Unit of General Pathology, Aging Research Center (Ce.S.I.), G. d'Annunzio University Foundation, Via Colle dell'Ara, 66100, Chieti, Italy.
Acta Neuropathol. 2013 Oct;126(4):575-94. doi: 10.1007/s00401-013-1165-y. Epub 2013 Aug 18.
Head and neck paragangliomas, rare neoplasms of the paraganglia composed of nests of neurosecretory and glial cells embedded in vascular stroma, provide a remarkable example of organoid tumor architecture. To identify genes and pathways commonly deregulated in head and neck paraganglioma, we integrated high-density genome-wide copy number variation (CNV) analysis with microRNA and immunomorphological studies. Gene-centric CNV analysis of 24 cases identified a list of 104 genes most significantly targeted by tumor-associated alterations. The "NOTCH signaling pathway" was the most significantly enriched term in the list (P = 0.002 after Bonferroni or Benjamini correction). Expression of the relevant NOTCH pathway proteins in sustentacular (glial), chief (neuroendocrine) and endothelial cells was confirmed by immunohistochemistry in 47 head and neck paraganglioma cases. There were no relationships between level and pattern of NOTCH1/JAG2 protein expression and germline mutation status in the SDH genes, implicated in paraganglioma predisposition, or the presence/absence of immunostaining for SDHB, a surrogate marker of SDH mutations. Interestingly, NOTCH upregulation was observed also in cases with no evidence of CNVs at NOTCH signaling genes, suggesting altered epigenetic modulation of this pathway. To address this issue we performed microarray-based microRNA expression analyses. Notably 5 microRNAs (miR-200a,b,c and miR-34b,c), including those most downregulated in the tumors, correlated to NOTCH signaling and directly targeted NOTCH1 in in vitro experiments using SH-SY5Y neuroblastoma cells. Furthermore, lentiviral transduction of miR-200s and miR-34s in patient-derived primary tympano-jugular paraganglioma cell cultures was associated with NOTCH1 downregulation and increased levels of markers of cell toxicity and cell death. Taken together, our results provide an integrated view of common molecular alterations associated with head and neck paraganglioma and reveal an essential role of NOTCH pathway deregulation in this tumor type.
头颈部副神经节瘤是由神经分泌细胞和神经胶质细胞巢组成的副神经节肿瘤，嵌入富含血管的基质中，为器官样肿瘤结构提供了一个显著的例子。为了鉴定头颈部副神经节瘤中常见的失调基因和途径，我们将高密度全基因组拷贝数变异 (CNV) 分析与 microRNA 和免疫形态学研究相结合。对 24 例病例的基因中心 CNV 分析确定了一组受肿瘤相关改变靶向的 104 个基因。“NOTCH 信号通路”是列表中最显著富集的术语（经 Bonferroni 或 Benjamini 校正后 P = 0.002）。在 47 例头颈部副神经节瘤病例中，通过免疫组织化学证实了相关 NOTCH 通路蛋白在支持细胞（神经胶质）、主细胞（神经内分泌）和内皮细胞中的表达。NOTCH1/JAG2 蛋白表达水平和模式与 SDH 基因中的种系突变状态（与副神经节瘤易感性相关）或 SDHB 的免疫染色（SDH 突变的替代标志物）之间没有关系。有趣的是，在没有 NOTCH 信号基因 CNV 证据的情况下，也观察到 NOTCH 上调，提示该途径的表观遗传调控改变。为了解决这个问题，我们进行了基于微阵列的 microRNA 表达分析。值得注意的是，包括在肿瘤中下调最明显的 5 个 microRNA（miR-200a，b，c 和 miR-34b，c），与 NOTCH 信号相关，并在体外实验中直接靶向 SH-SY5Y 神经母细胞瘤细胞中的 NOTCH1。此外，miR-200s 和 miR-34s 的慢病毒转导在源自患者的原发性鼓室颈副神经节瘤细胞培养物中与 NOTCH1 下调和细胞毒性和细胞死亡标志物水平升高相关。总之，我们的结果提供了与头颈部副神经节瘤相关的常见分子改变的综合视图，并揭示了 NOTCH 通路失调在这种肿瘤类型中的重要作用。
