携带TRAIL的骨髓间充质干细胞联合吉西他滨对人胰腺癌细胞Panc-1的杀伤作用  

Killing Effect of Mesenchymal Stem Cells Modified with TRAIL Gene Combined with Gemcitabine on Pancreatic Cancer Cells

作　　者：殷涛[1] 魏洪吉[1] 胡晓青[2] 田园[3] 

机构地区：[1]华中科技大学同济医学院附属协和医院胰腺外科中心,武汉430022 [2]华中科技大学同济医学院附属协和医院超声科,武汉430022 [3]华中科技大学同济医学院附属协和医院普外科实验室,武汉430022

出　　处：《华中科技大学学报（医学版）》2012年第2期127-131,共5页Acta Medicinae Universitatis Scientiae et Technologiae Huazhong

基　　金：国家自然科学基金资助项目(No.30801100);高等学校博士学科点专项科研基金资助课题(No.200804871112);华中科技大学同济医学院附属协和医院院基金资助项目

摘　　要：目的研究TRAIL基因修饰的骨髓间充质干细胞(MSC)联合吉西他滨对人胰腺癌细胞的杀伤作用。方法构建重组腺病毒Ad-CMV-TRAIL,体外转染人骨髓间充质干细胞。通过观察绿色荧光的表达强度,确定最佳转染条件。MTT法检测Ad-CMV-TRAIL转染对MSC细胞生长状态的影响。Transwell小室共培养携带TRAIL的MSC和胰腺癌细胞Panc-1,联合应用吉西他滨,AnnexinⅤ-FITC双标法检测凋亡的发生。Western blot法检测Caspase凋亡相关蛋白的表达变化。结果 Ad-CMV-TRAIL以MOI=10体外感染MSC 24h,显微镜下观察80%以上的MSC表达绿色荧光,转染组细胞高表达TRAIL。Ad-CMV-TRAIL转染之后的MSC仍然可以增殖,活力无明显改变。携带TRAIL的MSC与Panc-1细胞共培养之后,可诱导部分Panc-1细胞发生凋亡,凋亡率达(13.38±3.42)%。而携带TRAIL的MSC与Panc-1细胞共培养之后可以增强吉西他滨的杀伤效果,与单独吉西他滨处理组相比差异显著[(52.10±3.41)%vs.(29.68±1.71)%,P=0.002]。Western blot检测结果显示MSC-TRAIL和吉西他滨共同处理可以诱导凋亡相关的Caspase-8和Caspase-3蛋白活化。结论 MSC可能作为TRAIL的肿瘤靶向载体在胰腺癌的治疗中发挥作用。Objective To study the killing effect of mesenchymal stem cells（MSCs）modified by TRAIL gene combined with gemcitabine on the pancreatic cancer cells. Methods The adenovirus expressing TRAIL was constructed and infected into the human MSCs in vitro. The intensity of GFP was observed to determine the optimal transfection conditions. MTT assay was used to investigate the influence of Ad-CMV-TRAIL on the growth of MSC. The MSC-TRAIL and Panc 1 cells were co-cultured in Transwell inserts, and gemcitabine was used together to treat the Panc-1 cells. The apoptosis was detected by using AnnexinV-FITC methods. The apoptosis related proteins Caspase 3 and Caspase 8 were detected by using Western blot. Results After being transfected with Ad-CMV-TRAIL（MOI= 10） for 24 h, over 80 % MSCs expressed GFP, and the TRAIL was detec- ted in the transected group. The MSCs could still proliferate after Ad-CMV-TRAIL transfection and there were no significant changes about viability. MSCs TRAIL could induce apoptosis in some Panc-1 cellsE（13.38± 3.42）% ]. Gemcitabine could sig nificantly enhance the killing effects with MSC-TRAIL, as compared with solely gemcitabine-treated group[（52.10 ± 3.41 ） vs. （29.68 ± 1.71） %, P = 0. 002]. The apoptosis related proteins Caspase 8 and Caspase 3 were activated in the combined treatment group. Conclusion MSCs can act as an ideal vector for targeted gene therapy of pancreatic cancer.

关 键 词：胰腺癌 骨髓间充质干细胞 肿瘤坏死因子相关凋亡诱导配体 吉西他滨 

分 类 号：R735.9[医药卫生—肿瘤]