Role Of Monocytes In AIDS And As Targets For Antiviral Therapy

对艾滋病毒的先天防御 粘液相关淋巴组织是HIV早期感染和疾病进展的主要靶点，也可以在高效抗逆转录病毒治疗期间提供病毒安全港。在这些组织中，扁桃体作为逆转录病毒感染的主要和/或次要部位的地位仍然是个谜，特别是因为口服HIV传播似乎很罕见。要剖析的机制，潜在的可访问性和易感性，艾滋病毒在这个隔间，我们解剖的上皮激光捕获显微切割（LCM），并进行微阵列基因表达分析。我们的研究表明扁桃体上皮是一个独特的部位，与其他口腔上皮相比，扁桃体上皮显示出更高的HIV传播潜力，这可能部分是由于HIV辅助受体CXCR 4增加和抗病毒SLPI减少。为了确定有利于扁桃体淋巴细胞作为病毒库的选择性因素，我们比较了分离的扁桃体细胞与血液淋巴细胞的易感性。在这些研究中，我们确定了扁桃体微环境的几个独特的方面，可能支持其允许的性质，包括与PBMC相比，Th 2细胞因子水平升高和Th 1细胞因子水平有限。尽管抗病毒IFN水平升高，扁桃体在体外暴露于HIV后不能产生有效的抗病毒应答，也不能明显地在体内产生，这可能反映了IFN信号传导的负调节因子（如SOCS）的组成型表达。SOCS似乎有助于阻断扁桃体中的IFN信号级联，这通过Stat 1的磷酸化减少而明显，这可能是IFN信号的局部脱离和抗病毒活性降低的原因。在循环模式中，SOCS 3也可以促进Th 2极化。这种免疫调节的特征与控制免疫激活的必要性是一致的，但在不断受到抗原轰击的环境中保持准备状态。在正在进行的研究中，正在分离和研究产生大量HIV的扁桃体多核和单核细胞，以及Treg作为病毒调节剂和靶点的潜在参与。进一步了解这些人群和免疫调节途径，影响他们的易感性，艾滋病病毒的发病机制是重要的保护脆弱的粘膜隔室作为水库的病毒。 在早期的研究中，我们证明了一种合成的三萜类化合物和过氧化物酶体增殖物激活受体γ（PPARgamma）配体，2-氰基-3，12-二氧代齐墩果-1，9-二烯-28-酸（CDDO），已知会影响p21激酶抑制剂表达（与病毒生命周期相关），抑制巨噬细胞和PBMC中的病毒复制。最近，一种新的甲基酯衍生物的CDDO（CDDO-Me），这是唯一的口服生物利用度，已被合成，并在初步的I期试验中，癌症患者，它已被证明没有显着的毒性，与测试这种药剂在HIV患者的可能性一致。相当令人感兴趣的是这种甲酯衍生物靶向组织而不是血液的证据，因此我们专注于定义其在扁桃体组织衍生的CD 4 + HIV靶细胞中抑制HIV的能力。由于抗逆转录病毒疗法通常以高毒性为特征，并且经常导致耐药性病毒株的出现，因此鉴定出靶向宿主细胞分子的新的抗病毒剂，其可以独立使用或与当前的抗病毒药物联合使用，这将为治疗HIV感染提供额外的资源。 在其他临床研究中，我们一直与NIAID的研究人员合作，用IFN治疗艾滋病患者，试图确定IFN抑制HIV负荷的抗病毒机制。与我们的体外数据一致，IFN有效地上调胞苷脱氨酶家族的先天性细胞内蛋白质的成员，我们表明，这也发生在治疗患者的PBMC。载脂蛋白B mRNA编辑酶催化多肽样3G（APOBEC 3G）是一种对HIV具有致死活性的胞苷脱氨酶，被包装到子代病毒体中并导致HIV cDNA降解。作为反击，HIV病毒体感染因子（Vif）靶向APOBEC 3G进行蛋白酶体蛋白水解，从而排除其掺入出芽病毒体。我们发现IFN治疗增加了APOBEC 3G和其他家族成员，同时降低了患者的病毒载量。病毒下降的最大幅度发生在治疗后的第一周，特别是在病毒载量较低的患者中，与早期蛋白酶抑制剂相当。因此，可能需要进一步考虑IFN在低病毒载量情况下作为诱导治疗的作用，并且正在进行其他临床研究。此外，通过使用微阵列，我们已经确定了其他分子的干扰素，可能有助于其抗病毒作用的目标，并可能被视为额外的干预措施。

Innate defense against HIV Mucosal associated lymphoid tissues are major targets of HIV during early infection and disease progression, and can also provide a viral safe haven during highly active antiretroviral therapy. Among these tissues, the tonsils remain enigmatic regarding their status as primary and/or secondary sites of retroviral infection, particularly since oral HIV transmission appears rare. To dissect the mechanisms underlying accessibility and susceptibility to HIV in this compartment, we dissected the epithelium with laser capture microdissection (LCM) and performed microarray gene expression analyses. Our studies demonstrated that the tonsil epithelium is a unique site and exhibits heightened potential for HIV transmission compared to other oral epithelia, likely due in part, to increased HIV co-receptor CXCR4 and reduced antiviral SLPI. To identify selective factors that favor the tonsil lymphoid cells as a viral reservoir, we compared isolated tonsil cell susceptibility to that of blood lymphocytes. In these studies, we identified several unique aspects of the tonsil micromilieu that may support its permissive nature, including elevated levels of Th2 cytokines and limited Th1 cytokines compared with PBMC. Despite elevated levels of antiviral IFN, the tonsil was not able to mount an effective antiviral response after exposure to HIV in vitro, nor apparently in vivo, which may reflect constitutive expression of negative regulators of IFN signaling, such as SOCS. SOCS appear to contribute to the blockade of IFN signaling cascades in the tonsil, evident by the reduced phosphorylation of Stat1, which may account for the local disengagement of IFN signaling and reduced antiviral activity. In a cyclic pattern, SOCS3 may also promote Th2 polarization. Such a profile of immune regulation is consistent with the necessity to control immune activation, yet maintain a state of readiness in an environment constantly bombarded with antigens. In ongoing studies, the tonsil multinucleated and mononuclear cells that produce striking amounts of HIV are being isolated and studied, as is the potential involvement of Treg as regulators and targets for the virus. Further understanding of these populations and the immunoregulatory pathways that influence their susceptibility to HIV pathogenesis are important for protecting vulnerable mucosal compartments from serving as reservoirs for the virus. In earlier studies, we demonstrated that a synthetic triterpenoid and peroxisome proliferator-activated receptor gamma (PPARgamma) ligand, 2-cyano-3,12-dioxooleana-1,9-dien-28-oic acid (CDDO), known to influence p21 kinase inhibitor expression (linked to viral life cycle), suppressed viral replication in macrophages and PBMC. Recently, a new methyl ester derivative of CDDO (CDDO-Me), which is uniquely orally bioavailable, has been synthesized and in preliminary phase I trials in cancer patients, it has been shown to not have significant toxicities, consistent with the possibility of testing this agent in HIV patients. Of considerable interest is the evidence that this methyl ester derivative targets the tissues, rather than the blood, and therefore we have focused on defining its ability to inhibit HIV in tonsil tissue derived CD4+ HIV target cells. As anti-retroviral therapy is often characterized by high toxicity and frequently results in the emergence of drug resistant virus strains, the identification of new anti-viral agents targeting host cell molecules that can be used independently or in conjunction with current anti-viral drugs will provide additional resources in the treatment of HIV infection. In additional clinical studies, we have been collaborating with investigators at NIAID in the treatment of AIDS patients with IFN in an attempt to define the antiviral mechanisms by which IFN suppresses HIV burden. In concordance with our in vitro data that IFN potently upregulates members of the cytidine deaminase family of innate intracellular proteins, we show that this also occurs in the PBMC of treated patients. Apolipoprotein B mRNA-editing enzyme-catalytic polypeptide-like 3G (APOBEC3G), a cytidine deaminase with lethal activity against HIV is packaged into progeny virions and leads to HIV cDNA degradation. As a counterattack, HIV virion infectivity factor (Vif) targets APOBEC3G for proteasomal proteolysis, thus excluding its incorporation into budding virions. We found that IFN treatment increased APOBEC3G and other family members, while reducing patient viral load. The maximal magnitude of viral decline occurred during the first week after treatment, especially in patients with lower viral load, comparable to that of early protease inhibitors. Thus, further consideration of a role for IFN as an inductive therapy in the context of low viral loads may be warranted and additional clinical studies are in progress. Moreover, through the use of microarrays, we have identified additional molecular targets of IFN that may contribute to its antiviral effects and may be considered as additional intervention approaches.