RESUMO
HIV-1 infection in the era of combined antiretroviral therapy has been associated with premature aging. Among the various features of HIV-1 associated neurocognitive disorders, astrocyte senescence has been surmised as a potential cause contributing to HIV-1-induced brain aging and neurocognitive impairments. Recently, lncRNAs have also been implicated to play essential roles in the onset of cellular senescence. Herein, using human primary astrocytes (HPAs), we investigated the role of lncRNA TUG1 in HIV-1 Tat-mediated onset of astrocyte senescence. We found that HPAs exposed to HIV-1 Tat resulted in significant upregulation of lncRNA TUG1 expression that was accompanied by elevated expression of p16 and p21, respectively. Additionally, HIV-1 Tat-exposed HPAs demonstrated increased expression of senescence-associated (SA) markers-SA-ß-galactosidase (SA-ß-gal) activity and SA-heterochromatin foci-cell-cycle arrest, and increased production of reactive oxygen species and proinflammatory cytokines. Intriguingly, gene silencing of lncRNA TUG1 in HPAs also reversed HIV-1 Tat-induced upregulation of p21, p16, SA-ß gal activity, cellular activation, and proinflammatory cytokines. Furthermore, increased expression of astrocytic p16 and p21, lncRNA TUG1, and proinflammatory cytokines were observed in the prefrontal cortices of HIV-1 transgenic rats, thereby suggesting the occurrence of senescence activation in vivo. Overall, our data indicate that HIV-1 Tat-induced astrocyte senescence involves the lncRNA TUG1 and could serve as a potential therapeutic target for dampening accelerated aging associated with HIV-1/HIV-1 proteins.
Assuntos
Infecções por HIV , HIV-1 , RNA Longo não Codificante , Animais , Humanos , Ratos , Envelhecimento/metabolismo , Astrócitos/metabolismo , Senescência Celular , Citocinas/metabolismo , Infecções por HIV/metabolismo , HIV-1/fisiologia , Ratos Transgênicos , RNA Longo não Codificante/metabolismo , Produtos do Gene tat do Vírus da Imunodeficiência HumanaRESUMO
Substance use disorder (SUD) is a growing health problem that affects several millions of people worldwide, resulting in negative socioeconomic impacts and increased health care costs. Emerging evidence suggests that extracellular vesicles (EVs) play a crucial role in SUD pathogenesis. EVs, including exosomes and microvesicles, are membrane-encapsulated particles that are released into the extracellular space by most types of cells. EVs are important players in mediating cell-to-cell communication through transfer of cargo such as proteins, lipids and nucleic acids. The EV cargo can alter the status of recipient cells, thereby contributing to both physiological and pathological processes; some of these play critical roles in SUD. Although the functions of EVs under several pathological conditions have been extensively reviewed, EV functions and potential applications in SUD remain less studied. In this review, we provide an overview of the current knowledge of the role of EVs in SUD, including alcohol, cocaine, heroin, marijuana, nicotine and opiate abuse. The review will focus on the biogenesis and cargo composition of EVs as well as the potential use of EVs as biomarkers of SUD or therapeutic targets in SUD.
Assuntos
Vesículas Extracelulares/metabolismo , Transtornos Relacionados ao Uso de Substâncias/patologia , Animais , Biomarcadores/metabolismo , Comunicação Celular , Citocromo P-450 CYP2E1/metabolismo , Complexos Endossomais de Distribuição Requeridos para Transporte/metabolismo , Vesículas Extracelulares/transplante , Humanos , MicroRNAs/metabolismo , Transtornos Relacionados ao Uso de Substâncias/metabolismo , Transtornos Relacionados ao Uso de Substâncias/terapiaRESUMO
Neuroinflammation associated with HIV-1 infection is a problem affecting â¼50% of HIV-infected individuals. NLR family pyrin domain containing 3 (NLRP3) inflammasome has been implicated in HIV-induced microglial activation, but the mechanism(s) remain unclear. Because HIV-1 Transactivator of Transcription (Tat) protein continues to be present despite antiretroviral therapy and activates NF-kB, we hypothesized that Tat could prime the NLRP3 inflammasome. We found a dose- and time-dependent induction of NLRP3 expression in microglia exposed to Tat compared with control. Tat exposure also time-dependently increased the mature caspase-1 and IL-1ß levels and enhanced the IL-1ß secretion. These in vitro findings were validated in archival brain tissues from Simian Immunodeficiency Virus (SIV)-infected and uninfected rhesus macaques. Further validation of NLRP3 priming in vivo involved administration of lipopolysaccharide (LPS) to HIV transgenic (Tg) rats followed by assessment of IL-1ß mRNA expression and inflammasome activation (ASC oligomers and mature IL-1ß). Intriguingly, LPS potentiated upregulation of IL-1ß mRNA and inflammasome activation in HIV-Tg rats compared with the wild-type controls. Interestingly, we found an inverse relationship in the expression of NLRP3 and its negative regulator, miR-223, suggesting a miR-223-mediated mechanism for Tat-induced NLRP3 priming. Furthermore, blockade of NLRP3 resulted in decreased IL-1ß secretion. Collectively, these findings suggest a novel role of Tat in priming and activating the NLRP3 inflammasome. Therefore, NLRP3 can be envisioned as a therapeutic target for ameliorating Tat-mediated neuroinflammation.SIGNIFICANCE STATEMENT Despite successful suppression of viremia with increased longevity in the era of combined antiretroviral therapy, chronic inflammation with underlying neurocognitive impairment continues to afflict almost 50% of infected individuals. Viral, bacterial, and cellular products have all been implicated in promoting the chronic inflammation found in these individuals. Understanding the molecular mechanism(s) by which viral proteins such as HIV-1 Transactivator of Transcription (Tat) protein can activate microglia is thus of paramount importance. Herein, we demonstrate a novel role of Tat in priming and activating NLR family pyrin domain containing 3 (NLRP3) inflammasomes in microglial cells and in HIV-Tg rats administered lipopolysaccharide. Targeting NLRP3 inflammasome pathway mediators could thus be developed as therapeutic interventions to alleviate or prevent neuroinflammation and subsequent cognitive impairment in HIV-positive patients.
Assuntos
Encéfalo/imunologia , Encefalite Viral/imunologia , Inflamassomos/imunologia , Microglia/imunologia , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Produtos do Gene tat do Vírus da Imunodeficiência Humana/imunologia , Animais , Citocinas/imunologia , Feminino , Mediadores da Inflamação/imunologia , Macaca mulatta , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microglia/patologia , Ratos , Ratos TransgênicosRESUMO
Despite major advances in HIV-1 treatment, the prevalence of HIV-associated neurocognitive disorders (HAND) remains a problem, particularly as individuals on suppressive treatment continue to live longer. To facilitate discussion on emerging and future directions in HAND research, a meeting was held in Durban, South Africa in March 2015 as part of the Society of Neuroscientists of Africa (SONA) conference. The objective of the meeting was to assess the impact of HIV subtype diversity on HAND and immunological dysfunction. The meeting brought together international leaders in the area of neurological complications of HIV-1 infection with special focus on the African population. Research presentations indicated that HAND was highly prevalent and that inflammatory cytokines and immune-activation played important roles in progression of neurocognitive impairment. Furthermore, children on antiretroviral therapy were also at risk for developing neurocognitive impairment. With respect to the effect of HIV-1 subtype diversity, analyses of HIV-1 clade C infection among South Africans revealed that clade C infection induced cognitive impairment that was independent of the substitution in HIV-1 Trans-Activator of Transcription (Tat; C31S). At the cellular level, a Zambian study showed that clade C infection resulted in reduced brain cell death compared with clade B infection suggesting clade specific variations in mediating brain cell injury. Furthermore, ex vivo Tat protein from clade CRF02_AG, prevalent in West/ Central Africa, exhibited reduced disruption of brain endothelium compared with clade B Tat protein. Discussions shed light on future research directions aimed at understanding biomarkers and disease mechanisms critical for HAND.
Assuntos
Complexo AIDS Demência/diagnóstico , Encéfalo/patologia , Citocinas/imunologia , HIV-1/patogenicidade , Interações Hospedeiro-Patógeno , Complexo AIDS Demência/imunologia , Complexo AIDS Demência/patologia , Complexo AIDS Demência/virologia , Encéfalo/irrigação sanguínea , Encéfalo/imunologia , Encéfalo/virologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/patologia , Linfócitos T CD4-Positivos/virologia , Citocinas/biossíntese , Progressão da Doença , Endotélio Vascular/imunologia , Endotélio Vascular/patologia , Endotélio Vascular/virologia , Feminino , HIV-1/classificação , HIV-1/fisiologia , Humanos , Masculino , Tipagem Molecular , Testes Neuropsicológicos , Produtos do Gene tat do Vírus da Imunodeficiência Humana/biossíntese , Produtos do Gene tat do Vírus da Imunodeficiência Humana/imunologiaRESUMO
Human pegivirus (HPgV; originally called GB virus C/hepatitis G virus) is an RNA virus within the genus Pegivirus of the family Flaviviridae that commonly causes persistent infection. Worldwide, ~750 million people are actively infected (viraemic) and an estimated 0.75-1.5 billion people have evidence of prior HPgV infection. No causal association between HPgV and disease has been identified; however, several studies described a beneficial relationship between persistent HPgV infection and survival in individuals infected with human immunodeficiency virus. The beneficial effect appeared to be related to a reduction in host immune activation. HPgV replicates well in vivo (mean plasma viral loads typically >1×107 genome copies ml-1); however, the virus grows poorly in vitro and systems to study this virus are limited. Consequently, mechanisms of viral persistence and host immune modulation remain poorly characterized, and the primary permissive cell type(s) has not yet been identified. HPgV RNA is found in liver, spleen, bone marrow and PBMCs, including T- and B-lymphocytes, NK-cells, and monocytes, although the mechanism of cell-to-cell transmission is unclear. HPgV RNA is also present in serum microvesicles with properties of exosomes. These microvesicles are able to transmit viral RNA to PBMCs in vitro, resulting in productive infection. This review summarizes existing data on HPgV cellular tropism and the effect of HPgV on immune activation in various PBMCs, and discusses how this may influence viral persistence. We conclude that an increased understanding of HPgV replication and immune modulation may provide insights into persistent RNA viral infection of humans.
Assuntos
Vírus GB C/imunologia , Vírus GB C/fisiologia , Imunomodulação , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/virologia , Tropismo Viral , Exossomos/virologia , Infecções por Flaviviridae/virologia , Infecções por HIV/complicações , Infecções por HIV/mortalidade , Hepatite Viral Humana/virologia , Humanos , RNA Viral/isolamento & purificaçãoRESUMO
Viruses enter into complex interactions within human hosts, leading to facilitation or suppression of each other's replication. Upon coinfection, GB virus C (GBV-C) suppresses HIV-1 replication in vivo and in vitro, and GBV-C coinfection is associated with prolonged survival in HIV-infected people. GBV-C is a lymphotropic virus capable of persistent infection. GBV-C infection is associated with reduced T cell activation in HIV-infected humans, and immune activation is a critical component of HIV disease pathogenesis. We demonstrate that serum GBV-C particles inhibited activation of primary human T cells. T cell activation inhibition was mediated by the envelope glycoprotein E2, because expression of E2 inhibited TCR-mediated activation of Lck. The region on the E2 protein was characterized and revealed a highly conserved peptide motif sufficient to inhibit TCR-mediated signaling. The E2 region contained a predicted Lck substrate site, and substitution of an alanine or histidine for the tyrosine reversed TCR-signaling inhibition. GBV-C E2 protein and a synthetic peptide representing the inhibitory amino acid sequence were phosphorylated by Lck in vitro. The synthetic peptide also inhibited TCR-mediated activation of primary human CD4(+) and CD8(+) T cells. Extracellular microvesicles from GBV-C E2-expressing cells contained E2 protein and inhibited TCR signaling in bystander T cells not expressing E2. Thus, GBV-C reduced global T cell activation via competition between its envelope protein E2 and Lck following TCR engagement. This novel inhibitory mechanism of T cell activation may provide new approaches for HIV and immunoactivation therapy.
Assuntos
Vírus GB C/imunologia , Ativação Linfocitária/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Transdução de Sinais/imunologia , Linfócitos T/imunologia , Proteínas do Envelope Viral/imunologia , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Vírus GB C/metabolismo , Hepatite Viral Humana/imunologia , Hepatite Viral Humana/metabolismo , Humanos , Immunoblotting , Células Jurkat , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Linfócitos T/metabolismoRESUMO
Human pegivirus (HPgV; previously called GB virus C/hepatitis G virus) has limited pathogenicity, despite causing persistent infection, and is associated with prolonged survival in human immunodeficiency virus-infected individuals. Although HPgV RNA is found in and produced by T- and B-lymphocytes, the primary permissive cell type(s) are unknown. We quantified HPgV RNA in highly purified CD4(+) and CD8(+) T-cells, including naïve, central memory and effector memory populations, and in B-cells (CD19(+)), NK cells (CD56(+)) and monocytes (CD14(+)) using real-time reverse transcription-PCR. Single-genome sequencing was performed on viruses within individual cell types to estimate genetic diversity among cell populations. HPgV RNA was present in CD4(+) and CD8(+) T-lymphocytes (nine of nine subjects), B-lymphocytes (seven of ten subjects), NK cells and monocytes (both four of five). HPgV RNA levels were higher in naïve (CD45RA(+)) CD4(+) cells than in central memory and effector memory cells (P<0.01). HPgV sequences were highly conserved among subjects (0.117±0.02 substitutions per site; range 0.58-0.14) and within subjects (0.006±0.003 substitutions per site; range 0.006-0.010). The non-synonymous/synonymous substitution ratio was 0.07, suggesting a low selective pressure. Carboxyfluorescein succinimidyl ester (CFSE)-labelled HPgV RNA-containing particles precipitated by a commercial exosome isolation reagent delivered CSFE to uninfected monocytes, NK cells and T- and B-lymphocytes, and HPgV RNA was transferred to PBMCs with evidence of subsequent virus replication. Thus, HPgV RNA-containing serum particles including microvesicles may contribute to delivery of HPgV to PBMCs in vivo, explaining the apparent broad tropism of this persistent human RNA virus.
Assuntos
Vírus GB C/isolamento & purificação , Vírus GB C/patogenicidade , Leucócitos Mononucleares/virologia , RNA Viral/sangue , Adulto , Sequência de Aminoácidos , Linfócitos B/virologia , Sequência de Bases , Linfócitos T CD4-Positivos/virologia , Linfócitos T CD8-Positivos/virologia , Sequência Conservada , Feminino , Infecções por Flaviviridae/complicações , Infecções por Flaviviridae/virologia , Vírus GB C/genética , Infecções por HIV/complicações , Hepatite Viral Humana/complicações , Hepatite Viral Humana/virologia , Humanos , Células Matadoras Naturais/virologia , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Monócitos/virologia , Filogenia , RNA Helicases/genética , RNA Viral/genética , Serina Endopeptidases/genética , Proteínas não Estruturais Virais/genética , Virulência , Adulto JovemRESUMO
Hepatitis C virus (HCV) and GB virus type C (GBV-C) are associated with impaired T cell function despite the fact that HCV replicates in hepatocytes and GBV-C in a small proportion of lymphocytes. Recently, we showed that HCV and GBV-C E2-envelope proteins reduce T cell activation via the T cell receptor (TCR) by competing for phosphorylation with a critical kinase in the TCR signaling cascade (Lck). E2 interfered with TCR signaling in E2 expressing cells and in bystander cells. The bystander effect was mediated by virus particles and extracellular microvesicular particles (exosomes). Multiple kinase substrate sites are predicted to reside on viral structural proteins and based on bioinformatic predictions, many RNA virus pathogens may interfere with TCR signaling via a similar mechanism. Identification of T cell inhibitory effects of virus structural proteins may provide novel approaches to enhance the immunogenicity and memory of viral vaccines.
Assuntos
Vírus GB C/imunologia , Hepacivirus/imunologia , Hepatite/imunologia , Evasão da Resposta Imune , Ativação Linfocitária , Linfócitos T/imunologia , Proteínas do Envelope Viral/imunologia , Animais , Vírus GB C/metabolismo , Vírus GB C/patogenicidade , Hepacivirus/metabolismo , Hepacivirus/patogenicidade , Hepatite/epidemiologia , Hepatite/história , Hepatite/metabolismo , Hepatite/virologia , História do Século XX , História do Século XXI , Interações Hospedeiro-Patógeno , Humanos , Receptores de Antígenos de Linfócitos T/imunologia , Receptores de Antígenos de Linfócitos T/metabolismo , Transdução de Sinais , Linfócitos T/metabolismo , Linfócitos T/virologia , Proteínas do Envelope Viral/metabolismoRESUMO
Double-negative T cells (DNTCs; ie, CD3(+)CD4(-)CD8(-) T cells) play a role in limiting chronic immune activation. GB virus C (GBV-C) infection is associated with reduced T-cell activation in human immunodeficiency virus (HIV)-infected individuals. T-cell activation and DNTCs were measured in HIV-infected subjects with a nondetectable HIV load. GBV-C-viremic subjects had significantly reduced CD4(+) and CD8(+) T-cell activation (P = .003 and .034, respectively) and significantly increased DNTCs (P = .038), compared with nonviremic subjects. GBV-C load correlated with DNTC percentage (P = .004). Thus, GBV-C infection is associated with an increase in DNTCs, which may contribute to reduced immune activation during HIV infection.
Assuntos
Terapia Antirretroviral de Alta Atividade/métodos , Vírus GB C/isolamento & purificação , Infecções por HIV/complicações , Infecções por HIV/tratamento farmacológico , Hepatite Viral Humana/complicações , Subpopulações de Linfócitos T/imunologia , Viremia/complicações , Complexo CD3/análise , Antígenos CD4/análise , Linfócitos T CD4-Positivos/química , Linfócitos T CD4-Positivos/imunologia , Antígenos CD8/análise , Linfócitos T CD8-Positivos/química , Linfócitos T CD8-Positivos/imunologia , Vírus GB C/patogenicidade , Infecções por HIV/imunologia , Hepatite Viral Humana/imunologia , Humanos , Ativação Linfocitária , Masculino , Pessoa de Meia-Idade , Subpopulações de Linfócitos T/química , Viremia/imunologiaRESUMO
Despite the ability of combination antiretroviral therapy (cART) to suppress viremia, there is persistence low levels of HIV proteins such as Transactivator of transcription (Tat) in the central nervous system (CNS), contributing to glial activation and neuroinflammation. Accumulating evidence also implicates the role of drugs of abuse in exacerbating neurological complications associated with HIV-1. The combined effects of HIV Tat, drugs of abuse, and cART can thus create a toxic milieu in the CNS. The present study investigated the combinatorial effects of HIV-Tat, cocaine, and cART on autophagy and NLRP3 inflammasome activation. We selected a combination of three commonly used cART regimens: tenofovir, emtricitabine, and dolutegravir. Our results demonstrated that exposure of mouse primary microglia (MPMs) to these agents-HIV Tat (25 ng/ml), cocaine (1 µM), and cART (1 µM each) resulted in upregulation of autophagy markers: Beclin1, LC3B-II, and SQSTM1 with impaired lysosomal functioning involving increased lysosomal pH, decreased LAMP2 and cathepsin D, ultimately leading to dysregulated autophagy. Our findings also demonstrated activation of the NLRP3 signaling in microglia exposed to these agents. We further demonstrated that gene silencing of key autophagy protein BECN1 significantly blocked NLRP3-mediated activation of microglia. Silencing of NLRP3, however, failed to block HIV Tat, cocaine, and cART-mediated dysregulation of the autophagy-lysosomal axis; these in vitro phenomena were also validated in vivo using iTat mice administered cocaine and cART. This study thus underscores the cooperative effects of HIV Tat, cocaine, and cART in exacerbating microglial activation involving dysregulated autophagy and activation of the NLRP3 inflammasome signaling.
Assuntos
Cocaína , Infecções por HIV , Camundongos , Animais , Microglia , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Cocaína/farmacologia , Inflamassomos/metabolismo , Transativadores/metabolismo , Transativadores/farmacologia , Autofagia , Produtos do Gene tat do Vírus da Imunodeficiência Humana/metabolismo , Infecções por HIV/metabolismoRESUMO
Accumulating evidence from several studies has shown association between substance use, dysregulation of the microbiome and psychiatric disorders such as depression, anxiety, and psychosis. Many of the abused substances such as cocaine and alcohol have been shown to alter immune signaling pathways and cause inflammation in both the periphery and the central nervous system (CNS). In addition, these substances of abuse also alter the composition and function of the gut microbiome which is known to play important roles such as the synthesis of neurotransmitters and metabolites, that affect the CNS homeostasis and consequent behavioral outcomes. The emerging interactions between substance use, microbiome and CNS neurochemical alterations could contribute to the development of psychiatric disorders. This review provides an overview of the associative effects of substance use such as alcohol, cocaine, methamphetamine, nicotine and opioids on the gut microbiome and psychiatric disorders involving anxiety, depression and psychosis. Understanding the relationship between substance use, microbiome and psychiatric disorders will provide insights for potential therapeutic targets, aimed at mitigating these adverse outcomes.
Assuntos
Cocaína , Microbioma Gastrointestinal , Transtornos Mentais , Microbiota , Transtornos Relacionados ao Uso de Substâncias , Cocaína/farmacologia , Etanol/farmacologia , Humanos , Transtornos Relacionados ao Uso de Substâncias/psicologiaRESUMO
Cocaine abuse is known to cause inflammation, oxidative injury and alterations in the gut microbiota. Although emerging studies have demonstrated the role of gut microbiota in modulating neurological complications and behavior, the mechanism(s) underlying these processes remain unclear. In the present study, we investigated the protective effect of Lactobacillus rhamnosus probiotic on cocaine-induced oxidative stress, glial activation, and locomotion in mice. In this study, groups of male C56BL6 mice were administered gut-resident commensal bacteria L. rhamnosus probiotic (oral gavage) concurrently with cocaine (20 mg/kg, i.p.) or saline for 28 days and assessed for oxidative stress and cellular activation in both the gut and brain as well as alterations in locomotion behavior. Cocaine-induced gut dysregulation was associated with increased formation of 4-hydroxynonenal (4-HNE) adducts, increased expression of pERK-1/2, pNF-kB-p65 and antioxidant mediators (SOD1, GPx1). In cocaine administered mice, there was increased activation of both microglia and astrocytes in the striatum and cortex of the brain as shown by enhanced expression of CD11b and GFAP, respectively. Cocaine administration also resulted in increased locomotor activity in the open field test in these mice. Administration of L. rhamnosus attenuated cocaine-induced gut oxidative stress and inflammation as well as glial activation and locomotion. These results suggest the potential of microbial-based interventions to attenuate cocaine-mediated behavioral responses and neuroinflammation, in addition to systemic inflammation and oxidative damage.
Assuntos
Cocaína , Lacticaseibacillus rhamnosus , Masculino , Animais , Camundongos , Cocaína/toxicidade , Antígeno CD11b , Locomoção , Estresse OxidativoRESUMO
Microglia, the primary immunocompetent cells of the brain, are suggested to play a role in the development of drug addiction. Previous studies have identified the microglia-derived pro-inflammatory factor IL1ß can promote the progression of cocaine addiction. Additionally, the activation status of microglia and "two-hit hypothesis" have been proposed in the field of drug addiction to explain how early life stress (ELS) could significantly increase the incidence of drug addiction in later life. However, the mechanisms underlying microglia prime and full activation and their roles in drug addiction remain greatly unexplored. Here, we employed CX3CR1-GFP mice (CX3CR1 functional deficiency, CX3CR1-/-) to explore whether primed microglia could potentiate cocaine-mediated behavioral changes and the possible underlying mechanisms. CX3CR1-/- mice revealed higher hyperlocomotion activity and conditional place preference than wild-type (WT) mice did under cocaine administration. In parallel, CX3CR1-/- mice showed higher activity of NLR family pyrin domain-containing 3 (NLRP3) inflammasome than WT mice. Interestingly, CX3CR1 deficiency itself could prime NLRP3 signaling by increasing the expression of NLPR3 and affect lysosome biogenesis under basal conditions. Taken together, our findings demonstrated that the functional status of microglia could have an impact on cocaine-mediated reward effects, and NLRP3 inflammasome activity was associated with this phenomenon. This study was consistent with the two-hit hypothesis and provided solid evidence to support the involvement of microglia in drug addiction. Targeting the NLRP3 inflammasome may represent a novel therapeutic approach for ameliorating or blocking the development of drug addiction.
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While opiates like morphine play a major role in the pharmacotherapy for the control of pain associated with various diseases, paradoxically, their long-term use is associated with cognitive impairments. Furthermore, morphine administration has been shown to result in neuronal synaptodendritic injury in rodent brains, leading to neurodegeneration. We recently reported the role of astrocytes as contributors of amyloidosis associated with HIV-associated neurological disorders. Herein we hypothesize that morphine could induce astrocytic amyloidosis, which, in turn, could be disseminated to various regions in the brain by astrocyte-derived EVs (ADEVs). In this study we demonstrate brain region-specific up-regulation of astrocytic amyloids in morphine dependendent rhesus macaques. In addition, herein we also demonstrate increased expression of ß-site cleaving enzyme (BACE1), APP, and Aß in human primary astrocytes (HPAs) exposed to morphine. Mechanisms involved in this process included the up-regulation of hypoxia-inducible factor (HIF-1α), its translocation and binding to the promoter of BACE1, leading to increased BACE1 activity and, generation of Aß 1-42. Gene silencing approaches confirmed the regulatory role of HIF-1α in BACE1 mediated amyloidosis leading to astrocyte activation and neuroinflammation. We next sought to assess whether morphine-stimulated ADEVs could carry amyloid cargoes. Results showed that morphine exposure induced the release of morphine-ADEVs, carrying amyloids. Interestingly, silencing HIF-1α in astrocytes not only reduced the numbers of ADEV released, but also the packaging of amyloid cargos in the ADEVs. These findings were further validated in brain derived EVs (BEVs) isolated from macaques, wherein it was shown that BEVs from morphine-dependent macaques, carried varieties of amyloid cargoes including the cytokine IL-1ß. This is the first report implicating the role of HIF-1α-BACE1 axis in morphine-mediated induction of astrocytic amyloidosis, leading, in turn, to neuroinflammation and release of the amyloid cargoes via ADEVs. These findings set the groundwork for the future development of therapeutic strategies for targeting cognitive deficits in chronic opiate users.
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Cocaine use disorder is a major health crisis that is associated with increased oxidative stress and neuroinflammation. While the role of NLRP3 inflammasome in mediating neuroinflammation is well-recognized, whether cocaine induces this response remains unexplored. Based on the premise that cocaine induces both reactive oxygen species (ROS) as well as microglial activation, we hypothesized that cocaine-mediated microglial activation involves both ROS and NLRP3 signaling pathways. We examined activation of the NLRP3 pathway in microglia exposed to cocaine, followed by validation in mice administered either cocaine or saline for 7 days, with or without pretreatment with the NLRP3 inhibitor, MCC950, and in postmortem cortical brain tissues of chronic cocaine-dependent humans. We found that microglia exposed to cocaine exhibited significant induction of NLRP3 and mature IL-1ß expression. Intriguingly, blockade of ROS (Tempol) attenuated cocaine-mediated priming of NLRP3 and microglial activation (CD11b). Blockade of NLRP3 by both pharmacological (MCC950) as well as gene silencing (siNLRP3) approaches underpinned the critical role of NLRP3 in cocaine-mediated activation of inflammasome and microglial activation. Pretreatment of mice with MCC950 followed by cocaine administration for 7 days mitigated cocaine-mediated upregulation of mature IL-1ß and CD11b, in both the striatum and the cortical regions. Furthermore, cortical brain tissues of chronic cocaine-dependent humans also exhibited upregulated expression of the NLRP3 pathway mediators compared with non-cocaine dependent controls. Collectively, these findings suggest that cocaine activates microglia involving the NLRP3 inflammasome pathway, thereby contributing to neuroinflammation. NLRP3 can thus be considered as a potential therapeutic target for alleviating cocaine-mediated neuroinflammation.
Assuntos
Cocaína/farmacologia , Inibidores da Captação de Dopamina/farmacologia , Lobo Frontal/efeitos dos fármacos , Inflamassomos/efeitos dos fármacos , Microglia/efeitos dos fármacos , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Animais , Caspase 1/metabolismo , Linhagem Celular , Lobo Frontal/metabolismo , Humanos , Inflamassomos/metabolismo , Interleucina-1beta/metabolismo , Camundongos , Microglia/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/genéticaRESUMO
Various research studies that have investigated the association between HIV infection and addiction underpin the role of various drugs of abuse in impairing immunological and non-immunological pathways of the host system, ultimately leading to augmentation of HIV infection and disease progression. These studies have included both in vitro and in vivo animal models wherein investigators have assessed the effects of various drugs on several disease parameters to decipher the impact of drugs on both HIV infection and progression of HIV-associated neurocognitive disorders (HAND). However, given the inherent limitations in the existing animal models of HAND, these investigations only recapitulated specific aspects of the disease but not the complex human syndrome. Despite the inability of HIV to infect rodents over the last 30 years, multiple strategies have been employed to develop several rodent models of HAND. While none of these models can accurately mimic the overall pathophysiology of HAND, they serve the purpose of modeling some unique aspects of HAND. This review provides an overview of various animal models used in the field and a careful evaluation of methodological strengths and limitations inherent in both the model systems and study designs to understand better how the various animal models complement one another.
Assuntos
Encéfalo/fisiopatologia , Modelos Animais de Doenças , Infecções por HIV/complicações , Transtornos Neurocognitivos/epidemiologia , Transtornos Relacionados ao Uso de Substâncias/epidemiologia , Animais , Encéfalo/virologia , Comorbidade , Infecções por HIV/psicologia , HIV-1/patogenicidade , Humanos , Camundongos , Transtornos Neurocognitivos/etiologia , Transtornos Neurocognitivos/fisiopatologia , Transtornos Neurocognitivos/psicologia , Ratos , Transtornos Relacionados ao Uso de Substâncias/fisiopatologia , Transtornos Relacionados ao Uso de Substâncias/psicologiaRESUMO
The advent of combined antiretroviral treatment (cART) as a treatment for HIV-1 infection has not only resulted in a dramatic decrease in the peripheral viral load but has also led to increased life expectancy of the infected individuals. Paradoxically, increased lifespan is accompanied with higher prevalence of age-related comorbidities, including HIV-associated neurocognitive disorders (HAND). Present study was aimed at exploring the role of HIV TAT protein in mediating microglial mitochondrial oxidative stress, ultimately resulting in neuroinflammation and microglial senescence. Our findings demonstrated that exposure of mouse primary microglial cells (mPMs) to HIV TAT protein resulted in a senescence-like phenotype, that was characterized by elevated expression of both p16 and p21 proteins, increased numbers of senescence-associated-ß-galactosidase positive cells, augmented cell-cycle arrest, increased release of proinflammatory cytokines and decreased telomerase activity. Additionally, exposure of mPMs to HIV TAT also resulted downregulation of SIRT3 with a concomitant increase in mitochondrial oxidative stress. Dual luciferase reporter assay identified miR-505 as a novel target of SIRT3, which was upregulated in mPMs exposed to HIV TAT. Furthermore, transient transfection of mPMs with either the SIRT3 plasmid or miRNA-505 inhibitor upregulated the expression of SIRT3 and mitochondrial antioxidant enzymes, with a concomitant decrease in microglial senescence. These in vitro findings were also validated in the prefrontal cortices and striatum of HIV transgenic rats as well as cART-treated HIV-infected individuals. In summary, this study underscores a yet undiscovered novel mechanism(s) underlying HIV TAT-mediated induction of senescence phenotype in microglia, involving the miR-505-SIRT3 axis-mediated induction of mitochondrial oxidative stress.
Assuntos
Infecções por HIV , Sirtuína 3 , Animais , Camundongos , Microglia/metabolismo , Estresse Oxidativo , Ratos , Sirtuína 3/genética , Produtos do Gene tat do Vírus da Imunodeficiência Humana/genética , Produtos do Gene tat do Vírus da Imunodeficiência Humana/metabolismoRESUMO
Neurological diseases are multifactorial, devastating diseases that are causative for various neurodegenerative disorders. Emerging evidence points that accumulation of unfolded, misfolded, insoluble, and damaged proteins inside the CNS cells such as microglia, astrocytes, neurons, oligodendrocytes, pericytes, and endothelial cells, leads to endoplasmic reticulum (ER) stress and dysregulated autophagy, which, in turn, sets the stage for ensuing neuropathogenesis. Studies have also demonstrated that chronic ER stress/unfolded protein response (UPR) activates autophagy, and conversely, that blockade of autophagy aggravates ER stress with ensuing cell death, in turn, leading to the development and progression of neurodegeneration. ER stress and autophagy signaling pathways are thus of particular interest as target(s) for pharmacological intervention for the development of therapeutic strategies for various neurological diseases. Herein, we summarized the current knowledge of chronic ER stress/UPR and autophagy signaling pathways and their regulation in CNS cells such as microglia, astrocytes, neurons, oligodendrocytes, pericytes, and endothelial cells. We also reviewed various neurological diseases wherein ER stress/UPR, and autophagy play key roles and also discussed possible pharmacological interventions involving these processes.
Assuntos
Autofagia/efeitos dos fármacos , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Doenças Neurodegenerativas/tratamento farmacológico , Fármacos Neuroprotetores/farmacologia , Animais , Humanos , Doenças Neurodegenerativas/metabolismoRESUMO
Although cocaine exposure has been shown to potentiate neuroinflammation by upregulating glial activation in the brain, the role of mitophagy in this process remains an enigma. In the present study, we sought to examine the role of impaired mitophagy in cocaine-mediated activation of microglia and to determine the ameliorative potential of superoxide dismutase mimetics in this context. Our findings demonstrated that exposure of mouse primary microglial cells (mPMs) to cocaine resulted in decreased mitochondrial membrane potential, that was accompanied by increased expression of mitophagy markers, PINK1 and PRKN. Exposure of microglia to cocaine also resulted in increased expression of DNM1L and OPTN with a concomitant decrease in the rate of mitochondrial oxygen consumption as well as impaired mitochondrial functioning. Additionally, in the presence of cocaine, microglia also exhibited upregulated expression of autophagosome markers, BECN1, MAP1LC3B-II, and SQSTM1. Taken together, these findings suggested diminished mitophagy flux and accumulation of mitophagosomes in the presence of cocaine. These findings were further confirmed by imaging techniques such as transmission electron microscopy and confocal microscopy. Cocaine-mediated activation of microglia was further monitored by assessing the expression of the microglial marker (ITGAM) and the inflammatory cytokine (Tnf, Il1b, and Il6) mRNAs. Pharmacological, as well as gene-silencing approaches aimed at blocking both the autophagy/mitophagy and SIGMAR1 expression, underscored the role of impaired mitophagy in cocaine-mediated activation of microglia. Furthermore, superoxide dismutase mimetics such as TEMPOL and MitoTEMPO were shown to alleviate cocaine-mediated impaired mitophagy as well as microglial activation.Abbreviations: 3-MA: 3-methyladenine; Δψm: mitochondrial membrane potential; ACTB: actin, beta; AIF1: allograft inflammatory factor 1; ATP: adenosine triphosphate; BAF: bafilomycin A1; BECN1: beclin 1, autophagy related; CNS: central nervous system; DNM1L: dynamin 1 like; DMEM: Dulbecco modified Eagle medium; DAPI: 4,6-Diamidino-2-phenylindole; DRD2: dopamine receptor D2; ECAR: extracellular acidification rate; FBS: fetal bovine serum; FCCP: Trifluoromethoxy carbonylcyanide phenylhydrazone; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; IL1B: interleukin 1, beta; IL6: interleukin 6; ITGAM: integrin subunit alpha M; MAP1LC3B: microtubule-associated protein 1 light chain 3 beta; mPMs: mouse primary microglial cells; MRC: maximal respiratory capacity; NFKB: nuclear factor kappa B; NLRP3: NLR family pyrin domain containing 3; NTRK2: neurotrophic receptor tyrosine kinase 2; OCR: oxygen consumption rate; OPTN: optineurin; PBS: phosphate buffered saline; PINK1: PTEN induced putative kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; ROS: reactive oxygen species; siRNA: small interfering RNA; SQSTM1: sequestosome 1; TNF: tumor necrosis factor.
Assuntos
Cocaína/efeitos adversos , Microglia/patologia , Mitocôndrias/patologia , Mitofagia , Superóxido Dismutase/metabolismo , Animais , Autofagia , Proteína Beclina-1/metabolismo , Biomarcadores/metabolismo , Células Cultivadas , Óxidos N-Cíclicos , Citocinas/metabolismo , Regulação para Baixo/efeitos dos fármacos , Inativação Gênica , Mediadores da Inflamação/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Microglia/efeitos dos fármacos , Microglia/metabolismo , Microglia/ultraestrutura , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/ultraestrutura , Mitofagia/efeitos dos fármacos , Modelos Biológicos , Fagossomos/efeitos dos fármacos , Fagossomos/metabolismo , Fagossomos/ultraestrutura , Proteínas Quinases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Receptores sigma/metabolismo , Transdução de Sinais , Regulação para Cima/efeitos dos fármacosRESUMO
Despite the ability of combination antiretroviral therapy to dramatically suppress viremia, the brain continues to be a reservoir of HIV-1 low-level replication. Adding further complexity to this is the comorbidity of drug abuse with HIV-1 associated neurocognitive disorders and neuroHIV. Among several abused drugs, the use of opiates is highly prevalent in HIV-1 infected individuals, both as an abused drug as well as for pain management. Opioids and their receptors have attained notable attention owing to their ability to modulate immune functions, in turn, impacting disease progression. Various cell culture, animal and human studies have implicated the role of opioids and their receptors in modulating viral replication and virus-mediated pathology both positively and negatively. Further, the combinatorial effects of HIV-1/HIV-1 proteins and morphine have demonstrated activation of inflammatory signaling in the host system. Herein, we summarized the current knowledge on the role of opioids on peripheral immunopathogenesis, viral immunopathogenesis, epigenetic profiles of the host and viral genome, neuropathogenesis of SIV/SHIV-infected non-human primates, blood-brain-barrier, HIV-1 viral latency, and viral rebound. Overall, this review provides recent insights into the role of opioids in HIV-1 immunopathogenesis. Graphical abstract.