Exploring the involvement of NOX4 in hypoxia adaptation of papillary thyroid cancer

Hypoxic stress is a feature of rapidly growing thyroid tumors. Cancer progression is thought to be driven by reactive oxygen species (ROS) induced hypoxia adaptation. NADPH oxidases (NOXs), which produce ROS as their primary and sole function, has become of particular interest in thyroid malignancy. NOX4 was demonstrated to be upregulated in papillary thyroid cancers, functioning as a mitochondrial energetic sensor to modulate ATP levels, mediating overproduction of ROS induced by IL-δ, inversely correlating to thyroid differentiation. In this study, we analyzed hypoxia-treated BCPAP cells transfected with siRNA against NOX4. Results provides insight into the role of NOX4 in hypoxia adaptation. BCPAP cells were transfected with scrambled siRNA or siRNA against NOX4. Subsequently the siRNA-transfected cells were cultured within 1% oxygen incubator. Three independent transfections were performed cells with control siRNA and for cells with siRNA specific to NOX4 where each transfection constitutes biological replicate. Knock-down of NOX4 in all samples was confirmed by quantitative real-time PCR.

低氧应激是快速增殖性甲状腺肿瘤的典型特征。目前学界普遍认为，肿瘤进展由活性氧（reactive oxygen species，ROS）介导的低氧适应所驱动。烟酰胺腺嘌呤二核苷酸磷酸氧化酶（NADPH oxidases，NOXs）的核心且唯一功能即为产生活性氧，目前在甲状腺恶性肿瘤研究中受到广泛关注。已有研究证实，NOX4在甲状腺乳头状癌中表达上调，其可作为线粒体能量传感器调控三磷酸腺苷（adenosine triphosphate，ATP）水平，介导白细胞介素δ（IL-δ）诱导的活性氧过度产生，且与甲状腺分化程度呈负相关。本研究针对转染了NOX4靶向小干扰RNA（small interfering RNA，siRNA）的低氧处理BCPAP细胞展开分析，研究结果为阐明NOX4在低氧适应中的作用提供了新的见解。实验中将BCPAP细胞分别转染阴性对照siRNA（scrambled siRNA）与NOX4靶向siRNA，随后将转染后的细胞置于氧浓度为1%的培养箱中进行培养。本研究针对阴性对照siRNA组与NOX4靶向siRNA组细胞分别开展了三次独立转染，每次转染均视为一个生物学重复。所有样本中NOX4的敲低效果均通过实时荧光定量聚合酶链反应（quantitative real-time PCR）得以验证。