METHODS: Expression of FOXD3, miR-335 and disheveled-associated activator of morphogenesis 1 (DAAM1) was detected in OC cells and tissues. The regulatory network of FOXD3/miR-335/DAAM1 was validated by dual-luciferase reporter and ChIP assays in vitro. After ectopic expression and depletion experiments in carboplatin/paclitaxel (CP)-resistant (A2780CP) or sensitive (A2780S) OC cells, cell viability, colony formation and apoptosis were tested by CCK-8 assay, colony formation assay and flow cytometry respectively. Effects of FOXD3 on the chemoresistance of OC cells in vivo were evaluated in OC xenografts in nude mice.
RESULTS: Overexpression of FOXD3 impaired the proliferation and chemoresistance of OC cells, which was related to the promotion of the miR-335 expression. Functionally, DAAM1 was a putative target of miR-335. Silencing of DAAM1 was responsible for the inhibition of myosin II activation, consequently leading to suppressed OC cell proliferation and chemoresistance. In vivo results further showed that FOXD3 weakened the chemoresistance of OC cells to CP.
CONCLUSIONS: Taken together, we unveil a novel FOXD3/miR-335/DAAM1/myosin II axis that regulates the chemoresistance of OC both in vitro and in vivo.
方法:在OC细胞和组织中检测FOXD3、miR-335和结构异常相关激活剂1(DAAM1)的表达。FOXD3/miR-335/DAAM1的调控网络通过体外双荧光素酶报告基因和ChIP测定进行验证。在卡铂/紫杉醇(CP)耐药(A2780CP)或敏感(A2780S)OC细胞中进行异位表达和耗竭实验后,细胞活力,CCK-8法检测集落形成和细胞凋亡,分别进行集落形成实验和流式细胞术。在裸鼠OC异种移植物中评估FOXD3对OC细胞体内化学抗性的影响。
结果:FOXD3的过表达损害了OC细胞的增殖和化学抗性,这与miR-335表达的促进有关。功能上,DAAM1是miR-335的推定靶标。沉默DAAM1负责抑制肌球蛋白II的激活,因此导致抑制OC细胞增殖和化疗耐药。体内结果进一步显示FOXD3削弱了OC细胞对CP的化学抗性。
结论:综合来看,我们揭示了一种新的FOXD3/miR-335/DAAM1/肌球蛋白II轴,它在体外和体内调节OC的化学抗性.