Microglial annexin A3 downregulation alleviates bone cancer-induced pain through inhibiting the Hif-1α/vascular endothelial growth factor signaling pathway
Microglial annexin A3 downregulation alleviates bone cancer-induced pain through inhibiting the Hif-1α/vascular endothelial growth factor signaling pathway
复制标题
小胶质细胞膜联蛋白 A3 下调通过抑制 Hif-1α/血管内皮生长因子信号通路减轻骨癌引起的疼痛
DOI：
10.1097/j.pain.0000000000001962
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发表时间：
2020-12-01
期刊：
影响因子：
7.4
通讯作者：
Zou, Wangyuan
中科院分区：
文献类型：
--
作者：
Zhang, Zengli;Deng, Meiling;Zou, Wangyuan
骨癌引起的疼痛（BCP）是一个具有挑战性的临床问题，因为传统的治疗方法往往只有部分有效。Annexin A3 （ANXA3）在脊髓小胶质细胞中高表达，BCP时其表达上调。然而，小胶质细胞ANXA3在BCP发生和维持中的作用及其潜在的分子机制尚不清楚。本研究使用转移性肺BCP模型对雄性小鼠进行了研究。在骨癌诱导前14天和诱导后7天鞘内注射腺相关病毒shan -shANXA3 (AAV-shANXA3)，通过测量足退缩机械阈值、足退缩热潜伏期和后肢自发抬起来评估相关疼痛行为。慢病毒转染（LV-shANXA3）可下调N9小胶质细胞中ANXA3蛋白的表达。测定ANXA3、缺氧诱导因子-1 α (Hif-1 α)、血管内皮生长因子（VEGF）表达水平以及ANXA3调控的Hif-1 α反活化活性。结果显示，ANXA3在小胶质细胞中表达，并且在BCP期间其表达显著增加。ANXA3基因敲除可以逆转疼痛行为，但不能阻止疼痛的发展。此外，在体外和体内，ANXA3敲低显著降低了Hif-1 α和VEGF的表达水平。而过表达Hif-1 α或VEGF可阻断AAV-shANXA3对BCP的作用。在N9细胞中敲低ANXA3可显著降低共培养神经元中p-PKC蛋白的表达。最后，过表达ANXA3可显著提高293T细胞中Hif-1 α的活化活性。因此，小胶质细胞ANXA3下调可通过抑制Hif-1 α /VEGF信号通路缓解BCP，提示ANXA3可能是治疗BCP的潜在靶点。
Bone cancer-induced pain (BCP) is a challenging clinical problem because traditional therapies are often only partially effective. Annexin A3 (ANXA3) is highly expressed in microglia in the spinal cord, and its expression is upregulated during BCP. However, the roles of microglial ANXA3 in the development and maintenance of BCP and the underlying molecular mechanisms remain unclear. This study was performed on male mice using a metastatic lung BCP model. Adeno-associated virus shANXA3 (AAV-shANXA3) was injected intrathecally 14 days before and 7 days after bone cancer induction, and relevant pain behaviors were assessed by measuring the paw withdrawal mechanical threshold, paw withdrawal thermal latency, and spontaneous hind limb lifting. ANXA3 protein expression was downregulated in microglial N9 cells by lentiviral transfection (LV-shANXA3). ANXA3, hypoxia-inducible factor-1 alpha (Hif-1 alpha), vascular endothelial growth factor (VEGF) expression levels, and Hif-1 alpha transactivation activity regulated by ANXA3 were measured. As a result, ANXA3 was expressed in microglia, and its expression significantly increased during BCP. ANXA3 knockdown reversed pain behaviors but did not prevent pain development. Moreover, ANXA3 knockdown significantly reduced Hif-1 alpha and VEGF expression levels in vitro and in vivo. And overexpression of Hif-1 alpha or VEGF blocked the effects of AAV-shANXA3 on BCP. ANXA3 knockdown in N9 cells significantly decreased the p-PKC protein expression in the cocultured neurons. Finally, ANXA3 overexpression significantly increased Hif-1 alpha transactivation activity in 293T cells. Therefore, microglial ANXA3 downregulation alleviates BCP by inhibiting the Hif-1 alpha/VEGF signaling pathway, which indicates that ANXA3 may be a potential target for the treatment of BCP.