Activation of Sonic Hedgehog Signaling Pathway in Olfactory Neuroblastoma
Activation of Sonic Hedgehog Signaling Pathway in Olfactory Neuroblastoma
复制标题
嗅神经母细胞瘤中 Sonic Hedgehog 信号通路的激活
作者：
Mao, Ling;Xia, Yuan-peng;Hu, Bo
目的：Sonic hedgehog(Shh)信号通路与肿瘤的发生发展有关，但Shh信号在嗅神经母细胞瘤(ONB)发生发展中的作用尚不清楚。本研究旨在探讨Shh信号调控与ONB发病机制的关系。方法：用免疫组织化学方法检测Shh信号元件在人非肿瘤嗅觉上皮和ONB标本中的表达，用RT-PCR和免疫印迹方法检测人ONB细胞系中Shh信号元件的表达。采用四甲基偶氮唑盐比色法、软琼脂集落形成法和流式细胞术分别检测Shh途径的选择性抑制剂环丙胺和/或外源性Shh对ONB细胞增殖、周期和细胞凋亡的影响。免疫印迹法检测Shh信号对Shh信号成分表达的影响；定量RT-PCR法检测Shh信号对细胞周期调控因子表达的影响。结果：Pacthed1、Gli1和Gli2分别在70%、70%和65%的人ONB组织中表达，且在ONB细胞系中呈阳性表达，而在非肿瘤嗅觉上皮细胞中未见表达。在体外，环丙胺可抑制ONB细胞的增殖和集落形成，诱导ONB细胞周期停滞和凋亡，下调Pacthed1、Gli1和Cyclin D1的表达，上调p21的表达。外源性Shh可部分或完全消除环多巴胺的这些调节作用。结论：Shh信号通路对ONB的生长至关重要。
Objectives: Sonic hedgehog (Shh) signaling pathway is associated with tumor development; however, the role of Shh signaling in the development of olfactory neuroblastoma (ONB) is unknown. This study aimed to investigate the relationship between the regulation of Shh signaling and the pathogenesis of ONB. Methods: The expression of Shh signaling components was characterized by immunohistochemistry in human non-tumor olfactory epithelium and ONB specimens, and by RT-PCR and immunoblotting in human ONB cell lines. The impact of the treatment with cyclopamine (a selective inhibitor of the Shh pathway) and/or exogenous Shh on ONB cell proliferation, cycle and apoptosis was examined by MTT, soft agar colony formation and flow cytometry assays, respectively. The influence of Shh signaling on the expression of Shh signaling components and cell cycle-related regulators was determined by immunoblotting and quantitative RT-PCR, respectively. Results: The expression of Pacthed1, Gli1 and Gli2 was detected in 70, 70, and 65% of human ONB specimens, respectively, and in proportion of ONB cell lines, but not in non-tumor olfactory epithelium. Treatment with cyclopamine inhibited the proliferation and colony formation of ONB cells, induced ONB cell cycle arrest and apoptosis, and down-regulated the expression of Pacthed1, Gli1 and cyclin D1, but up-regulated p21 expression in vitro. These regulatory effects of cyclopamine were partially or completely erased by exogenous Shh. Conclusion: These data suggest that the Shh signaling pathway is crucial for the growth of ONB.