Effects of saliva on herpes simplex virus infection of oralcells

描述(由申请人提供)：牙周病(PD)是一种常见的口腔疾病，也是导致牙齿脱落的主要原因。它是一种多菌感染，其特征是由细菌定植引起的牙周炎，导致软组织破坏和牙槽骨形成。 骨吸收。有趣的是，单纯疱疹病毒1型(HSV-1)经常与牙周病变有关。它被认为是一个辅助因素，可能会通过破坏牙龈组织和维持局部炎症来增加个人患帕金森病的易感性。已知唾液中含有抗病毒和抗微生物药物，包括作用于HSV病毒粒子以防止感染的成分。我们第一次观察到，一些人的唾液中含有一种成分，可以增加牙龈成纤维细胞对HSV-1感染的易感性。这种作用有利于病毒的传播和扩大HSV对暴露在牙周病变中的细胞的趋向性。对一种新的唾液效应的意外观察促使我们在这一应用中进一步探索它。因此，这项研究的目的是确定这一因素及其影响，以了解HSV-1如何在口腔内有效传播，以及它如何加剧PD的进展。为了实现这些目标，我们提出了两个目标：在目标1中，我们将确定增加细胞感染易感性的唾液蛋白。我们将结合生化分离和功能感染试验对活性物质进行纯化。将通过质谱学、2D-Gel电泳法和生化分析对活性纯化组分进行鉴定。新的初步数据显示，雅加林会消耗人的唾液 它的活性表明碳水化合物对糖蛋白、粘蛋白和/或IgA的作用。因此，我们将研究碳水化合物的作用，并使用这种凝集素来改善纯化。我们还将通过研究分泌型IgA的作用来确定先前感染HSV是否与唾液的刺激作用有关。在目标2中，我们将首先定义唾液暴露如何影响口腔来源的各种细胞中的基因表达。我们已经聘请了专家来协助分析微阵列数据。我们还将使用模型细胞系来定义唾液如何在受体相互作用、进入途径和膜融合的水平上刺激HSV-1进入。最后，为了将这种影响与体内感染联系起来，我们将调查原代人类口腔细胞和HSV临床分离株。通过宾夕法尼亚大学的合作者，我们可以接触到各种这样的细胞和病毒。一种作用于口腔细胞以促进HSV-1感染的唾液剂的鉴定开创了一个新的先例。这将开辟新的研究领域，促进对HSV-1在帕金森病进展过程中作用的研究。由于这种活性并不存在于所有个体的唾液中，我们将能够进一步测试它作为口腔疱疹和可能的帕金森病易感性的生物标记物。该制剂将为预防HSV在口腔内传播和限制其在牙周炎中的潜在加重活动的治疗干预提供一个可接近的靶点。 公共卫生相关性：牙周病(PD)是一种由口腔细菌的炎症和增殖引起的多菌疾病，大约30%的美国成年人受到影响。单纯疱疹病毒1型(HSV-1)与帕金森病的皮损有关，被认为是该病发展的辅助因素。通过确定增加口腔细胞对HSV-1感染的易感性的唾液成分，以及通过表征细胞对唾液的反应，这项研究将：1)确定可能的口腔疱疹易感性的生物标志物；2)发现新的靶点 用于HSV-1感染和可能的牙周病的新治疗方法。

DESCRIPTION (provided by applicant): Periodontal disease (PD) is a common oral condition and a leading cause of tooth loss. It is a polymicrobial infection characterized by gingival inflammation stimulated by bacterial colonization resulting in soft tissue destruction and alveolar bone resorption. Interestingly, herpes simplex virus 1 (HSV-1) is frequently associated with periodontal lesions. It is thought to be a co-factor that may increase an individual's susceptibiliy to PD by damaging gingival tissue and sustaining local inflammation. Saliva is known to contain antiviral and antimicrobial agents, including components that act on the HSV virion to prevent infection. For the first time, we observed that saliva from some individuals but not others contains a component that increases the susceptibility of gingival fibroblasts to HSV-1 infection. This effect can favor viral spread and broaden HSV tropism to cells that are exposed in periodontal lesions. This unexpected observation of a novel salivary effect prompted us to explore it further in this application. Thus, the goals of this study are to identify this factor ad its effects in order to understand how HSV-1 efficiently spreads in the oral cavity and how it may exacerbate the progression of PD. Two aims are proposed to achieve these objectives: In aim 1, we will identify the salivary protein that increases cell susceptibility to infection. We will combine biochemical fractionation and functional infection assays to purify the active agent. Identification will be performed by mass spectrometry, 2D-gel electrophoresis and biochemical assays on active purified fractions. Novel preliminary data showing that jacalin depletes saliva of its activity suggest a role for carbohydrates on glycoproteins, mucins and/or IgA. Thus we will examine the role of carbohydrates and use this lectin to improve purification. We will also determine if prior infection by HSV relates to the stimulatory effect of saliva by investigating th role of secretory IgA. In aim 2, we will first define how gene expression is affected by exposure to saliva in various cells of oral origin. We have enlisted experts to aid in analyzing microarray data. We will also use model cell lines to define how saliva stimulates HSV-1 entry at the level of receptor interaction, entry pathways and membrane fusion. Finally, to relate this effect to in vivo infection, we will survey primary human oral cells and HSV clinical isolates. We have access to a variety of such cells and viruses by virtue of collaborators here at PENN. The identification of a salivary agent that acts on oral cells to favor HSV-1 infection sets a new precedent. It will open new areas of investigation and facilitate studies of the role of HSV-1 in the progression of PD. Since the activity is not present in the saliva from all individuals, we wil be able to further test it as a biomarker for susceptibility to oral herpes and possibly PD. This agent will provide an accessible target for therapeutic interventions to prevent HSV spread in the oral cavity and limit its potential aggravating activity in periodontitis. PUBLIC HEALTH RELEVANCE: Periodontal disease (PD) is a polymicrobial disease caused by inflammation and proliferation of oral bacteria that affects around 30% of American adults. Herpes simplex virus 1 (HSV-1) is associated with PD lesions and is thought to be a cofactor in the development of the disease. By identifying salivary components that increase susceptibility of oral cells to HSV-1 infection and by characterizing the cellular response to saliva, this study will: 1) identify possible biomarkers for susceptibility to oral herpes; and 2) uncover new targets for novel therapeutic approaches to HSV-1 infection and possibly periodontal diseases.