Carboxy-fluorescein diacetate, succinimidyl ester labeled papillomavirus virus-like particles fluoresce after internalization and interact with heparan sulfate for binding and entry
Carboxy-fluorescein diacetate, succinimidyl ester labeled papillomavirus virus-like particles fluoresce after internalization and interact with heparan sulfate for binding and entry
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DOI：
10.1016/s0042-6822(03)00114-4
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发表时间：
2003-05-25
期刊：
影响因子：
3.7
通讯作者：
Evander, M
中科院分区：
文献类型：
--
作者：
Drobni, P;Mistry, N;Evander, M
人乳头瘤病毒（hpv）感染上皮细胞并与生殖器癌相关。大多数上皮细胞系表达细胞表面糖胺聚糖（GAGs），通常附着在蛋白聚糖的蛋白核心上。我们的目的是研究gag如何影响HPV的进入。使用人角化细胞细胞系（HaCaT），将HPV病毒样颗粒（VLPs）与gag预孵育显示出剂量依赖性的结合抑制。肝素的IC50（50%抑制）仅为0.5马克杯/毫升，硫酸葡聚糖为1马克杯/毫升，粘膜来源的硫酸肝素为5-10马克杯/毫升。缺乏硫酸肝素或所有gag的突变中国鼠卵巢（CHO）细胞系不能结合HPV VLPs。在这里，我们还报告了一种研究内化的方法，通过使用标记有羧基荧光素二乙酸琥珀酰氨基酯的VLPs，这是一种只在细胞进入后激活的荧光染料。用肝素预处理标记的HPV VLPs抑制摄取，提示HPV和细胞表面硫酸肝素之间的主要相互作用。(C) 2003 Elsevier Science（美国）版权所有。
Human papillomaviruses (HPVs) infect epithelial cells and are associated with genital carcinoma. Most epithelial cell lines express cell-surface glycosaminoglycans (GAGs) usually found attached to the protein core of proteoglycans. Our aim was to study how GAGs influenced HPV entry. Using a human keratinocyte cell line (HaCaT), preincubation of HPV virus-like particles (VLPs) with GAGs showed a dose-dependent inhibition of binding. The IC50 (50% inhibition) was only 0.5 mug/ml for heparin, 1 mug/ml for dextran sulfate, and 5-10 mug/ml for heparan sulfate from mucosal origin. Mutated chinese hamster ovary (CHO) cell lines lacking heparan sulfate or all GAGs were unable to bind HPV VLPs. Here we also report a method to study internalization by using VLPs labeled with carboxy-fluorescein diacetate, succinimidyl ester, a fluorochrome that is only activated after cell entry. Pretreatment of labeled HPV VLPs with heparin inhibited uptake, suggesting a primary interaction between HPV and cell-surface heparan sulfate. (C) 2003 Elsevier Science (USA). All rights reserved.