Functional Studies of HIV-Tat Effects on Endothelium

描述（由申请人提供）：最初的提案侧重于HIV-1与大血管脐静脉和骨髓内皮的相互作用。虽然在实现目标方面取得了进展，但很明显，HIV-1的病理生理是在微血管内皮中，特别是淋巴起源。这是因为艾滋病的原发性内皮改变发生在卡波西肉瘤（KS）的发生过程中，其细胞起源是淋巴细胞。因此，我们将工作重点转移到富含淋巴细胞的真皮微血管内皮（DMEC）上。另一个焦点转移是KSHV/HHV8。这种伽玛疱疹病毒似乎是KS的主要病原。它的包膜糖蛋白gB与DMEC表面的alpha3beta1整合素结合。为了建立更接近模拟HIV-1和KSHV/HHV8共同感染的体内发病机制的体外系统，我们开始研究tat和gB对DMEC的影响。我们发现，淋巴内皮中标志性的VEGF受体VEGFR3/FLT4与gB连接alpha3beta1整合素相关，并在缺乏同源VEGF配体的情况下被激活。此外，gB或通过VEGFR3介导DMEC迁移和增殖。这些关于alpha3beta1整合素与VEGFR3关联的数据表明，VEGFR3可能作为KSHV的辅助受体，或者其信号传导促进KSHV进入。新的特异性目标解决了这一假设，即它和gB可能在改变内皮细胞功能和KSHV感染方面具有协同作用：(1)进一步表征KSHV gB激活alpha3beta1整合素如何激活VEGFR3；(2)研究VEGFR3是否参与KSHV进入微血管内皮细胞；(3)表征HIV- 1与KSHV gB的功能相关的合作相互作用。这些研究旨在提供艾滋病共感染宿主的新信息，从而为设计KS的预防或治疗策略提供基础。

DESCRIPTION (provided by applicant): The initial proposal focused on interactions of HIV-1 tat with large vessel umbilical vein and marrow endothelium. While progress was made in fulfilling the aims, it became clear that the pathophysiology of HIV-1 tat was in microvascular endothelium, particularly that of lymphatic origin. This was because the primary endothelial alteration in AIDS is in the genesis of Kaposi's sarcoma (KS), whose cell of origin is lymphatic. Thus, we shifted the focus of our work to dermal microvascular endothelium (DMEC) that are enriched with cells of lymphatic origin. The other shift in focus was to KSHV/HHV8. This gamma herpes virus appears to be the key etiological agent in KS. Its envelope glycoprotein, gB, binds to the alpha3beta1 integrin on the surface of DMEC. To develop in vitro systems that more closely model in vivo pathogenesis in coinfection with HIV-1 and KSHV/HHV8, we began to study the effects of tat and gB on DMEC. We found that VEGFR3/FLT4, the signature VEGF receptor in lymphatic endothelium, associated with, and was activated upon, alpha3beta1 integrin ligation by gB or tat in the absence of cognate VEGF ligands. Furthermore, gB or tat caused DMEC migration and proliferation via VEGFR3. These data on the association of the alpha3beta1 integrin with VEGFR3 suggested that VEGFR3 may either act as a coreceptor for KSHV or that its signaling facilitates KSHV entry. The new specific aims address the hypothesis that tat and gB may have cooperative effects on altering endothelial cell function and KSHV infection: (1) to further characterize how alpha3beta1 integrin activation by KSHV gB can activate VEGFR3; (2) to investigate whether VEGFR3 is involved in entry of KSHV into microvascular endothelial cells; and (3) to characterize functionally relevant cooperative interactions of HIV- 1 tat and KSHV gB. These studies are designed to provide new information on the coinfected host with AIDS and thereby provide a foundation for the design of preventive or therapeutic strategies against KS.