叶酸耦联白蛋白Cypate纳米粒的构建及体外膀胱癌靶向光热杀伤效应的研究  

Preparation of folic acid-bovine serum albumin-cypate nanaparticles for targeted photothermal therapy on bladder cancer cells in vitro

作　　者：朱伟[1] 李亚伟[1] 汪良[1] 蒋国松[1] 曾甫清[1] 

机构地区：[1]华中科技大学同济医学院附属协和医院泌尿外科,武汉430022

出　　处：《中华实验外科杂志》2016年第4期1026-1029,共4页Chinese Journal of Experimental Surgery

基　　金：国家自然科学基金（81272816）

摘　　要：目的 构建一种新型光热纳米材料——叶酸耦联白蛋白Cypate （FA-BSA-Cypate）纳米粒,探讨其介导的靶向光热疗法对人膀胱癌5637细胞的体外杀伤效应及抗肿瘤机制.方法 透析法制备FA-BSA-Cypate纳米粒并进行电镜表征观察;观察不同浓度FA-BSA-Cypate纳米粒溶液（1、10、100 μg/ml）在808 nm激光（1.5 W/cm^2）照射下温度随时间变化;噻唑蓝（MTT）法检测不同浓度FA-BSA-Cypate纳米粒（0、25、50、100、250、500 μg/ml）对人膀胱癌5637细胞的毒性大小以及不同浓度FA-BSA-Cypate纳米粒（0、25、50、100、250、500 μg/ml）在808 nm激光（1.5 W/cm^2,5 min）照射下对人膀胱癌5637细胞的靶向光热杀伤效应;将细胞分为4组：空白对照组、激光组、BSA-Cypate＋激光组、FA-BSA-Cypate＋激光组,采用Western blot法检测各组B细胞淋巴瘤/白血病-2（bcl-2）、B细胞淋巴瘤/白血病-2相关X蛋白（bax）、半胱氨酰天冬氨酸特异性蛋白酶（Caspase）-3、生存素（Survivin）表达水平.结果 成功制备了具有良好的光热特性的FA-BSA-Cypate纳米粒,粒径为（95.0±8.7） nm;808 nm激光（1.5 W/cm^2,5 min）照射下,纳米粒溶液最高温度可达59 ℃,且具有浓度依赖性;不同浓度的FA-BSA-Cypate纳米粒作用于5637细胞24h后细胞存活率在95％以上,无明显生物毒性;在联合808 nm激光照射下（1.5 W/cm^2,5 min）,含不同浓度FA-BSA-Cypate纳米粒（0、25、50、100、250、500μg/ml）的细胞存活率分别为（98.60±5.25）％、（87.85±4.37）％、（80.72±3.61）％、（69.43±4.28）％、（59.28 ±4.51）％和（51.15±3.79）％,细胞存活率随着纳米粒浓度增加而下降,差异有统计学意义（P＜0.05）.Western blot结果显示,FA-BSA-Cypate＋激光组及BSA-Cypate＋激光组bax、Caspase-3蛋白表达水平显著增高,而bcl-2、Survivin蛋白表达水平明显降低.结论 FA-BSA-Cypate纳米粒介导的靶向光热疗法对人膀胱癌5637细胞具有良好的杀伤Objective To investigate the targeted photothermal effects and the potential antitumor mechanism of folic acid-bovine serum albumin-cypate （FA-BSA-Cypate） nanaparticles on bladder cancer cells in vitro.Methods Dialysis method was developed to prepare the FA-BSA-Cypate nanaparticles,and transmission electron microscopy was used to characterize the nanoparticles.Temperature time curve was observed in different concentrations （1,10,and 100 μg/ml） of nanoparticles.Methyl thiazol tetrazolium （MTT） method was used to observe the different concentrations （0,25,50,100,250,500 μg/ml） of FA-BSA-Cypate nanoparticles for 5637 cell toxicity.The bladder cancer 5637 cells were incubated with the FA-BSA-Cypate nanaparticles of various concentrations （0,25,50,100,250,500 μg/ml） and exposured to 808 nm infrared laser irradiation of the same energy density and irradiation time （1.5 W/cm^2,5 min）.Cell survival rate of each group was detected by MTT assay.The cells were divided into four groups：control group,laser group,BSA-Cypate ＋ Laser group,FA-BSA-Cypate ＋ Laser group.The protein expression levels of B-cell lymphoma/leukemia-2 （bcl-2）,B cell lymphoma/leukemia-2 associated X protein （bax）,Cysteinyl aspartate-specific protease-3 （Caspase-3） and Survivin in each group were detected by Western blotting.Results FA-BSA-Cypate nanaparticles with a mean size of （95.0 ± 8.7） nm was successfully achieved.Upon 808 nm infared laser irradiation （1.5 W/cm^2,5 min）,the maximum temperature of FA-BSA-Cypate nanaparticles increased to 59 ℃ in a dose-dependent manner.The FA-BSA-Cypate nanaparticles of different concentrations had no obvious cytotoxicity to 5637 cells after 24 h incubation.When cells incubated with FA-BSA-Cypate nanaparticles for 12 h underwent laser irradiation （1.5 W/cm^2,5 min）,the cellular survival rate at different concentrations （0,25,50,100,250 and 500 μg/ml） of nanoparticles was （98.60 ±5.25）%,（87.85 ± 4.37） %,（80.72 ±3.61） %,（69.43 ±4.28） %

关 键 词：纳米技术 光热疗法 膀胱癌 靶向治疗 

分 类 号：R730.51[医药卫生—肿瘤]