免疫组化分析鼠疫疫苗免疫和鼠疫菌攻毒后恒河猴脾组织中T和B淋巴细胞增殖

摘要

In our previous study, more B and T cells were observed in the splenic tissues from the immunized Rhesus macaques with plague vaccines than from normal animals. However, whether these cells represent proliferating B and T cells elicited by plague vaccines is unclear. To answer this question, the proliferation of T and B cells was examined by means of double immunohistochemistry in the spleens from the Rhesus macaques immunized with subunit vaccine SV1 (20 μg F1 + 10 μg rV270), and those both immunized with SV1, SV2 (200 μg F1 + 100 μg rV270), live attenuated vaccine EV or alhydrogel and challanged with Y. pestis, respectively. Double staining results showed that higher B cell proliferation in germinal centers and more resting B cells in marginal zone (MZ) were observed in the tested animals than in the normal animal, and that more resting T cells in periarterial lymphatic sheath (PALS) of the spleen tissues from the tested animals than from the normal animal, earlier stages of the immunization. Germinal centers in the control spleen tissues were incomplete, which was attributed to the histopathological lesions caused by virulent Y. pestis. The size of germinal centers in the spleen tissues from the animals both immunized with SV1, SV2 or EV and infected with Y. pestis was larger than that from the animal only immunized with SV1. In conclusion, B cell proliferation, more resting B cells, more resting T cells and expanded germinal centers are the signs of eliciting specific humoral immunity and keeping immune memory response. These results were also consistent with results of our previous study that the immunized animals with SV1, SV2 or EV elicited higher antibody and IL-4 production.%目的 在我们先前的研究中发现,鼠疫疫苗免疫的猕猴脾组织中B细胞和T细胞数量明显增加.然而,是否这些细胞是由鼠疫疫苗引起的增殖性B细胞和T细胞还不得而知.方法 为回答这个问题,本研究使用免疫组化双标记方法检测了猕猴脾组织中T细胞和B细胞的增殖.应用Ki67抗体以及T细胞和B细胞特异性单克隆抗体的免疫组化双标法,对脾组织T细胞和B细胞增殖进行检测.脾组织分别来自于亚单位疫苗SV1(20 μg F1 + 10 μg rV270)免疫的猕猴以及分别通过SV2 (200 μg F1 + 100 μg rV270)、减毒活疫苗EV和铝佐剂免疫并分别攻毒的猕猴.结果 与正常动物相比,受试动物脾组织生发中心有较多的B细胞增殖,边缘区有较多的静止B细胞,在动脉周围淋巴鞘区有较多的静止T细胞,提示原始T细胞可能在免疫早期发生增殖.经SV1、SV2或EV76免疫并攻毒的动物脾组织生发中心较仅用SV1免疫动物的生发中心扩大.此外,铝佐剂免疫并攻毒的猕猴脾组织生发中心不完整,这可能归因于强毒株鼠疫菌感染引起的病理损伤.B细胞增殖、静止B细胞增加、静止T细胞增多及生发中心扩大是诱导特异性体液免疫和保持免疫记忆反应的标志.结论 这些结果与我们先前的发现的SV1、SV2或者EV76免疫动物激发较高的抗体和IL-4分泌相一致.