FIV/HIV IMMUNOPATHOGENESIS--A CASE FOR THE CD8+ CELL

CD8+细胞数量的增加是HIV和 FIV感染。在艾滋病毒的情况下，这种不正常的动态平衡被标记为 通过进行性丧失L-选择素(LS)阳性的初始CD8+细胞和 伴随的LS阴性CD8+效应细胞的扩增，从而 CD8+LSneg细胞可能占总数的80%-90% 无症状晚期感染者外周血中CD8+细胞的变化FIV 感染诱导CD8+亚群的进行性扩张 以β链显著减少为特征的。这个CD8+的Betalo 表型可能占全血CD8+细胞总数的90% 在无症状感染的晚期。我们已经利用了测试版 链下调FACStar Sort高度富含CD8+Betalo和 CD8+Betahi表型，并已显示CD8+Betalo表型 合成高水平的IL10和IFNGamma mRNA，并具有强大的抗 FIV活动。此外，我们已经通过2色和3色FAC显示， CD8+Betalo细胞是一种效应表型(CD8+Betalo LSneg CD8+Betahi是一种幼稚的表型(CD8+BetahiLSneg CD44Hi)。这项提议将检验FIV诱导的 CD8+β-LSneg CD44hi效应表型的慢性扩增 不仅具有强大的抗病毒活性，而且显示出异常的组织 贩运和对继发感染的免疫抑制反应。 实验将被设计来进一步表征表型 CD8+β-LSneg亚群及其在淋巴组织中的分布 FIV感染不同阶段的非淋巴组织。荧光 将进行染料标记以评估CD8+的运输潜力 血与LN、血与肺之间的β-LSneg细胞 以应对弓形虫的感染。将设计RT-qcPCR研究 确定CD8+BetaloLSneg表型是否在 以及它是否与病毒载量有关。纯净的 CD8+BetaloLSneg(效应器)和CD8+BetahiLSpos(幼稚)的子集将 用RT-qcPCR法检测细胞因子和趋化因子mRNA以解决 CD8+β-LSneg具有细胞因子/趋化因子的假说 CTL和/或T抑制细胞的轮廓。体外实验将会 确定CD8+β-Sneg细胞是CTL细胞还是病毒抑制细胞 以及它们是否抑制有丝分裂原和抗原特异性回忆反应 由PBMC提供。研究还将解决CD8+BetaloLSneg 细胞对肺巨噬细胞的免疫抑制作用导致 弓形虫细胞因子反应降低和复制失控 速殖子。这些实验将共同检验这一假设 FIV诱导新的CD8+β-LSneg T- 抑制子样活化表型不仅介导免疫 FIV，但由于其改变了组织运输(丢失L-选择素) 并且选择性地招募细胞因子表达谱(例如，IL10) 转移到炎症部位并抑制对继发性 病原体。

Elevation in the number of CD8+ cells is a consistent feature of HIV and FIV infections. In the case of HIV, this abnormal homeostasis is marked by a progressive loss of L-selectin (LS) positive naive CD8+ cells and a concomitant expansion of LS negative CD8+ effector cells, such that the CD8+ LSneg cells may represent 80-90 percent of the total circulating CD8+ cells in the late asymptomatic stage infection. FIV infection induces the progressive expansion of a CD8+ subset characterized by a marked reduction of the beta chain. This CD8+ betalo phenotype may comprise as much as 90 percent of total blood CD8+ cells at late-stage asymptomatic infection. We have taken advantage of beta chain down-regulation to FACStar sort highly enriched CD8+ betalo and CD8+ betahi phenotypes, and have shown that the CD8+ betalo phenotype synthesizes high levels of IL10 and IFNgamma mRNA, and has potent anti- FIV activity. In addition, we have shown by 2- and 3-color FACS that the CD8+ betalo cells is an effector phenotype (CD8+ betaloLSneg CD44hi) and the CD8+ betahi is a naive phenotype (CD8+ betahiLSneg CD44hi). This proposal will test the hypothesis that FIV induces a chronic expansion of a CD8+betaloLSneg CD44hi effector phenotype that not only has potent antiviral activity but exhibits abnormal tissue trafficking and immunosuppressive responses to secondary infections. Experiments will be designed to further characterize the phenotype of this CD8+betaloLSneg subset and its distribution in lymphoid and nonlymphoid tissues at different stages of FIV infection. Fluorescent dye labeling will be done to assess the trafficking potential of CD8+ betaloLSneg cells between blood and LN, and between blood and the lung in response to T. gondii infection. RT-qcPCR studies will be designed to determine if the CD8+betaloLSneg phenotype is regulated at the level of gene transcription and if it is related to virus load. Purified subsets of CD8+betaloLSneg (effector) and CD8+betahiLSpos (naive) will be assayed by RT-qcPCR assays for cytokine and chemokine mRNA to address the hypothesis that the CD8+ betaloLSneg has the cytokine/chemokine profile of a CTL and/or T suppressor cell. In vitro experiments will determine if CD8+ betaloSneg cells are CTL's or virus suppressor cells and whether they suppress mitogen and antigen-specific recall responses by PBMC. Studies will also address the hypothesis that CD8+betaloLSneg cells exert immunosuppressive effects on lung macrophages resulting in decreased cytokine responses and uncontrolled replication of T. gondii tachyzoites. These experiments will collectively test the hypothesis that FIV induces the chronic expansion of a novel CD8+betaloLSneg T- suppressor-like activation phenotype that not only mediates immunity to FIV, but because of its altered tissue trafficking (loss of L-selectin) and cytokine expression profile (e.g. IL10) is selectively recruited into inflammatory sites and suppresses immune responses to secondary pathogens.