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| 体外皮肤移植和TUNEL实验评价猕猴间充质干细胞输注对细胞免疫的影响 |
| 黄雅静,孙琪云,刘丽辉,范传波,胡恺勋,边莉,郭梅,艾辉胜,HUANGYa-jing,SUNQi-yun,LIULi-hui,FANChuan-bo,HUKai-xun,BIANLi,GUOMei,AIHui-sheng |
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| 摘要: |
| 目的:通过给猕猴单次无预处理输注猕猴白细胞抗原(RhLA)不相合供者的骨髓来源的间充质干细胞(MSCs),观察能否抑制供受者之间的免疫反应.方法:无菌抽取MSCs供者猕猴骨髓,分离单个核细胞,用2％FCS的DMEM/F12培养液培养骨髓MSCs,用流式细胞术检测其表型,不同培养条件诱导MSCs多向分化以鉴定;单次无预处理输注MSCs给RhLA不相合受者,分别于MSCs输注前及输注后20、40、60 d取受者外周血淋巴细胞分别与供者及非供者皮肤体外共培养72 h,观察培养皮肤病理损伤情况,同时TUNEL法检测培养皮肤石蜡切片中凋亡细胞的比率.结果:输注MSCs后20 d,受者淋巴细胞与供者皮肤呈现减低的免疫反应,皮肤病理损伤程度较输注前减轻,凋亡细胞比例由(23.0±4.9)％下降到(12.2±1.9)％;MSCs受者与非供者之间体外皮肤病理片中凋亡细胞比例也由输注前的(22.0±2.0)％下降至(11.8±1.4)％,具有显著差异(P<0.05);输注后40 d和60 d,受者淋巴细胞无论对供者还是非供者的皮肤均产生较强的免疫反应,凋亡细胞比例增至(22.0±3.2)％、(22.0±2.5)％(供者)及(23.2±2.4)％、(24.0±2.6)％(非供者).结论:猕猴输注同种异基因MSCs后,通过体外实验可以检测到供受者之间降低的免疫反应,但是这种免疫抑制作用是短期的、可逆的. |
| 关键词: 间充质干细胞、免疫,细胞、皮肤移植、TUNEL法 |
| DOI：10.3724/SP.J.1008.2006.00465 |
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| 基金项目:国家高技术研究发展计划/863计划（2002AA216081）. |
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| In vitro skin transplantation and TUNEL assay in evaluation of cellular immunity caused by infusion of rhesus mesenchymal stem cells |
| 黄雅静,孙琪云,刘丽辉,范传波,胡恺勋,边莉,郭梅,艾辉胜,HUANG Ya-jing,SUN Qi-yun,LIU Li-hui,FAN Chuan-bo,HU Kai-xun,BIAN Li,GUO Mei,AI Hui-sheng |
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| Abstract: |
| Objective：To transfuse rhesus bone marrow derived mesenchymal stem cells （MSCs） to rhesus leucocytes antigen （RhLA） mismatched recipients, in attempt to study the immunological reaction between recipients＇ T lymphocytes and the skin of donor or a third party. Methods： Bone marrow aspirates were collected from femurs of donor rhesus, and the mononuclear cells were isolated and resuspended in low glucose Dulbecco＇s Modified Eagles Medium （DMEM） containing 2% fetal bovine serum. Phenotype of MSCs was analyzed by flow cytometry and the differentiated cells were identified by relevant specific staining. Allogeneic MSCs were given to recipients （single infusion） without preconditioning. Peripheral blood lymphocytes from recipients were co-cultured in vitro with the donor＇s skin or a third rhesus for 72 h before and 20 days, 40 days, and 60 days after infusion. The pathological lesions were evaluated and the keratinocyte apoptosis of the skin sections was determined by TUNEL assay. Results： Twenty days after infusion, the pathological lesion in in vitro skin explants from MSCs donor was less and the apoptosis rate was declined to （12.2± 1.9）% from a pre-infusion level of （23.0±4.9）%. The apoptosis of cells in skin explants from the third party also declined to （11.8±1.4）% from a pre-infusion level of （22.0±2.0）% （P〈0.05）. Intensive immunological reaction was observed between recipient and donor/the third party on day 40 and day 60 after transplantation, with the apoptosis rate being （22.0±3.2）%, （22.0±2.5）% （donor） and （23.2±2.4）%, （24.0±2.6）% （the third party）. Conclusion： Allogenic MSCs transfusion can decrease the immunological reaction between donors and recipients in rhesus, but this decreased reaction is temporary and reversible. |
| Key words: mesenehymal stem cells immunity,cellular skin transplantation TUNEL assay |
