Thermal hyperalgesia and mechanical allodynia produced by intrathecal administration of the human immunodeficiency virus-1 (HIV-1) envelope glycoprotein, gp120

Astrocytes and microglia in the spinal cord have recently been reported to contribute to the development of peripheral inflammation-induced exaggerated pain states. Both lowering of thermal pain threshold (thermal hyperalgesia) and lowering of response threshold to light tactile stimuli (mechanical allodynia) have been reported. The notion that spinal cord glia are potential mediators of such effects is based on the disruption of these exaggerated pain states by drugs thought to preferentially affect glial function. Activation of astrocytes and microglia can release many of the same substances that are known to mediate thermal hyperalgesia and mechanical allodynia. The aim of the present series of studies was to determine whether exaggerated pain states could also be created in rats by direct, intraspinal immune activation of astrocytes and microglia. The immune stimulus used was peri-spinal (intrathecal, i.t.) application of the Human Immunodeficiency Virus type 1 (HIV-1) envelope glycoprotein, gp120. This portion of HIV-1 is known to bind to and activate microglia and astrocytes. Robust thermal hyperalgesia (tail-flick, TF, and Hargreaves tests) and mechanical allodynia (von Frey and touch-evoked agitation tests) were observed in response to i.t. gp120. Heat denaturing of the complex protein structure of gp120 blocked gp120-induced thermal hyperalgesia. Lastly, both thermal hyperalgesia and mechanical allodynia to i.t. gp120 were blocked by spinal pretreatment with drugs (fluorocitrate and CNI-1493) thought to preferentially disrupt glial function.     

Characteristics of HIV-1 gp120 glycoprotein binding to glycolipids

We examined the binding of the gp120 envelope glycoprotein (gp120) of the human immunodeficiency virus (HIV-1) to sulfatide (GalS), galactocerebroside (GalC), and GMI-ganglioside (GMI). The gp120 glycoprotein bound to GalS but not to GalC or GMI by enzyme-linked immunosorbent assay (ELISA) and by an immunospot assay on nitrocellulose paper. However, it bound to all three glycolipids by an immunospot assay on thin layer chromotography (TLC) plates. In studies to determine whether GalS could be a receptor for gp120 on the surface of cells, gp120 bound to GalS incorporated into the plasma membrane of lymphoid cells as determined by cytofluorometric analysis and immunofluorescence microscopy. These studies indicate that GalS may function as a receptor for gp120 and HIV-1. © 1994 Wiley-Liss, Inc.     

Curcumin improves synaptic plasticity impairment induced by HIV-1gp120 V3 loop

Curcumin has been shown to significantly improve spatial memory impairment induced by HIV-1 gp120 V3 in rats, but the electrophysiological mechanism remains unknown. Using extracellular microelectrode recording techniques, this study confirmed that the gp120 V3 loop could suppress long-term potentiation in the rat hippocampal CA1 region and synaptic plasticity, and that curcumin could antagonize these inhibitory effects. Using a Fura-2/AM calcium ion probe, we found that curcumin resisted the effects of the gp120 V3 loop on hippocampal synaptosomes and decreased Ca²⁺ concentration in synaptosomes. This effect of curcumin was identical to nimodipine, suggesting that curcumin improved the inhibitory effects of gp120 on synaptic plasticity, ameliorated damage caused to the central nervous system, and might be a potential neuroprotective drug.     

The gp120 glycoprotein of HIV-1 binds to sulfatide and to the myelin associated glycoprotein.

We investigated the binding of the gp120 glycoprotein of the human immunodeficiency virus (HIV-1) to neural glycolipids and glycoproteins by ELISA. The gp120 protein bound to sulfatide (GalS), a sulfated glycolipid autoantigen implicated in sensory neuritis, and to the myelin associated glycoprotein (MAG), an autoantigen in demyelinating neuropathy. Binding of gp120 to MAG was inhibited by the HNK-1 antibody, which recognizes a sulfated glucuronic acid epitope, suggesting that the interaction involves carbohydrate determinants. Sulfatide and MAG are potential receptors for gp120 in peripheral nerve and may have a role in the neuropathy associated with HIV-1 infection.     

Involvement of brain cytokines in the neurobehavioral disturbances induced by HIV-1 glycoprotein120

Intracerebroventricular (i.c.v.) administration of HIV-1 glycoprotein 120 (gp120), the envelope protein used by the virus to gain access into immune cells, induces neurobehavioral alterations in rats. To examine the role of proinflammatory cytokines in mediating these effects, we measured the effects of gp120 on brain proinflammatory cytokine expression and the effects of anti-inflammatory agents, including interleukin-1 receptor antagonist (IL-1ra), pentoxifylline (a TNFalpha synthesis blocker) and IL-10, on gp120-induced sickness behavior. I.c.v. administration of gp120 induced the expression of IL-1beta, but not TNFalpha, mRNA in the hypothalamus, 3 h after the injection. Pretreatment of rats with IL-1ra, but not with pentoxifylline, significantly attenuated gp120-induced anorexia and loss in body weight, whereas both agents had no effect on gp120-induced reduction in locomotor activity in the open field. Pretreatment with either IL-1ra and pentoxifylline simultaneously, or with IL-10, produced effects that were similar to the effects of IL-1ra alone. Together, these findings indicate that IL-1, but not TNFalpha, mediates some of the behavioral effects of acute gp120 administration, suggesting that brain IL-1 may be involved in some of the neurobehavioral abnormalities evident in AIDS patients.     

17beta-estradiol reduces neuronal apoptosis induced by HIV-1 gp120 in the neocortex of rat

The human immunodeficiency virus type 1 (HIV-1) coat glycoprotein gp120 represents a likely contributor to the development of HIV-1 associated dementia (HAD), a neurological syndrome often observed in AIDS patients and characterised by significant neuronal loss in the neocortex. Since recent studies have highlighted that female sex hormones represent potential neuroprotective agents against damage produced by acute and chronic injuries in the adult brain, we have investigated whether estrogens exert protection in a rat model of gp120 neurotoxicity. Our results demonstrate that systemic administration of 17beta-estradiol (E2, 0.02-0.2 mg/kg) significantly reduces apoptotic cell death observed in the neocortex of rat following subchronic i.c.v. administration of gp120 (100 ng/rat/day). Furthermore, both tamoxifen and ICI182,780, two selective antagonists of estrogen receptors (ER) in the brain, reverted the neuroprotective effect of E2. The molecular mechanism of estrogenic neuroprotection does not appear to involve modulation of the antiapoptotic Bcl-2 or the proapoptotic Bax since we failed to observe changes in the levels of the two proteins in the neocortical tissue after gp120 and/or E2 treatment. However, we detected increased levels of IL-1beta in the neocortex of rats injected with gp120, as early as 6h after drug administration, and this effect was potentiated following pretreatment with E2. Taken together, our results demonstrate that E2 exerts neuroprotection against gp120 neurotoxicity in vivo through a mechanism involving ER activation and, possibly, via modulation of neocortical levels of IL-1beta.     

Requirement for macrophages in neuronal injury induced by HIV envelope protein gp120.

HIV-1-related neuronal injury may involve a complex web of viral proteins and cytokines, but neurons themselves are not infected. The HIV envelope protein gp120 has been shown to engender an early increase in neuronal free calcium followed by delayed excitotoxic-like damage, which is prevented by N-methyl-D-aspartate (NMDA) antagonists. In the present study, we found that the injurious effects of gp120 on retinal ganglion cell neurons require the presence of macrophages in mixed neuronal glial cultures of postnatal retina. Within 24 hours of incubation, 20 pM gp120 injured nearly 40% of retinal ganglion cells in cultures containing macrophages and other glial cells, whereas no deleterious effects of gp120 were noted on retinal ganglion cells in cultures depleted of macrophages. Thus, the toxic effect of gp120 on neurons appears to be an indirect one, mediated by activation of macrophages and perhaps other glial cells.     

Galantamine and nicotine have a synergistic effect on inhibition of microglial activation induced by HIV-1 gp120

Chronic brain inflammation is the common final pathway in the majority of neurodegenerative diseases and central to this phenomenon is the immunological activation of brain mononuclear phagocyte cells, called microglia. This inflammatory mechanism is a central component of HIV-associated dementia (HAD). In the healthy state, there are endogenous signals from neurons and astrocytes, which limit excessive central nervous system (CNS) inflammation. However, the signals controlling this process have not been fully elucidated. Studies on the peripheral nervous system suggest that a cholinergic anti-inflammatory pathway regulates systemic inflammatory response by way of acetylcholine acting at the alpha7 nicotinic acetylcholine receptor (alpha7nAChR) found on blood-borne macrophages. Recent data from our laboratory indicates that cultured microglial cells also express this same receptor and that microglial anti-inflammatory properties are mediated through it and the p44/42 mitogen-activated protein kinase (MAPK) system. Here we report for the first time the creation of an in vitro model of HAD composed of cultured microglial cells synergistically activated by the addition of IFN-gamma and the HIV-1 coat glycoprotein, gp120. Furthermore, this activation, as measured by TNF-alpha and nitric oxide (NO) release, is synergistically attenuated through the alpha7 nAChR and p44/42 MAPK system by pretreatment with nicotine, and the cholinesterase inhibitor, galantamine. Our findings suggest a novel therapeutic combination to treat or prevent the onset of HAD through this modulation of the microglia inflammatory mechanism.     

Adsorptive endocytosis of HIV-1gp120 by blood-brain barrier is enhanced by lipopolysaccharide

Previous work suggests that gp120 mediates the passage of HIV-1 and infected immune cells across the blood-brain barrier (BBB) by induction of adsorptive endocytosis (AE) in brain endothelial cells. Other work has suggested that cytokines may increase the permeability of the BBB to free virus or infected immune cells. Here, we investigated the ability of lipopolysaccharide (LPS), a bacterial wall toxin that stimulates the release of cytokines, to increase gp120 passage across the BBB by enhancement of AE and/or induction of BBB disruption. We found that LPS enhanced the passage of gp120 radioactively labeled with 125I (I-gp120) in a reversible, time-dependent, prostaglandin-independent manner that was not completely explained by disruption of the BBB. LPS also enhanced wheatgerm agglutinin mediated uptake of I-gp120 almost exclusively through the potentiation of AE. These results show that LPS or cytokines released by LPS can have a major effect on the permeability of the BBB to HIV-1gp120 both by stimulating AE and by inducing a disruption of the BBB. This suggests that bacterial infection or other inflammatory states could facilitate invasion of the CNS by HIV-1.     

Transgenic mice expressing HIV-1 envelope protein gp120 in the brain as an animal model in neuroAIDS research

HIV-1 infection causes injury to the central nervous system (CNS) and is often associated with neurocognitive disorders. One model for brain damage seen in AIDS patients is the transgenic (tg) mouse expressing a soluble envelope protein gp120 of HIV-1 LAV in the brain in astrocytes under the control of the promoter of glial fibrillary acidic protein. These GFAP-gp120tg mice manifest several key neuropathological features observed in AIDS brains, such as decreased synaptic and dendritic density, increased numbers of activated microglia, and pronounced astrocytosis. Several recent studies show that brains of GFAP-gp120tg mice and neurocognitively impaired HIV patients share also a significant number of differentially regulated genes, activation of innate immunity and other cellular signaling pathways, disturbed neurogenesis, and learning deficits. These findings support the continued relevance of the GFAP-gp120tg mouse as a useful model to investigate neurodegenerative mechanisms and develop therapeutic strategies to mitigate the consequences associated with HIV infection of the CNS, neuroAIDS, and HAND.     

HIV-1 gp120-Mediated Mitochondrial Dysfunction and HIV-Associated Neurological Disorders

The treatment of HIV infection presents a great challenge among the patients who develop various forms of cognitive impairments. Particularly, the neurotoxicity associated with HIV is attributed to different viral proteins present in HIV, and is a root cause for HIV-associated neurocognitive disorders (HAND). A common characteristic among several neurodegenerative disorders including HAND is mitochondrial dysfunction in various brain cells. However, there has been very little effort to explore the possibility of exploiting mitochondrial dynamics in HAND treatment. A recent study by Avdoshina and colleagues has reported the role of mitochondrial dysfunction in HIV-1 gp120-mediated neuronal dysfunction, which presents a novel mechanism for the development of adjunct therapy to treat HAND.     

Erythropoietin protects cerebrocortical neurons from HIV-1/gp120-induced damage

Infection with human immunodeficiency virus (HIV)-1 can lead to neurological complications that range from mild cognitive and motor impairment to HIV-associated dementia (HAD). The mechanism of brain injury and dementia remains poorly understood. Interestingly, post mortem brain specimen from HAD patients and transgenic mice expressing the viral envelope protein gp120 present with similar neuropathological signs. The cytokine erythropoietin (EPO) is clinically used to treat anemia but has also been found to prevent neuronal death due to inflammation or excitotoxicity. Here we show that EPO protects cerebrocortical neurons against apoptosis induced by HIV-1/gp120.     

Transmission routes of HIV-1 gp120 from brain to lymphoid tissues

The blood-brain barrier (BBB) restricts the entry of antiviral agents into the CNS thereby facilitating the creation of a reservoir of HIV that could potentially reinfect peripheral tissues. We characterized the efflux from brain of radioactively labeled viral coat HIV-1 gp120 (I-gp120) after intracerebroventricular (i.c.v.) injection. The half-time disappearance rate of I-gp120 from brain was 12.6 min, which was faster than could be explained by the reabsorption of cerebrospinal fluid into blood but could not be explained by a saturable transporter. After i.c.v. injection, I-gp120 appeared in the serum and was sequestered by spleen and the cervical nodes, demonstrating a potential for virus within the CNS to reinfect peripheral tissues. However, the amount of I-gp120 appearing in serum was less than that expected based on the efflux rate, whereas uptake by the cervical nodes was much greater after i. c.v. than after i.v. injection of I-gp120. These findings were explained by drainage from the brain directly to the cervical lymph nodes through the brain's primitive lymphatic system. These lymphatics potentially provide a pathway through which CNS reservoirs of HIV-1 could directly reinfect lymphoid tissue without being exposed to circulating antiviral agents.     

Suppression of PrP(Sc)- and HIV-1 gp120 induced neuronal cell death by sulfated colominic acid.

The scrapie prion protein (PrP(Sc)) has been shown to induce apoptosis of rat cortical neurons in vitro. Here we demonstrate that the toxic effect displayed by PrP(Sc) can be blocked by sulfated colominic acid (polymer of N-acetylneuraminic acid). This compound acts neuroprotectively at a concentration of > or = 0.3 microg/ml when preincubated with the neurons or PrP(Sc). Rat cortical cells also undergo apoptosis after incubation with the HIV-1 coat protein gpl20 in vitro. This effect was abolished also by sulfated colominic acid when preincubated with the cells or gpl20. Addition of 0.3 microg/ml of compound resulted in an increase in cell viability by about 1.6-1.9-fold compared to cultures incubated for 18 h with 30 ng/ml of PrP(Sc) or 20 ng/ml of gpl20 alone (containing about 40% viable cells). Sulfated colominic acid does not act as antagonist of NMDA receptor channels at concentrations of up to 3 microg/ml when co-administered with 100 microg/ml of NMDA. It displayed a strong cytoprotective effect on human T lymphoblastoid CEM cells exposed to HIV-1; a 50% protection occurred after preincubation of the cells with 0.43 microg/ml of compound. At the same concentration, the compound caused an inhibition of HIV-1-induced syncytium formation. Sulfated colominic acid may be a promising compound for treatment of dementia caused by PrP(Sc) and HIV-1 infections.     

Reduction of cerebral glucose utilization by the HIV envelope glycoprotein Gp-120

Gp-120 is a glycoprotein constituent of the human immunodeficiency virus (HIV) envelope. The effects of gp-120 on cerebral glucose utilization in rats were studied by the quantitative 2-deoxy-D-[1-14C] glucose method. Intracerebroventricular injection of gp-120 significantly reduced glucose utilization in the lateral habenula and the suprachiasmatic nucleus and decreased the global cerebral metabolic rate for glucose. The findings suggest that gp-120 and closely related peptides can alter neuronal function, thereby contributing to the sequelae of HIV infection.     

Schwann cell chemokine receptors mediate HIV-1 gp120 toxicity to sensory neurons

Human immunodeficiency virus (HIV)-associated sensory neuropathy (HIV-SN) is the most common neurological complication of HIV infection. Currently, the pathogenesis of HIV-SN is unknown. Because there is no convincing evidence of neuronal infection, HIV neurotoxicity is likely to be effected either by secreted viral proteins such as the envelope glycoprotein gp120 or by neurotoxic cytokines released from infected/activated glial cells. We describe a model of gp120 toxicity to primary sensory neurons, in which gp120 induces neuritic degeneration and neuronal apoptosis. We show that Schwann cells, the cells that ensheath peripheral nerve axons, and which traditionally have been viewed as having a passive, supporting role, mediate this neurotoxicity. Ligation of the chemokine receptor CXCR4 on Schwann cells by gp120 resulted in the release of RANTES, which induced dorsal root ganglion neurons to produce tumor necrosis factor-alpha and subsequent TNFR1-mediated neurotoxicity in an autocrine fashion. This newly described Schwann cell-neuron interaction may be pathogenically relevant not only in HIV-SN but also in other peripheral neuropathies.     

Neurotoxicity of HIV-1 proteins gp120 and Tat in the rat striatum

HIV-associated dementia complex is a serious disabling disease characterized by cognitive, behavioral and motor dysfunction. Basal ganglia involvement in HIV-1 infection may be responsible for some of the psychomotor symptoms associated with HIV dementia. The objectives of the present study were to determine: (1) whether gp120 and Tat produce striatal toxicity, and (2) whether gp120 and Tat show synergistic toxicity in the striatum. In these studies, the recombinant proteins gp120, Tat, or saline (0.9%) were stereotaxically injected in the striatum of adult male rats. The striatal sections were evaluated for area of tissue loss (Cresyl-violet stained sections) and the number of GFAP immunoreactive cells 7 days after the injections. Doses of gp120 250 ng/microl or higher and Tat 5 microg/microl or higher produced a significant area of tissue loss and significantly increased the number of GFAP reactive cells. We found no toxicity in animals treated with immunoabsorbed gp120 or Tat. Combined gp120 (100 ng/microl)+Tat (1 microg/microl) injections into the rat striatum significantly increased the area of tissue loss and altered morphology and increased number of GFAP reactive cells, as compared to controls. Thus, the present results suggest the involvement of gp120 and Tat in striatal toxicity and provide a model for further studies to fully characterize their role in HIV-1 toxicity and to develop therapeutic strategies for HIV-1 associated dementia complex.     

Neurotrophic activity of monomeric glucophosphoisomerase was blocked by human immunodeficiency virus (HIV-1) and peptides from HIV-1 envelope glycoprotein

Glucophosphoisomerase (GPI), a glycolytic enzyme, was recently described to share 90% sequence homology with neuroleukin, a recently discovered growth factor which promotes motor neuron regeneration in vivo, survival of peripheral and central neurons in vitro, and affects B cell immunoglobulin synthesis. Interestingly, neuroleukin activity was described to be antagonized by the human immunodeficiency virus (HIV-1) envelope glycoprotein (gp120), with which neuroleukin was found to share partial sequence homology. In this study, reduced GPI demonstrated similar activity to neuroleukin in a novel bioassay using human and rat neuroblastoma cell lines. In the presence of reduced GPI, these cells were found to differentiate, in terms of enhanced neurite extension at a reduced proliferation rate. These results demonstrate the existence of a novel growth factor activity of an evolutionary ancient enzyme. The nonreduced commercial form of GPI, probably the dimer, was found to be inactive in this bioassay. Using the neuroblastoma cells model system, we further investigated the significance of the region of homology to HIV-1 envelope glycoprotein (gp120) as the putative binding site of GPI to its receptor on neuronal cells.     

Methamphetamine potentiates HIV-1gp120-induced microglial neurotoxic activity by enhancing microglial outward K+ current

Methamphetamine (Meth) abuse not only increases the risk of human immunodeficiency virus-1 (HIV-1) infection, but exacerbates HIV-1-associated neurocognitive disorders (HAND) as well. The mechanisms underlying the co-morbid effect are not fully understood. Meth and HIV-1 each alone interacts with microglia and microglia express voltage-gated potassium (K_V) channel K_V1.3. To understand whether K_V1.3 functions an intersecting point for Meth and HIV-1, we studied the augment effect of Meth on HIV-1 glycoprotein 120 (gp120)-induced neurotoxic activity in cultured rat microglial cells. While Meth and gp120 each alone at low (subtoxic) concentrations failed to trigger microglial neurotoxic activity, Meth potentiated gp120-induced microglial neurotoxicity when applied in combination. Meth enhances gp120 effect on microglia by enhancing microglial K_V1.3 protein expression and K_V1.3 current, leading to an increase of neurotoxin production and resultant neuronal injury. Pretreatment of microglia with a specific K_V1.3 antagonist 5-(4-Phenoxybutoxy)psoralen (PAP) or a broad spectrum K_V channel blocker 4-aminopyridine (4-AP) significantly attenuated Meth/gp120-treated microglial production of neurotoxins and resultant neuronal injury, indicating an involvement of K_V1.3 in Meth/gp120-induced microglial neurotoxic activity. Meth/gp120 activated caspase-3 and increased caspase-3/7 activity in microglia and inhibition of caspase-3 by its specific inhibitor significantly decreased microglial production of TNF-α and iNOS and attenuated microglia-associated neurotoxic activity. Moreover, blockage of K_V1.3 by specific blockers attenuated Meth/gp120 enhancement of caspase-3/7 activity. Taking together, these results suggest an involvement of microglial K_V1.3 in the mediation of Meth/gp120 co-morbid effect on microglial neurotoxic activity via caspase-3 signaling.