Analysis of host factors on neurotropism of SSPE virus

我们的研究结果表明，SSPE菌株的第三种受体可能不同于CD 150（SLAM）和CD 46。为了进一步证实这种新受体的存在并克隆这种分子的cDNA，我们用VSV假型系统进行了病毒进入试验，并用抗CD 150或CD 46的特异性单克隆抗体进行了进入阻断。SSPE菌株除CD46外，还可利用CD150，并可利用除CD46外的某种细胞表面分子。在人神经母细胞瘤细胞和人胶质瘤细胞中表达了这种新分子。作为病毒进入细胞的另一种估计，通过用细胞松弛素D处理感染细胞制备的SSPE假病毒粒子的合胞体形成监测，也获得了与VSV假型系统相同的结果.在上述证实的基础上，尝试用嗜亲性逆转录病毒载体从Vero细胞cDNA文库中克隆第3受体的cDNA，该分子似乎在我们所检测的可用细胞中表达最多。此外，还尝试制备阻断SSPE菌株进入Vero细胞的小鼠单克隆抗体以捕获新分子。目前，我们还没有成功，可能是因为它的表达量太少。这些工作目前仍在进行中。作为一种不同的方法，正在考虑一些更敏感和有效的克隆方法。

Our results obtained so far suggested a possibility of the 3rd receptor for SSPE strains different from CD150 (SLAM) and CD46. The purpose of this project is to further confirm existence of this novel receptor and to clone cDNA of this molecule.To confirm its existence, virus entry assay was performed using VSV pseudotype system together with entry blocking by the specific monoclonal antibody against CD150 or CD46. SSPE strains were found to use CD150 but not CD46, in addition, a certain cell surface molecule except for the both ones. This novel molecule was expressed human neuroblastoma cells and human glioma cells. As another estimation on virus entry, syncytium formation by SSPE psedovirions prepared by treatment of the infected cells with cytochalasin D was monitored and the same results as VSV pseudotype system were also obtained.Based on the above confirmation, cDNA cloning of the 3rd receptor was tried using ecotropic retroviral vector from cDNA library of Vero cells, which seemed to express the molecule most among our available cells examined. In addition to it, preparation of mouse monoclonal antibody blocking entry of SSPE strains into Vero cells was tried to catch the novel molecule. At present, we have not yet succeeded probably because of too little amount of its expression. These are now still in progress. As a different approach, some more sensitive and effective cloning methods are under consideration.