Modeling KSHV Infection and Pathogenesis in SCID Mice

描述（由申请人提供）：体内模型对于理解病毒性疾病中的致病机制以及评价对抗这些疾病的治疗方法至关重要。KSHV已从AIDS患者中发现的B细胞来源的淋巴瘤中分离出来，称为PEL和MCD。目前对KSHV感染的初始细胞靶点以及导致病毒在体内传播和潜伏的细胞还不完全了解。我们推测KSHV感染前体B细胞是肿瘤发生启动的关键过程。已在KS患者的造血祖细胞（CD 34+）中检测到KSHV基因组序列。我们已经证明，KSHV感染人CD 34+细胞和病毒基因表达维持在CFU-GM克隆形成的克隆来自这些细胞在体外分化。用感染的人CD 34+细胞重建淋巴细胞生成后，在人（CD 14+）单核细胞和NOD/SCID-hu嵌合小鼠的B淋巴细胞中检测到KSHV基因组DNA和拉娜-1表达。尽管该模型系统不能完全代表正常人造血和/或KSHV感染的所有方面，但NOD/SCID-hu小鼠中KSHV持续存在和CD 34+细胞感染的组合提供了一种实验系统，其中解决了体内KSHV复制、潜伏期和病理生物学的许多问题。本研究将利用此新的动物模型来检测AZT和IFN-α在体内抑制KSHV感染的功效。该建议旨在进一步完善和发展KSHV感染的NOD/SCID-hu小鼠模型，并更好地了解KSHV感染在CD 34+造血祖细胞中的作用。具体目的是：（1）评价、优化和评估KSHV和rKSHV.219体外从头感染CD 34+细胞的效果。确定KSHV是否在人CD 34+造血祖细胞中建立潜伏感染，并表征病毒感染对造血抑制的作用。(2)通过用KSHV和rKSHV感染的CD 34 + HPC对NOD/SCID小鼠进行免疫重建，确定KSHV细胞嗜性和致瘤潜力。评价和表征KSHV/HHV-8感染对体内造血的影响。(3)确定AZT和IFN α是否诱导rKSHV.219感染的原代造血细胞凋亡。开发NOD/SCID-hu小鼠作为临床前治疗干预模型，以抑制KSHV复制。

DESCRIPTION (provided by applicant): In vivo models are critical to the understanding of pathogenic mechanisms operating in viral diseases, and for evaluation of therapeutic approaches to combat these diseases. KSHV has been isolated from B cell derived lymphomas found in patients with AIDS, termed PEL and MCD. There is an incomplete understanding of the initial cellular targets of KSHV infection and cells that contribute to viral dissemination and latency in vivo. We speculate that KSHV infection of progenitor B cells is key process in the initiation of oncogenesis. KSHV genomic sequences have been detected in hematopoietic progenitor (CD34+) cells from KS patients. We have demonstrated that KSHV infects human CD34+ cells and that viral gene expression is maintained in CFU-GM clonogenic colonies derived from the differentiation of these cells in vitro. KSHV genomic DNA and LANA-1 expression was detected in human (CD14+) monocytes and in B lymphocytes from NOD/SCID-hu chimeric mice following reconstitution of lymphopoiesis with infected human CD34+ cells. Although this model system is not perfectly representative of normal human hematopoiesis and/or all aspects of KSHV infection, the combination of KSHV persistence in NOD/SCID-hu mice and infection of CD34+ cells provides an experimental system in which to address many of the issues of KSHV replication, latency and pathobiology in vivo. This novel animal model will be employed to assay the efficacy of AZT and IFN-alpha in suppressing KSHV infection in vivo. This proposal aims to further refine and develop the NOD/SCID-hu mouse model of KSHV infection and to better understand the role of KSHV infection in CD34+ hematopoietic progenitor cells. Specific Aims are: (1) Evaluate, optimize and assess the effects of KSHV and rKSHV.219 de novo infection of CD34+ cells in vitro. Determine if KSHV establishes a latent infection in human CD34+ hematopoietic progenitor cells and characterize the role of viral infection on suppression of hematopoiesis. (2) Determine KSHV cell tropism and tumorigenic potential by immune reconstitution of NOD/SCID mice with CD34+ HPCs infected with KSHV and rKSHV.219. Evaluate and characterize the effect of KSHV/HHV-8 infection on hematopoiesis in vivo. (3) Determine if AZT and IFNalpha induces apoptosis in primary hematopoietic cells infected with rKSHV.219. Develop the NOD/SCID-hu mouse as a pre-clinical model for therapeutic intervention to inhibit KSHV replication.