Evaluation of thymosin β4 in the regulation of epithelial-mesenchymal transformation in urothelial carcinoma.
OBJECTIVE
To study the underlying alteration in the expression of epithelial markers involved in epithelial-mesenchymal transition (EMT), and elucidate the potential mechanism(s) for Tβ4-induced EMT-like phenotypic changes in bladder cancer cells.


METHODS
All tissue samples in this study were obtained from clinical patients of the Union Hospital of Tongji Medical College, and were confirmed by surgery and pathology. Of these, normal bladder tissues (control), primary urothelial carcinoma of different grades (Stage pTa, Stage pT3), bladder paracancerous tissues, accompanied with 2 bladder cancer cell lines (BIU-87 and T24), were divided into 6 groups. Quantitative RT-PCR, Western blotting, and immunohistochemical study of adhesion molecules Tβ4, ILK, E-cadherin, and β-catenin involved in EMT were carried out. A lentiviral gene transferring vector containing the RNA polymerase III-dependent U6 promoter to express short hairpin RNA (shRNA) directed against Tβ4 was also applied. In the present study, all agents were evaluated using commercial kits.


RESULTS
A strong correlation between the expression levels of Tβ4, ILK, E-cadherin, and β-catenin was found in the bladder transitional cell carcinoma (TCC) patients. In the BIU-87 and T24 bladder cancer cells overexpressing Tβ4, which were accompanied by a loss of E-cadherin as well as a cytosolic accumulation of β-catenin, up-regulation of ILK was also revealed. The inhibition of the Tβ4 expression with lentiviral shRNA vector could raise EMT-like phenotypic changes, significantly depressed motility, and subsequent invasiveness of bladder cancer cells.


CONCLUSIONS
Our results imply that the Tβ4 is likely to play a crucial role in EMT progression, and that inhibition of the Tβ4 expression or interactions with other genes should be novel therapeutic targets for bladder cancers with high invasive and metastatic potential.
Wang
Zhi Yu
ZY
Department of Urology, Union Hospital of Tongji Medical College, Huazhong University of Science and Technology, Hubei, Republic of China.
Zeng
Fu Qing
FQ
Zhu
Zhao Hui
ZH
Jiang
Guo Song
GS
Lv
Lei
L
Wan
Feng
F
Dong
Rui
R
Xiao
Xing Yuan
XY
Xing
Shi An
SA
D000368:Aged; D015153:Blotting, Western; D015820:Cadherins / Q000235:genetics / Q000378:metabolism; D002295:Carcinoma, Transitional Cell / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D016022:Case-Control Studies; D002465:Cell Movement; D049109:Cell Proliferation; D058750:Epithelial-Mesenchymal Transition; D005260:Female; D005455:Fluorescent Antibody Technique; D005500:Follow-Up Studies; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007124:Immunoenzyme Techniques; D008297:Male; D008875:Middle Aged; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D013947:Thymosin / Q000235:genetics / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D001743:Urinary Bladder / Q000378:metabolism; D001749:Urinary Bladder Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D014945:Wound Healing; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism*; D000099201:Thymosin beta(4)
D015820:Cadherins; D012333:RNA, Messenger; D013947:Thymosin; D051176:beta Catenin; D000099201:Thymosin beta(4)