低频超声联合微泡造影剂通过激活内质网应激逆转前列腺癌紫杉醇耐药  

Low-frequency ultrasound with ultrasound contrast agent reverses paclitaxel resistance in prostate cancer cells by activating endoplasmic reticulum stress

作　　者：武雨琦 王祥卫 WU Yuqi;WANG Xiangwei(Department of Urology, Second Affiliated Hospital, Army Medical University （Third Military Medical University）, Chongqing, 400037, Chin)

机构地区：[1]陆军军医大学(第三军医大学)第二附属医院泌尿外科,重庆400037

出　　处：《第三军医大学学报》2018年第8期673-678,共6页Journal of Third Military Medical University

基　　金：国家自然科学基金面上项目(81172443)~~

摘　　要：目的探讨低频超声联合微泡造影剂辅助紫杉醇逆转前列腺癌耐药的作用和机制。方法筛选超声辐照紫杉醇耐药前列腺癌细胞株PC-3R的非毒性照射时间;将PC-3R细胞随机分为对照组、紫杉醇组(PTX组)、低频超声组(LFUS组)、紫杉醇和低频超声组(PTX+LFUS组),以及紫杉醇和低频超声联合微泡剂组(PTX+LFUS+UCA组),分别进行干预。Annexin V-FITC/PI双染法检测各组细胞凋亡;q-PCR法检测各组细胞Bcl-2和多耐药基因表达变化;Western blot检测各组细胞GRP78和c-jun蛋白表达。GRP78 siRNA转染PC-3R细胞,微泡和低频超声联合处理后,Western blot检测GRP78蛋白表达,Annexin V-FITC/PI双染法检测凋亡情况。结果低频超声照射频率1 MHz,强度1.2 W/cm2,非毒性照射时间为10 s。与单纯应用紫杉醇相比低频超声辅助紫杉醇处理前列腺耐药细胞可显著提高细胞凋亡率(P<0.01),联合微泡剂照射后,凋亡率进一步提高(P<0.01),并下调Bcl-2和多耐药基因的表达。PTX+LFUS+UCA组内质网应激相关蛋白GRP78的表达较PTX组和PTX+LFUS组升高,c-jun的表达下降(P<0.01)。应用siRNA干扰GRP78基因表达后,细胞凋亡率下降(P<0.01)。结论低频超声联合微泡造影剂通过激活内质网应激,抑制其下游JNK/c-jun通路下调多耐药基因及凋亡抑制基因的表达,提高化疗药物诱导细胞凋亡的作用,逆转肿瘤细胞的耐药性。Objective To investigate the effect of low-frequency ultrasound（ LFUS） combined with ultrasound contrast agent（ UCA） on reversing paclitaxel（ PTX） resistance in prostate cancer cells and explore the mechanism. Methods The non-toxic ultrasound irradiation time was determined in PTX-resistant prostate cancer PC-3（ PC-3 R） cells. The cells were then divided into control group,paclitaxel（ PTX） group,LFUS group,PTX ＋ LFUS group,and PTX ＋ LFUS ＋ UCA group for corresponding treatments. After the treatments,the apoptotic rates of PC-3 R cells were determined using flow cytometry,and the protein levels of GRP78 and c-jun and the mRNA levels of Bcl-2 and multidrug resistance-associated proteins（ MRPs） were detected using Western blotting and q-PCR,respectively. A lentiviral vector expressing siRNA targeting human GRP78 was transfected in the cells prior to exposure to LFUS with UCA,and the protein expression level of GRP78 was determined by Western blotting and cell apoptosis was analyzed using flow cytometry. Results Under the optimal settings（ 1 MHz,1. 2 W/cm2,50% duty cycle,and 10 s）,LFUS exposure combined with PTX treatment significantly increased the apoptotic rate of prostate cancer cells compared with PTX treatment alone（ P〈0. 01）; in the presence of UCA,LFUS plus PTX treatment further increased the cell apoptosis rate（ P〈0. 01） and down-regulated the expressions of Bcl-2 and MRPs. LFUS with UCA plus PTX treatment resulted in significantly increased expression of endoplasmic reticulum stress-related protein GRP78 and lowered c-jun expression as compared LFUS plus PTX treatment without UCA. Transfection of the cells with siRNA targeting GRP78 caused a significant decrease of the cell apoptosis rate following LFUS with UCA plus PTX treatment（ P〈0. 01）. Conclusion LFUS combined with UCA can inhibit the expression of MRPs and the apoptosissuppressing gene by down-regulating the JNK/c-jun pathway in the endoplasmic reticulum to enhance the effect of PTX-induced apoptosi

关 键 词：低频超声 微泡剂 前列腺癌 耐药 

分 类 号：R445.1[医药卫生—影像医学与核医学] R737.25[医药卫生—诊断学]