摘要
[目的]分析微小RNA (microRNA,miR)-142-5p靶向DEAD box p68 RNA解旋酶(DEAD-box RNA helicase 5,DDX5)调控卵巢癌顺铂(cisplatin,DDP)耐药机制。[方法 ]收集2019年4月至2020年11月45例卵巢癌手术切除的癌组织及癌旁组织,采用RT-qPCR测定miR-142-5p表达,Western blot法测定DDX5表达。实验分4组:SKOV3组、SKOV3/DDP组、阴性对照组(感染阴性对照慢病毒LV-miR-142-5p-NC的SKOV3/DDP细胞)、病毒感染组(感染沉默miR-142-5p的慢病毒LV-miR-142-5p-IN的SKOV3/DDP细胞),对比各组miR-142-5p、DDX5表达、细胞生长抑制率、细胞凋亡率和细胞OD值。双荧光素酶报告实验验证miR-142-5p和DDX5的靶向关系。[结果]卵巢癌组织miR-142-5p mRNA表达、DDX5蛋白表达比癌旁组织高(P<0.05);SKOV3/DDP组、阴性对照组miR-142-5p mRNA表达和DDX5蛋白表达比SKOV3组高(P<0.05);病毒感染组miR-142-5p mRNA表达、DDX5蛋白表达相比SKOV3/DDP组、阴性对照组低(P<0.05)。不同浓度顺铂(1.25、2.5、5、10、20μg/mL)作用后,SKOV3/DDP组、阴性对照组的细胞生长抑制率比SKOV3组低(P<0.05);病毒感染组细胞生长抑制率比SKOV3/DDP组及阴性对照组高(P<0.05)。DDP培养48 h后,SKOV3/DDP组、阴性对照组细胞凋亡率比SKOV3组低,OD值比SKOV3组高(P<0.05);相比SKOV3/DDP组和阴性对照组,病毒感染组凋亡率高,OD值低(P<0.05)。DDX5 WT+miR-142-5p mimic组荧光素酶活性比DDX5 WT组高(P<0.05)。[结论] mi R-142-5p下调可能通过抑制DDX5表达促进细胞凋亡、抑制细胞增殖,从而逆转卵巢癌细胞对顺铂的耐药性。
[Objective] To analyze the mechanism of micro RNA(mi R)-142-5 p targeting DEAD box P68 RNA helicase(DDX5) in regulating cisplatin resistance in ovarian cancer. [Methods] From April 2019 to November2020, the cancer tissues and adjacent tissues of 45 cases of ovarian cancer were collected. The expression of mi R-142-5 p was measured by RT-q PCR and the expression of DDX5 was measured by Western blot. Human ovarian carcinoma cell line and cisplatin resistant cell line SKOV3/DDP were used for in vitro experiments(SKOV3 group and SKOV3/DDP group), the SKOV3/DDP cells were infected with negative control lentivirus LV-mi R-142-5 p-NC(negative control group) or with lentivirus LV-mi R-142-5 p-in silencing mi R-142-5 p(mi R-142-5 p positive group). The expression of mi R-142-5 p and DDX5, cell growth inhibition rate, apoptosis rate and cell opitical density(OD) were compared among groups. Double luciferase report assay was used to verify the targeting relationship between mi R-142-5 p and DDX5. [Results] The expressions of mi R-142-5 p m RNA and DDX5 protein in ovarian cancer were higher than those in adjacent tissues(P<0.05). The expression of mi R-142-5 p m RNA, DDX5 protein in SKOV3/DDP group and negative control group were higher than those in SKOV3 group(P<0.05). The expression of mi R-142-5 p m RNA, DDX5 protein in mi R-142-5 p positive group were lower than those in SKOV3/DDP group and negative control group(P<0.05). After treated with DDP(1.25,2.5, 5, 10 and 20 μg/m L respectively), the cell growth inhibition rate in SKOV3/DDP group and negative control group was lower than that of SKOV3 group(P<0.05). The cell growth inhibition rate of mi R-142-5 p positive group was higher than that of SKOV3/DDP group and negative control group(P<0.05). The apoptosis rate of SKOV3/DDP group and negative control group at 48 h was lower than that of SKOV3 group, and the OD value was higher than that of SKOV3 group(P<0.05). The apoptosis rate in mi R-142-5 p positive group was higher than that in SKOV3/DDP group and negative control group at 48 h, and the OD value was lower than that in SKOV3/DDP group and negative control group(P<0.05). The luciferase activity of DDX5 WT + mi R-142-5 p mimic group was higher than that of DDX5 WT group(P<0.05). [Conclusion] The down-regulation of mi R-142-5 p can promote apoptosis and inhibit cell proliferation by inhibiting the expression of DDX5, to reverse the resistance of ovarian cancer cells to cisplatin.
作者
刘渤娜
杜成
郑振东
LIU Bo-na;DU Cheng;ZHENG Zhen-dong(General Hospital of the Northern Theater of the Chinese People’s Liberation Army,Shenyang 110013,China)
出处
《肿瘤学杂志》
CAS
2022年第7期539-544,共6页
Journal of Chinese Oncology
基金
辽宁省科学技术计划项目书(2019-ZD-1050)。
