Meyer K, Basu A, Ray R
Department of Internal Medicine, Saint Louis University, St. Louis, Missouri 63110, USA.
Virology. 2000 Oct 10;276(1):214-26. doi: 10.1006/viro.2000.0547.
We have previously reported the generation of pseudotype virus from chimeric gene constructs encoding the ectodomain of the E1 or E2 glycoprotein of hepatitis C virus (HCV) genotype 1a appended to the trans membrane domain and cytoplasmic tail of the vesicular stomatitis virus (VSV) G protein. Sera derived from chimpanzees immunized with homologous HCV glycoproteins neutralized pseudotype virus infectivity (L. M. Lagging et al., J. Virol. 72, 3539-3546, 1998). We have now extended this study to further understand the role of HCV glycoproteins in pseudotype virus entry. Although a number of mammalian epithelial cells were susceptible to VSV/HCV pseudotype virus infection, plaquing efficiency was different among host cell lines. Pseudotype virus adsorption at low temperature decreased plaque numbers. Treatment of E1 or E2 pseudotype virus in media between pH 5 and 8 before adsorption on cells did not significantly reduce plaque numbers. On the other hand, treatment of cells with lysosomotropic agents or inhibitors of vacuolar H(+) ATPases had an inhibitory role on virus entry. Concanavalin A, a plant lectin, exhibited neutralization of both HCV E1 and E2 pseudotype virus infectivity. However, mannose binding protein, a C-type mammalian lectin, did not neutralize virus in the absence or presence of serum complement. Pseudotype virus infectivity was only partially inhibited by heparin, a highly sulfated glycosaminoglycan, in a saturable manner. Additional studies suggested that low-density lipoprotein receptor related molecules partially inhibit E1 pseudotype virus infectivity, while CD81 related molecules interfere with E2 pseudotype virus infectivity. A further understanding of HCV entry and strategies appropriate for mimicking cell surface molecules may help in the development of new therapeutic modalities against HCV infection.
我们之前报道过，从嵌合基因构建体中产生了假型病毒，该构建体编码丙型肝炎病毒（HCV）1a基因型E1或E2糖蛋白的胞外域，并附加到水疱性口炎病毒（VSV）G蛋白的跨膜结构域和胞质尾上。用同源HCV糖蛋白免疫的黑猩猩血清可中和假型病毒的感染性（L.M.拉金等人，《病毒学杂志》72卷，3539 - 3546页，1998年）。我们现在扩展了这项研究，以进一步了解HCV糖蛋白在假型病毒进入过程中的作用。尽管许多哺乳动物上皮细胞对VSV/HCV假型病毒感染敏感，但在宿主细胞系中蚀斑形成效率有所不同。低温下假型病毒的吸附会减少蚀斑数量。在吸附到细胞上之前，将E1或E2假型病毒在pH 5至8的培养基中处理，并不会显著减少蚀斑数量。另一方面，用溶酶体促渗剂或液泡H(+)ATP酶抑制剂处理细胞对病毒进入有抑制作用。植物凝集素伴刀豆球蛋白A可中和HCV E1和E2假型病毒的感染性。然而，C型哺乳动物凝集素甘露糖结合蛋白在有无血清补体的情况下都不能中和病毒。高硫酸化糖胺聚糖肝素只能以饱和方式部分抑制假型病毒的感染性。进一步的研究表明，低密度脂蛋白受体相关分子部分抑制E1假型病毒的感染性，而CD81相关分子干扰E2假型病毒的感染性。对HCV进入过程以及适合模拟细胞表面分子的策略的进一步了解，可能有助于开发针对HCV感染的新治疗方法。
