miRNA-155/PU.1信号通路阻滞对大鼠骨髓来源树突状细胞成熟及移植免疫的影响  

Influences of miRNA-155/PU.1 Signaling Pathway Blockade on Rat Bone Marrow-Derived Dendritic Cell Maturation and Transplantation Immunity

作　　者：徐兴伟[1] 丁博文 朱传荣[3] 郑鹏[1] 冯涛[1] 嵇武[1] 

机构地区：[1]南京大学医学院临床学院,南京军区南京总医院普通外科研究所,江苏南京210002 [2]徐州市中医院普外科,江苏徐州221000 [3]淮安市第一人民医院普外科,江苏淮安223001

出　　处：《中国普外基础与临床杂志》2014年第2期168-172,共5页Chinese Journal of Bases and Clinics In General Surgery

基　　金：江苏省自然科学基金项目(编号:BK2008237);南京市科技发展计划项目(编号:201104027)~~

摘　　要：目的探讨miRNA-155/PU.1信号通路阻滞对骨髓来源树突状细胞(DCs)成熟及其对应用于大鼠小肠移植中的免疫功能的影响。方法利用贴壁法培养诱导DCs,针对miRNA-155/PU.1信号通路中关键转录因子PU.1基因设计并合成3对PU.1 siRNA(PU.1沉默组、阴性对照组及对照组),用脂质体法转染细胞,并筛选一对高沉默效率PU.1 siRNA。流式细胞术分析PU.1沉默组、阴性对照组及对照组细胞表面CD80、CD86及主要组织相容性复合体(MHC)-Ⅱ的表达;ELISA法检测上清液中IL-10及IL-12p70的水平;混合淋巴细胞反应检测对同种异体T淋巴细胞的增殖作用;在大鼠小肠移植前7 d经受体尾静脉等量注射3组细胞,移植后观察各组受体存活情况及移植物病理情况。结果①DCs表面分子CD80、CD86和MHC-Ⅱ表达率在PU.1沉默组分别为(27.0±5.6)%、(23.6±4.8)%及(36.8±6.8)%,阴性对照组分别为(74.0±9.4)%、(76.5±8.7)%及(87.8±11.3)%,PU.1沉默组均分别明显低于阴性对照组(P<0.05)。②PU.1沉默组IL-10水平较阴性对照组明显升高(P<0.05),IL-12p70则明显降低(P<0.05)。③PU.1沉默组DCs刺激同种异体T淋巴细胞增殖也较阴性对照组明显降低(P<0.05)。④将PU.1沉默组DCs术前注入受体行大鼠小肠移植后,受体平均存活时间为(14.3±3.3)d,明显长于阴性对照组的(7.8±1.5)d(P<0.05)和对照组的(8.0±2.5)d(P<0.05),且术后5 d时移植物病理显示移植肠组织轻度淋巴细胞浸润和绒毛水肿。结论体内外实验表明,miRNA-155/PU.1信号通路阻滞的DCs成熟度明显受到抑制,并能稳定发挥诱导受体特异性免疫耐受的作用。Objective To explore the influence of miRNA-155/PU. 1 signaling pathway blockade on bone marrow- derived dendritic cells （DCs） maturation and immune function of rat small intestinal transplantation. Methods The DCs were induced by adherent culture. The critical transcription factor gene PU. 1 was designed and PU. 1 siRNA was synthe- sized. The DCs were transfected by liposome transfection and a pair of PU.1 siRNA was screened according to the high silencing efficiency. The expressions of DCs surface markers CD80, CD86, and MHC- l] among three groups （PU.1 silent group, negative control group, and control group） were analyzed by flow cytometry. The IL-IO andIL-12p70 secretion levels in the supematant were tested by ELISA method. The allogeneic T lymphocyte proliferation was tested by mixed lymphocyte reaction. The transfected cells were intravenously injected into the recipient rat on day 7 before intestinal transplantation. The survival conditions as well as pathological changes were observed in each group recipients. Results OThe surface molecules CD80, CD86, and MHC-I] in the PU.1 silent group were （27. 0＋5.6）%, （23.6 ＋4. 8） %, and （36.8 q-6.8） %, respectively; versus （74. 0 ＋ 9.4） %, （76. 5 ＋ 8.7） %, and （87.8 ＋ 1 I. 3） % in the negative control group, respectively, which were significantly lower in former and showing an in creased trend （P〈0. 05）. （~） Compared with the negative control group, IL-10 secretion level was significantly increased （P〈0.05）, IL-12p70 secretion level significantly decreased （P 〈 0.05） in the PU.1 silent group. （~） The proliferation ofT lymphocytes in the PU.1 silent group was significantly lower than that in the negative control group （P〈0. 05）. （~）When the transfected DCs were injected into the intestinal transplantation rats on day 7 before operation, the survival time was （14. 3 ＋3.3） d, （7. 8＋ 1.5） d, and （8.0＋2.5） d in the PU.1 silent group, negative control group, and control group,

关 键 词：树突状细胞 miRNA-155 PU 1 免疫耐受 小肠移植 

分 类 号：R392[医药卫生—免疫学]