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HIV-1 Replication in the Central Nervous System Occurs in Two Distinct Cell Types - May Explain Persistent HIV Brain Damage
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"Less severe neurological problems associated with HIV-1 infection such as minor cognitive impairments may also be increasing, indicating that neurological disorders will remain a problem for HIV-1-infected subjects in the future.......The initiation of antiretroviral therapy results in rapid decay of virus in the blood, which is associated with virus replicating in activated CD4+ T cells; however, HIV-1 in the cerebrospinal fluid (CSF) can decay slowly with the initiation of therapy in some subjects with HAD, suggesting a longer-lived cell type as the origin of this virus.....suggesting that autonomous viral replication is occurring in the CNS of subjects with more severe neurological disease.......Individuals diagnosed with HAD commonly have genetically distinct HIV-1 variants in their cerebrospinal fluid (CSF) that are not detected in the blood virus population, suggesting that independent viral replication is occurring in the CNS of HIV-1-infected subjects with severe neurological disease.....Finally, macrophage-tropic viruses were detected in the CSF but poorly represented in the blood virus population. Our results suggest that HIV-1 variants in the CSF can provide information about independent viral replication in the CNS during the course of HIV-1 infection......Here we show that genetically compartmentalized CCR5-tropic (R5) T cell-tropic and macrophage-tropic HIV-1 populations are independently found in the CSF of subjects diagnosed with HIV-1-associated dementia. Our results demonstrate that HIV-1 can replicate in at least two cell types within the CNS in association with the development of dementia. Macrophage-tropic viruses were poorly represented in the blood population, highlighting the restricted, tissue-specific host range of these variants."
"HIV replication in the CNS can contribute to neurocognitive decline, so the ability to detect features of the CSF viral population associated with viral replication in the CNS may provide new opportunities to guide interventions prior to the development of overt neurological disease"
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To explore these and other questions, Swanstrom and Price of UCSF again will collaborate under a 5-year, $3 million grant from the National Institute of Mental Health to expand the research in HIV patients who don't have dementia and are starting therapy. The new study will look for biomarkers in the CSF in the form of HIV variants or other immune protein information that may predict improvement, stability or decrease in cognitive capacity during therapy.
In the new project, Swanstrom's UNC team will include Joseph J. Eron, MD, professor of medicine and Director of the UNC Center for AIDS Research Clinical Core, Kevin Robertson, PhD, clinical psychologist in the department of neurology, and Angela Kashuba, PharmD, associate professor, Eshelman School of Pharmacy and director, UNC Center for AIDS Research Clinical Pharmacology and Analytical Chemistry Core.
People infected with HIV sometimes delay going on HAART, Swanstrom said. "Our research will help further understand what's going on in the central nervous system of patients who are still alive and in tissue that's accessible in the clinical setting, i.e. CSF. If these individuals knew there was an AIDS virus replicating independently in their CNS, it might affect their decision when to start treatment with HAART."
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HIV-1 Replication in the Central Nervous System Occurs in Two Distinct Cell Types
http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002286
Oct 6 2011 http://www.plospathogens.org
Gretja Schnell1a, Sarah Joseph2, Serena Spudich3b, Richard W. Price3, Ronald Swanstrom1,2,4*
1 Department of Microbiology and Immunology, University of North Carolina at Chapel Hill, School of Medicine, Chapel Hill, North Carolina, United States of America, 2 Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, School of Medicine, Chapel Hill, North Carolina, United States of America, 3 Department of Neurology, University of California at San Francisco, San Francisco, California, United States of America, 4 UNC Center for AIDS Research, University of North Carolina at Chapel Hill, School of Medicine, Chapel Hill, North Carolina, United States of America
Abstract
Human immunodeficiency virus type 1 (HIV-1) infection of the central nervous system (CNS) can lead to the development of HIV-1-associated dementia (HAD). We examined the virological characteristics of HIV-1 in the cerebrospinal fluid (CSF) of HAD subjects to explore the association between independent viral replication in the CNS and the development of overt dementia. We found that genetically compartmentalized CCR5-tropic (R5) T cell-tropic and macrophage-tropic HIV-1 populations were independently detected in the CSF of subjects diagnosed with HIV-1-associated dementia. Macrophage-tropic HIV-1 populations were genetically diverse, representing established CNS infections, while R5 T cell-tropic HIV-1 populations were clonally amplified and associated with pleocytosis. R5 T cell-tropic viruses required high levels of surface CD4 to enter cells, and their presence was correlated with rapid decay of virus in the CSF with therapy initiation (similar to virus in the blood that is replicating in activated T cells). Macrophage-tropic viruses could enter cells with low levels of CD4, and their presence was correlated with slow decay of virus in the CSF, demonstrating a separate long-lived cell as the source of the virus. These studies demonstrate two distinct virological states inferred from the CSF virus in subjects diagnosed with HAD. Finally, macrophage-tropic viruses were largely restricted to the CNS/CSF compartment and not the blood, and in one case we were able to identify the macrophage-tropic lineage as a minor variant nearly two years before its expansion in the CNS. These results suggest that HIV-1 variants in CSF can provide information about viral replication and evolution in the CNS, events that are likely to play an important role in HIV-associated neurocognitive disorders.
Author Summary
Human immunodeficiency virus type 1 (HIV-1) infection of the central nervous system (CNS) can lead to the development of a severe neurological disease termed HIV-1-associated dementia (HAD). Individuals diagnosed with HAD commonly have genetically distinct HIV-1 variants in their cerebrospinal fluid (CSF) that are not detected in the blood virus population, suggesting that independent viral replication is occurring in the CNS of HIV-1-infected subjects with severe neurological disease. We examined HIV-1 variants in the blood plasma and CSF of HAD subjects to determine the viral characteristics associated with the development of dementia during HIV-1 infection. We found that genetically distinct HIV-1 variants in the CSF of HAD subjects were either R5 T cell-tropic or macrophage-tropic. The R5 T cell-tropic viruses required high levels of the cellular surface receptor CD4 to enter cells, while macrophage-tropic viruses could enter cells with low levels of CD4, suggesting that HIV-1 can replicate in at least two cell types within the CNS during the course of dementia. Finally, macrophage-tropic viruses were detected in the CSF but poorly represented in the blood virus population. Our results suggest that HIV-1 variants in the CSF can provide information about independent viral replication in the CNS during the course of HIV-1 infection.
Introduction
Human immunodeficiency virus type 1 (HIV-1) infects CD4+ T cells in the blood and lymphoid organs. In addition, infection of the central nervous system (CNS) can result in mild to severe neurological disease, including HIV-1-associated dementia (HAD) [1]. Although the incidence of HAD and minor cognitive motor disorder have been significantly reduced following the introduction of highly active antiretroviral therapy (HAART), these disorders continue to affect a substantial proportion of the HIV-1-infected population [2], [3]. The insufficient CNS penetration of some antiretroviral drugs or viral resistance may allow HIV-1 to persist in the CNS during the course of therapy [4], [5], [6], [7]. The success of HAART has led to an increased lifespan and an older demographic of HIV-infected subjects, and these subjects in particular have an increased risk of developing HAD due to their enhanced age [8], [9]. Less severe neurological problems associated with HIV-1 infection such as minor cognitive impairments may also be increasing [10], [11], indicating that neurological disorders will remain a problem for HIV-1-infected subjects in the future. Finally, unequal access to HAART and the potential of CNS involvement prior to the initiation of HAART makes the question of HIV replication in the CNS relevant to many infected people.
Several lines of evidence suggest that some HAD subjects can harbor macrophage-tropic HIV-1 variants [12], [13], [14], [15], [16], [17], a distinct phenotype associated with the ability to infect cells with low surface expression of CD4. The initiation of antiretroviral therapy results in rapid decay of virus in the blood, which is associated with virus replicating in activated CD4+ T cells [18], [19]; however, HIV-1 in the cerebrospinal fluid (CSF) can decay slowly with the initiation of therapy in some subjects with HAD, suggesting a longer-lived cell type as the origin of this virus [20], [21], [22]. Macrophage tropism does not appear to be a feature of the transmitted variants of HIV-1 [23], [24], leaving open the question of when and where macrophage-tropic variants of HIV arise and their role in HIV-1-associated pathogenesis.
Previous studies have reported that HIV-1 populations in the CSF of HAD subjects have increased viral genetic compartmentalization compared to virus in the blood [25], [26], and genetically distinct HIV-1 variants have been detected at autopsy in the CNS of some subjects with HAD [27], [28], [29], suggesting that autonomous viral replication is occurring in the CNS of subjects with more severe neurological disease. We examined HIV-1 variants in the CSF of HAD subjects to determine the viral genotypes and phenotypes associated with the development of HAD. Here we show that genetically compartmentalized CCR5-tropic (R5) T cell-tropic and macrophage-tropic HIV-1 populations are independently found in the CSF of subjects diagnosed with HIV-1-associated dementia. Our results demonstrate that HIV-1 can replicate in at least two cell types within the CNS in association with the development of dementia. Macrophage-tropic viruses were poorly represented in the blood population, highlighting the restricted, tissue-specific host range of these variants.
Discussion
We have demonstrated that genetically compartmentalized R5 T cell-tropic and macrophage-tropic HIV-1 populations are independently found in the CSF of subjects diagnosed with HIV-1-associated dementia. Our study was limited to a cohort of eight subjects with neurological disease who were receiving lumbar punctures at the time of HAD diagnosis and while initiating therapy, with samples available prior to therapy in two subjects followed longitudinally. In spite of a relatively small sample size we were able to link several features that distinguish two different virological states associated with severe neurological dysfunction. Compartmentalized HIV-1 populations in the CSF with R5 T cell-tropic entry phenotypes were separated phylogenetically from plasma virus populations, and were associated with clonal amplification of the CSF viral population. Replication in CD4+ T cells is consistent with both the rapid decay of compartmentalized virus in the CSF after the initiation of therapy in these subjects, and the migration of immune cells into the CNS/CSF as indicated by the presence of elevated CSF pleocytosis. Compartmentalized macrophage-tropic HIV-1 populations were associated with more genetically diverse viral populations in the CSF, and the slow decay of virus in the CSF after the initiation of therapy, indicative of viral replication in a long-lived cell. However, given the small sample size we cannot provide an accurate estimate of the relative frequency of each type of virologic state other than to note that they appeared with similar frequency in this cohort of eight subjects (three with compartmentalized R5 T cell-tropic virus, five with compartmentalized macrophage-tropic virus). Overall we detected a significantly compartmentalized CSF population in seven of these eight subjects suggesting that virus outgrowth in the CNS, whether macrophage-tropic or R5 T cell-tropic, will be a feature in a majority of HAD cases.
HIV-1 replication in the CNS is thought to occur in perivascular macrophages and/or microglia within the brain parenchyma [13], [35], [36]. We found that both R5 T cell-tropic and macrophage-tropic HIV-1 populations are independently associated with clinical dementia. This indicates a more complex interaction between HIV-1 and the CNS since the genetically compartmentalized R5 T-cell tropic viruses are unlikely to be replicating in macrophages or microglia given their requirement for high levels of CD4 to enter target cells. During simian immunodeficiency virus (SIV) infection of macaques, CNS infection is associated with the presence of infiltrating SIV-specific CD8+ T cells in the brain, but infiltrating CD4+ T cells have not been detected [37]. Trafficking of CD4+ T cells has been reported in the CNS during infection of other neurotropic viruses [38], [39]. We propose that the presence of viral antigen, especially during periods of increased HIV-1 replication in the CNS/CSF compartment, could drive the migration of both CD8+ and CD4+ T cells into the CNS/CSF (resulting in elevated CSF pleocytosis) and lead to persistence of compartmentalized virus through replication in the CD4+ T cells, and thus the apparent loss of this cell type. Consistent with the loss of these cells is the rapid decay of virus in the CSF during the initiation of therapy, which is considered a marker of viral replication in activated T cells [18], [19].
The pathological determinants of HAD are poorly understood. Some subjects with dementia exhibit HIV-1 encephalitis (HIVE) characterized by the presence of multinucleated giant cells of the macrophage/microglia origin and immunohistochemical evidence of viral replication [40], [41]. Although the incidence of HIVE has decreased during the HAART era, neuropathological changes in brain tissue, including glial activation and monocyte/macrophage infiltration [3], [42], [43], are still common. Future studies examining HIV-1 populations in paired blood, CSF, and brain tissue from HAD subjects with and without neuropathological findings will help determine whether there are physiological differences in brain pathology between subjects with R5 T cell-tropic versus macrophage-tropic HIV-1 variants as the predominant CSF population. Also, the appearance of macrophage-tropic viruses largely restricted to the CNS/CSF compartment is most consistent with the appearance of these viruses late in the infection time course of HIV-1, representing an expanded host range of the virus that is initially replicating in activated T cells. Although severe neurological disease associated with HIV-1 infection has declined in the HAART era, milder forms of neurological disease are increasing. In this study we detected a significantly compartmentalized macrophage-tropic HIV-1 population in the CSF of one subject with more mild neurological dysfunction (subject 4013; Table 1), illustrating the potential importance of understanding the correlates of HIV-1-associated neurological dysfunction with CNS/CSF viral population phenotypes.
HIV replication in the CNS can contribute to neurocognitive decline, so the ability to detect features of the CSF viral population associated with viral replication in the CNS may provide new opportunities to guide interventions prior to the development of overt neurological disease. In our study, one subject with longitudinal sampling (subject 7115) had macrophage-tropic variants present as a minor CSF population prior to the diagnosis of severe dementia (Figure 5). The application of sequencing technologies with greater capacity to sample a large number of viral genomes would allow the identification of minor CSF population variants, but this approach would rely on genotypic determinants of macrophage tropism rather than the phenotypic determinants used in our study. Several sequence determinants in env have been reported to be associated with macrophage tropism [27], [44], [45]; however, none of these determinants distinguishes the CSF-derived macrophage-tropic viruses from the paired blood-derived T cell-tropic viruses for the subjects in our study. Thus, the evolution of macrophage-tropic virus likely occurs through multiple pathways that will require a larger catalog of env sequences to allow reliable genotypic identification. It remains a possibility that the clonal amplification of R5 T cell-tropic viruses we detected in three HAD subjects is obscuring a smaller population of macrophage-tropic CNS virus, a question that could be addressed using more sensitive sampling methods. Developing a more complete understanding of the virological markers of CNS replication, and utilizing deep sequencing technologies to find minor populations, will provide opportunities to examine the use of CSF for information about viral replication in the CNS as a potential predictor of neurological involvement in the pathogenic process.
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