BBB Protection in HIV Infection: Barrier-shielding effects of PARP inhibition

描述(由申请人提供)：尽管引入了联合抗逆转录病毒治疗，但艾滋病毒相关神经认知障碍(HAND)的流行率仍然很高。HAND与血液中促炎症因子和激活的感染单核细胞亚群的升高有关，两者都被证明会导致血脑屏障(BBB)损害，从而导致HAND。干扰单核细胞迁移的治疗策略可以减缓艾滋病毒感染和血脑屏障损伤的进展，从而改善手。在之前的资助期间，我们主要研究了(1)血脑屏障损伤的分子机制和(2)糖原合成酶激酶(GSK)3？抑制艾滋病毒感染引起的神经炎症。我们揭示了血脑屏障功能障碍的分子机制[紧密连接(TJ)磷酸化，血脑屏障CD40/CD40配体相互作用，以及HIV对脑内皮细胞直接影响背后的信号事件]。我们演示了GSK3之后的屏障收紧？TJ稳定对脑内皮细胞的抑制作用。我们发现了GSK3的有效抗炎作用？在体外和体内抑制脑内皮细胞(抑制单核细胞迁移、减少炎症因子产生和保护血脑屏障)。葛兰素史克？抑制单核细胞的黏附和迁移，通过抑制小的GTPase rac1，下调整合素的活性表达，保护血脑屏障。然而，在体外或体内，血脑屏障的屏蔽特性或抑制单核细胞迁移/黏附并未完全实现，这表明补充GSK3？的其他途径。是恢复血脑屏障功能所必需的。为了寻找这样的分子，我们将注意力转向聚(ADP-核糖)聚合酶-1(PARP-1)及其抑制剂，它们最近被认为是具有显著神经保护作用的抗炎/免疫调节剂。根据初步数据，我们认为抑制PARP将通过影响单核细胞、脑内皮细胞、激活的小胶质细胞和HIV-1感染的巨噬细胞来减轻HIV-1引起的BBB损伤。事实上，初步数据表明，抑制原代人脑微血管内皮细胞(BMVEC)中的PARP可以改善血脑屏障的完整性，并增强TJ蛋白的表达。PARP抑制剂可阻止炎症因子引起的屏障破坏，减少单核细胞在血脑屏障模型中的黏附/迁移，下调黏附分子/促炎分子，并降低RhoA/rac1的活性。在单核细胞中，PARP抑制剂下调了与RhoA/rac1抑制平行的活性β-整合素。PARP抑制剂减少了人巨噬细胞中促炎分子的表达，并减少了艾滋病毒的复制。虽然PARP抑制剂对免疫细胞的调节作用已有一定的研究，但对其在HIV中枢神经系统感染中的作用尚不清楚。PARP抑制剂现在已经达到了癌症治疗的临床测试阶段，确保了快速过渡到免疫/炎症疾病的治疗。

DESCRIPTION (provided by applicant): Prevalence of HIV-associated neurocognitive disorders (HAND) remains high despite the introduction of combined antiretroviral therapy. HAND is associated with elevation of pro-inflammatory factors in blood and subsets of activated infected monocytes, both shown to cause blood brain barrier (BBB) impairment that contributes to HAND. Therapeutic strategies that disrupt monocyte migration can slow progression of HIV infection and BBB injury, thereby ameliorating HAND. During the previous period of funding, we focused on studies of (1) molecular mechanisms of BBB injury and (2) anti-inflammatory and barrier protective properties of glycogen synthase kinase (GSK) 3? inhibition in neuroinflammation driven by HIV infection. We uncovered molecular mechanisms of BBB dysfunction [tight junction (TJ) phosphorylation, CD40/CD40 ligand interactions at BBB and signaling events behind the direct effects of HIV on brain endothelium]. We demonstrated barrier tightening following GSK3? suppression in brain endothelium due to TJ stabilization. We uncovered potent anti-inflammatory effects of GSK3? inhibition in brain endothelium (suppression of monocyte migration, diminution of inflammatory factor production, and BBB protection) in vitro and in vivo. GSK? inhibition in monocytes attenuated their adhesion/migration across the BBB, down regulated active integrin expression via suppression of the small GTPase, Rac1, and protected the BBB. Yet, BBB shielding properties or inhibition of monocyte migration/adhesion were not fully attained in vitro or in vivo, suggesting that additional pathways complimentary to GSK3? are necessary for the restitution of BBB function. In search of such molecules, we turned our attention to poly(ADP-ribose) polymerase-1 (PARP-1) and its inhibitors, recently recognized as anti-inflammatory/immune modulating agents with significant neuroprotective properties. Based upon preliminary data, we propose that PARP inhibition will attenuate BBB injury caused by HIV-1 via effects on monocytes, brain endothelium, activated microglia and HIV-1 infected macrophages. Indeed, preliminary data indicate that PARP suppression in primary human brain microvascular endothelial cells (BMVEC) improved BBB integrity and augmented expression of TJ proteins. PARP inhibitors prevented barrier disruption caused by inflammatory factors, diminished monocyte adhesion/migration across a BBB model, down regulated adhesion molecules/pro-inflammatory molecules and decreased activity of RhoA/Rac1. In monocytes, PARP inhibitors down regulated the active ?-integrin that paralleled RhoA/Rac1 suppression. PARP inhibitors decreased expression of pro-inflammatory molecules and diminished HIV replication in human macrophages. Although the modulatory effects of PARP inhibitors on immune cells have been studied to some extent, nothing is known about their effects in the setting of HIV CNS infection. PARP inhibitors have now reached the stage of clinical testing for cancer treatment, assuring quick translation to therapy of immune/inflammatory disorders.