Moses R D, Beschorner W E, Singer D, Maguire J E, Orr K S, Bacher J D, Gress R E
Experimental Immunology Branch, National Cancer Institute, Bethesda, MD 20892.
Bone Marrow Transplant. 1989 Sep;4(5):475-81.
Our interest in MHC-mismatched allogeneic bone marrow transplantation (BMT) in the rhesus monkey prompted us to investigate restriction fragment length polymorphism analysis as a means for detecting lymphohematopoietic chimerism in the primate. A human MHC (HLA) class II DR beta gene cDNA probe was tested on rhesus peripheral blood mononuclear cell DNA digested with any of three restriction enzymes. We found that (1) the human DR beta probe hybridized to as many as 15 restriction fragments per rhesus DNA sample, suggesting cross-hybridization at more than one locus of rhesus MHC class II beta genes; (2) restriction fragment length polymorphisms were common among outbred monkeys as a result of class II beta gene polymorphisms and would be sufficient for chimerism detection in the majority of random pairs of outbred monkeys utilizing only a single restriction enzyme (Bgl II); and (3) sensitivity (5-10% chimerism) was comparable to that reported for restriction fragment length polymorphism assays utilizing non-MHC probes in clinical HLA-identical BMT. Utility of the assay was demonstrated in a preliminary series of experiments in rhesus monkeys conditioned with mixed T cell-depleted MHC-mismatched allogeneic plus T cell-depleted autologous BMT with or without cardiac allograft implantation.
我们对恒河猴中主要组织相容性复合体（MHC）不匹配的同种异体骨髓移植（BMT）感兴趣，这促使我们研究限制性片段长度多态性分析，以此作为检测灵长类动物淋巴细胞造血嵌合体的一种方法。用三种限制性内切酶中的任何一种消化恒河猴外周血单个核细胞DNA后，对人MHC（HLA）II类DRβ基因cDNA探针进行了检测。我们发现：（1）人DRβ探针与每个恒河猴DNA样本多达15个限制性片段杂交，这表明在恒河猴MHC II类β基因的多个位点存在交叉杂交；（2）由于II类β基因多态性，限制性片段长度多态性在远交系猴子中很常见，并且仅使用一种限制性内切酶（Bgl II）就足以在大多数随机配对的远交系猴子中检测嵌合体；（3）敏感性（5%-10%嵌合体）与临床HLA相同的BMT中使用非MHC探针的限制性片段长度多态性分析所报道的敏感性相当。在一系列初步实验中证明了该检测方法的实用性，这些实验使用混合T细胞去除的MHC不匹配的同种异体加T细胞去除的自体BMT对恒河猴进行预处理，同时伴有或不伴有心脏同种异体移植植入。
