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1.
Vaccine ; 41 Suppl 2: S53-S75, 2023 11 03.
Artigo em Inglês | MEDLINE | ID: mdl-37806805

RESUMO

Cytomegalovirus (CMV) is the most common infectious cause of congenital malformation and a leading cause of developmental disabilities such as sensorineural hearing loss (SNHL), motor and cognitive deficits. The significant disease burden from congenital CMV infection (cCMV) led the US National Institute of Medicine to rank CMV vaccine development as the highest priority. An average of 6.7/1000 live births are affected by cCMV, but the prevalence varies across and within countries. In contrast to other congenital infections such as rubella and toxoplasmosis, the prevalence of cCMV increases with CMV seroprevalence rates in the population. The true global burden of cCMV disease is likely underestimated because most infected infants (85-90 %) have asymptomatic infection and are not identified. However, about 7-11 % of those with asymptomatic infection will develop SNHL throughout early childhood. Although no licensed CMV vaccine exists, several candidate vaccines are in development, including one currently in phase 3 trials. Licensure of one or more vaccine candidates is feasible within the next five years. Various models of CMV vaccine strategies employing different target populations have shown to provide substantial benefit in reducing cCMV. Although CMV can cause end-organ disease with significant morbidity and mortality in immunocompromised individuals, the focus of this vaccine value profile (VVP) is on preventing or reducing the cCMV disease burden. This CMV VVP provides a high-level, comprehensive assessment of the currently available data to inform the potential public health, economic, and societal value of CMV vaccines. The CMV VVP was developed by a working group of subject matter experts from academia, public health groups, policy organizations, and non-profit organizations. All contributors have extensive expertise on various elements of the CMV VVP and have described the state of knowledge and identified the current gaps. The VVP was developed using only existing and publicly available information.


Assuntos
Infecções por Citomegalovirus , Vacinas contra Citomegalovirus , Perda Auditiva Neurossensorial , Lactente , Humanos , Pré-Escolar , Citomegalovirus , Infecções Assintomáticas , Estudos Soroepidemiológicos , Infecções por Citomegalovirus/epidemiologia , Infecções por Citomegalovirus/prevenção & controle , Perda Auditiva Neurossensorial/congênito , Perda Auditiva Neurossensorial/epidemiologia
2.
Sci Transl Med ; 15(706): eadh9917, 2023 07 26.
Artigo em Inglês | MEDLINE | ID: mdl-37494473

RESUMO

Severe acute hepatitis of unknown etiology in children is under investigation in 35 countries. Although several potential etiologic agents have been investigated, a clear cause for the liver damage observed in these cases remains to be identified. Using VirScan, a high-throughput antibody profiling technology, we probed the antibody repertoires of nine cases of severe acute hepatitis of unknown etiology treated at Children's of Alabama and compared their antibody responses with 38 pediatric and 470 adult controls. We report increased adeno-associated dependoparvovirus A (AAV-A) breadth in cases relative to controls and adeno-associated virus 2 (AAV-2) peptide responses that were conserved in seven of nine cases but rarely observed in pediatric and adult controls. These findings suggest that AAV-2 is a likely etiologic agent of severe acute hepatitis of unknown etiology.


Assuntos
Hepatite , Hepatopatias , Adulto , Humanos , Criança , Dependovirus
3.
Viruses ; 15(7)2023 07 04.
Artigo em Inglês | MEDLINE | ID: mdl-37515187

RESUMO

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which was responsible for the COVID-19 pandemic, efficiently spreads cell-to-cell through mechanisms facilitated by its membrane glycoprotein spike. We established a dual split protein (DSP) assay based on the complementation of GFP and luciferase to quantify the fusogenic activity of the SARS-CoV-2 spike protein. We provide several lines of evidence that the spike protein of SARS-CoV-2, but not SARS-CoV-1, induced cell-cell fusion even in the absence of its receptor, angiotensin-converting enzyme 2 (ACE2). This poorly described ACE2-independent cell fusion activity of the spike protein was strictly dependent on the proteasomal cleavage of the spike by furin while TMPRSS2 was dispensable. Previous and current variants of concern (VOCs) differed significantly in their fusogenicity. The Delta spike was extremely potent compared to Alpha, Beta, Gamma and Kappa, while the Omicron spike was almost devoid of receptor-independent fusion activity. Nonetheless, for all analyzed variants, cell fusion was dependent on furin cleavage and could be pharmacologically inhibited with CMK. Mapping studies revealed that amino acids 652-1273 conferred the ACE2-independent fusion activity of the spike. Unexpectedly, residues proximal to the furin cleavage site were not of major relevance, whereas residue 655 critically regulated fusion. Finally, we found that the spike's fusion activity in the absence of ACE2 could be inhibited by antibodies directed against its N-terminal domain (NTD) but not by antibodies targeting its receptor-binding domain (RBD). In conclusion, our BSL-1-compatible DSP assay allowed us to screen for inhibitors or antibodies that interfere with the spike's fusogenic activity and may therefore contribute to both rational vaccine design and development of novel treatment options against SARS-CoV-2.


Assuntos
COVID-19 , Humanos , Enzima de Conversão de Angiotensina 2/genética , Anticorpos Monoclonais , Fusão Celular , Furina/metabolismo , Pandemias , SARS-CoV-2/genética , SARS-CoV-2/metabolismo , Glicoproteína da Espícula de Coronavírus/metabolismo
4.
PLoS Pathog ; 19(4): e1011316, 2023 04.
Artigo em Inglês | MEDLINE | ID: mdl-37058447

RESUMO

The presence of human cytomegalovirus (HCMV) in glioblastoma (GBM) and improved outcomes of GBM patients receiving therapies targeting the virus have implicated HCMV in GBM progression. However, a unifying mechanism that accounts for the contribution of HCMV to the malignant phenotype of GBM remains incompletely defined. Here we have identified SOX2, a marker of glioma stem cells (GSCs), as a key determinant of HCMV gene expression in gliomas. Our studies demonstrated that SOX2 downregulated promyelocytic leukemia (PML) and Sp100 and consequently facilitated viral gene expression by decreasing the amount of PML nuclear bodies in HCMV-infected glioma cells. Conversely, the expression of PML antagonized the effects of SOX2 on HCMV gene expression. Furthermore, this regulation of SOX2 on HCMV infection was demonstrated in a neurosphere assay of GSCs and in a murine xenograft model utilizing xenografts from patient-derived glioma tissue. In both cases, SOX2 overexpression facilitated the growth of neurospheres and xenografts implanted in immunodeficient mice. Lastly, the expression of SOX2 and HCMV immediate early 1 (IE1) protein could be correlated in tissues from glioma patients, and interestingly, elevated levels of SOX2 and IE1 were predictive of a worse clinical outcome. These studies argue that HCMV gene expression in gliomas is regulated by SOX2 through its regulation of PML expression and that targeting molecules in this SOX2-PML pathway could identify therapies for glioma treatment.


Assuntos
Glioma , Proteínas Imediatamente Precoces , Animais , Humanos , Camundongos , Citomegalovirus/fisiologia , Regulação para Baixo , Expressão Gênica , Glioma/genética , Glioma/patologia , Proteínas Imediatamente Precoces/metabolismo , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
5.
JCI Insight ; 8(5)2023 03 08.
Artigo em Inglês | MEDLINE | ID: mdl-36719764

RESUMO

Reactivation of human cytomegalovirus (HCMV) from latency is a frequent complication following hematopoietic stem cell transplantation (HSCT). The development of acute graft-versus-host disease (GVHD) is a significant risk factor for HCMV disease. Using a murine GVHD model in animals latently infected with murine CMV (MCMV), we studied preventive and therapeutic interventions in this high-risk scenario of HSCT. Mice latently infected with MCMV experienced reactivated MCMV and developed disseminated MCMV infection concomitant with the manifestations of GVHD. Dissemination was accompanied by accelerated mortality. We demonstrate that MCMV reactivation and dissemination was modulated by MCMV-specific antibodies, thus demonstrating in vivo protective activity of antiviral antibodies. However, the efficacy of serum therapy required repetitive doses of high-titer immune serum secondary to the shortened serum half-life of IgG in animals with GVHD. In a complementary approach, treatment of GVHD by adoptive transfer of donor-derived Tregs facilitated production of MCMV-specific antibodies from newly developing donor-derived B cells. Together, our findings strongly suggest that antibodies play a major role in controlling recurrent MCMV infection that follows GVHD, and they argue for reassessing the potential of antibody treatments as well as therapeutic strategies that enhance de novo antibody development against HCMV.


Assuntos
Infecções por Citomegalovirus , Doença Enxerto-Hospedeiro , Transplante de Células-Tronco Hematopoéticas , Muromegalovirus , Camundongos , Humanos , Animais , Citomegalovirus/fisiologia , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Anticorpos Antivirais
6.
Viruses ; 14(2)2022 01 28.
Artigo em Inglês | MEDLINE | ID: mdl-35215877

RESUMO

Human cytomegalovirus (HCMV) can cause severe clinical disease in immunocompromised individuals, such as allograft recipients and infants infected in utero. Neutralizing activity of antibodies, measured as the ability to prevent the entry of cell-free virus, has been correlated with the reduction in HCMV transmission and the severity of HCMV-associated disease. However, in vivo HCMV amplification may occur mainly via cell-to-cell spread. Thus, quantifying the inhibition of cell-to-cell transmission could be important in the evaluation of therapeutic antibodies and/or humoral responses to infection or immunization. Here, we established a quantitative plaque reduction assay, which allowed for the measurement of the capacity of antibodies to limit HCMV spread in vitro. Using an automated fluorescence spot reader, infection progression was assayed by the expansion of viral plaques during the course of infection with various GFP-expressing viruses. We found that in contrast to non-neutralizing monoclonal antibodies (mAbs), neutralizing mAbs against both glycoprotein B and H (gB and gH) could significantly inhibit viral plaque expansion of different HCMV strains and was equally efficient in fibroblasts as in epithelial cells. In contrast, an anti-pentamer mAb was active only in epithelial cells. Taken together, our data demonstrate that specific anti-HCMV mAbs can significantly limit cell-associated virus spread in vitro.


Assuntos
Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Citomegalovirus/imunologia , Citomegalovirus/fisiologia , Células Epiteliais/virologia , Fibroblastos/virologia , Anticorpos Monoclonais/imunologia , Linhagem Celular , Células Cultivadas , Humanos , Proteínas do Envelope Viral/imunologia , Ensaio de Placa Viral , Internalização do Vírus
7.
J Virol ; 96(5): e0182721, 2022 03 09.
Artigo em Inglês | MEDLINE | ID: mdl-35020472

RESUMO

Human cytomegalovirus (HCMV) has a large (∼235 kb) genome with more than 200 predicted open reading frames that exploits numerous cellular factors to facilitate its replication. A key feature of HCMV-infected cells is the emergence of a distinctive membranous cytoplasmic compartment termed the virion assembly compartment (vAC). Here, we report that host protein WD repeat domain 11 (WDR11) plays a key role in vAC formation and virion morphogenesis. We found that WDR11 was upregulated at both mRNA and protein levels during HCMV infection. At the late stage of HCMV replication, WDR11 relocated to the vAC and colocalized with markers of the trans-Golgi network (TGN) and vAC. Depletion of WDR11 hindered HCMV-induced membrane reorganization of the Golgi and TGN, altered vAC formation, and impaired HCMV secondary envelopment and virion morphogenesis. Further, motifs critical for the localization of WDR11 in TGN were identified by alanine-scanning mutagenesis. Mutation of these motifs led to WDR11 mislocation outside the TGN and loss of vAC formation. Taken together, these data indicate that host protein WDR11 is required for efficient viral replication at the stage of virion assembly, possibly by facilitating the remodeling of the endomembrane system for vAC formation and virion morphogenesis. IMPORTANCE During the late phase of human cytomegalovirus (HCMV) infection, the endomembrane system is dramatically reorganized, resulting in the formation of a unique structure termed the virion assembly compartment (vAC), which is critical for the assembly of infectious virions. The mechanism of HCMV-induced vAC formation is still not fully understood. In this report, we identified a host factor, WDR11, that plays an important role in vAC formation. Our findings argue that WDR11 contributes to the relocation of the Golgi and trans-Golgi network to the vAC, a membrane reorganization process that appears to be required for efficient virion maturation. The present work provides new insights into the vAC formation and HCMV virion morphogenesis and a potential novel target for antiviral treatment.


Assuntos
Infecções por Citomegalovirus , Citomegalovirus , Interações entre Hospedeiro e Microrganismos , Repetições WD40 , Citomegalovirus/genética , Citomegalovirus/metabolismo , Infecções por Citomegalovirus/fisiopatologia , Infecções por Citomegalovirus/virologia , Humanos , Morfogênese , Vírion/metabolismo , Montagem de Vírus/genética , Replicação Viral/genética , Repetições WD40/genética , Rede trans-Golgi/metabolismo
8.
JCI Insight ; 7(1)2022 01 11.
Artigo em Inglês | MEDLINE | ID: mdl-35014624

RESUMO

Congenital cytomegalovirus (cCMV) infection is the leading infectious cause of neurodevelopmental disorders. However, the neuropathogenesis remains largely elusive due to a lack of informative animal models. In this study, we developed a congenital murine CMV (cMCMV) infection mouse model with high survival rate and long survival period that allowed long-term follow-up study of neurodevelopmental disorders. This model involves in utero intracranial injection and mimics many reported clinical manifestations of cCMV infection in infants, including growth restriction, hearing loss, and impaired cognitive and learning-memory abilities. We observed that abnormalities in MRI/CT neuroimaging were consistent with brain hemorrhage and loss of brain parenchyma, which was confirmed by pathological analysis. Neuropathological findings included ventriculomegaly and cortical atrophy associated with impaired proliferation and migration of neural progenitor cells in the developing brain at both embryonic and postnatal stages. Robust inflammatory responses during infection were shown by elevated inflammatory cytokine levels, leukocyte infiltration, and activation of microglia and astrocytes in the brain. Pathological analyses and CT neuroimaging revealed brain calcifications induced by cMCMV infection and cell death via pyroptosis. Furthermore, antiviral treatment with ganciclovir significantly improved neurological functions and mitigated brain damage as shown by CT neuroimaging. These results demonstrate that this model is suitable for investigation of mechanisms of infection-induced brain damage and long-term studies of neurodevelopmental disorders, including the development of interventions to limit CNS damage associated with cCMV infection.


Assuntos
Infecções por Citomegalovirus , Modelos Animais de Doenças , Neuroimagem , Animais , Infecções por Citomegalovirus/congênito , Infecções por Citomegalovirus/diagnóstico por imagem , Infecções por Citomegalovirus/fisiopatologia , Infecções por Citomegalovirus/terapia , Feminino , Seguimentos , Camundongos , Camundongos Endogâmicos ICR , Gravidez
9.
J Infect Dis ; 224(11): 1807-1809, 2021 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-34647593
10.
Viruses ; 13(6)2021 06 04.
Artigo em Inglês | MEDLINE | ID: mdl-34200083

RESUMO

Human cytomegalovirus (HCMV) is a highly prevalent herpesvirus that can cause severe disease in immunocompromised individuals and immunologically immature fetuses and newborns. Most infected newborns are able to resolve the infection without developing sequelae. However, in severe cases, congenital HCMV infection can result in life-threatening pathologies and permanent damage of organ systems that possess a low regenerative capacity. Despite the severity of the problem, HCMV infection of the central nervous system (CNS) remains inadequately characterized to date. Cytomegaloviruses (CMVs) show strict species specificity, limiting the use of HCMV in experimental animals. Infection following intraperitoneal administration of mouse cytomegalovirus (MCMV) into newborn mice efficiently recapitulates many aspects of congenital HCMV infection in CNS. Upon entering the CNS, CMV targets all resident brain cells, consequently leading to the development of widespread histopathology and inflammation. Effector functions from both resident cells and infiltrating immune cells efficiently resolve acute MCMV infection in the CNS. However, host-mediated inflammatory factors can also mediate the development of immunopathologies during CMV infection of the brain. Here, we provide an overview of the cytomegalovirus infection in the brain, local immune response to infection, and mechanisms leading to CNS sequelae.


Assuntos
Encéfalo/virologia , Sistema Nervoso Central/virologia , Infecções por Citomegalovirus/complicações , Citomegalovirus/patogenicidade , Inflamação/virologia , Animais , Encéfalo/imunologia , Encéfalo/patologia , Sistema Nervoso Central/imunologia , Sistema Nervoso Central/patologia , Citomegalovirus/imunologia , Infecções por Citomegalovirus/imunologia , Infecções por Citomegalovirus/patologia , Modelos Animais de Doenças , Humanos , Inflamação/patologia , Camundongos , Muromegalovirus , Tropismo Viral
11.
Emerg Infect Dis ; 27(9): 2454-2458, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34193339

RESUMO

Not all persons recovering from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection develop SARS-CoV-2-specific antibodies. We show that nonseroconversion is associated with younger age and higher reverse transcription PCR cycle threshold values and identify SARS-CoV-2 viral loads in the nasopharynx as a major correlate of the systemic antibody response.


Assuntos
COVID-19 , Formação de Anticorpos , COVID-19/imunologia , Teste Sorológico para COVID-19 , Humanos , Nasofaringe , SARS-CoV-2 , Soroconversão
12.
J Virol ; 95(18): e0065721, 2021 08 25.
Artigo em Inglês | MEDLINE | ID: mdl-34160252

RESUMO

Human cytomegalovirus (HCMV) is a ubiquitous pathogen that causes severe clinical disease in immunosuppressed patients and congenitally infected newborn infants. Viral envelope glycoproteins represent attractive targets for vaccination or passive immunotherapy. To extend the knowledge of mechanisms of virus neutralization, monoclonal antibodies (MAbs) were generated following immunization of mice with HCMV virions. Hybridoma supernatants were screened for in vitro neutralization activity, yielding three potent MAbs, 6E3, 3C11, and 2B10. MAbs 6E3 and 3C11 blocked infection of all viral strains that were tested, while MAb 2B10 neutralized only 50% of the HCMV strains analyzed. Characterization of the MAbs using indirect immunofluorescence analyses demonstrated their reactivity with recombinantly derived gH. While MAbs 6E3 and 3C11 reacted with gH when expressed alone, 2B10 detected gH only when it was coexpressed with gB and gL. Recognition of gH by 3C11 was dependent on the expression of the entire ectodomain of gH, whereas 6E3 required residues 1 to 629 of gH. The strain-specific determinant for neutralization by Mab 2B10 was identified as a single Met→Ile amino acid polymorphism within gH, located within the central part of the protein. The polymorphism is evenly distributed among described HCMV strains. The 2B10 epitope thus represents a novel strain-specific antibody target site on gH of HCMV. The dependence of the reactivity of 2B10 on the simultaneous presence of gB/gH/gL will be of value in the structural definition of this tripartite complex. The 2B10 epitope may also represent a valuable tool for diagnostics to monitor infections/reinfections with different HCMV strains during pregnancy or after transplantation. IMPORTANCE HCMV infections are life threatening to people with compromised or immature immune systems. Understanding the antiviral antibody repertoire induced during HCMV infection is a necessary prerequisite to define protective antibody responses. Here, we report three novel anti-gH MAbs that potently neutralized HCMV infectivity. One of these MAbs (2B10) targets a novel strain-specific conformational epitope on gH that only becomes accessible upon coexpression of the minimal fusion machinery gB/gH/gL. Strain specificity is dependent on a single amino acid polymorphism within gH. Our data highlight the importance of strain-specific neutralizing antibody responses against HCMV. The 2B10 epitope may also represent a valuable tool for diagnostics to monitor infections/reinfections with different HCMV strains during pregnancy or after transplantation. In addition, the dependence of the reactivity of 2B10 on the simultaneous presence of gB/gH/gL will be of value in the structural definition of this tripartite complex.


Assuntos
Anticorpos Monoclonais/imunologia , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Infecções por Citomegalovirus/imunologia , Citomegalovirus/imunologia , Epitopos/imunologia , Proteínas do Envelope Viral/imunologia , Animais , Citomegalovirus/classificação , Infecções por Citomegalovirus/virologia , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos BALB C
13.
Pediatrics ; 147(5)2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33622794

RESUMO

BACKGROUND AND OBJECTIVES: In children with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, virological characteristics and correlation with disease severity have not been extensively studied. The primary objective in this study is to determine the correlation between SARS-CoV-2 viral load (VL) in infected children with age, disease severity, and underlying comorbidities. METHODS: Children <21 years, screened for SARS-CoV-2 at the time of hospitalization, who tested positive by polymerase chain reaction were included in this study. VL at different sites was determined and compared between groups. RESULTS: Of the 102 children included in this study, 44% of the cohort had asymptomatic infection, and children with >1 comorbidity were the most at risk for severe disease. VL in children with symptomatic infection was significantly higher than in children with asymptomatic infection (3.0 × 105 vs 7.2 × 103 copies per mL; P = .001). VL in the respiratory tract was significantly higher in children <1 year, compared with older children (3.3 × 107 vs 1.3 × 104 copies per mL respectively; P < .0001), despite most infants presenting with milder illness. Besides the respiratory tract, SARS-CoV-2 RNA was also detectable in samples from the gastrointestinal tract (saliva and rectum) and blood. In 13 children for whom data on duration of polymerase chain reaction positivity was available, 12 of 13 tested positive 2 weeks after initial diagnosis, and 6 of 13 continued to test positive 4 weeks after initial diagnosis. CONCLUSIONS: In hospitalized children with SARS-CoV-2, those with >1 comorbid condition experienced severe disease. SARS-CoV-2 VL in the respiratory tract is significantly higher in children with symptomatic disease and children <1 year of age.


Assuntos
COVID-19/virologia , Hospitalização , Carga Viral , Adolescente , Infecções Assintomáticas , COVID-19/diagnóstico , Teste de Ácido Nucleico para COVID-19 , Criança , Pré-Escolar , Humanos , Lactente , Nasofaringe/virologia , Nariz/virologia , Reto/virologia , SARS-CoV-2 , Salvia/virologia , Índice de Gravidade de Doença , Fatores de Tempo , Eliminação de Partículas Virais , Adulto Jovem
14.
J Exp Med ; 218(5)2021 05 03.
Artigo em Inglês | MEDLINE | ID: mdl-33630019

RESUMO

Congenital human cytomegalovirus (cHCMV) infection of the brain is associated with a wide range of neurocognitive sequelae. Using infection of newborn mice with mouse cytomegalovirus (MCMV) as a reliable model that recapitulates many aspects of cHCMV infection, including disseminated infection, CNS infection, altered neurodevelopment, and sensorineural hearing loss, we have previously shown that mitigation of inflammation prevented alterations in cerebellar development, suggesting that host inflammatory factors are key drivers of neurodevelopmental defects. Here, we show that MCMV infection causes a dramatic increase in the expression of the microglia-derived chemokines CXCL9/CXCL10, which recruit NK and ILC1 cells into the brain in a CXCR3-dependent manner. Surprisingly, brain-infiltrating innate immune cells not only were unable to control virus infection in the brain but also orchestrated pathological inflammatory responses, which lead to delays in cerebellar morphogenesis. Our results identify NK and ILC1 cells as the major mediators of immunopathology in response to virus infection in the developing CNS, which can be prevented by anti-IFN-γ antibodies.


Assuntos
Encéfalo/imunologia , Infecções por Citomegalovirus/imunologia , Citomegalovirus/imunologia , Inflamação/imunologia , Células Matadoras Naturais/imunologia , Linfócitos/imunologia , Animais , Animais Recém-Nascidos , Encéfalo/patologia , Encéfalo/virologia , Quimiocina CXCL10/genética , Quimiocina CXCL10/imunologia , Quimiocina CXCL10/metabolismo , Quimiocina CXCL9/genética , Quimiocina CXCL9/imunologia , Quimiocina CXCL9/metabolismo , Citomegalovirus/fisiologia , Infecções por Citomegalovirus/virologia , Regulação da Expressão Gênica/imunologia , Humanos , Imunidade Inata/imunologia , Inflamação/genética , Inflamação/virologia , Células Matadoras Naturais/metabolismo , Linfócitos/metabolismo , Camundongos da Linhagem 129 , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microglia/imunologia , Microglia/metabolismo , Microglia/virologia , Receptores CXCR3/genética , Receptores CXCR3/imunologia , Receptores CXCR3/metabolismo
15.
Methods Mol Biol ; 2244: 403-463, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33555597

RESUMO

Human cytomegalovirus is the largest human herpesvirus and shares many core features of other herpesviruses such as tightly regulated gene expression during genome replication and latency as well as the establishment of lifelong persistence following infection. In contrast to stereotypic clinical syndromes associated with alpha-herpesvirus infections, almost all primary HCMV infections are asymptomatic and acquired early in life in most populations in the world. Although asymptomatic in most individuals, HCMV is a major cause of disease in hosts with deficits in adaptive and innate immunity such as infants who are infected in utero and allograft recipients following transplantation. Congenital HCMV is a commonly acquired infection in the developing fetus that can result in a number of neurodevelopmental abnormalities. Similarly, HCMV is a major cause of disease in allograft recipients in the immediate and late posttransplant period and is thought to be a major contributor to chronic allograft rejection. Even though HCMV induces robust innate and adaptive immune responses, it also encodes a vast array of immune evasion functions that are thought aid in its persistence. Immune correlates of protective immunity that prevent or modify intrauterine HCMV infection remain incompletely defined but are thought to consist primarily of adaptive responses in the pregnant mother, thus making congenital HCMV a potentially vaccine modifiable disease. Similarly, HCMV infection in allograft recipients is often more severe in recipients without preexisting adaptive immunity to HCMV. Thus, there has been a considerable effort to modify HCMV specific immunity in transplant recipient either through active immunization or passive transfer of adaptive effector functions. Although efforts to develop an efficacious vaccine and/or passive immunotherapy to limit HCMV disease have been underway for nearly six decades, most have met with limited success at best. In contrast to previous efforts, current HCMV vaccine development has relied on observations of unique properties of HCMV in hopes of reproducing immune responses that at a minimum will be similar to that following natural infection. However, more recent findings have suggested that immunity following naturally acquired HCMV infection may have limited protective activity and almost certainly, is not sterilizing. Such observations suggest that either the induction of natural immunity must be specifically tailored to generate protective activity or alternatively, that providing targeted passive immunity to susceptible populations could be prove to be more efficacious.


Assuntos
Vacinas contra Citomegalovirus/imunologia , Citomegalovirus/imunologia , Vacinação/métodos , Imunidade Adaptativa/imunologia , Anticorpos Antivirais/imunologia , Citomegalovirus/genética , Infecções por Citomegalovirus/imunologia , Infecções por Citomegalovirus/prevenção & controle , Suscetibilidade a Doenças , Feminino , Humanos , Imunidade Humoral/imunologia , Imunidade Inata/imunologia , Lactente , Masculino , Gravidez , Vacinas/imunologia , Vacinas/metabolismo , Vacinas/farmacologia
16.
J Virol ; 95(8)2021 03 25.
Artigo em Inglês | MEDLINE | ID: mdl-33504601

RESUMO

We previously reported that human cytomegalovirus (HCMV) utilizes the cellular protein WD repeat-containing protein 5 (WDR5) to facilitate capsid nuclear egress. Here, we further show that HCMV infection results in WDR5 localization in a juxtanuclear region, and that its localization to this cellular site is associated with viral replication and late viral gene expression. Furthermore, WDR5 accumulated in the virion assembly compartment (vAC) and co-localized with vAC markers of gamma-tubulin (γ-tubulin), early endosomes, and viral vAC marker proteins pp65, pp28, and glycoprotein B (gB). WDR5 co-immunoprecipitated with multiple virion proteins, including MCP, pp150, pp65, pIRS1, and pTRS1, which may explain WDR5 accumulation in the vAC during infection. WDR5 fractionated with virions either in the presence or absence of Triton X-100 and was present in purified viral particles, suggesting that WDR5 was incorporated into HCMV virions. Thus, WDR5 localized to the vAC and was incorporated into virions, raising the possibility that in addition to capsid nuclear egress, WDR5 could also participate in cytoplasmic HCMV virion morphogenesis.Importance Human cytomegalovirus (HCMV) has a large (∼235-kb) genome that contains over 170 ORFs and exploits numerous cellular factors to facilitate its replication. In the late phase of HCMV infection cytoplasmic membranes are reorganized to establish the virion assembly compartment (vAC), which has been shown to necessary for efficient assembly of progeny virions. We previously reported that WDR5 facilitates HCMV nuclear egress. Here, we show that WDR5 is localized to the vAC and incorporated into virions, perhaps contributing to efficient virion maturation. Thus, findings in this study identified a potential role for WDR5 in HCMV assembly in the cytoplasmic phase of virion morphogenesis.

17.
J Perinatol ; 41(2): 315-323, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-32884104

RESUMO

OBJECTIVES: To determine the incidence and risk factors of hearing loss (HL) in Brazilian neonates. STUDY DESIGN: 11,900 neonates were screened for hearing and congenital CMV (cCMV). Low and high-risk babies who did not pass their hearing screening and infants with cCMV were scheduled for a diagnostic audiologic evaluation. RESULTS: The incidence of HL was 2 per 1000 live-born infants (95% CI: 1-3). HL was higher in high-risk neonates than in low risk babies (18.6 vs. 0.3/1000 live births, respectively). Among infants exposed to isolated risk factors, association of HL with craniofacial abnormalities/syndromes (RR = 24.47; 95% CI: 5.9-100.9) and cCMV (RR = 9.54; 95% CI: 3.3-27.7) were observed. HL was 20 to 100-fold more likely in neonates exposed to ototoxic drugs in combination with cCMV or craniofacial/congenital anomalies. CONCLUSIONS: Strategies for the prevention of cCMV and exposure to ototoxic drugs may decrease the incidence of HL in this population.


Assuntos
Infecções por Citomegalovirus , Perda Auditiva , Perda Auditiva/diagnóstico , Perda Auditiva/epidemiologia , Perda Auditiva/etiologia , Testes Auditivos , Humanos , Lactente , Recém-Nascido , Triagem Neonatal , Fatores de Risco
18.
Nat Cancer ; 2(10): 1018-1038, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-35121884

RESUMO

Expanding the utility of immune-based cancer treatments is a clinical challenge due to tumor-intrinsic factors that suppress the immune response. Here we report the identification of tumoral ring finger protein 2 (RNF2), the core subunit of polycomb repressor complex 1, as a negative regulator of antitumor immunity in various human cancers, including breast cancer. In syngeneic murine models of triple-negative breast cancer, we found that deleting genes encoding the polycomb repressor complex 1 subunits Rnf2, BMI1 proto-oncogene, polycomb ring finger (Bmi1), or the downstream effector of Rnf2, remodeling and spacing factor 1 (Rsf1), was sufficient by itself to induce durable tumor rejection and establish immune memory by enhancing infiltration and activation of natural killer and CD4+ T cells, but not CD8+ T cells, into the tumor and enabled their cooperativity. These findings uncover an epigenetic reprogramming of the tumor-immune microenvironment, which fosters durable antitumor immunity and memory.


Assuntos
Neoplasias , Complexo Repressor Polycomb 1/metabolismo , Microambiente Tumoral , Ubiquitina-Proteína Ligases/metabolismo , Animais , Linfócitos T CD4-Positivos , Linfócitos T CD8-Positivos , Humanos , Camundongos , Neoplasias/genética , Proteínas Nucleares , Complexo Repressor Polycomb 1/genética , Proteínas do Grupo Polycomb , Transativadores , Microambiente Tumoral/genética
19.
Clin Infect Dis ; 72(7): 1253-1255, 2021 04 08.
Artigo em Inglês | MEDLINE | ID: mdl-32596725

RESUMO

Testing of paired midturbinate (MT) nasal and nasopharyngeal (NP) swabs, collected by trained personnel from 40 patients with coronavirus disease 2019 (COVID-19), showed that more NP (76/95 [80%]) than MT swabs tested positive (61/95 [64%]) (P = .02). Among samples collected a week after study enrollment, fewer MT than NP samples were positive (45% vs 76%; P = .001).


Assuntos
COVID-19 , SARS-CoV-2 , Testes Diagnósticos de Rotina , Humanos , Nasofaringe , Manejo de Espécimes
20.
mBio ; 11(5)2020 09 29.
Artigo em Inglês | MEDLINE | ID: mdl-32994323

RESUMO

The human cytomegalovirus (HCMV) UL132 open reading frame encodes a 270-amino-acid type I envelope glycoprotein, gpUL132. The deletion of UL132 (ΔUL132) from the HCMV genome results in a pronounced deficit in virus yield, with an approximately 2-log decrease in the production of infectious virus compared to the wild-type (WT) virus. Characterization of the ΔUL132 mutant virus indicated that it was less infectious with a high particle-to-infectious unit ratio and an altered composition of virion proteins compared to the WT virus. In addition, the viral assembly compartment (AC) failed to form in cells infected with the ΔUL132 mutant virus. The expression of gpUL132 in trans rescued the defects in the morphogenesis of the AC in cells infected with the ΔUL132 mutant virus and in infectious virus production. Furthermore, using cell lines expressing chimeric proteins, we demonstrated that the cytosolic domain of gpUL132 was sufficient to rescue AC formation and WT levels of virus production. Progeny virions from ΔUL132-infected cells expressing the cytosolic domain of gpUL132 exhibited particle-to-infectious unit ratios similar to those of the WT virus. Together, our findings argue that gpUL132 is essential for HCMV AC formation and the efficient production of infectious particles, thus highlighting the importance of this envelope protein for the virus-induced reorganization of intracellular membranes and AC formation in the assembly of infectious virus.IMPORTANCE Following infection of permissive cells, human cytomegalovirus (HCMV) induces the reorganization of intracellular membranes resulting in the formation of a distinctive membranous compartment in the cytoplasm of infected cells. This compartment has been designated the viral assembly compartment (AC) and is thought to be a site for cytoplasmic virion assembly and envelopment. In this study, we have demonstrated that a single virion envelope glycoprotein is essential for AC formation in infected cells, and in its absence, there is a significant decrease in the production of infectious virions. These findings are consistent with those from other studies that have demonstrated the importance of host cell proteins in the formation of the AC and demonstrate a critical role of a single virion protein in AC formation and the efficient assembly of infectious virus.


Assuntos
Citomegalovirus/genética , Citomegalovirus/fisiologia , Regulação Viral da Expressão Gênica , Glicoproteínas de Membrana/genética , Proteínas do Envelope Viral/genética , Montagem de Vírus , Linhagem Celular , Células Cultivadas , Fibroblastos/virologia , Prepúcio do Pênis/citologia , Humanos , Masculino , Proteínas Virais/metabolismo , Vírion/genética
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