中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
CHINESE JOURNAL OF EXPERIMENTAL SURGERY
2014年
4期
737-739
,共3页
姜雪明%赵娜%高昊鹏%郭昊%何向辉%章志翔
薑雪明%趙娜%高昊鵬%郭昊%何嚮輝%章誌翔
강설명%조나%고호붕%곽호%하향휘%장지상
白细胞介素-35%间充质干细胞%免疫耐受%调节性T细胞
白細胞介素-35%間充質榦細胞%免疫耐受%調節性T細胞
백세포개소-35%간충질간세포%면역내수%조절성T세포
Interleukin-35%Mesenchymal stem cells%Immune tolerance%Regulatory T cells
目的 探讨白细胞介素-35(IL-35)基因修饰小鼠骨髓间充质干细胞(BMSCs)的可行性,并通过体内和体外实验评价其诱导调节性T细胞(Tregs)增殖的能力.方法 体外培养的小鼠BMSCs经过鉴定后,采用脂质体转染IL-35表达质粒,采用酶联免疫吸附试验(ELISA)法检测培养上清中IL-35含量,评价IL-35-MSCs分泌IL-35的能力.采用流式细胞方法检测淋巴细胞亚群的变化评价IL-35-MSCs体外混合淋巴细胞培养和静脉注射后诱导Tregs增殖的能力.结果 利用全骨髓贴壁法可成功获得小鼠BMSCs,IL-35表达质粒转染后能表达具有生物活性的IL-35.接受静脉注射IL-35-MSCs组小鼠外周CD4+ CD25+ Tregs的比例为(5.24±0.61)％,明显高于注射转染对照质粒的MSCs组的(4.21±0.64)％和单纯MSCs组的(4.32±0.76)％,差异有统计学意义(P＜0.01),而后两者之间差异无统计学意义(P＞0.05).IL-35-MSCs静脉注射组小鼠脾脏淋巴细胞在单向淋巴细胞混合培养中CD4+T淋巴细胞和CD4+ CD25+ Tregs比例分别为(14.47±3.86)％、(4.35±0.57)％,而注射转染对照质粒的MSCs组分别为(20.89±2.32)％、(3.29±0.78)％,单纯MSCs组分别为(18.81±2.84)％、(3.43±1.02)％,差异有统计学意义(P＜0.05)而后两者之间差异无统计学意义(P＞0.05).结论 IL-35基因修饰的MSCs能促进CD4+ CD25+ Tregs增殖,其作用明显强于转染对照质粒和未转染的MSCs.IL-35和BMSCs发挥协同作用,放大各自的免疫调节和免疫抑制功能.
目的 探討白細胞介素-35(IL-35)基因脩飾小鼠骨髓間充質榦細胞(BMSCs)的可行性,併通過體內和體外實驗評價其誘導調節性T細胞(Tregs)增殖的能力.方法 體外培養的小鼠BMSCs經過鑒定後,採用脂質體轉染IL-35錶達質粒,採用酶聯免疫吸附試驗(ELISA)法檢測培養上清中IL-35含量,評價IL-35-MSCs分泌IL-35的能力.採用流式細胞方法檢測淋巴細胞亞群的變化評價IL-35-MSCs體外混閤淋巴細胞培養和靜脈註射後誘導Tregs增殖的能力.結果 利用全骨髓貼壁法可成功穫得小鼠BMSCs,IL-35錶達質粒轉染後能錶達具有生物活性的IL-35.接受靜脈註射IL-35-MSCs組小鼠外週CD4+ CD25+ Tregs的比例為(5.24±0.61)％,明顯高于註射轉染對照質粒的MSCs組的(4.21±0.64)％和單純MSCs組的(4.32±0.76)％,差異有統計學意義(P＜0.01),而後兩者之間差異無統計學意義(P＞0.05).IL-35-MSCs靜脈註射組小鼠脾髒淋巴細胞在單嚮淋巴細胞混閤培養中CD4+T淋巴細胞和CD4+ CD25+ Tregs比例分彆為(14.47±3.86)％、(4.35±0.57)％,而註射轉染對照質粒的MSCs組分彆為(20.89±2.32)％、(3.29±0.78)％,單純MSCs組分彆為(18.81±2.84)％、(3.43±1.02)％,差異有統計學意義(P＜0.05)而後兩者之間差異無統計學意義(P＞0.05).結論 IL-35基因脩飾的MSCs能促進CD4+ CD25+ Tregs增殖,其作用明顯彊于轉染對照質粒和未轉染的MSCs.IL-35和BMSCs髮揮協同作用,放大各自的免疫調節和免疫抑製功能.
목적 탐토백세포개소-35(IL-35)기인수식소서골수간충질간세포(BMSCs)적가행성,병통과체내화체외실험평개기유도조절성T세포(Tregs)증식적능력.방법 체외배양적소서BMSCs경과감정후,채용지질체전염IL-35표체질립,채용매련면역흡부시험(ELISA)법검측배양상청중IL-35함량,평개IL-35-MSCs분비IL-35적능력.채용류식세포방법검측림파세포아군적변화평개IL-35-MSCs체외혼합림파세포배양화정맥주사후유도Tregs증식적능력.결과 이용전골수첩벽법가성공획득소서BMSCs,IL-35표체질립전염후능표체구유생물활성적IL-35.접수정맥주사IL-35-MSCs조소서외주CD4+ CD25+ Tregs적비례위(5.24±0.61)％,명현고우주사전염대조질립적MSCs조적(4.21±0.64)％화단순MSCs조적(4.32±0.76)％,차이유통계학의의(P＜0.01),이후량자지간차이무통계학의의(P＞0.05).IL-35-MSCs정맥주사조소서비장림파세포재단향림파세포혼합배양중CD4+T림파세포화CD4+ CD25+ Tregs비례분별위(14.47±3.86)％、(4.35±0.57)％,이주사전염대조질립적MSCs조분별위(20.89±2.32)％、(3.29±0.78)％,단순MSCs조분별위(18.81±2.84)％、(3.43±1.02)％,차이유통계학의의(P＜0.05)이후량자지간차이무통계학의의(P＞0.05).결론 IL-35기인수식적MSCs능촉진CD4+ CD25+ Tregs증식,기작용명현강우전염대조질립화미전염적MSCs.IL-35화BMSCs발휘협동작용,방대각자적면역조절화면역억제공능.
Objective To investigate the feasibility of interleukin (IL)-35 gene transfection of mice bone marrow mesenchymal stem cells (BMSCs) and the effect of IL-35 gene modified MSCs (IL-35-MSCs) on the the proliferation of regulatory T cells (Tregs).Methods MSCs were cultured in vitro and transfected with IL-35 expressing plasmid by lipofectamine.The IL-35 expression of IL-35-MSCs was detected by enzyme linked immunosorbent assay (ELISA).The lymphocytes subsets after one-way mixed lymphocyte culture and in vivo transplantation were analyzed by flow cytometry to evaluate the effect of IL-35-MSCs on the proliferation of Tregs.Results The mice BMSCs can be isolated and cultured by the whole bone marrow adherent method.IL-35 is expressed in the MSCs supernatant and serum after IL-35 in vitro and in vivo transfection.Percentage of CD4 + CD25 + Tregs in mice treated with IL-35-MSCs increased significantly than control plasmid transfected MSCs and non-transfected MSCs groups(P ＜ 0.01).IL-35-MSCs up-regulated the CD4 + CD25 + Tregs level in the allogeneic mixed lymphocyte reaction system,lowered the percentage of CD4 + cells compared with the other two control groups (P ＜ 0.05).Conclusion IL-35-MSCs enhanced the proliferation of CD4 + CD25 + Tregs in vitro and in vivo compared to control MSCs.IL-35 and MSCs have synergic effects on the negative immune regulation and the induction of immune tolerance.
