消岩汤对A549/DDP细胞自噬及耐药蛋白的影响研究

Study on the autophagy and resistance protein affected by Xiaoyan decoction in A549/DDP cells

关键词:  消岩汤  细胞自噬  耐药  分子通路

Key words:  Xiaoyan decoction  autophagy  resistance  molecular pathway

摘要:

[目的] 通过消岩汤对耐顺铂人肺腺癌细胞A549/DDP多药耐药及自噬蛋白的影响,探究消岩汤对耐顺铂人肺腺癌A549/DDP的耐药逆转作用及探讨相关分子通路。[方法] 建立稳定细胞耐药模型,连续灌胃不同浓度消岩汤10 d后,通过血清药理学的实验方法,提取药物血清,用含药血清作用于肺腺癌细胞耐药细胞A549/DDP,通过 实时荧光定量逆转录聚合酶链反应(QPT-PCR)和蛋白免疫印迹法(Werstern Blot)检测A549/DDP耐药、自噬相关基因的表达及相关通路蛋白的变化。[结果] 高剂量消岩汤,低剂量消岩汤和生理盐水作用于肺腺癌耐药细胞A549/DDP,检测A549/DDP细胞中耐药相关蛋白P-糖蛋白(P-gp)、肺抗药性相关蛋白(LRP)及自噬相关蛋白Beclin1,HMGB1 mRNA和蛋白表达变化,发现高剂量组消岩汤处理细胞株后P-gp、LRP及自噬相关蛋白Beclin1,HMGB1 mRNA和蛋白均发生降低(P<0.05),进一步发现消岩汤可影响AKT1-mTOR相关基因,消岩汤明显降低AKT1,mTOR蛋白的表达。[结论] 消岩汤通过影响AKT1-mTOR分子通路进而影响耐药相关蛋白P-gp、LRP及自噬相关蛋白Beclin1,HMGB1基因的变化。

Abstract:

[Objective] Through the cisplatin resistance of the human lung adenocarcinoma cells A549/DDP multi-resistant and influence of autophagy protein by Xiaoyan decoction, finding out the influence of elimination of cisplatin resistance of lung adenocarcinoma A549/DDP resistance reversal agents and explore relevant molecular pathways. [Methods] Establish a stable cell resistance model, continuous lavaging Xiaoyan decoction with different concentrations in 10 days, by serum pharmacology experiment method with drug-containing serum on lung adenocarcinoma cells resistant cells A549/DDP, through the QRT-PCR and Werstern Blot western Blot method to detect A549/DDP resistant, autophagy protein expression of related genes and related pathways. [Results] Resistance effect on lung adenocarcinoma cells A549/DDP with high dose Xiaoyan decoction, low dose Xiaoyan Decoction and normal saline, elimination saline resistance effect on lung adenocarcinoma cells A549/DDP, detecting drug-resistant related proteins in A549/DDP cell P-gp, LRP and Beclin1 protein involved in autophagy, HMGB1mRNA and protein expression changes,through the detection of A549/DDP cells drug-resistant related proteins P-gp, LRP and Beclin1 protein involved in autophagy, HMGB1mRNA and protein expression changes, high dose group of rock soup elimination processing cell lines after P-gp, LRP and Beclin1 protein involved in autophagy, both the HMGB1 mRNA and protein were lower (P<0.05). Further found that Xiaoyan decoction can affect the AKT1-elimination mTOR related genes and significantly reduce the AKT1, mTOR protein expression. [Conclusion] We hypothesized that Xiaoyan decoction influence elimination drug-resistant related proteins P-gp, LRP and Beclin1 protein involved in autophagy, HMGB1 genetic changes by AKT1- mTOR molecular pathways.