Deep Sequencing Reveals Central Nervous System Compartmentalization in Multiple Transmitted/Founder Virus Acute HIV-1 Infection.

HIV-1 disseminates to a broad range of tissue compartments during acute HIV-1 infection (AHI). The central nervous system (CNS) can serve as an early and persistent site of viral replication, which poses a potential challenge for HIV-1 remission strategies that target the HIV reservoir. CNS compartmentalization is a key feature of HIV-1 neuropathogenesis. Thus far, the timing of how early CNS compartmentalization develops after infection is unknown. We examined whether HIV-1 transmitted/founder (T/F) viruses differ between CNS and blood during AHI using single-genome sequencing of envelope gene and further examined subregions in pol and env using next-generation sequencing in paired plasma and cerebrospinal fluid (CSF) from 18 individuals. Different proportions of mostly minor variants were found in six of the eight multiple T/F-infected individuals, indicating enrichment of some variants in CSF that may lead to significant compartmentalization in the later stages of infection. This study provides evidence for the first time that HIV-1 compartmentalization in the CNS can occur within days of HIV-1 exposure in multiple T/F infections. Further understanding of factors that determine enrichment of T/F variants in the CNS, as well as potential long-term implications of these findings for persistence of HIV-1 reservoirs and neurological impairment in HIV, is needed.

Effects of HIV-1 Tat on oligodendrocyte viability are mediated by CaMKIIß-GSK3ß interactions.

Myelin disruptions are frequently reported in human immunodeficiency virus (HIV)-infected individuals and can occur in the CNS very early in the disease process. Immature oligodendrocytes (OLs) are quite sensitive to toxic increases in [Ca2+ ]i caused by exposure to HIV-1 Tat (transactivator of transcription, a protein essential for HIV replication and gene expression), but sensitivity to Tat-induced [Ca2+ ]i is reduced in mature OLs. Tat exposure also increased the activity of Ca2+ /calmodulin-dependent kinase IIß (CaMKIIß), the major isoform of CaMKII expressed by OLs, in both immature and mature OLs. Since CaMKIIß is reported to interact with glycogen synthase kinase 3ß (GSK3ß), and GSK3ß activity is implicated in OL apoptosis as well as HIV neuropathology, we hypothesized that disparate effects of Tat on OL viability with maturity might be because of an altered balance of CaMKIIß-GSK3ß activities. Tat expression in vivo led to increased CaMKIIß and GSK3ß activity in multiple brain regions in transgenic mice. In vitro, immature murine OLs expressed higher levels of GSK3ß, but much lower levels of CaMKIIß, than did mature OLs. Exogenous Tat up-regulated GSK3ß activity in immature, but not mature, OLs. Tat-induced death of immature OLs was rescued by the GSK3ß inhibitors valproic acid or SB415286, supporting involvement of GSK3ß signaling. Pharmacologically inhibiting CaMKIIß increased GSK3ß activity in Tat-treated OLs, and genetically knocking down CaMKIIß promoted death in mature OL cultures treated with Tat. Together, these results suggest that the effects of Tat on OL viability are dependent on CaMKIIß-GSK3ß interactions, and that increasing CaMKIIß activity is a potential approach for limiting OL/myelin injury with HIV infection.

Productive infection of human neural progenitor cells by R5 tropic HIV-1: opiate co-exposure heightens infectivity and functional vulnerability.

OBJECTIVE: HIV type-1 (HIV-1) causes a spectrum of central nervous system (CNS) complications; many are worsened by opiate co-exposure. Human neural progenitor cells (hNPCs) give rise to all CNS neurons and macroglia. We tested the hypothesis that hNPC maturation and fate are altered by HIV and opiates, contributing to HIV-1-related neuropathology. Reports of hNPC infection remain controversial. We rigorously examined this question, testing whether hNPCs propogated infection, and whether HIV affected hNPCs absent their infection. DESIGN AND METHODS: Primary hNPCs were characterized over multiple passages. Following R5 HIV-1BaL exposure, p24, Nef, and tat assays monitored infection; a serial dilution approach tested infection transfer to naive hNPCs. Bromodeoxyuridine uptake, population doubling time, and immunostaining assessed proliferation and differentiation. Morphine co-exposure assessed opiate interactions. Supernatant from HIV-1BaL-infected PBMCs (HIVsup), HIV-1BaL, and ultraviolet light-inactivated HIVsup were compared to test effects of inflammatory milieu versus virus or infection per se. RESULTS: The hNPCs (CD4/CD8/Iba/CXC3CL1/CD11b) were infectable and could transfer infection to naive hNPCs. Infection was partly blocked by maraviroc, implicating CCR5. HIVsup reduced hNPC proliferation and caused premature differentiation into neurons/astroglia. Effects on proliferation were due to soluble factors/viral proteins, not infection per se. Morphine co-exposure exacerbated certain functional consequences of HIVsup, and sustained the infection of hNPCs. CONCLUSION: hNPCs can be infected and propagate virus in vitro. hNPCs or their progeny may represent an underappreciated viral reservoir. Factors from infected cells alter hNPC proliferation and neural cell maturation, which likely compromises CNS structure and function. Morphine-HIV interactions may worsen dysfunction and sustain infection.

Differential expression of the alternatively spliced OPRM1 isoform µ-opioid receptor-1K in HIV-infected individuals.

OBJECTIVE: We previously examined the expression of specific C-terminal µ-opioid receptor (MOR) splice variants in human central nervous system cell types and HIV-infected brain tissue from individuals with neurocognitive impairment ±â€ŠHIV encephalitis (HIVE). In the present study, we examined the N-terminal splice variant MOR-1K, which mediates excitatory cellular signaling. METHODS AND RESULTS: We found segregation of expression ranging from undetectable to seemingly exclusive across nervous system cell types compared to the pool of C-terminal MOR splice variants using the real-time polymerase chain reaction (RT-PCR). Expression of MOR-1K mRNA was also increased in HIV-infected individuals with combined neurocognitive impairment and HIVE compared with the other groups. MOR-1K expression correlated with the level of patient neurocognitive impairment, whereas the pool of C-terminal MOR splice variants did not. HIVE was also associated with increased expression of the inflammatory mediators MCP-1, MCP-2, and RANTES, but not the host HIV coreceptors CXCR4 and CCR5 or the CD4 receptor using qRT-PCR. Network analysis of microarray data from these same patients revealed filamin A (FLNA) as a possible interaction partner with MOR-1K, and FLNA gene expression was also found to be upregulated in HIVE using qRT-PCR. Overexpression of FLNA in HEK293 cells redistributed MOR-1K from intracellular compartments to the cell surface. CONCLUSION: These results suggest that HIVE, and neurocognitive impairment depending on its severity, are associated with enhanced MOR-1K signaling through both increased expression and trafficking to the cell surface, which may alter the contribution of MOR receptor isoforms and exacerbate the effects of MOR activation in neuroAIDS.