人胚神经干细胞体外免疫原性测定

Detection of immunogenecity of human embryonic neural stem cells in vitro

目的探讨人胚神经干细胞体外免疫原性情况。方法取孕14周流产人胚侧脑室下区细胞,进行分离、培养、鉴定。提取人胚神经干细胞和正常人外周血单核细胞中的RNA,采用RT-PCR法反转录,扩增,电泳,观察目的条带。根据外周血单核细胞培养分4组阴性对照组以外周血单核细胞单独培养,实验组为B、C组分别以103ml-1、105ml-1神经干细胞加入外周血单核细胞中共培养,阳性对照组将植物血凝素以0.1mg/L加入外周血单核细胞中共培养。用MTT比色法测定刺激后外周血单核细胞增殖能力。结果RNA反转录后电泳条带显示神经干细胞表达HLA-A、HLA-DRA,MTT比色法细胞活力测定显示B组、C组D(490nm)分别为0.4450±0.0265、0.5675±0.0781,C组与阳性对照组比较差异无统计学意义(P>0.05),B组、C组与阴性对照组比较差异有统计学意义(P<0.05)。结论人胚侧脑室下区的神经干细胞在体外有一定的免疫原性,可促进外周血单核细胞增殖,移植时需注意免疫排斥问题。

immunogenecity ; neural stem cell ; embryo ; human ; in vitro Abstract Aim ： To explore the immunogeneeity of human embryonic neural stem cells in vitro. Methods：Tissue from 14 week human subventrieular zone （SVZ） was separated, cultured and identified in vitro. Total RNA was isolated from neural stem cells and peripheral blood mononuelear cells, and reversely transcribed into eDNA, which was then used for multiplex PCR. The PCR reaction product was analyzed by gel eleetrophoresis. Different numbers of NSCs were added into different experimental groups,and proliferation ability of PBMCs were measured by MTT methods. Results： Gel eleetrophoresis indicated the expression of HLA-A and HLA-DRA by NSCs and PBMCs. MTT showed that there was no significant difference between group C and positive-control group （ P 〉 0.05 ）, and there was significant difference between group B （ D（490 nm） ：0. 445 0± 0. 026 5 ） and negative-control group and between group C （ D（490 nm ） ：0. 567 5± 0. 078 1 ） and negative-control group（ P 〈 0.05 ）. Conclusion ： NSCs from SVZ have immunogeneeity in vitro, and can stimulate the proliferation of PBMCs. Immunologic rejection should be paid attention to in transplantation.