Mechanism of Respiratory Syncytial Virus Entry in Normal Human Bronchial Epitheli

呼吸道合胞病毒侵入正常人支气管上皮的机制

• 批准号: 9231389
• 负责人: Homero Gabriel San juan Vergara
• 金额: $ 11.05万
• 依托单位: UNIVERSITY DEL NORTE
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-03-15 至 2019-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9231389
• 关键词: Alpha Cell; Apoptosis; Bayesian Analysis; Biological Assay; Cell Culture Techniques; Cell Density; Cell Line; Cells; Characteristics; Cholesterol; Cilia; Clathrin; Colombia; DNA cassette; Dynamin; Endocytosis; Epithelial Cells; Etiology; Gene Silencing; Goals; Guidelines; Hela Cells; Heterogeneity; Human; Infant; Infection; Intervention; Label; Lectin; Lentivirus Vector; Link; Lower respiratory tract structure; Mediating; Mitosis; Molecular; Pathway interactions; Pharmacology; Phenotype; Population; Population Sizes; Positioning Attribute; Principal Investigator; Publications; Recombinants; Research; Respiration; Respiratory Process; Respiratory Syncytial Virus Infections; Respiratory syncytial virus; Route; Staining method; Stains; Testing; Tetanus Helper Peptide; Time; Undifferentiated; Virus; Virus Diseases; design; experimental study; genetic approach; inducible gene expression; inhibitor/antagonist; pathogen; prevent; programs; public health relevance; screening; small molecule; therapy development; vaccine development

DESCRIPTION (provided by applicant): The long term goal of our research is to find new ways to prevent or to treat the infection caused by respiratory syncytial virus (RSV). In order to reach this goal, we will examine the molecular aspects by which the virus enters its natural targets, in particular, human bronchial epithelial cells. The route used by RSV to infect its primary target cells has not been clearly established. The current proposal would help to clarify the influence of the cell's population context and cellular state during RSV infection and take them into account when we test endocytic pathways using approaches providing "down" and "up" phenotypes regarding RSV infection in NHBE cells and well-differentiated bronchial epithelial cell cultures. The specific aims of this project are: (1) Determine if the cell's populaion context exerts an influence on the RSV infection and entry in undifferentiated primary cultures of normal human bronchial epithelial (NHBE) cells. Using high cell content screening, the influence that several features associated with a cell's population context and cellular state have on RSV infection and entry will be evaluated. (2) Determine the entry pathway that RSV hijacks to infect undifferentiated NHBE cells by taking cell's population context into account. Experiments will be designed using pharmacological (small molecules) and genetic approaches that generate either "down" or "up" phenotypes in each of the different endocytic pathways. Modulator-induced changes on cell's population context will be characterized to correct their effects when evaluating their positive or negative impact on RSV entry, fusion or infection in order to identify those interventions, and consequent endocytic pathway, involved in RSV infection. (3) Determine RSV entry pathway in well-differentiated bronchial epithelial cell (BEC) cultures. Using the general "down" or "up" assay approach, several endocytic pathways will be probed through pharmacological interventions to determine RSV entry route in well-differentiated BEC. For the pharmacological interventions, we will follow the general guidelines described in Aim 2, and the impact of these interventions on RSV infection and entry will be evaluated taking into consideration the potential effect on the state of the cells (i.e. cilia status and apoptosis). In Colombia, RSV is the most important pathogen agent causing 41.8% of the new cases of lower tract respiratory illness in infants. Understanding the mechanistic process of the RSV infection will help in the development of the vaccines and therapies. PHS 398/2590 (Rev. 06/09) Page Continuation Format Page

描述（由申请人提供）：我们研究的长期目标是找到预防或治疗由呼吸道合胞病毒（RSV）引起的感染的新方法。为了实现这一目标，我们将检查病毒进入其自然靶标的分子方面，特别是人支气管上皮细胞。 RSV用来感染其主要靶细胞的途径尚未清楚地确​​定。当前的提案将有助于阐明RSV感染期间细胞种群环境和细胞状态的影响，并在我们使用提供有关NHBE细胞中RSV感染的“向下”和“ UP”表型测试内吞途径时考虑到它们的影响。该项目的具体目的是：（1）确定细胞的人口上下文是否对正常人支气管上皮（NHBE）细胞的未分化原发性培养物的RSV感染和进入。使用高细胞含量筛选，将评估与细胞群体上下文和细胞状态有关的几个特征对RSV感染和进入的影响。 （2）确定RSV劫持通过考虑细胞的种群上下文来感染未分化的NHBE细胞的进入途径。实验将使用药理学（小分子）和遗传方法设计，这些方法在每个不同的内吞途径中都会产生“向下”或“向上”表型。调节器引起的细胞种群环境的变化将被表征以纠正其对RSV进入，融合或感染的正面或负面影响，以识别其效果 这些干预措施以及随之而来的内吞途径涉及RSV感染。 （3）确定良好差异的支气管上皮细胞（BEC）培养物中的RSV进入途径。使用一般的“向下”或“向上”测定方法，将通过药理学干预措施探测几种内吞途径，以确定分化良好的BEC中的RSV进入途径。对于药理学干预措施，我们将遵循AIM 2中描述的一般指南，这些干预措施对RSV感染和进入的影响将考虑到对细胞状态的潜在影响（即纤毛状态和凋亡）。在哥伦比亚，RSV是最重要的病原体剂，导致婴儿较低呼吸道疾病的新病例中有41.8％。了解RSV感染的机械过程将有助于疫苗和疗法的开发。 PHS 398/2590（修订版06/09）页面延续格式页面