Dambaeva S V, Bondarenko G I, Grendell R L, Kravitz R H, Durning M, Golos T G
Department of Obstetrics and Gynecology, Wisconsin National Primate Research Center, University of Wisconsin-Madison, Madison, WI 53715, USA.
Placenta. 2008 Jan;29(1):58-70. doi: 10.1016/j.placenta.2007.10.001. Epub 2007 Nov 9.
The aim of this study was to characterize the expression of the rhesus HLA-E ortholog Mamu-E, particularly at the maternal-fetal interface. Mamu-E expression was confirmed by locus-specific RT-PCR in the placenta as well as in peripheral blood mononuclear cells (PBMC) and other organs. We evaluated the utility of antibodies recognizing HLA-E (MEM-E/06 against native HLA-E, MEM-E/02 against denatured HLA-E) to detect Mamu-E by flow cytometry/immunofluorescence, Western blot, and immunohistochemistry (IHC). Western blot analysis of cells and selected transfectants confirmed the recognition of Mamu-E but not Mamu-AG by antibodies MEM-E/06 and HC10 but not MEM-E/02. Immunohistochemical staining of frozen sections of rhesus placenta with the MEM-E/06 antibody demonstrated expression in most populations of rhesus monkey trophoblast cells, including villous cytotrophoblasts (strong positive staining), apical membrane of syncytiotrophoblasts (light to moderate staining) and extravillous cytotrophoblasts (moderate to strong staining, especially endovascular trophoblasts in early pregnancy). Expression was not trophoblast cell-specific, especially at term, when endothelial cells in both the chorionic plate and placental villi showed strong staining for Mamu-E. Staining of rhesus extravillous trophoblast cells suggested the co-expression of Mamu-E and Mamu-AG (the rhesus HLA-G homolog) on these cells. MEM-E/06 was shown also to react with differentiating rhesus placental syncytiotrophoblasts in primary culture, detecting intracellular and weak surface expression of Mamu-E. We conclude that the gestation-dependent co-expression of Mamu-E with Mamu-AG in villous and extravillous trophoblast cells suggests important and perhaps complementary but distinct roles of these two non-classical MHC class I loci in pregnancy at the maternal-fetal interface. In addition, the MEM-E/06 antibody will be useful for the detection of Mamu-E at the maternal-fetal interface in the rhesus monkey.
本研究的目的是表征恒河猴HLA-E直系同源物Mamu-E的表达情况，尤其是在母胎界面处的表达。通过位点特异性逆转录聚合酶链反应（RT-PCR）在胎盘、外周血单个核细胞（PBMC）及其他器官中证实了Mamu-E的表达。我们评估了识别HLA-E的抗体（针对天然HLA-E的MEM-E/06、针对变性HLA-E的MEM-E/02）通过流式细胞术/免疫荧光、蛋白质免疫印迹及免疫组织化学（IHC）检测Mamu-E的效用。对细胞和选定转染子进行的蛋白质免疫印迹分析证实，抗体MEM-E/06和HC10可识别Mamu-E而非Mamu-AG，而MEM-E/02则不能。用MEM-E/06抗体对恒河猴胎盘冰冻切片进行免疫组织化学染色，结果显示在大多数恒河猴滋养层细胞群体中均有表达，包括绒毛细胞滋养层细胞（强阳性染色）、合体滋养层细胞顶膜（轻度至中度染色）和绒毛外细胞滋养层细胞（中度至强染色，尤其是早孕时的血管内滋养层细胞）。这种表达并非滋养层细胞特异性的，特别是在足月时，绒毛板和胎盘绒毛中的内皮细胞均显示出对Mamu-E的强染色。对恒河猴绒毛外滋养层细胞的染色表明，这些细胞上Mamu-E和Mamu-AG（恒河猴HLA-G同源物）共表达。还显示MEM-E/06与原代培养中正在分化的恒河猴胎盘合体滋养层细胞发生反应，检测到Mamu-E的细胞内表达及微弱的表面表达。我们得出结论，绒毛和绒毛外滋养层细胞中Mamu-E与Mamu-AG的妊娠依赖性共表达表明这两个非经典MHC I类基因座在母胎界面妊娠过程中发挥着重要作用，可能是互补但又不同的作用。此外，MEM-E/06抗体将有助于在恒河猴母胎界面检测Mamu-E。
