Smad3 siRNA对树突状细胞疫苗抗乳腺癌作用的影响  

Effect of anti-breast cancer of vaccine of dentritic cells by Smad3 siRNA

作　　者：马楠[1] 祁馨卉[1] 崔慧霞[1] 李妍[1] 刘云鹏[1] 姜又红[1] 

机构地区：[1]中国医科大学附属第一医院肿瘤研究所二室,辽宁沈阳110001

出　　处：《现代肿瘤医学》2013年第6期1209-1213,共5页Journal of Modern Oncology

基　　金：辽宁省科学技术计划项目(编号:2011415052-3)

摘　　要：目的:研究在通过siRNA(small interference,RNA)干扰技术沉默外周单核细胞来源的树突细胞(dentritic cells,DC)的Smad3,阻断TGF-β1/Smad3传导通路,并在外源性TGF-β1(transforming growth factorbeta-1,TGF-β1)的作用下对树突状细胞疫苗的影响。方法:针对设计Smad3特异性siRNA利用RNAIMAX转入DC细胞并用Westen-blot法检测Smad3的沉默效果。应用重组人粒细胞-单核细胞集落刺激因子(hmGM-CSF)和重组人白介素-4(hmIL-4),刺激DC成熟。利用反复冻融的方法提取乳腺癌细胞MCF-7的全抗原。实验分组:Control-DC-冻融抗原(Ag)组、TGF-β1-Control-DC-冻融抗原(Ag)组、TGF-β1-Negtive siRNA-Smad3-DC-冻融抗原(Ag)组和TGF-β1-siRNA-Smad3-DC-冻融抗原(Ag)组。流式检测各组DC成熟表型的变化,ELISA检测各组DC上清IL-12的水平,CCK-8法检测DC诱导细胞毒性T淋巴细胞(CTL)杀瘤活性。结果:TGF-β1-siRNA-Smad3-DC-冻融抗原(Ag)组表面成熟标志CD80、CD83、CD86、HLA-DR(P<0.05)及IL-12分泌水平(P<0.05)明显高于TGF-β1-Control-DC-冻融抗原(Ag)组和TGF-β1-Negtive siRNA-Smad3-冻融抗原(Ag)-DC组。且TGF-βI-siRNA-Smad3-DC-冻融抗原(Ag)组诱导细胞毒性T淋巴细胞(CTL)杀瘤活性较TGF-β1-Control-DC-冻融抗原(Ag)组、TGF-β1-Negtive siRNA-Smad3-DC-冻融抗原(Ag)组明显增强。结论:通过沉默DC的Smad3的方法,阻断TGF-β1/Smad3传导通路,能逆转TGF-β1对DC分化发育和递呈乳腺癌相关抗原的作用。Objective：To investigate the effect of vaccine of dentritic by small interference RNA targeting Smad3 on the maturity and function of dentrictic cell（DC） to block the signal pathway of TGF-β1/Smad3 under the effect of exogenous TGF-β1（transforming growth factor beta-1,TGF-β1）.Methods：The siRNA sequences targeting Smad3 gene were designed and transfected to DC by RNAIMAX,further to detect the transfection effect by Westen-blot.Using hmGM-CSF and hmIL-4 to stumulate the maturation of DC.Using repeated freezing and thawing method to extract the breast cancer cell MCF-7 antigen.The experimental classification was performed as follow：group of Control-DC-Ag,group of TGF-β1-Control-DC-Ag,group of TGF-β1-Negtive siRNA Smad3-DC-Ag and group of TGF-βI-siRNA-Smad3-DC-Ag.The maturity of DC was investigated by FCM.Detect the level of IL-12 by ELISA.Detection of cytotoxic Tlymphcyto（CTL） by CCK-8 assays.Results：The expression of CD80,CD83,CD86 and CD11c and the level of IL-12 in group of TGF-β1-siRNA-Smad3-DC-Ag was higher than TGF-β1-Control-DC-Ag group and TGF-β1-Negtive siRNA-Smad3-DC-Ag group（P0.05）.Compared with the TGF-β1-Control-DC-Ag group and TGF-β1-Negtive siRNA-Smad3-DC-Ag group,the proliferation of CTL induced by TGF-β1-siRNA-Smad3-DC-Ag was different statistically.Conclusion： The DC defect induced by TGF-β1 which is immature and poor ability of presenting breast cancer associated antigen will be reversed by blocking the TGF-β1/Smad3 signal pathing in DC.

关 键 词：树突状细胞 TGF-Β1 SMAD3 肿瘤疫苗 

分 类 号：R730.51[医药卫生—肿瘤]