二十二碳六烯酸逆转食管癌Eca109/DDP细胞对顺铂耐药的机制

Mechanisms underlying reversal effect of docosahexaenoic acid on cisplatin resistance in human esophageal carcinoma cell line Eca109/DDP

目的:构建食管癌耐药细胞株,探讨二十二碳六烯酸(docosahexaenoic acid,DHA)逆转人食管癌Eca109/顺铂(cisplatin,DDP)细胞对DDP的耐药作用及其机制.方法:采用递增顺铂药物质量浓度持续作用诱导法建立耐顺铂细胞株;MTT法检测DHA对Eca109/DDP细胞增殖影响,计算耐药逆转作用,Western blot检测Eca109/DDP细胞膜P-糖蛋白(P-glycoprotein,P-gp)表达水平.结果:历经6 mo的诱导,Eca109细胞能在含1 g/mL DDP的培养基中稳定生长,耐药指数(resistant index,RI)为15.9;DHA浓度在1.560 g/mL以下,对Eca109/DDP细胞的生长无显著抑制作用(P>0.05),但与DDP联合应用能增加DDP对Eca109/DDP细胞的毒性且呈浓度依赖性(P<0.05);1 g/mL DDP对Eca109/DDP细胞的半数抑制浓度(IC50)为3.50g/mL±1.04 g/mL,当分别与0.195、0.390、0.780、1.560 g/mL的DHA联合应用时,对Eca109/DDP细胞的半数抑制浓度(IC50)从1.99g/mL±0.11 g/mL降低到0.83 g/mL±0.22g/mL(P<0.05),提高逆转指数,并能显著抑制Eca109/DDP细胞P-gp表达水平,P-gp相对表达量从0.99±0.12降至0.52±0.08(P<0.05).结论:DHA与DDP联合应用通过下调Eca109/DDP细胞P-gp的表达水平逆转对DDP的耐药,促进DDP对耐药细胞的杀伤作用.

AIM： To establish a cisplatin （DDP）-resistant human esophageal carcinoma cell line （Eca109/ DDP） and to explore the mechanisms respon- sible for the reversal effect of docosahexaenoicacid （DHA） on DDP resistance.METHODS： A DDP-resistant esophageal cancer cell line （Eca109/DDP） was established by exposure of Eca109 cells to gradually increasing doses of DDP. The effects of DHA on cell proliferation and DDP resistance in Eca109/DDP cells were determined by MTT assay. The expression level of P-glycoprotein （P-gp） was detected by Western blot. RESULTS： After 6 mo of induction, Ecal09 cells in culture medium containing 1 μg/mL DDP grew stably and the resistant index was 15.9. DHA at concentrations 〈 1.560 μg/mL had no obvious inhibition on Eca109/DDP cells （P 〉 0.05）; however, DHA could enhance the cyto- toxicity of DDP to Eca109/DDP cells in a dose- dependent manner （P 〈 0.05）. The half inhibitory concentration （IC50） for DDP in Eca109/DDP cells was 3.50 μg/mL± 1.04 μg/mL. When com- bined with 0.195 μg/mL, 0.390 μg/mL, 0.780 μg/mL, or 1.560 μg/mL of DHA, the IC50 for DDP decreased from 1.99 μg/mL ± 0.11 μg/mL to 0.83 μg/mL ±0.22 μg/mL （P 〈 0.05）. DHA significantly increased the reversal index and down-regulate the expression of P-gp in Ecal09/ DDP cells （0.99 ±0.12 vs 0.52 ± 0.08, P 〈 0.05）. CONCLUSION： DHA possesses a reversal effect on DDP resistance in Ecal09/DDP cells by down- regulating the expression of P-gp and enhancing the killing effect of DDP on drug-resistant cells.