猪肠黏膜PP结中树突状细胞的分离与鉴定

Isolation and idetification of dendritic cells from porcine intestinal mucosal PP

[目的]本试验旨在建立体外分离猪肠黏膜派尔集合淋巴结(Peyer's patch,PP)中树突状细胞(dendritic cell,DC)的方法。[方法]应用密度梯度离心的方法获得单个核细胞,然后分别采用免疫磁珠法、流式细胞术根据细胞表面标志MHC-Ⅱ和SWC3a分选猪肠黏膜PP结DC,并进行DC形态学和免疫学性质鉴定。[结果]磁珠分选(magnetic cell sorting,MACS)富集SWC3a+细胞纯度达90.2%,而SWC3a+/MHC-Ⅱ+细胞纯度达50.5%。流式细胞术分选猪肠黏膜PP结中SWC3a+/MHC-Ⅱ+DC纯度高达95.6%,将该DC培养2 d后,细胞有聚集现象,细胞表面不规则,略有突起,具有刺激同种异体淋巴细胞增殖的能力。[结论]成功建立了体外分离猪肠黏膜PP结中DC的方法,分离到的DC具有体内DC的生物学特性,为进一步研究猪肠黏膜PP结中DC的功能奠定了基础。

[Objectives]The paper was aimed to create a method for the isolation of dendritic cells(DC)from porcine intestinal mucosal Peyer’s patch(PP)in vitro.[Methods]The mononuclear cells were obtained by density gradient centrifugation and then DC was sorted from porcine intestinal mucosal PP by magnetic activated cell sorting(MACS)and fluorescence activated cell sorting(FACS),and their morphological and immunological characteristics were investigated.[Results]The purity of SWC3a^+cells enriched by MACS was 90.2%,while the purity of SWC3a^+/MHC-Ⅱ+cells was 50.5%.The purity of SWC3a^+/MHC-Ⅱ+DC in porcine intestinal mucosal PP was 95.6%when it was sorted by flow cytometry.After 2 days of culture,the cells had aggregation.The surface of the cells was irregular and slightly protuberant,and had the ability to stimulate the proliferation of allogeneic lymphocytes.[Conclusions]The method to isolate DC from porcine intestinal mucosal PP is established,which might contribute to further study on the role of DC in porcine intestinal mucosal PP.