Sonic hedgehog (Shh) signaling promotes tumorigenicity and stemness via activation of epithelial-to-mesenchymal transition (EMT) in bladder cancer.
Activation of the sonic hedgehog (Shh) signaling pathway controls tumorigenesis in a variety of cancers. Here, we show a role for Shh signaling in the promotion of epithelial-to-mesenchymal transition (EMT), tumorigenicity, and stemness in the bladder cancer. EMT induction was assessed by the decreased expression of E-cadherin and ZO-1 and increased expression of N-cadherin. The induced EMT was associated with increased cell motility, invasiveness, and clonogenicity. These progression relevant behaviors were attenuated by treatment with Hh inhibitors cyclopamine and GDC-0449, and after knockdown by Shh-siRNA, and led to reversal of the EMT phenotype. The results with HTB-9 were confirmed using a second bladder cancer cell line, BFTC905 (DM). In a xenograft mouse model TGF-β1 treated HTB-9 cells exhibited enhanced tumor growth. Although normal bladder epithelial cells could also undergo EMT and upregulate Shh with TGF-β1 they did not exhibit tumorigenicity. The TGF-β1 treated HTB-9 xenografts showed strong evidence for a switch to a more stem cell like phenotype, with functional activation of CD133, Sox2, Nanog, and Oct4. The bladder cancer specific stem cell markers CK5 and CK14 were upregulated in the TGF-β1 treated xenograft tumor samples, while CD44 remained unchanged in both treated and untreated tumors. Immunohistochemical analysis of 22 primary human bladder tumors indicated that Shh expression was positively correlated with tumor grade and stage. Elevated expression of Ki-67, Shh, Gli2, and N-cadherin were observed in the high grade and stage human bladder tumor samples, and conversely, the downregulation of these genes were observed in the low grade and stage tumor samples. Collectively, this study indicates that TGF-β1-induced Shh may regulate EMT and tumorigenicity in bladder cancer. Our studies reveal that the TGF-β1 induction of EMT and Shh is cell type context dependent. Thus, targeting the Shh pathway could be clinically beneficial in the ability to reverse the EMT phenotype of tumor cells and potentially inhibit bladder cancer progression and metastasis.
Developmental and Stem Cell Biology, The Hospital for Sick Children, Toronto, ON, Canada.|Division of Urology, The Hospital for Sick Children, Toronto, ON, Canada.
S S
SS
Islam
Developmental and Stem Cell Biology, The Hospital for Sick Children, Toronto, ON, Canada.
R B
RB
Mokhtari
Department of Biochemistry and Molecular Biology, University of Chittagong, Chittagong, Bangladesh.
A S
AS
Noman
Genetics and Genome Biology, The Hospital for Sick Children, Toronto, ON, Canada.
M
M
Uddin
Department of Pathology, Chittagong Medical College, Chittagong, Bangladesh.
M Z
MZ
Rahman
Faculty of Biological Sciences, University of Chittagong, Chittagong, Bangladesh.
M A
MA
Azadi
Department of Uro-Oncology, Mount Sinai Hospital, Toronto, ON, Canada.
A
A
Zlotta
Department of Pathology, Laboratory Medicine Program, University Health Network, Toronto, ON, Canada.
T
T
van der Kwast
Developmental and Stem Cell Biology, The Hospital for Sick Children, Toronto, ON, Canada.
H
H
Yeger
Developmental and Stem Cell Biology, The Hospital for Sick Children, Toronto, ON, Canada.|Division of Urology, The Hospital for Sick Children, Toronto, ON, Canada.
W A
WA
Farhat
D000818:Animals; D014408:Biomarkers, Tumor / Q000378:metabolism; D045744:Cell Line, Tumor; D002465:Cell Movement; D058750:Epithelial-Mesenchymal Transition / Q000187:drug effects; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D053823:Hedgehog Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051379:Mice; D060787:Neoplasm Grading; D009367:Neoplasm Staging; D009368:Neoplasm Transplantation; D014411:Neoplastic Stem Cells / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D015398:Signal Transduction / Q000187:drug effects; D053773:Transforming Growth Factor beta1 / Q000494:pharmacology*; D001749:Urinary Bladder Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology*
D014408:Biomarkers, Tumor; D053823:Hedgehog Proteins; C509239:SHH protein, human; D053773:Transforming Growth Factor beta1
Bladder cancer; EMT; Shh