缺氧条件下人卵巢癌细胞对顺铂耐药与TLR9的关系研究

发布时间：2018-06-14 16:21

  本文选题：人卵巢细胞 + 缺氧　； 参考：《中国药房》2017年13期

【摘要】：目的:研究缺氧条件下人卵巢癌细胞(SKOV3)对顺铂耐药与Toll样受体9(TLR9)的关系。方法:采用免疫荧光法检测破膜和未破膜SKOV3中TLR9的表达。取SKOV3,加入TLR9特异性激动剂Cp G-ODN 2006(A组)及其同型对照Cp G-ODN 2006control(B组)作用6、12、24 h后,加入顺铂,以CCK-8法检测细胞抑制率。取SKOV3或经0(未加)、1、10、102、103μmol/L TLR9特异性拮抗剂氯喹预处理3 h后的SKOV3,加入SKOV3常氧(21%O2)、缺氧(1%O2)孵育6、12、24 h的细胞上清作用24 h后,加入顺铂,检测细胞增殖抑制率,计算预处理细胞缺氧时药物抑制率的降低倍数(HICR);并采用Western blot法检测常氧24 h细胞上清(C组)、缺氧24 h细胞上清(D组)、缺氧24 h细胞上清+10μmol/L氯喹(E组)条件下SKOV3中多药耐药相关蛋白(MRP)的表达。结果:TLR9在SKOV3的细胞膜和细胞质中均有表达。与A组比较,B组细胞增殖抑制率降低(P0.01)。与常氧细胞上清比较,缺氧细胞上清作用后细胞增殖抑制率降低(P0.05)。与未加氯喹比较,加入10、102μmol/L氯喹能明显降低HICR(P0.01),其中缺氧24 h细胞上清+10μmol/L氯喹降低最明显。与C组比较,D组细胞中MRP表达增强(P0.01);与D组比较,E组细胞中MRP表达减弱(P0.01)。结论:TLR9受体在缺氧下可被激活,并可导致SKOV3对顺铂耐药,该作用可能与上调MRP表达有关。
[Abstract]:Objective: To study the relationship between cisplatin resistance and Toll like receptor 9 (TLR9) in human ovarian cancer cells (SKOV3) under hypoxic conditions. Methods: the expression of TLR9 in the rupture and unbroken membrane SKOV3 was detected by immunofluorescence. SKOV3, TLR9 specific agonist Cp G-ODN 2006 (A group) and the same type control Cp G-ODN (Group) were added. The inhibition rate of cisplatin was detected by CCK-8. SKOV3 or 0 (not added), chloroquine, a specific antagonist of 1,10102103 mu mol/L TLR9, was pretreated with chloroquine, SKOV3, SKOV3 normal oxygen (21%O2), and hypoxia (1%O2) incubated for 24 of the cell supernatant of 6,12,24 h, and cisplatin was added to detect cell proliferation inhibition rate and to calculate the drug pretreated cells when hypoxia. The inhibition rate was reduced (HICR), and Western blot method was used to detect the expression of multidrug resistance associated protein (C), hypoxia 24 h cell supernatant (group C), hypoxia 24 h cell supernatant (group D), and +10 micron mol/L chloroquine (E group) under the condition of hypoxia (E group). The inhibitory rate of cell proliferation decreased (P0.01). Compared with the supernatant of normoxic cells, the inhibition rate of cell proliferation decreased (P0.05). Compared with chloroquine, 10102 mol/L chloroquine could obviously decrease HICR (P0.01), and the decrease of +10 u mol/L chloroquine in 24 h cell supernatant was the most obvious. Compared with group C, MRP expression in the D group was enhanced. P0.01); compared with group D, the expression of MRP in E group was weakened (P0.01). Conclusion: TLR9 receptor can be activated under hypoxia and may lead to SKOV3 resistance to cisplatin, which may be related to the up regulation of MRP expression.
【作者单位】： 第三军医大学第三附属医院药剂科;
【基金】：国家自然科学基金面上项目(No.30973493)
【分类号】：R737.31
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本文编号：2018123