RESUMEN
The novel coronavirus SARS-CoV-2 emerged in late 2019, rapidly reached pandemic status, and has maintained global ubiquity through the emergence of variants of concern. Efforts to develop animal models have mostly fallen short of recapitulating severe disease, diminishing their utility for research focusing on severe disease pathogenesis and life-saving medical countermeasures. We tested whether route of experimental infection substantially changes COVID-19 disease characteristics in two species of nonhuman primates (Macaca mulatta; rhesus macaques; RM, Chlorocebus atheiops; African green monkeys; AGM). Species-specific cohorts were experimentally infected with SARS-CoV-2 by either direct mucosal (intratracheal + intranasal) instillation or small particle aerosol in route-discrete subcohorts. Both species demonstrated analogous viral loads in all compartments by either exposure route although the magnitude and duration of viral loading was marginally greater in AGMs than RMs. Clinical onset was nearly immediate (+1dpi) in the mucosal exposure cohort whereas clinical signs and cytokine responses in aerosol exposure animals began +7dpi. Pathologies conserved in both species and both exposure modalities include pulmonary myeloid cell influx, development of pleuritis, and extended lack of regenerative capacity in the pulmonary compartment. Demonstration of conserved pulmonary pathology regardless of species and exposure route expands our understanding of how SARS-CoV-2 infection may lead to ARDS and/or functional lung damage and demonstrates the near clinical response of the nonhuman primate model for anti-fibrotic therapeutic evaluation studies.
Asunto(s)
COVID-19 , Aerosoles , Animales , Chlorocebus aethiops , Modelos Animales de Enfermedad , Humanos , Pulmón/patología , Macaca mulatta , SARS-CoV-2RESUMEN
BACKGROUND: Lyme neuroborreliosis, caused by the bacterium Borrelia burgdorferi affects both the central and peripheral nervous systems (CNS, PNS). The CNS manifestations, especially at later stages, can mimic/cause many other neurological conditions including psychiatric disorders, dementia, and others, with a likely neuroinflammatory basis. The pathogenic mechanisms associated with Lyme neuroborreliosis, however, are not fully understood. METHODS: In this study, using cultures of primary rhesus microglia, we explored the roles of several fibroblast growth factor receptors (FGFRs) and fibroblast growth factors (FGFs) in neuroinflammation associated with live B. burgdorferi exposure. FGFR specific siRNA and inhibitors, custom antibody arrays, ELISAs, immunofluorescence and microscopy were used to comprehensively analyze the roles of these molecules in microglial neuroinflammation due to B. burgdorferi. RESULTS: FGFR1-3 expressions were upregulated in microglia in response to B. burgdorferi. Inhibition of FGFR 1, 2 and 3 signaling using siRNA and three different inhibitors showed that FGFR signaling is proinflammatory in response to the Lyme disease bacterium. FGFR1 activation also contributed to non-viable B. burgdorferi mediated neuroinflammation. Analysis of the B. burgdorferi conditioned microglial medium by a custom antibody array showed that several FGFs are induced by the live bacterium including FGF6, FGF10 and FGF12, which in turn induce IL-6 and/or CXCL8, indicating a proinflammatory nature. To our knowledge, this is also the first-ever described role for FGF6 and FGF12 in CNS neuroinflammation. FGF23 upregulation, in addition, was observed in response to the Lyme disease bacterium. B. burgdorferi exposure also downregulated many FGFs including FGF 5, 7, 9, 11, 13, 16, 20 and 21. Some of the upregulated FGFs have been implicated in major depressive disorder (MDD) or dementia development, while the downregulated ones have been demonstrated to have protective roles in epilepsy, Parkinson's disease, Alzheimer's disease, spinal cord injury, blood-brain barrier stability, and others. CONCLUSIONS: In this study we show that FGFRs and FGFs are novel inducers of inflammatory mediators in Lyme neuroborreliosis. It is likely that an unresolved, long-term (neuro)-Lyme infection can contribute to the development of other neurologic conditions in susceptible individuals either by augmenting pathogenic FGFs or by suppressing ameliorative FGFs or both.
Asunto(s)
Borrelia burgdorferi , Demencia , Trastorno Depresivo Mayor , Enfermedad de Lyme , Neuroborreliosis de Lyme , Humanos , Microglía/patología , Enfermedades Neuroinflamatorias , Marco Interseccional , Receptores de Factores de Crecimiento de Fibroblastos , Factores de Crecimiento de Fibroblastos , ARN Interferente PequeñoRESUMEN
The novel coronavirus SARS-CoV-2, the causative agent of COVID-19 disease, has killed over five million people worldwide as of December 2021 with infections rising again due to the emergence of highly transmissible variants. Animal models that faithfully recapitulate human disease are critical for assessing SARS-CoV-2 viral and immune dynamics, for understanding mechanisms of disease, and for testing vaccines and therapeutics. Pigtail macaques (PTM, Macaca nemestrina) demonstrate a rapid and severe disease course when infected with simian immunodeficiency virus (SIV), including the development of severe cardiovascular symptoms that are pertinent to COVID-19 manifestations in humans. We thus proposed this species may likewise exhibit severe COVID-19 disease upon infection with SARS-CoV-2. Here, we extensively studied a cohort of SARS-CoV-2-infected PTM euthanized either 6- or 21-days after respiratory viral challenge. We show that PTM demonstrate largely mild-to-moderate COVID-19 disease. Pulmonary infiltrates were dominated by T cells, including CD4+ T cells that upregulate CD8 and express cytotoxic molecules, as well as virus-targeting T cells that were predominantly CD4+. We also noted increases in inflammatory and coagulation markers in blood, pulmonary pathologic lesions, and the development of neutralizing antibodies. Together, our data demonstrate that SARS-CoV-2 infection of PTM recapitulates important features of COVID-19 and reveals new immune and viral dynamics and thus may serve as a useful animal model for studying pathogenesis and testing vaccines and therapeutics.
Asunto(s)
COVID-19 , Modelos Animales de Enfermedad , Macaca nemestrina , Enfermedades de los Monos/virología , Animales , COVID-19/inmunología , COVID-19/patología , COVID-19/fisiopatología , COVID-19/virología , Humanos , Inmunidad Humoral , Pulmón/inmunología , Pulmón/virología , Masculino , Enfermedades de los Monos/inmunología , Enfermedades de los Monos/patología , Enfermedades de los Monos/fisiopatología , Linfocitos T/inmunologíaRESUMEN
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) induces a wide range of disease severity, ranging from asymptomatic infection to a life-threating illness, particularly in the elderly population and individuals with comorbid conditions. Among individuals with serious coronavirus 2019 (COVID-19) disease, acute respiratory distress syndrome (ARDS) is a common and often fatal presentation. Animal models of SARS-CoV-2 infection that manifest severe disease are needed to investigate the pathogenesis of COVID-19-induced ARDS and evaluate therapeutic strategies. We report two cases of ARDS in two aged African green monkeys (AGMs) infected with SARS-CoV-2 that had pathological lesions and disease similar to severe COVID-19 in humans. We also report a comparatively mild COVID-19 phenotype characterized by minor clinical, radiographic, and histopathologic changes in the two surviving, aged AGMs and four rhesus macaques (RMs) infected with SARS-CoV-2. Notable increases in circulating cytokines were observed in three of four infected, aged AGMs but not in infected RMs. All the AGMs had increased levels of plasma IL-6 compared with baseline, a predictive marker and presumptive therapeutic target in humans infected with SARS-CoV-2. Together, our results indicate that both RMs and AGMs are capable of modeling SARS-CoV-2 infection and suggest that aged AGMs may be useful for modeling severe disease manifestations, including ARDS.
Asunto(s)
COVID-19/etiología , Pulmón/virología , SARS-CoV-2/patogenicidad , Envejecimiento , Animales , Chlorocebus aethiops/virología , Infecciones por Coronavirus/tratamiento farmacológico , Citocinas/metabolismo , Humanos , Pulmón/patología , Macaca mulatta/virología , Carga Viral/métodosRESUMEN
Preclinical mouse models that recapitulate some characteristics of coronavirus disease (COVID-19) will facilitate focused study of pathogenesis and virus-host responses. Human agniotensin-converting enzyme 2 (hACE2) serves as an entry receptor for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to infect people via binding to envelope spike proteins. Herein we report development and characterization of a rapidly deployable COVID-19 mouse model. C57BL/6J (B6) mice expressing hACE2 in the lung were transduced by oropharyngeal delivery of the recombinant human adenovirus type 5 that expresses hACE2 (Ad5-hACE2). Mice were infected with SARS-CoV-2 at Day 4 after transduction and developed interstitial pneumonia associated with perivascular inflammation, accompanied by significantly higher viral load in lungs at Days 3, 6, and 12 after infection compared with Ad5-empty control group. SARS-CoV-2 was detected in pneumocytes in alveolar septa. Transcriptomic analysis of lungs demonstrated that the infected Ad5-hACE mice had a significant increase in IFN-dependent chemokines Cxcl9 and Cxcl10, and genes associated with effector T-cell populations including Cd3 g, Cd8a, and Gzmb. Pathway analysis showed that several Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were enriched in the data set, including cytokine-cytokine receptor interaction, the chemokine signaling pathway, the NOD-like receptor signaling pathway, the measles pathway, and the IL-17 signaling pathway. This response is correlative to clinical response in lungs of patients with COVID-19. These results demonstrate that expression of hACE2 via adenovirus delivery system sensitized the mouse to SARS-CoV-2 infection and resulted in the development of a mild COVID-19 phenotype, highlighting the immune and inflammatory host responses to SARS-CoV-2 infection. This rapidly deployable COVID-19 mouse model is useful for preclinical and pathogenesis studies of COVID-19.
Asunto(s)
Células Epiteliales Alveolares/inmunología , COVID-19/inmunología , Expresión Génica , SARS-CoV-2/inmunología , Transducción de Señal/inmunología , Adenoviridae/genética , Adenoviridae/metabolismo , Células Epiteliales Alveolares/metabolismo , Células Epiteliales Alveolares/virología , Enzima Convertidora de Angiotensina 2/biosíntesis , Enzima Convertidora de Angiotensina 2/genética , Enzima Convertidora de Angiotensina 2/inmunología , Animales , COVID-19/genética , COVID-19/metabolismo , COVID-19/patología , Citocinas/genética , Citocinas/inmunología , Modelos Animales de Enfermedad , Humanos , Ratones , Ratones Transgénicos , SARS-CoV-2/genética , SARS-CoV-2/metabolismo , Transducción de Señal/genética , Transducción GenéticaRESUMEN
Antiretroviral therapy in HIV patients has lengthened lifespan but led to an increased risk for secondary comorbidities, such as pulmonary complications characterized by vascular dysfunction. In the lung, PDGFRß+ mesenchymal cells known as pericytes intimately associate with endothelial cells and are key for their survival both structurally and through the secretion of prosurvival factors. We hypothesize that in HIV infection there are functional changes in pericytes that may lead to destabilization of the microvasculature and ultimately to pulmonary abnormalities. Our objective in this study was to determine whether lung pericytes could be directly infected with HIV. We leveraged lung samples from macaque lungs with or without SIV infection and normal human lung for in vitro experiments. Pericytes were isolated based on the marker platelet-derived growth factor receptor-ß (PDGFRß). We determined that lung PDGFRß-positive (PDGFRß+) pericytes from both macaques and humans express CD4, the primary receptor for SIV/HIV, as well as the major coreceptors CXCR4 and CCR5. We found cells positive for both PDGFRß and SIV in lungs from infected macaques. Lung pericytes isolated from these animals also harbored detectable SIV. To confirm relevance to human disease, we demonstrated that human lung pericytes are capable of being productively infected by HIV in vitro, with the time course of infection suggesting development of viral latency. In summary, we show for the first time that SIV/HIV directly infects lung pericytes, implicating these cells as a novel target and potential reservoir for the virus in vivo.
Asunto(s)
Células Endoteliales/virología , Infecciones por VIH/virología , Pulmón/virología , Macrófagos/virología , Linfocitos T CD4-Positivos/virología , Humanos , Pulmón/inmunología , Macrófagos/inmunología , Receptores CXCR4/inmunología , Virus de la Inmunodeficiencia de los Simios/patogenicidad , Latencia del Virus/fisiología , Replicación ViralRESUMEN
Recent studies have identified circular RNAs (circRNAs) expressed from the Epstein-Barr virus (EBV) and Kaposi's sarcoma herpesvirus (KSHV) human DNA tumor viruses. To gain initial insights into the potential relevance of EBV circRNAs in virus biology and disease, we assessed the circRNAome of the interspecies homologue rhesus macaque lymphocryptovirus (rLCV) in a naturally occurring lymphoma from a simian immunodeficiency virus (SIV)-infected rhesus macaque. This analysis revealed rLCV orthologues of the latency-associated EBV circular RNAs circRPMS1_E4_E3a and circEBNA_U. Also identified in two samples displaying unusually high lytic gene expression was a novel rLCV circRNA that contains both conserved and rLCV-specific RPMS1 exons and whose backsplice junctions flank an rLCV lytic origin of replication (OriLyt). Analysis of a lytic infection model for the murid herpesvirus 68 (MHV68) rhadinovirus identified a cluster of circRNAs near an MHV68 lytic origin of replication, with the most abundant of these, circM11_ORF69, spanning the OriLyt. Lastly, analysis of KSHV latency and reactivation models revealed the latency associated circRNA originating from the vIRF4 gene as the predominant viral circRNA. Together, the results of this study broaden our appreciation for circRNA repertoires in the Lymphocryptovirus and Rhadinovirus genera of gammaherpesviruses and provide evolutionary support for viral circRNA functions in latency and viral replication.IMPORTANCE Infection with oncogenic gammaherpesviruses leads to long-term viral persistence through a dynamic interplay between the virus and the host immune system. Critical for remodeling of the host cell environment after the immune responses are viral noncoding RNAs that modulate host signaling pathways without attracting adaptive immune recognition. Despite the importance of noncoding RNAs in persistent infection, the circRNA class of noncoding RNAs has only recently been identified in gammaherpesviruses. Accordingly, their roles in virus infection and associated oncogenesis are unknown. Here we report evolutionary conservation of EBV-encoded circRNAs determined by assessing the circRNAome in rLCV-infected lymphomas from an SIV-infected rhesus macaque, and we report latent and lytic circRNAs from KSHV and MHV68. These experiments demonstrate utilization of the circular RNA class of RNAs across 4 members of the gammaherpesvirus subfamily, and they identify orthologues and potential homoplastic circRNAs, implying conserved circRNA functions in virus biology and associated malignancies.
Asunto(s)
Gammaherpesvirinae/genética , ARN/genética , Animales , Línea Celular , Regulación Viral de la Expresión Génica/genética , Herpesvirus Humano 4/genética , Herpesvirus Humano 8/genética , Humanos , Lymphocryptovirus/genética , Macaca mulatta , Masculino , ARN Circular , ARN Viral/genética , Rhadinovirus/genética , Virus de la Inmunodeficiencia de los Simios/genética , Latencia del Virus/genética , Replicación Viral/genéticaRESUMEN
BACKGROUND: The BTB domain of B-cell lymphoma 6 (BCL6) protein was identified as a therapeutic target for B-cell lymphoma. This study compared the pharmacokinetics (PK) of the BCL6 BTB inhibitor (FX1) between mice and macaques, as well as evaluating its lymphoid suppressive effect in uninfected macaques with lymphoid hyperplasia. MATERIALS AND METHODS: Eight uninfected adult Indian rhesus macaques (Macaca mulatta) were used in the study, four animals carrying lymphoid tissue hyperplasia. Plasma FX1 levels were measured by HPLC-MS/MS. Lymph node biopsies were used for H&E and immunohistochemistry staining, as well as mononuclear cell isolation for flow cytometry analysis. RESULTS: Inhibition of the BCL6 BTB domain with FX1 led to a reduction in the frequency of GC, Tfh CD4+ , and Tfh precursor cells, as well as resolving lymphoid hyperplasia, in rhesus macaques. CONCLUSIONS: B-cell lymphoma 6 inhibition may represent a novel strategy to reduce hyperplastic lymphoid B-cell follicles and decrease Tfh cells.
Asunto(s)
Hiperplasia/tratamiento farmacológico , Indoles/farmacología , Proteínas Proto-Oncogénicas c-bcl-6/antagonistas & inhibidores , Células T Auxiliares Foliculares/efectos de los fármacos , Tiazolidinedionas/farmacología , Animales , Linfocitos/efectos de los fármacos , Linfocitos/patología , Masculino , Ratones , Células T Auxiliares Foliculares/fisiologíaRESUMEN
Background: Tuberculosis (TB) and human immunodeficiency virus (HIV)/acquired immune deficiency syndrome (AIDS) profoundly affect the immune system and synergistically accelerate disease progression. It is believed that CD4+ T-cell depletion by HIV is the major cause of immunodeficiency and reactivation of latent TB. Previous studies demonstrated that blood monocyte turnover concurrent with tissue macrophage death from virus infection better predicted AIDS onset than CD4+ T-cell depletion in macaques infected with simian immunodeficiency virus (SIV). Methods: In this study, we describe the contribution of macrophages to the pathogenesis of Mycobacterium tuberculosis (Mtb)/SIV coinfection in a rhesus macaque model using in vivo BrdU labeling, immunostaining, flow cytometry, and confocal microscopy. Results: We found that increased monocyte and macrophage turnover and levels of SIV-infected lung macrophages correlated with TB reactivation. All Mtb/SIV-coinfected monkeys exhibited declines in CD4+ T cells regardless of reactivation or latency outcomes, negating lower CD4+ T-cell levels as a primary cause of Mtb reactivation. Conclusions: Results suggest that SIV-related damage to macrophages contributes to Mtb reactivation during coinfection. This also supports strategies to target lung macrophages for the treatment of TB.
Asunto(s)
Tuberculosis Latente/inmunología , Macrófagos Alveolares/inmunología , Monocitos/inmunología , Síndrome de Inmunodeficiencia Adquirida del Simio/inmunología , Virus de la Inmunodeficiencia de los Simios/inmunología , Tuberculosis/inmunología , Animales , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/microbiología , Linfocitos T CD4-Positivos/virología , Coinfección/inmunología , Coinfección/microbiología , Coinfección/virología , Modelos Animales de Enfermedad , Tuberculosis Latente/microbiología , Tuberculosis Latente/virología , Depleción Linfocítica/métodos , Macaca mulatta , Macrófagos Alveolares/microbiología , Macrófagos Alveolares/virología , Masculino , Monocitos/microbiología , Monocitos/virología , Mycobacterium tuberculosis/inmunología , Síndrome de Inmunodeficiencia Adquirida del Simio/microbiología , Síndrome de Inmunodeficiencia Adquirida del Simio/virología , Tuberculosis/microbiología , Tuberculosis/virología , Carga Viral/inmunologíaRESUMEN
To our knowledge, this study demonstrates for the first time that the AIDS virus differentially impacts two distinct subsets of lung macrophages. The predominant macrophages harvested by bronchoalveolar lavage (BAL), alveolar macrophages (AMs), are routinely used in studies on human lung macrophages, are long-lived cells, and exhibit low turnover. Interstitial macrophages (IMs) inhabit the lung tissue, are not recovered with BAL, are shorter-lived, and exhibit higher baseline turnover rates distinct from AMs. We examined the effects of SIV infection on AMs in BAL fluid and IMs in lung tissue of rhesus macaques. SIV infection produced massive cell death of IMs that contributed to lung tissue damage. Conversely, SIV infection induced minimal cell death of AMs, and these cells maintained the lower turnover rate throughout the duration of infection. This indicates that SIV produces lung tissue damage through destruction of IMs, whereas the longer-lived AMs may serve as a virus reservoir to facilitate HIV persistence.
Asunto(s)
Enfermedades Pulmonares/inmunología , Pulmón/inmunología , Macrófagos Alveolares/inmunología , Síndrome de Inmunodeficiencia Adquirida del Simio/inmunología , Virus de la Inmunodeficiencia de los Simios/inmunología , Animales , Muerte Celular/inmunología , Humanos , Pulmón/patología , Enfermedades Pulmonares/patología , Macaca mulatta , Macrófagos Alveolares/patología , Síndrome de Inmunodeficiencia Adquirida del Simio/patologíaRESUMEN
UNLABELLED: Like varicella-zoster virus (VZV), simian varicella virus (SVV) reactivates to produce zoster. In the present study, 5 rhesus macaques were inoculated intrabronchially with SVV, and 5 months later, 4 monkeys were immunosuppressed; 1 monkey was not immunosuppressed but was subjected to the stress of transportation. In 4 monkeys, a zoster rash developed 7 to 12 weeks after immunosuppression, and a rash also developed in the monkey that was not immunosuppressed. Analysis at 24 to 48 h after zoster revealed SVV antigen in the lung alveolar wall, in ganglionic neurons and nonneuronal cells, and in skin and in lymph nodes. In skin, SVV was found primarily in sweat glands. In lymph nodes, the SVV antigen colocalized mostly with macrophages, dendritic cells, and, to a lesser extent, T cells. The presence of SVV in lymph nodes, as verified by quantitative PCR detection of SVV DNA, might reflect the sequestration of virus by macrophages and dendritic cells in lymph nodes or the presentation of viral antigens to T cells to initiate an immune response against SVV, or both. IMPORTANCE: VZV causes varicella (chickenpox), becomes latent in ganglia, and reactivates to produce zoster and multiple other serious neurological disorders. SVV in nonhuman primates has proved to be a useful model in which the pathogenesis of the virus parallels the pathogenesis of VZV in humans. Here, we show that SVV antigens are present in sweat glands in skin and in macrophages and dendritic cells in lymph nodes after SVV reactivation in monkeys, raising the possibility that macrophages and dendritic cells in lymph nodes serve as antigen-presenting cells to activate T cell responses against SVV after reactivation.
Asunto(s)
Herpes Zóster/patología , Herpes Zóster/virología , Ganglios Linfáticos/virología , Varicellovirus/fisiología , Activación Viral/fisiología , Animales , Chlorocebus aethiops , ADN Viral/análisis , Células Dendríticas/virología , Técnica del Anticuerpo Fluorescente , Inmunohistoquímica , Terapia de Inmunosupresión , Ganglios Linfáticos/citología , Macaca mulatta , Macrófagos/virología , Reacción en Cadena en Tiempo Real de la Polimerasa , Piel/patología , Piel/virología , Linfocitos T/virología , Células VeroRESUMEN
UNLABELLED: Deletion of Gly-720 and Tyr-721 from a highly conserved GYxxØ trafficking signal in the SIVmac239 envelope glycoprotein cytoplasmic domain, producing a virus termed ΔGY, leads to a striking perturbation in pathogenesis in rhesus macaques (Macaca mulatta). Infected macaques develop immune activation and progress to AIDS, but with only limited and transient infection of intestinal CD4(+) T cells and an absence of microbial translocation. Here we evaluated ΔGY in pig-tailed macaques (Macaca nemestrina), a species in which SIVmac239 infection typically leads to increased immune activation and more rapid progression to AIDS than in rhesus macaques. In pig-tailed macaques, ΔGY also replicated acutely to high peak plasma RNA levels identical to those for SIVmac239 and caused only transient infection of CD4(+) T cells in the gut lamina propria and no microbial translocation. However, in marked contrast to rhesus macaques, 19 of 21 pig-tailed macaques controlled ΔGY replication with plasma viral loads of <15 to 50 RNA copies/ml. CD4(+) T cells were preserved in blood and gut for up to 100 weeks with no immune activation or disease progression. Robust antiviral CD4(+) T cell responses were seen, particularly in the gut. Anti-CD8 antibody depletion demonstrated CD8(+) cellular control of viral replication. Two pig-tailed macaques progressed to disease with persisting viremia and possible compensatory mutations in the cytoplasmic tail. These studies demonstrate a marked perturbation in pathogenesis caused by ΔGY's ablation of the GYxxØ trafficking motif and reveal, paradoxically, that viral control is enhanced in a macaque species typically predisposed to more pathogenic manifestations of simian immunodeficiency virus (SIV) infection. IMPORTANCE: The pathogenesis of human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) reflects a balance between viral replication, host innate and adaptive antiviral immune responses, and sustained immune activation that in humans and Asian macaques is associated with persistent viremia, immune escape, and AIDS. Among nonhuman primates, pig-tailed macaques following SIV infection are predisposed to more rapid disease progression than are rhesus macaques. Here, we show that disruption of a conserved tyrosine-based cellular trafficking motif in the viral transmembrane envelope glycoprotein cytoplasmic tail leads in pig-tailed macaques to a unique phenotype in which high levels of acute viral replication are followed by elite control, robust cellular responses in mucosal tissues, and no disease. Paradoxically, control of this virus in rhesus macaques is only partial, and progression to AIDS occurs. This novel model should provide a powerful tool to help identify host-specific determinants for viral control with potential relevance for vaccine development.
Asunto(s)
Secuencias de Aminoácidos , Linfocitos T CD4-Positivos/inmunología , Inmunidad Mucosa , Macaca nemestrina/virología , Eliminación de Secuencia , Síndrome de Inmunodeficiencia Adquirida del Simio/inmunología , Proteínas del Envoltorio Viral/inmunología , Animales , Linfocitos T CD4-Positivos/virología , Progresión de la Enfermedad , Femenino , Expresión Génica , Intestinos/inmunología , Intestinos/virología , Macaca mulatta/virología , Masculino , Datos de Secuencia Molecular , Membrana Mucosa/inmunología , Membrana Mucosa/virología , Señales de Clasificación de Proteína , Estructura Terciaria de Proteína , Transporte de Proteínas , Síndrome de Inmunodeficiencia Adquirida del Simio/patología , Síndrome de Inmunodeficiencia Adquirida del Simio/virología , Virus de la Inmunodeficiencia de los Simios/genética , Virus de la Inmunodeficiencia de los Simios/inmunología , Especificidad de la Especie , Proteínas del Envoltorio Viral/deficiencia , Proteínas del Envoltorio Viral/genética , Carga Viral/genética , Carga Viral/inmunología , Viremia/inmunología , Viremia/patología , Replicación Viral/genética , Replicación Viral/inmunologíaRESUMEN
Macrophage recruitment to the central nervous system (CNS) during AIDS pathogenesis is poorly understood. We measured the accumulation of brain perivascular (CD163(+)) and inflammatory (MAC387(+)) macrophages in SIV-infected monkeys. Monocyte progenitors were 5-bromo-2'-deoxyuridine (BrdU) labeled in bone marrow, and CNS macrophages were labeled serially with fluorescent dextrans injected into the cisterna magna. MAC387(+) macrophages accumulated in the meninges and choroid plexus in early inflammation and in the perivascular space and SIV encephalitis (SIVE) lesions late. CD163(+) macrophages accumulated in the perivascular space and SIVE lesions with late inflammation. Most of the BrdU(+) cells were MAC387(+); however, CD163(+)BrdU(+) macrophages were present in the meninges and choroid plexus with AIDS. Most (81.6% ± 1.8%) of macrophages in SIVE lesions were present in the CNS before SIVE lesion formation. There was a 2.9-fold increase in SIVp28(+) macrophages entering the CNS late compared with those entering early (P < 0.05). The rate of CD163(+) macrophage recruitment to the CNS inversely correlated with time to death (P < 0.03) and increased with SIVE. In SIVE animals, soluble CD163 correlated with CD163(+) macrophage recruitment (P = 0.02). Most perivascular macrophages that comprise SIVE lesions and multinucleated giant cells are present in the CNS early, before SIVE lesions are formed. Most SIV-infected macrophages traffic to the CNS terminally with AIDS.
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Encéfalo/patología , Encefalitis/patología , Macrófagos/patología , Síndrome de Inmunodeficiencia Adquirida del Simio/patología , Virus de la Inmunodeficiencia de los Simios , Animales , Encéfalo/metabolismo , Encéfalo/virología , Encefalitis/metabolismo , Encefalitis/virología , Células Gigantes/metabolismo , Células Gigantes/patología , Células Gigantes/virología , Macaca mulatta , Macrófagos/metabolismo , Macrófagos/virología , Masculino , Síndrome de Inmunodeficiencia Adquirida del Simio/metabolismo , Síndrome de Inmunodeficiencia Adquirida del Simio/virologíaRESUMEN
Mycobacterium tuberculosis (Mtb) must counter hypoxia within granulomas to persist. DosR, in concert with sensor kinases DosS and DosT, regulates the response to hypoxia. Yet Mtb lacking functional DosR colonize the lungs of C57Bl/6 mice, presumably owing to the lack of organized lesions with sufficient hypoxia in that model. We compared the phenotype of the Δ-dosR, Δ-dosS, and Δ-dosT mutants to Mtb using C3HeB/FeJ mice, an alternate mouse model where lesions develop hypoxia. C3HeB/FeJ mice were infected via aerosol. The progression of infection was analyzed by tissue bacterial burden and histopathology. A measure of the comparative global immune responses was also analyzed. Although Δ-dosR and Δ-dosT grew comparably to wild-type Mtb, Δ-dosS exhibited a significant defect in bacterial burden and pathology in vivo, accompanied by ablated proinflammatory response. Δ-dosS retained the ability to induce DosR. The Δ-dosS mutant was also attenuated in murine macrophages ex vivo, with evidence of reduced expression of the proinflammatory signature. Our results show that DosS, but not DosR and DosT, is required by Mtb to survive in C3HeB/FeJ mice. The attenuation of Δ-dosS is not due to its inability to induce the DosR regulon, nor is it a result of the accumulation of hypoxia. That the in vivo growth restriction of Δ-dosS could be mimicked ex vivo suggested sensitivity to macrophage oxidative burst. Anoxic caseous centers within tuberculosis lesions eventually progress to cavities. Our results provide greater insight into the molecular mechanisms of Mtb persistence within host lungs.
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Proteínas Bacterianas/genética , Granuloma del Sistema Respiratorio/microbiología , Mycobacterium tuberculosis/patogenicidad , Protamina Quinasa/genética , Tuberculosis Pulmonar/microbiología , Animales , Proteínas Bacterianas/metabolismo , Hipoxia de la Célula , Células Cultivadas , Regulación Bacteriana de la Expresión Génica , Macrófagos/microbiología , Masculino , Ratones Endogámicos C3H , Viabilidad Microbiana , Mycobacterium tuberculosis/genética , Protamina Quinasa/metabolismo , Regulón , VirulenciaRESUMEN
Here we show that simian immunodeficiency virus (SIV) infection of rhesus macaques results in rapid upregulation of tetherin (BST-2 or CD317) on peripheral blood lymphocytes, including the CD4(+) CCR5(+) T cell targets of virus infection, with a peak of induction that coincides with peak alpha interferon (IFN-α) levels in plasma, and that tetherin remains above baseline levels throughout chronic infection. These observations are consistent with a role for tetherin in innate immunity to immunodeficiency virus infection.
Asunto(s)
Antígenos CD/genética , Síndrome de Inmunodeficiencia Adquirida del Simio/genética , Virus de la Inmunodeficiencia de los Simios/fisiología , Regulación hacia Arriba , Animales , Antígenos CD/inmunología , Macaca , Síndrome de Inmunodeficiencia Adquirida del Simio/inmunología , Síndrome de Inmunodeficiencia Adquirida del Simio/virología , Virus de la Inmunodeficiencia de los Simios/genéticaRESUMEN
A hallmark of pathogenic simian immunodeficiency virus (SIV) and human immunodeficiency virus (HIV) infections is the rapid and near-complete depletion of mucosal CD4(+) T lymphocytes from the gastrointestinal tract. Loss of these cells and disruption of epithelial barrier function are associated with microbial translocation, which has been proposed to drive chronic systemic immune activation and disease progression. Here, we evaluate in rhesus macaques a novel attenuated variant of pathogenic SIVmac239, termed ΔGY, which contains a deletion of a Tyr and a proximal Gly from a highly conserved YxxØ trafficking motif in the envelope cytoplasmic tail. Compared to SIVmac239, ΔGY established a comparable acute peak of viremia but only transiently infected lamina propria and caused little or no acute depletion of mucosal CD4(+) T cells and no detectable microbial translocation. Nonetheless, these animals developed T-cell activation and declining peripheral blood CD4(+) T cells and ultimately progressed with clinical or pathological features of AIDS. ΔGY-infected animals also showed no infection of macrophages or central nervous system tissues even in late-stage disease. Although the ΔGY mutation persisted, novel mutations evolved, including the formation of new YxxØ motifs in two of four animals. These findings indicate that disruption of this trafficking motif by the ΔGY mutation leads to a striking alteration in anatomic distribution of virus with sparing of lamina propria and a lack of microbial translocation. Because these animals exhibited wild-type levels of acute viremia and immune activation, our findings indicate that these pathological events are dissociable and that immune activation unrelated to gut damage can be sufficient for the development of AIDS.
Asunto(s)
Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/virología , Síndrome de Inmunodeficiencia Adquirida del Simio/inmunología , Síndrome de Inmunodeficiencia Adquirida del Simio/virología , Virus de la Inmunodeficiencia de los Simios/patogenicidad , Proteínas del Envoltorio Viral/metabolismo , Factores de Virulencia/metabolismo , Animales , Sistema Nervioso Central/virología , Progresión de la Enfermedad , Femenino , Tracto Gastrointestinal/inmunología , Mucosa Intestinal/inmunología , Macaca mulatta , Macrófagos/virología , Masculino , Datos de Secuencia Molecular , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Transporte de Proteínas , ARN Viral/genética , Análisis de Secuencia de ADN , Eliminación de Secuencia , Síndrome de Inmunodeficiencia Adquirida del Simio/patología , Proteínas del Envoltorio Viral/genética , Factores de Virulencia/genéticaRESUMEN
Virus-induced accelerated aging has been proposed as a potential mechanism underlying the persistence of HIV-associated neurocognitive disorders (HAND) despite advances in access and adherence to combination antiretroviral therapies (cART). While some studies have demonstrated evidence of accelerated aging in PLWH, studies examining acute infection, and cART intervention are limited, with most studies being in vitro or utilizing small animal models. Here, we utilized FFPE tissues from Simian immunodeficiency virus (SIV) infected rhesus macaques to assess the levels of two proteins commonly associated with aging - the cellular senescence marker p16 INK4a (p16) and the NAD-dependent deacetylase sirtuin 1 (SIRT1). Our central hypothesis was that SIV infection induces accelerated aging phenotypes in the brain characterized by increased expression of p16 and altered expression of SIRT1 that correlate with increased neurodegeneration, and that cART inhibits this process. We found that SIV infection induced increased GFAP, p16, SIRT1, and neurodegeneration in multiple brain regions, and treatment with cART reduced GFAP expression in SIV-infected animals and thus likely decreases inflammation in the brain. Importantly, cART reversed SIV-induced accelerated aging (p16 and SIRT1) and neurodegeneration in the frontal lobe and hippocampus. Combined, these data suggest that cART is both safe and effective in reducing neuroinflammation and age-associated alterations in astrocytes that contribute to neurodegeneration, providing possible therapeutic targets in the treatment of HAND.
RESUMEN
Chimeric antigen receptor (CAR)-T cells have demonstrated clinical potential, but current receptors still need improvements to be successful against chronic HIV infection. In this study, we address some requirements of CAR motifs for strong surface expression of a novel anti-HIV CAR by evaluating important elements in the extracellular, hinge, and transmembrane (TM) domains. When combining a truncated CD4 extracellular domain and CD8α hinge/TM, the novel CAR did not express extracellularly but was detectable intracellularly. By shortening the CD8α hinge, CD4-CAR surface expression was partially recovered and addition of the LYC motif at the end of the CD8α TM fully recovered both intracellular and extracellular CAR expression. Mutation of LYC to TTA or TTC showed severe abrogation of CAR expression by flow cytometry and confocal microscopy. Additionally, we determined that CD4-CAR surface expression could be maximized by the removal of FQKAS motif at the junction of the extracellular domain and the hinge region. CD4-CAR surface expression also resulted in cytotoxic CAR T cell killing of HIV Env+ target cells. In this study, we identified elements that are crucial for optimal CAR surface expression, highlighting the need for structural analysis studies to establish fundamental guidelines of CAR designs.
Asunto(s)
Antígenos CD4 , Receptores Quiméricos de Antígenos , Humanos , Receptores Quiméricos de Antígenos/inmunología , Receptores Quiméricos de Antígenos/metabolismo , Receptores Quiméricos de Antígenos/genética , Antígenos CD4/metabolismo , Antígenos CD4/inmunología , Infecciones por VIH/inmunología , Dominios Proteicos , Células HEK293 , Antígenos CD8/inmunología , Antígenos CD8/metabolismo , Membrana Celular/metabolismo , Secuencias de Aminoácidos , VIH-1/inmunologíaRESUMEN
Obesity is a risk factor for developing severe COVID-19. However, the mechanism underlying obesity-accelerated COVID-19 remains unclear. Here, we report results from a study in which 2-3-month-old K18-hACE2 (K18) mice were fed a western high-fat diet (WD) or normal chow (NC) over 3 months before intranasal infection with a sublethal dose of SARS-CoV2 WA1 (a strain ancestral to the Wuhan variant). After infection, the WD-fed K18 mice lost significantly more body weight and had more severe lung inflammation than normal chow (NC)-fed mice. Bulk RNA-seq analysis of lungs and adipose tissue revealed a diverse landscape of various immune cells, inflammatory markers, and pathways upregulated in the infected WD-fed K18 mice when compared with the infected NC-fed control mice. The transcript levels of IL-6, an important marker of COVID-19 disease severity, were upregulated in the lung at 6-9 days post-infection in the WD-fed mice when compared to NC-fed mice. Transcriptome analysis of the lung and adipose tissue obtained from deceased COVID-19 patients found that the obese patients had an increase in the expression of genes and the activation of pathways associated with inflammation as compared to normal-weight patients (n = 2). The K18 mouse model and human COVID-19 patient data support a link between inflammation and an obesity-accelerated COVID-19 disease phenotype. These results also indicate that obesity-accelerated severe COVID-19 caused by SARS-CoV-2 WA1 infection in the K18 mouse model would be a suitable model for dissecting the cellular and molecular mechanisms underlying pathogenesis.
Asunto(s)
COVID-19 , Pulmón , Obesidad , SARS-CoV-2 , Regulación hacia Arriba , COVID-19/genética , COVID-19/virología , COVID-19/metabolismo , COVID-19/patología , Animales , Obesidad/genética , Obesidad/metabolismo , Obesidad/complicaciones , Ratones , Humanos , Pulmón/metabolismo , Pulmón/patología , Pulmón/virología , Dieta Alta en Grasa/efectos adversos , Inflamación/genética , Inflamación/patología , Inflamación/metabolismo , Modelos Animales de Enfermedad , Interleucina-6/genética , Interleucina-6/metabolismo , Masculino , Tejido Adiposo/metabolismo , Tejido Adiposo/patología , Índice de Severidad de la Enfermedad , Enzima Convertidora de Angiotensina 2/genética , Enzima Convertidora de Angiotensina 2/metabolismoRESUMEN
Hyperglycemia, and exacerbation of pre-existing deficits in glucose metabolism, are manifestations of the post-acute sequelae of SARS-CoV-2. Our understanding of metabolic decline after acute COVID-19 remains unclear due to the lack of animal models. Here, we report a non-human primate model of metabolic post-acute sequelae of SARS-CoV-2 using SARS-CoV-2 infected African green monkeys. Using this model, we identify a dysregulated blood chemokine signature during acute COVID-19 that correlates with elevated and persistent hyperglycemia four months post-infection. Hyperglycemia also correlates with liver glycogen levels, but there is no evidence of substantial long-term SARS-CoV-2 replication in the liver and pancreas. Finally, we report a favorable glycemic effect of the SARS-CoV-2 mRNA vaccine, administered on day 4 post-infection. Together, these data suggest that the African green monkey model exhibits important similarities to humans and can be utilized to assess therapeutic candidates to combat COVID-related metabolic defects.