Immune checkpoints in the CNS and HIV-associated neurocognitive disorder

摘要 免疫检查点（ICP）对免疫防御、监视、免疫应答和免疫应答发挥抑制或刺激作用， 调节和通过其配体-受体相互作用的自身耐受性。ICP存在于两种膜结合的 和可溶性形式。抑制性和刺激性膜结合ICP（mICP）之间的不平衡 外周血和组织中的恶性细胞和免疫细胞已被充分证明。封锁 抑制性mICP，如CTLA-4，PD-1和PD-L1，已成为一种革命性的治疗先进的 恶性肿瘤如黑色素瘤的阶段。然而，两者的起源，调节和生物学意义 中枢神经系统（CNS）中的mICP和可溶性ICP（sICP）尚未被系统地 研究了我们使用多重免疫测定同时定量16个sICP的浓度， 来自33名HIV阴性个体（HNI）和105名HIV感染者的脑脊液（CSF）样本 （PLWH），其中伴HAND者80例，不伴HAND者25例。 我们发现10种sICP（sHVEM、sCD 27、sGITR、sICOS、sLAG-3、sPD-1、sTIM-3、sBTLA、sCD 28和sCD 40） 在所有CSF样本中一致检测到，并且其中大多数在PLWH中高度升高。引人注目的是， 与无HAND的PLWH相比，HAND的CSF sHVEM显著增加。这10个sICP 在恒河猴（RM）的CSF样品中也可检测到。免疫组织化学（IHC）分析 来自有和没有猴免疫缺陷病毒的RM的脑组织（额叶皮质和海马） (SIV)感染表明mHVEM在神经元上以基础水平表达，但在星形胶质细胞或 在SIV感染的RM（SIV/RM）中，mHVEM表达高度增加。而且我们 最近报道，PD-L1在U87 MG细胞（一种人星形胶质细胞系）中的表达可以被稳健地抑制， IFN-γ/IL-1β/TNF-α的混合物诱导，诱导的PD-L1控制MCP-1的产生（最多 调节单核细胞/巨噬细胞迁移的重要趋化因子）。因此，在本发明中， 我们假设sICP和mICP在PLWH的CNS中均失调，这与 手神经发病机制。我们有两个具体的目的来检验这个中心假设：（1）确定sICP PLWH和SIV/RM在CNS中的概况和生物学功能，以及（2）表征细胞小生境 ICP受体/配体阳性神经细胞和浸润性免疫细胞在脑中的空间联系， PLWH和SIV/RM。我们的研究结果将为深入了解ICP在CNS神经免疫中的作用提供参考。 有和没有HAND的PLHIV的失调和神经炎症。sICP的CSF水平和特征 例如sHVEM可以潜在地用作HAND严重性和进展的生物标志物。

Abstract Immune checkpoints (ICPs) exert inhibitory or stimulatory effects on immune defense, surveillance, regulation, and self-tolerance through their ligand-receptor interactions. ICPs exist in both membrane-bound and soluble forms in vivo. Imbalances between inhibitory and stimulatory membrane-bound ICPs (mICPs) in malignant cells and immune cells in the peripheral blood and tissues have been well documented. Blockade of inhibitory mICPs such as CTLA-4, PD-1, and PD-L1 has become a revolutionary treatment for advanced stages of malignancies such as melanoma. However, the origin, regulation, and biological significance of both mICPs and soluble ICPs (sICPs) in the central nervous system (CNS) have not yet been systematically studied. We used a multiplex immunoassay to simultaneously quantify the concentrations of 16 sICPs in cerebrospinal fluid (CSF) samples from 33 HIV-negative individuals (HNIs) and 105 people living with HIV (PLWH), including 80 cases with HIV-associated neurocognitive disorder (HAND) and 25 cases without HAND. We found that 10 sICPs (sHVEM, sCD27, sGITR, sICOS, sLAG-3, sPD-1, sTIM-3, sBTLA, sCD28, & sCD40) were consistently detected in all CSF samples, and most of them were highly elevated in PLWH. Strikingly, CSF sHVEM was significantly increased in HAND when compared to PLWH without HAND. These 10 sICPs were also detectable in the CSF samples from rhesus macaques (RMs). Immunohistochemistry (IHC) analysis of brain tissues (frontal cortex and hippocampus) from RMs with and without simian immunodeficiency virus (SIV) infection showed that mHVEM was expressed at a basal level on neurons, but not on astrocytes or microglia, and mHVEM expression was highly increased in SIV-infected RMs (SIV/RMs). Furthermore, we have recently reported that PD-L1 expression in U87MG cells (a human astrocyte cell line) can be robustly induced by a mixture of IFN-γ/IL-1β/TNF-α, and induced PD-L1 controls production of MCP-1 (the most important chemokine that regulates migration of monocytes/macrophages) via PD-L1 intrinsic signaling. Thus, we hypothesize that both sICPs and mICPs are dysregulated in the CNS of PLWH, which are associated with HAND neuropathogenesis. We have two Specific Aims to test this central hypothesis: (1) To determine sICP profiles and biological functions in the CNS of PLWH and SIV/RMs, and (2) To characterize the cellular niches and spatial associations of ICP receptor/ligand-positive neural cells and infiltrating immune cells in the brain of PLWH and SIV/RMs. Our results will provide insights into the roles of ICPs in the CNS neuroimmune dysregulation and neuroinflammation of PLHIV with and without HAND. The CSF levels and profiles of sICPs such as sHVEM can potentially be used as a biomarker for HAND severity and progression.