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Research for a retrovirus in myasthenic thymus and study for the pathogenesis

肌无力胸腺逆转录病毒的研究及发病机制研究

基本信息

批准号：
02454248
负责人：
ONO Akira
金额：
$ 2.24万
依托单位：
The Kitasato Institute
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1990
资助国家：
日本
起止时间：
1990 至 1992
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-02454248/
关键词：
myasthenia gravis thymus thymoma retrovirus 遺伝子クロ-ニングレトロウィルス

项目摘要

By electron microscopy, we have demonstrated in retrovirus particles in cultured thymus cells (thymomas and thymus hyperplasias) of patients with myasthenia gravis. In most cases, these particles were detected in thymus epithelial cells when cultured with mitomycin C-treated human B-cells. For further characterization of these particles, virus-producer cell lines were established by co-culturing human T-lymphoblastoid cells with B-cell-stimulated human thymus cells. Mn^<++>-dependent RNA- directed DNA polymerase activity was detected at a density of 1.15-1.17 g/cm^3 on sucrose density gradients in the culture fluid of these co-cultured T-cells. We molecularly cloned the cDNA prepared by the detergent-activated endogenous polymerase reaction in the culture fluid of KK-ll cells, one of the virus-producer T-cell lines. The ^<32>p-labelled cloned cDNA hybridized to DNAs from freshly prepared hyman thymoma cells, in addition to KK-ll cell DNA. We sequenced about 400-1200 bp of seven cDNA clones, and the sequence of each cDNA was compared with that of all genes with known sequences present in EMBL data base. No striking similarity was observed when compared to other sequences. The nucleotide sequence of seven cloned cDNAs constantly revealed high homology (50-70%) with that of retroviruses such as HIV-l and equine infectious anemia virus (EIAV). These results indicate that cDNAs obtained here are new viral genes, possibly retroviral genes, derived from human thymus cells.

通过电子显微镜，我们在培养的重症肌无力患者的胸腺细胞（胸腺瘤和胸腺增生）中证实了逆转录病毒颗粒。在大多数情况下，当与丝裂霉素c处理的人b细胞一起培养时，在胸腺上皮细胞中检测到这些颗粒。为了进一步表征这些颗粒，通过将人t淋巴母细胞样细胞与b细胞刺激的人胸腺细胞共培养，建立了病毒产生细胞系。在这些共培养t细胞的培养液中，在1.15-1.17 g/cm^3的密度梯度上检测了Mn^<++>依赖性RNA定向DNA聚合酶的活性。我们在产毒t细胞系之一KK-ll细胞的培养液中分子克隆了由洗涤剂激活内源性聚合酶反应制备的cDNA。^<32>p标记的克隆cDNA与新鲜制备的hyman胸腺瘤细胞的DNA杂交，除了KK-ll细胞的DNA。我们对7个cDNA克隆进行了400- 1200bp的测序，并将每个cDNA的序列与EMBL数据库中已知序列的所有基因的序列进行了比较。与其他序列相比，没有观察到惊人的相似性。7个克隆cdna的核苷酸序列不断显示出与hiv - 1和马传染性贫血病毒（EIAV）等逆转录病毒的高度同源性（50-70%）。这些结果表明，这里获得的cdna是新的病毒基因，可能是来自人胸腺细胞的逆转录病毒基因。