Role of normal cervical stem cells in the HPV induced initiation of cervical can

正常宫颈干细胞在HPV诱导的宫颈癌发生中的作用

• 批准号: 7592964
• 负责人: John Niederhuber
• 金额: $ 29.77万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7592964
• 关键词: Apical; Architecture; Basal Cell; Basement membrane; Binding; Binding Sites; Cell Count; Cell Fraction; Cell Separation; Cell physiology; Cells; Cervical; Cervix Uteri; Condition; Cooperative Human Tissue Network; Data; Development; Dissociation; Dysplasia; Environment; Epithelial; Epithelial Cells; Epithelium; Fresh Tissue; Goals; HPV-High Risk; Heterogeneity; Human; Human Papillomavirus; Immune; In Situ; In Vitro; Infection; Laboratories; Lead; Lesion; Malignant Neoplasms; Malignant neoplasm of cervix uteri; Modeling; Monitor; Natural regeneration; Neoplasms; Numbers; Organ Culture Techniques; Papillomavirus; Population; Premalignant; Property; Protocols documentation; Research; Risk; Role; Stem cells; Stratum Basale; Testing; Time; Tissues; Virus; Virus Diseases; Woman; carcinogenesis; design; improved; in vivo; neoplastic; novel strategies; particle

Though HPV has been established as the etiologic agent for virtually all cervical cancers, the majority of cervical HPV infections do not lead to cervical cancer. Only 1% of cases progress to high grade dysplasia and cancer. The majority of cervical HPV-infections are transient non-neoplastic productive viral infections which disappear within a year. It is generally accepted that persistence of high risk HPV infection is an imperative state in the development of cervical neoplasia. The crucial factors determining persistence of infection and its correlation with an increased risk for cervical cancer in women with normal immune status are still largely unknown. Persistent infection may only occur upon targeted infection of specific cervical cells, possibly with stem cell properties. In order test this idea, we use fresh tissue cervical epithelial tissue obtained from the Cooperative Human Tissue Network. A former research fellow from our laboratory successfully developed a protocol to isolate cervical epithelial cells and to identify a phenotypically distinct subpopulation with functional properties expected for stem cells. The preliminary data show that this cell fraction, originating from the basal layer of the ectocervical epithelium possesses an 80 fold increased capacity to bind papillomavirus-like particles (VLP), thus implying that these cells may express an increased number of papillomavirus binding sites facilitating viral infection if targeted by the virus. In effort to optimize the in vitro environment that recapitulates the conditions under which stem cells function in situ, we have also developed an ectocervical organ culture model that allows us to establish and monitor HPV binding and infection in fresh cervical tissue. Cervical tissue maintained in culture undergoes rapid cell dissociation and exfoliation of the more differentiated, apical epithelial layers. In contrast, the basal cells remain attached to the basement membrane. The remaining cells, formerly quiescent, become proliferative and regenerate the entire epithelium, preserving the tissue architecture and cellular heterogeneity normally observed in vivo. In order to establish infection in our model, exposure of the basal cells is necessary. We were able to show that only a very limited number of cells within the basal layer remains infected over time. It is our plan to characterize the infected cells using FACs analysis to determine whether they are indeed cervical stem cells. We plan to further characterize the cervical stem cell population by searching for new markers using a microarray approach which will improve stem cell isolation and to investigate the role of these cells in progression of a productive viral infection to virus induced pre-malignant lesions and cervical cancer. This study aims to improve our understanding of HPV induced cervical carcinogenesis and contribute to the development of novel strategies designed to target cervical cancer.

虽然HPV已被确定为几乎所有宫颈癌的病原体， 宫颈癌是一种常见的恶性肿瘤。只有1%的 例进展为高度异型增生和癌症。大多数宫颈HPV感染是 短暂的非肿瘤性生产性病毒感染，在一年内消失。是 普遍认为，高风险HPV感染的持续性是一个必要的状态， 宫颈肿瘤的发展。决定感染持续性的关键因素 及其与患有宫颈癌的妇女患宫颈癌风险增加的相关性 正常的免疫状态在很大程度上仍然是未知的。持续性感染可能只 发生在特定宫颈细胞的靶向感染后，可能具有干细胞特性。 为了验证这一想法，我们使用了从子宫颈癌组织中获得的新鲜组织宫颈上皮组织。 人类组织合作网络。我们实验室的一位前研究员成功地 开发了一种分离宫颈上皮细胞的方案， 具有干细胞预期功能特性的独特亚群。初步 数据显示，这种细胞部分，起源于外宫颈的基底层， 上皮细胞结合乳头瘤病毒样颗粒的能力增加了80倍 (VLP)因此，这意味着这些细胞可能表达更多的乳头状瘤病毒， 如果被病毒靶向，则促进病毒感染的结合位点。为了优化 在体外环境中，概括了干细胞在体内发挥作用的条件， 原位，我们还开发了宫颈外器官培养模型，使我们能够建立 并监测新鲜宫颈组织中的HPV结合和感染。宫颈组织保存在 培养物经历快速的细胞解离和更分化的顶端细胞的剥落， 上皮层相反，基底细胞仍然附着在基底膜上。的 其余的细胞，以前静止，成为增殖和再生的整个 上皮，保留组织结构和细胞异质性，通常在 vivo.为了在我们的模型中建立感染，暴露基底细胞是必要的。 我们能够证明只有非常有限数量的细胞在基底层内仍然存在 随着时间的推移感染。我们计划使用FACs分析来表征受感染的细胞， 确定它们是否真的是宫颈干细胞。我们计划进一步描述 通过使用微阵列方法寻找新的标记物， 将改善干细胞分离，并研究这些细胞在进展中的作用， 对病毒引起的癌前病变和宫颈癌的生产性病毒感染。这 这项研究旨在提高我们对HPV诱导的宫颈癌发生的理解， 开发针对宫颈癌的新策略。