Characterization of NADPH oxidase 5 expression in human tumors and tumor cell lines with a novel mouse monoclonal antibody.
Characterization of NADPH oxidase 5 expression in human tumors and tumor cell lines with a novel mouse monoclonal antibody.
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DOI：
10.1016/j.freeradbiomed.2013.07.005
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发表时间：
2013-12
影响因子：
7.4
通讯作者：
Doroshow, James H.
中科院分区：
文献类型：
--
作者：
Antony, Smitha;Wu, Yongzhong;Hewitt, Stephen M.;Anver, Miriam R.;Butcher, Donna;Jiang, Guojian;Meitzler, Jennifer L.;Liu, Han;Juhasz, Agnes;Lu, Jiamo;Roy, Krishnendu K.;Doroshow, James H.
关键词：
NADPH 氧化酶 5 (Nox5) 产生的活性氧与生理和病理生理信号通路有关，包括癌症的发生和进展。然而，由于缺乏免疫学工具，对Nox5在肿瘤生物学中的作用的了解有限。 Nox5 蛋白在肿瘤和正常组织中的表达基本上是未知的。在此，我们报告了针对重组 Nox5 蛋白 (600-746) 的小鼠单克隆抗体的表征和用途，通过组织微阵列分析对人类肿瘤中的 Nox5 进行表达谱分析。使用我们的新型抗体，我们还报告了在人 UACC-257 黑色素瘤细胞中检测到内源性 Nox5 蛋白。采用免疫荧光、共聚焦显微镜和免疫组织化学技术来证明 Nox5 在整个 UACC-257 细胞中的定位，并具有核周增强。组织微阵列分析首次揭示了几种人类癌症中Nox5的大量过度表达，包括前列腺癌、乳腺癌、结肠癌、肺癌、脑癌和卵巢癌以及恶性黑色素瘤和非霍奇金淋巴瘤；大多数非恶性组织中的表达为阴性至弱。这种经过验证的小鼠单克隆抗体将促进进一步探索 Nox5 在人类病理生理学中的功能意义，包括肿瘤细胞的生长和增殖。
Reactive oxygen species generated by NADPH oxidase 5 (Nox5) have been implicated in physiological and pathophysiological signaling pathways, including cancer development and progression. However, because immunological tools are lacking, knowledge of the role of Nox5 in tumor biology has been limited; the expression of Nox5 protein across tumors and normal tissues is essentially unknown. Here, we report the characterization and use of a mouse monoclonal antibody against a recombinant Nox5 protein (600–746) for expression profiling of Nox5 in human tumors by tissue microarray analysis. Using our novel antibody, we also report the detection of endogenous Nox5 protein in human UACC-257 melanoma cells. Immunofluorescence, confocal microscopy, and immunohistochemical techniques were employed to demonstrate Nox5 localization throughout UACC-257 cells, with perinuclear enhancement. Tissue microarray analysis revealed, for the first time, substantial Nox5 overexpression in several human cancers including those of prostate, breast, colon, lung, brain, and ovary as well as in malignant melanoma and non-Hodgkin lymphoma; expression in most non-malignant tissues was negative to weak. This validated mouse monoclonal antibody will promote further exploration of the functional significance of Nox5 in human pathophysiology, including tumor cell growth and proliferation.
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