Activation of Sonic hedgehog signaling pathway in S-type neuroblastoma cell lines.
The effects of Sonic hedgehog (Shh) signaling pathway activation on S-type neuroblastoma (NB) cell lines and its role in NB tumorigenesis were investigated. Immunohistochemistry was used to detect the expression of Shh pathway components-Patched1 (PTCH1) and Gli1 in 40 human primary NB samples. Western blotting and RT-PCR were used to examine the protein expression and mRNA levels of PTCH1 and Gli1 in three kinds of S-type NB cell lines (SK-N-AS, SK-N-SH and SHEP1), respectively. Exogenous Shh was administrated to activate Shh signaling pathway while cyclopamine was used as a selective antagonist of Shh pathway. S-type NB cell lines were treated with different concentrations of Shh or/and cyclopamine for different durations. Cell viability was measured by using MTT method. Apoptosis rate and cell cycle were assayed by flow cytometry. The xenograft experiments were used to evaluate the role of Shh pathway in tumor growth in immunodeficient mice. High-level expression of PTCH1 and Gli1 was detected in both NB samples and S-type NB cell lines. Cyclopamine decreased the survival rate of the three cell lines while Shh increased it, and the inhibition effects of cyclopamine could be partially reversed by shh pre-treatment. Cyclopamine induced the cell apoptosis and the cell cycle arrest in G0/G1 phase, while Shh induced the reverse effects and could partially prevent effects of cyclopamine. Cyclopamine could also inhibit the growth of NB in vivo. Our studies revealed that activation of the Shh pathway is important for survival and proliferation of S-type NB cells in vivo and in vitro through affecting cell apoptosis and cell cycle, suggesting a new therapeutic approach to NB.
Zhou
Yunan
Y
Department of Neurology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China, 430022. talent62@hotmail.com
Dai
Ruolian
R
Mao
Ling
L
Xia
Yuanpeng
Y
Yao
Yufang
Y
Yang
Xue
X
Hu
Bo
B
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D002470:Cell Survival; D053823:Hedgehog Proteins / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051379:Mice; D009447:Neuroblastoma / Q000378:metabolism / Q000473:pathology*; D000072079:Patched Receptors; D000072081:Patched-1 Receptor; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000378:metabolism; D015398:Signal Transduction; D014157:Transcription Factors / Q000378:metabolism; D014704:Veratrum Alkaloids / Q000494:pharmacology; D000071676:Zinc Finger Protein GLI1
C400551:GLI1 protein, human; D053823:Hedgehog Proteins; C000606236:PTCH1 protein, human; D000072079:Patched Receptors; D000072081:Patched-1 Receptor; C000606240:Ptch1 protein, mouse; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; C509239:SHH protein, human; D014157:Transcription Factors; D014704:Veratrum Alkaloids; D000071676:Zinc Finger Protein GLI1; C000541:cyclopamine