Organotypic Culture as a Model for EBV Infection of Epithelial Cells

器官型培养作为上皮细胞 EBV 感染的模型

• 批准号: 8038004
• 负责人: Clare E Sample
• 金额: $ 23.27万
• 依托单位: PENNSYLVANIA STATE UNIV HERSHEY MED CTR
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-01 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8038004
• 关键词: Acquired Immunodeficiency Syndrome; Address; Antiviral Agents; B-Lymphocytes; Binding Proteins; Biological Assay; Biological Models; Biology; Biopsy; Boxing; Cell Line; Cell Proliferation; Cells; Coculture Techniques; Data; Detection; Development; Environment; Episome; Epithelial; Epithelial Cells; Epithelium; Epstein-Barr Virus Infections; Epstein-Barr Virus latency; Event; Factor X; Funding Mechanisms; Future; Gene Mutation; Genes; Glycoproteins; Hairy Leukoplakia; Head and Neck Cancer; Human; Human Herpesvirus 4; Human Papillomavirus; In Situ; Incidence; Infection; Knowledge; Life Cycle Stages; Light; Lytic; Maintenance; Malignant Neoplasms; Membrane Proteins; Modeling; Nasopharynx Carcinoma; Nature; Oral; Oral cavity; Oral mucous membrane structure; Patients; Play; Proliferating; Proteins; Recombinants; Research; Role; Site; Staging; Stem cells; Stomach Carcinoma; Stratum Basale; System; Testing; Tongue; Tonsil; Tropism; Viral; Viral Proteins; Virion; Virus; Virus Diseases; base; cell type; differentiated B cell; in vitro Model; in vivo; keratinocyte; latent infection; lytic replication; monolayer; oral cavity epithelium; public health relevance; research study; transcription factor; tumor

DESCRIPTION (provided by applicant): Epstein-Barr virus (EBV) is associated with a number of malignancies of B lymphocyte and epithelial origin. Many of these are specific to or increased in AIDS patients including epithelial cancers of the head and neck. This proposal focuses on the development of an in vitro model of EBV infection of epithelium using organotypic cultures. Organotypic cultures differ from monolayers in that the cells differentiate and stratify to resemble epithelium found in situ. Because EBV can replicate in differentiated epithelial cells in vivo, organotypic culture is an ideal system to study the role of epithelial cells in the viral life cycle. Our preliminary data demonstrate that we can efficiently infect primary oral keratinocytes in culture using a recombinant EBV that expresses GFP as a marker, and that we can generate organotypic cultures of primary oral keratinocytes that effectively mimic oral mucosal epithelium. Based on these preliminary data, we propose to characterize EBV infection in organotypic culture identifying the cells infected and the viral gene products expressed. Several studies have suggested that EBV may establish a latent infection in vivo. We will determine whether EBV can establish a latent infection and characterize the type of latency. The functions of the viral gene products expressed in epithelium will be examined. Thus, we will establish and characterize a model system for the study of EBV in epithelium and use this system to define the functions of the viral gene products as far as possible within the limited scope of the R21 funding mechanism. These studies will set the stage for future experiments that will examine several long standing questions concerning EBV infection of human oral epithelium. PUBLIC HEALTH RELEVANCE: Epstein-Barr virus (EBV) is associated with a variety of human malignancies that occur predominantly in B lymphocytes or epithelial cells, many of which have an increased incidence in AIDS patients. The proposed research is intended to develop an in vitro model of EBV infection of epithelium that will be invaluable in filling in the gaps in our knowledge about the role of epithelial cells in the EBV lifecycle. Because the oral mucosa is the site of entry and exit of the virus, epithelial cells play a major role in viral spread and thus, understanding their role in the viral life cycle will not only shed light on this important step but has the potential to impact all downstream events from EBV infection. This model is likely to be important not only in adding to our knowledge of the role of the epithelium in the viral life cycle and the contribution of the virus to the development of malignancies, but could also be used for the development and/or testing of antiviral strategies.

描述（由申请人提供）：Epstein-Barr病毒（EBV）与B淋巴细胞和上皮起源的许多恶性肿瘤有关。其中许多是针对艾滋病患者的特定或增加的，包括头部和颈部上皮癌。该提案的重点是使用器官型培养物的EBV感染的体外模型。细胞培养物与单层不同，因为细胞分化并分层以类似于原位发现的上皮。因为EBV可以在体内分化的上皮细胞中复制，所以器官培养是研究上皮细胞在病毒生命周期中作用的理想系统。我们的初步数据表明，我们可以使用将GFP表示为标记物的重组EBV在培养中有效地感染原发性口服角质形成细胞，并且我们可以生成原代口腔角质形成细胞的器官型培养物，从而有效地模仿口服粘膜上皮。基于这些初步数据，我们建议表征EBV感染在鉴定感染细胞和表达的病毒基因产物的器官型培养物中的EBV感染。几项研究表明，EBV可能在体内建立潜在感染。我们将确定EBV是否可以建立潜在感染并表征潜伏期的类型。将研究在上皮中表达的病毒基因产物的功能。因此，我们将在上皮中建立并表征用于研究EBV的模型系统，并使用该系统在R21资金机制的有限范围内尽可能地定义病毒基因产物的功能。这些研究将为将来的实验奠定阶段，这些实验将研究有关人口腔上皮的EBV感染的几个长期存在的问题。 公共卫生相关性：Epstein-Barr病毒（EBV）与B淋巴细胞或上皮细胞中主要发生的多种人类恶性肿瘤有关，其中许多人在AIDS患者中的发病率增加。拟议的研究旨在开发上皮细胞EBV感染的体外模型，这对于填补了关于上皮细胞在EBV生命周期中作用的知识的差距是无价的。由于口腔粘膜是病毒的进入和退出的位置，因此上皮细胞在病毒扩散中起主要作用，因此，了解它们在病毒生命周期中的作用不仅会阐明这一重要步骤，而且有可能影响EBV感染的所有下游事件。该模型不仅在增加我们对上皮在病毒生命周期中的作用以及病毒对恶性肿瘤发展的贡献的知识不仅重要，而且还可以用于开发和/或测试抗病毒药策略。