Resolving Central Nervous System Inflammation in Acquired Immunodeficiency Syndrome

Abstract

Understanding the persistence of viral reservoirs despite durable antiretroviral therapy (ART) is essential to addressing the challenge of viral clearance and chronic immune activation with human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV). It had not previously been demonstrated that HIV or SIV- infected macrophage traffic out of the CNS to reseed the periphery, potentially contributing to viral recrudescence. This thesis proposes the central hypothesis that persistent traffic of monocytes and macrophages out of the CNS and subsequent viral reseeding of the periphery plays a key role in viral dissemination, particularly in the context of acquired immunodeficiency syndrome (AIDS), with ART, and following ART interruption. In Chapter 2, utilizing Superparamagnetic iron oxide nanoparticles (SPION) as a novel in vivo method to label CNS macrophages, we demonstrate that under normal conditions, CNS macrophages migrate out to the deep cervical lymph nodes. However, during SIV infection, we observe an accumulation of macrophages within the CNS and a reduction in traffic out to the periphery. Importantly, with SIV-infection, we found that SIV-infected macrophages traffic out to deep cervical lymph nodes. From these, we find that under normal conditions, macrophages traffic out of the CNS. However, during SIVinfection, macrophages are retained within the CNS, contributing to inflammation in the brain, and those that do migrate out are virally infected. In Chapter 3, we hypothesized that ART restores CNS macrophage traffic and prevents viral dissemination from the CNS reservoir by eliminating the traffic of virally infected macrophage out of the CNS, as seen with AIDS and SIV-induced encephalitis (SIVE). We also hypothesized that following four weeks of ART interruption there would be expansion of the CNS viral reservoir with traffic out of virally infected macrophages to the deep cervical lymph node. Utilizing a rapid AIDS model with CD8 depletion to induce a high incidence of SIVE and intracisternal injection of SPION, we found that SIV-infected macrophages accumulate in the perivascular space, meninges, choroid plexus, and traffic out at a low rate to the deep cervical lymph node, spleen, and even to the dorsal root ganglia (DRG). With ART, we found clearance of virally infected macrophages in the brain perivascular space but not in the meninges or choroid plexus. Importantly following four weeks of ART interruption, the perivascular space remained clear of virus but there was a rebound in the meninges and scattered virally infected macrophages in the choroid plexus. With ART and following a brief ART interruption, there was no traffic of CNS virally infected macrophages out to the deep cervical lymph node, spleen, or DRGs. These data demonstrate that ART effectively clears virus-infected perivascular macrophages and eliminates the traffic of virus-infected macrophages out of the CNS to the deep cervical lymph node and spleen but does not eliminate virally infected macrophages in the meninges or choroid plexus. By using two differently colored SPION injected early and late, we observed an increase in early SPION+ macrophages within and outside the CNS with SIVE, ART, and ART interruption, indicating that SIV-infected perivascular macrophages establish an early viral reservoir with ongoing seeding in the meninges and choroid plexus throughout infection. These findings are consistent with the retention of CNS macrophages in the presence of inflammation and viral infection, as well as the potential for viral rebound in the CNS from sources such as the blood, meninges, and choroid plexus with ART and following ART interruption. In Chapter 4, we propose a novel pathway for virus-infected macrophages to traffic out of the CNS via cranial and spinal nerves. Due to the persistence of virally infected macrophages in the meninges with durable ART and continuity of the CNS meninges with peripheral nerves, we hypothesize that virally infected macrophage traffic out of the CNS via cranial and peripheral nerves with AIDS and SIVE, on ART, and following ART interruption. To test this hypothesis, we tracked SPION+ macrophages by quantifying them at central (spinal cord and cranial nerves) and peripheral sites (dorsal root ganglia, DRG). Similar to our previous findings in the brain, SIV infection increased the numbers of macrophages in the spinal cord and decreased them in peripheral sites. Staining for viral RNA and GP41 identified virus-infected SPION+ macrophages in cranial nerves and DRG, which were significantly reduced but not eliminated by ART. In animals with AIDS, late- and dual-labeled SPION+ macrophages decreased, suggesting reduced macrophage trafficking late in infection. ART appeared to restore traffic, as higher numbers of late- and dual-labeled macrophages were observed, though this reversed to levels seen in AIDS/SIVE upon ART interruption. Our findings reveal a previously understudied pathway that allows CNS macrophage viral reservoirs to reseed virus to the periphery, a process that persists despite ART. In Chapter 5, we performed a literature review to better understand the effects of HIV infection on aging, as age is a primary risk factor for the development of HIV-associated neurocognitive disorders and HIV-associated sensory neuropathy. With ART extending the lifespan of people living with HIV, they now also experience accelerated aging, leading to earlier onset of age-related conditions like cardiovascular disease and neurocognitive disorders. Evidence suggests this is due to chronic immune activation, co-infections, and possibly ART itself. HIV and aging both alter immune cell populations, increasing inflammatory markers and contributing to "inflamm-aging." While ART slows this acceleration, it cannot prevent aging or related comorbidities. This thesis explores the role of macrophage traffic from the CNS and its contribution to the spread of the virus to peripheral tissues. To investigate this, we utilized a novel in vivo labeling method to track CNS macrophages, identify migration out of the CNS, and evaluate how ART and ART interruption influence the traffic of virally infected macrophages to peripheral tissues. Our findings underscore the role of CNS macrophages in the resolution of inflammation by trafficking out of the CNS, viral rebound from blood- derived sources following ART interruption, and the role of perineural pathways in viral dissemination even with durable ART.