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T regulatory cell suppression of CD8+ cell responses during FIV Infection

FIV 感染期间 T 调节细胞抑制 CD8 细胞反应

基本信息

批准号：
8260417
负责人：
Jonathan Fogle
金额：
$ 12.29万
依托单位：
NORTH CAROLINA STATE UNIVERSITY RALEIGH
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-05-01 至 2013-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8260417
关键词：
AIDS/HIV problem Acquired Immunodeficiency Syndrome Acute Address Animal Model Animals Antibodies Antigens Autologous Binding Blood CD4 Positive T Lymphocytes CD8-Positive T-Lymphocytes CD8B1 gene Cell membrane Cell physiology Cell surface Cells Clinical Clonal Anergy Clonal Expansion Coculture Techniques Complement 3d Receptors Cyclin-Dependent Kinase Inhibitor Cyclin-Dependent Kinase Inhibitor Gene Cyclin-Dependent Kinases Data Disease Progression Doctor&apos s Degree Effector Cell Enzyme-Linked Immunosorbent Assay Epitopes Event Failure Family Felidae Feline Immunodeficiency Virus Felis catus Flow Cytometry Future G1 Phase Genes Goals HIV HIV Antigens HIV Infections Helper-Inducer T-Lymphocyte Human IL2RA gene Immune Immune System Diseases Immune response Immunology Immunosuppressive Agents Incubated Indium Infection Internal Medicine Laboratories Leishmania Measures Mediating Membrane Messenger RNA Mus Outcome Patients Peptides Peripheral Blood Mononuclear Cell Phenotype Phosphorylation Plasma Plasmodium Production Proliferating Regulatory T-Lymphocyte Research Research Personnel Residencies Reverse Transcriptase Polymerase Chain Reaction Sampling Signal Pathway Signal Transduction Staging Subfamily lentivirinae Surface T cell anergy T-Lymphocyte Testing Therapeutic Time Transforming Growth Factor beta Up-Regulation Veterinary Medicine Viral Viral Antigens Viral Load result Virus anergy arm design experience human subject in vivo pathogen programs receptor research study response

项目摘要

DESCRIPTION (provided by applicant): Candidate: Jonathan E. Fogle, DVM, has completed a residency in small animal internal medicine at NCSU and is board certified in Veterinary Internal Medicine. Dr. Fogle's short term goal is to obtain a doctoral degree from the Immunology program at NCSU. Dr. Fogle's long term goal is to continue independent lentiviral research that draws from both his clinical veterinary medicine and doctoral research experience. Proposal: A significant fraction of untreated HIV+ patients (progressors) are characterized by CD4+ and CD8+ T cell immune dysfunction. HIV-specific CD8+ effector cells are induced during the acute stage infection, but become non-responsive or "anergic" to antigenic stimulation shortly after induction. We propose that the CD8+ T cell immune dysfunction is the result of "clonal anergy" induced by membrane-bound TGF-beta (mTGF-beta) on virus activated CD4+CD25+ T reg cells that engage TGF-betaRII on the target cell. This hypothesis cannot be addressed in human subjects but rather requires a well-characterized experimental AIDS lentivirus animal model such as FIV. SPF cats will be infected with the NCSU1 isolate of FIV, plasma and PBMC/LN cells will be collected at various times post infection and quantitated for virus burden by ELISA and Real Time-PCR. PBMC/LN will be phenotyped for surface receptors by flow cytometry, and for Foxp3 mRNA by RT-PCR. Treg cells will be assessed for suppressor function by CD4+CD25+ cell depletion and by incubating FACS purified CD4+CD25+ cells with ConA and rFIVgag-stimulated CD8+ T cells and measuring IFNgamma by ELISpot. Ab-blocking experiments will be designed to test the hypothesis that Treg cells from FIV-infected cats anergize CD8+ T cells via mTGF-beta signaling. Purified CD4+CD25+ cells +/- anti-TGF-beta and purified CD8+ cells +/- anti-TGF-betaRII will be co-cultured and Treg suppressor function analyzed by measuring IFNgamma by ELISpot. Critical elements in the TGFbeta/ TGF-betaRII signaling pathway leading to suppression of CD8+ cell function will be analyzed, including Smad 2/3 phosphorylation, up-regulation of Foxp3 and Smad 7, and suppression of IFNgamma mRNA production. HIV is currently estimated to infect more than 37 million people world wide. FIV is a well established animal model of HIV/AIDS. The results of these experiments will have implications on future therapeutic strategies against HIV and FIV, and contribute to our current understanding of Treg cell function.

候选人：Jonathan E. Fogle， DVM，在NCSU完成了小动物内科的住院医师，并获得了兽医内科的委员会认证。Fogle博士的短期目标是获得NCSU免疫学项目的博士学位。Fogle博士的长期目标是从他的临床兽医和博士研究经验中继续进行独立的慢病毒研究。建议：很大一部分未经治疗的HIV+患者（进展者）以CD4+和CD8+ T细胞免疫功能障碍为特征。hiv特异性CD8+效应细胞在急性感染阶段被诱导，但在诱导后不久对抗原刺激无反应或“无能”。我们提出CD8+ T细胞的免疫功能障碍是由膜结合的tgf - β (mtgf - β)对病毒激活的CD4+CD25+ T细胞诱导的“克隆能”的结果，这些细胞与靶细胞上的tgf - β接合。这一假设不能在人类实验对象中得到解决，而是需要一个具有良好特征的实验性艾滋病慢病毒动物模型，如FIV。将SPF级猫感染FIV的NCSU1分离物，在感染后的不同时间收集血浆和PBMC/LN细胞，并通过ELISA和Real - Time-PCR定量检测病毒负荷。PBMC/LN表面受体的表型将通过流式细胞术检测，Foxp3 mRNA的表型将通过RT-PCR检测。通过CD4+CD25+细胞耗损和FACS纯化的CD4+CD25+细胞与ConA和rfivgag刺激的CD8+ T细胞孵育，并通过ELISpot测定IFNgamma，来评估Treg细胞的抑制功能。抗体阻断实验将被设计用于验证fiv感染猫的Treg细胞通过mtgf - β信号使CD8+ T细胞失能的假设。将纯化的CD4+CD25+细胞+/-抗tgf - β和纯化的CD8+细胞+/-抗tgf - β共培养，并通过ELISpot检测IFNgamma来分析Treg抑制功能。我们将分析TGFbeta/ TGF-betaRII信号通路中导致CD8+细胞功能抑制的关键因素，包括Smad 2/3磷酸化、Foxp3和Smad 7的上调以及IFNgamma mRNA产生的抑制。据估计，目前全世界有3700多万人感染艾滋病毒。FIV是一种成熟的艾滋病毒/艾滋病动物模型。这些实验的结果将对未来针对HIV和FIV的治疗策略产生影响，并有助于我们目前对Treg细胞功能的理解。