity in agar. To further establish the role of PTEN in tumor invasion, we evaluated vector- and PTEN-transfected T24T cells in an orthotopic in vivo assay that faithfully reproduces human disease. Microscopic examination of murine bladders at the completion of this experiment parallels the results obtained with the organotypic assay. Our results are the first demonstration: ( 1) that the inhibitory effects of PTEN on cell motility translate into suppression of in vivo invasion; ( 2) that PTEN can inhibit tumor invasion even in the absence of its lipid phosphatase activity; ( 3) how organotypic in vitro approaches can be used as surrogates of in vivo invasion allowing rapid dissection of molecular processes leading to this phenotype while reducing the number of animals used in research.