Cervical Tissue Derived Organ Culture to Test Microbicides

宫颈组织衍生器官培养以测试杀菌剂

• 批准号: 7681868
• 负责人: Phalguni GUPTA
• 金额: $ 29.26万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-01 至 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7681868
• 关键词: Antiviral Agents; Applications Grants; Bacteria; Biological Assay; Cell Culture System; Cell Culture Techniques; Cells; Cervical; Cervix Uteri; Clinical Trials; Collaborations; Complement; Data; Development; Drug Formulations; Environmental Risk Factor; Epithelium; Evaluation; Genital system; HIV; HIV-1; HIV-1 vaccine; Histocompatibility; Histology; IL8 gene; Immune; In Vitro; Infection; Inflammation; Inflammatory; Inflammatory Response Pathway; Interleukin-6; Measures; Memory; Modeling; Monitor; Monkeys; Mucous Membrane; Neisseria gonorrhoeae; Nucleosides; Organ Culture Techniques; Outcome; Outcome Study; Peptides; Physiological; Prevotella melaninogenica; Principal Investigator; Property; Reverse Transcriptase Inhibitors; Screening procedure; Seminal fluid; Sexually Transmitted Diseases; System; Testing; Time; Tissues; Topical application; Toxic effect; Vagina; Vaginal Ring; anti-HIV microbicide; antimicrobial peptide; base; cell injury; cytokine; cytotoxic; cytotoxicity; cytotoxicity test; genital secretion; in vitro Assay; in vivo; inhibitor/antagonist; microbicide; microorganism; non-nucleoside reverse transcriptase inhibitors; pinacolyl methylphosphonic acid; preclinical evaluation; prevent; programs; response; retrocyclin; simian human immunodeficiency virus; tool; transmission process; vaginal fluid; vaginal microbicide

The in vitro cell culture system has been widely used as a primary screening tool for evaluating anti-HIV-1 activity of microbicides. However, there is clearly a need to develop a vaginal and cervical tissue based in vitro system to test the cytotoxicity and antiviral activity in the context of the complete tissue matrix. We have recently developed a cervical tissue-derived organ culture model which mimics in vivo conditions and has been used to test microbicides for their ability to block HIV-1 transmission and measure inflammatory cytokines in response to microbicides and sexually transmitted infection-related bacteria. Our hypothesis is that a cervical tissue-based organ culture is an ideal system to test microbicides for toxicity in genital tissue and for its ability to block transmission of HIV-1 with varying phenotypic properties across the cervical epithelium in the presence of common environmental factors that are present in vagina, such as semen, vaginal fluid, and STI-related microorganisms. Specific aims of the project are: 1) Evaluation of -nucleoside reverse transcriptase inhibitor (NNRTI) 5-chloro-3-phenylsulfonylindole-2-carboxamide (CSIC) from Project 1 and the entry inhibitor antimicrobial peptide retrocyclin RC101 from Project 2 alone or in combination and their formulations for their ability to block HIV-1 transmission across the mucosa in a cervical tissue-based organ culture. 2) Assessment of cervical tissue inflammation and their changes in response to CSIC and RC101 from Projects 1 and 2, respectively, using the organ culture model. The expression of proinflammatory cytokines, such as 11-13, IL-6, IL-8 and TNF-a will be monitored by measuring their messages in the tissues by the real time PCR and secretion in the culture supernatant using the Luminex system; 3) Evaluation of anti-HIV activity of CSIC and RC101 under physiologically relevant conditions. CSIC and RC101 alone or in combination and their formulations will be evaluated for their antiviral activities in the presence of semen and vaginal fluid. In addition the organ culture model will be expanded to measure STIrelated microorganisms, such as Neisseria gonorrhoeae and Prevotella melaninogenica by measuring proinflammatory cytokine response. Microbicides will then be evaluated for their antiviral activity in the aresence of the proinflammatory cytokines induced by these microorganisms.The proposed studies in this Droject will complement various other projects in this U19 grant application by providing a valuable in vitro cervical tissue-based assay which will bridge between the microbicide development (Projects 1 and 2), monkey model (Project 4) and Formulation Core (Core B).

体外细胞培养系统已被广泛用作评估抗HIV-1的主要筛选工具 微生物的活性。但是，显然需要在 体外系统在完整的组织基质中测试细胞毒性和抗病毒活性。我们有 最近开发了一种宫颈组织衍生的器官培养模型，该模型在体内疾病中模仿，并具有 用于测试微生物的阻断HIV-1传播和测量炎症的能力 细胞因子响应菌皮和性传播感染相关的细菌。我们的假设是 基于宫颈组织的器官培养是测试生殖器组织毒性菌皮的理想系统 并能够通过颈部跨宫颈特性不同的表型特性阻断HIV-1的传播 在存在阴道中存在的常见环境因素的情况下，例如精液， 阴道液和与STI相关的微生物。该项目的具体目的是：1）-Nucleoside评估 来自项目1 以及仅项目2或组合和组合的入口抑制剂抗菌肽折叠蛋白RC101 他们的制剂能够阻止基于宫颈组织中粘膜的HIV-1传播的能力 器官文化。 2）评估宫颈组织炎症及其对CSIC和CSIC的变化 RC101分别来自项目1和2的RC101，使用器官培养模型。表达 促炎细胞因子，例如11-13，IL-6，IL-8和TNF-A，将通过测量其测量来监测 使用Luminex通过实时PCR和培养物中的分泌在组织中的消息和分泌 系统; 3）在生理相关条件下评估CSIC和RC101的抗HIV活性。 CSIC RC101仅在 存在精液和阴道液。此外，将扩展器官培养模型以测量搅拌 微生物，例如淋病奈瑟氏菌和prevotella黑色素生成症。 促炎性细胞因子反应。然后，将评估微生物的抗病毒活性 这些微生物诱导的促炎细胞因子的消除性。在此提出的研究 DROKEXT将通过提供有价值的体外来补充本U19赠款申请中的其他各种项目 基于宫颈组织的测定，将在杀菌剂发育之间桥接（项目1和2）， 猴子模型（项目4）和配方核心（核心B）。