Inhibition of CTL killing by HSV

描述：(改编自调查人员摘要)：单纯疱疹病毒 (HSV)编码多个基因，抑制感染细胞的凋亡。病毒 抑制细胞凋亡可能具有复制优势，通过确保 宿主细胞存活到病毒复制完成为止。一种机制，通过它 宿主试图诱导受感染细胞凋亡的作用是通过 细胞毒性T淋巴细胞(Cm)。CTH功能是控制HSV的关键 感染，CU功能受损的患者可能会有严重的HSV 复发和延迟愈合。因此，HSV进化出了 钝化CU的攻击，例如通过1CP47下调MHC I类分子的表达。我们 最近演示了HSV使用的第二种机制来钝化CU的攻击， 其中HSV保护感染细胞免受CTL诱导的DNA片段化。 然而，抑制DNA片段化并不意味着抑制其他 细胞死亡的几个方面(细胞裂解、半胱氨酸氨基转移酶激活等)。此外，CU 根据CU的不同，在目标细胞中触发不同的死亡路径 使用了效应器机构。因此，我们假设虽然单纯疱疹病毒抑制DNA 除碎裂外，铜诱导的细胞死亡的其他方面可能不受影响。 此外，我们假设并不是所有的细胞死亡指标都与 同样有效地防止了病毒在感染细胞中的复制。这个 因此，本项目的目的是评估HSV及其 抗细胞凋亡基因对细胞死亡的影响，经CTL诱导死亡或分离 CTL效应器机构。所生成的数据最终可能允许Rational 单纯疱疹病毒病的免疫调节途径。具体目标1.确定是否 单纯疱疹病毒保护感染细胞免受依赖死亡受体的杀伤机制 由CTL提供。我们的初步数据表明，单纯疱疹病毒感染保护被感染的 LCL或Jurkat细胞的凋亡是通过Fas受体触发的。我们会 因此，测试HSV或其单个抗凋亡基因是否对细胞具有保护作用 由Fas结扎、肿瘤坏死因子或TRAIL引起的死亡，由细胞测量 形态、DNA片段化、51Cr释放、半胱氨酸氨基转移酶激活、线粒体 跨膜电位、感染细胞的克隆性和病毒产量。 特定目标2.确定HSV是否保护受感染的细胞 CTL杀伤作用的溶粒依赖机制。颗粒介导 细胞毒性似乎是CTL杀伤的主要机制 非造血细胞，因此对这种细胞的保护可能对 有效的免疫逃避。具体目标3.确定单纯疱疹病毒是否具有保护作用 感染细胞不受整体CTL损伤和死亡的影响。保护您免受 整个CTL可能最能反映体内的情况。中生成的数据 这一目标将首次全面评估反恐战略的影响。 对成纤维细胞死亡的各个方面的攻击，并将提供一个了解 单纯疱疹病毒感染对这些参数的保护作用。

DESCRIPTION: (Adapted from the Investigator's abstract): Herpes simplex virus (HSV) encodes multiple genes which inhibit apoptosis of infected cells. Viruses which inhibit apoptosis may have a replicative advantage, by ensuring that the host cell lives until viral replication is complete. One mechanism by which the host attempts to induce apoptosis of infected cells is through the action of cytotoxic T lymphocytes (Cm). CTh function is critical for the control of HSV infections, and patients with impaired CU function can have severe HSV recurrences and delayed healing. Therefore, HSV has evolved mechanisms which blunt the attack of CU, such as downregulation of MHC class I via 1CP47. We recently demonstrated a second mechanism used by HSV to blunt the attack of CU, in which HSV protects infected cells from CTL-induced DNA fragmentation. However, inhibition of DNA fragmentation may not imply inhibition of other aspects of cell death (cell lysis, caspase activation, etc.). Furthermore, CU trigger different death pathways in their target cell, depending on the CU effector mechanism used. Therefore, we hypothesize that while HSV inhibits DNA fragmentation, other aspects of cell death induced by CU may remain unaffected. Furthermore, we hypothesize that not all measures of cell death will correlate equally well with the prevention of viral replication in the infected cell. The aims of this project, therefore, are to evaluate the effects of HSV and its anti-apoptosis genes on cell death, after induction of death by CTL or isolated CTL effector mechanisms. The data generated may ultimately allow rational immune modulatory approaches to HSV disease. Specific Aim 1. Determine whether HSV protects infected cells from death-receptor dependent mechanisms of killing by CTL. Our preliminary data demonstrate that HSV infection protects infected LCL or Jurkat cells from apoptosis triggered through the fas receptor. We will therefore test whether HSV or its individual anti-apoptosis genes protect cells from death induced by fas ligation, TNF, or TRAIL, as measured by cell morphology, DNA fragmentation, 51Cr release, caspase activation, mitochondrial transmembrane potential, clonogenicity of infected cells, and viral yield. Specific Aim 2. Determine whether HSV protects infected cells from lytic-granule dependent mechanisms of killing by CTL. Granule-mediated cytotoxicity appears to be the predominant mechanism for CTL killing of nonhematopoietic cells, and thus protection from this may be critical for effective immune evasion. Specific Aim 3. Determine whether HSV protects infected cells from damage and death induced by whole CTL. Protection from whole CTL may most closely reflect the in vivo situation. The data generated in this aim will provide the first comprehensive evaluation of the ramifications of CTL attack on the various aspects of fibroblast death, and will provide an understanding of the protective effect of HSV infection on each of these parameters.