EphA2经P-糖蛋白调控鼻咽癌紫杉醇敏感性的实验研究

EphA2 promotes paclitaxel resistance via Pgp in nasopharyngeal carcinoma in vitro

摘要:

摘要：目的探讨EphA2经P糖蛋白(Pglycoprotein,Pgp)调控鼻咽癌紫杉醇耐药性的分子机制。方法使用慢病毒载体在鼻咽癌细胞中沉默或过表达EphA2，Western blotting检测Pgp蛋白的表达变化； siRNA干扰沉默Pgp表达，CCK8法检测Pgp蛋白在EphA2介导的鼻咽癌紫杉醇耐药中的作用；小分子化合物LY294002阻断PI3K/Akt信号通路，观察该通路在EphA2调控Pgp表达中的作用。结果Pgp在EphA2沉默组中表达下调，过表达组中上升；紫杉醇作用于空白对照组、阴性对照组和Pgp沉默组的细胞存活率分别是(68.9±4.0)%、(72.4±6.2)%和(52.1±3.7)%，Pgp沉默组对紫杉醇的耐药性较空白和阴性对照组降低(P<0.05)；在过表达EphA2的鼻咽癌细胞中下调Pgp表达，紫杉醇刺激后，Pgp沉默组的存活率(36.1±5.1)%显著低于空白对照组(57.9±6.2)%和阴性对照组(58.2±5.8)%(P<0.05)；在EphA2过表达鼻咽癌细胞中，给予抑制剂LY294002作用后，Akt总量无明显改变而磷酸化的Akt表达下降，且Pgp表达下降。结论EphA2通过Pgp依赖性途径促进鼻咽癌紫杉醇耐药，且Pgp蛋白的表达增高与EphA2活化PI3K/Akt有关。

Abstract:

Abstract:ObjectiveTo investigate the role of Pglycoprotein (Pgp) in EphA2mediated paclitaxel resistance in nasopharyngeal carcinoma (NPC) in vitro.MethodsEphA2 expression in NPC cell lines was modulated by lentiviral vector, Pgp expression was detected by western blotting accordingly. Pgp was downregulated in normal or EphA2 overexpressed NPC cell lines and its survival rate after paclitaxel treatment was determined by CCK8. PI3K/Akt specific inhibitor LY294002 was added to EphA2 upregulated NPC cells, the expressions of Akt, pAkt and Pgp were determined by western blotting after 48h paclitaxel treatment.ResultsThe expression of Pgp was downregulated in the normal EphA2 group and upregulated in the overexpressed EphA2 group. After paclitaxel treatment, the survival rate of NPC cells in Pgp knock down group (52.1±3.7%) was significantly lower than those of the negative control group (72.4±6.2%) and blank control group (68.9±4.0%) (both P<0.05).Similar results were also observed in overexpressed EphA2 group ［(36.1±5.1%) vs (58.2±5.8%);(36.1±5.1%)vs(57.9±6.2%) (both P<0.05)］. In paclitaxel treated EphA2 overexpressed NPC cell line, LY294002 inhibited the expressions of pAkt and Pgp but not that of total Akt.ConclusionEphA2 modulates paclitaxel resistance in NPC cells via regulation of Pgp, and PI3K/Akt path way may be involved in this process.