Assuntos
Infecções por Coronavirus/patologia , Síndrome da Liberação de Citocina/patologia , Eritroblastos/patologia , Linfócitos/patologia , Pneumonia Viral/patologia , Anti-Inflamatórios/uso terapêutico , Aspirina/uso terapêutico , Betacoronavirus/imunologia , Betacoronavirus/patogenicidade , COVID-19 , Teste para COVID-19 , Criança , Técnicas de Laboratório Clínico/métodos , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Síndrome da Liberação de Citocina/tratamento farmacológico , Síndrome da Liberação de Citocina/imunologia , Síndrome da Liberação de Citocina/virologia , Diarreia/diagnóstico , Diarreia/fisiopatologia , Diarreia/virologia , Eritroblastos/imunologia , Eritroblastos/virologia , Feminino , Febre/diagnóstico , Febre/fisiopatologia , Febre/virologia , Humanos , Linfócitos/imunologia , Linfócitos/virologia , Náusea/diagnóstico , Náusea/fisiopatologia , Náusea/virologia , Pandemias , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/imunologia , Pneumonia Viral/virologia , Prednisona/uso terapêutico , SARS-CoV-2 , Resultado do Tratamento , Vômito/diagnóstico , Vômito/fisiopatologia , Vômito/virologiaRESUMO
Nucleated teleost red blood cells (RBCs) are known to express molecules from the major histocompatibility complex and peptide-generating processes such as autophagy and proteasomes, but the role of RBCs in antigen presentation of viruses have not been studied yet. In this study, RBCs exposed ex vivo to viral hemorrhagic septicemia virus (VHSV) were evaluated by means of transcriptomic and proteomic approaches. Genes and proteins related to antigen presentation molecules, proteasome degradation, and autophagy were up-regulated. VHSV induced accumulation of ubiquitinated proteins in ex vivo VHSV-exposed RBCs and showed at the same time a decrease of proteasome activity. Furthermore, induction of autophagy was detected by evaluating LC3 protein levels. Sequestosome-1/p62 underwent degradation early after VHSV exposure, and it may be a link between ubiquitination and autophagy activation. Inhibition of autophagosome degradation with niclosamide resulted in intracellular detection of N protein of VHSV (NVHSV) and p62 accumulation. In addition, antigen presentation cell markers, such as major histocompatibility complex (MHC) class I & II, CD83, and CD86, increased at the transcriptional and translational level in rainbow trout RBCs exposed to VHSV. In summary, we show that nucleated rainbow trout RBCs can degrade VHSV while displaying an antigen-presenting cell (APC)-like profile.
Assuntos
Apresentação de Antígeno/imunologia , Eritroblastos/imunologia , Eritroblastos/virologia , Septicemia Hemorrágica Viral/imunologia , Septicemia Hemorrágica Viral/virologia , Novirhabdovirus/imunologia , Oncorhynchus mykiss/imunologia , Oncorhynchus mykiss/virologia , Animais , Apresentação de Antígeno/genética , Células Apresentadoras de Antígenos/citologia , Células Apresentadoras de Antígenos/imunologia , Antígenos CD/análise , Antígenos CD/imunologia , Autofagossomos/efeitos dos fármacos , Autofagossomos/imunologia , Autofagossomos/virologia , Autofagia/efeitos dos fármacos , Autofagia/imunologia , Antígeno B7-2/análise , Antígeno B7-2/imunologia , Biomarcadores/análise , Septicemia Hemorrágica Viral/genética , Antígenos de Histocompatibilidade Classe I/análise , Antígenos de Histocompatibilidade Classe I/imunologia , Antígenos de Histocompatibilidade Classe II/análise , Antígenos de Histocompatibilidade Classe II/imunologia , Imunoglobulinas/análise , Imunoglobulinas/imunologia , Glicoproteínas de Membrana/análise , Glicoproteínas de Membrana/imunologia , Niclosamida/farmacologia , Proteínas do Nucleocapsídeo , Complexo de Endopeptidases do Proteassoma/efeitos dos fármacos , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteólise , Proteômica , Proteína Sequestossoma-1/metabolismo , Antígeno CD83RESUMO
Lymph- and blood-borne retroviruses exploit CD169/Siglec-1-mediated capture by subcapsular sinus and marginal zone metallophilic macrophages for trans-infection of permissive lymphocytes. However, the impact of CD169-mediated virus capture on retrovirus dissemination and pathogenesis in vivo is unknown. In a murine model of the splenomegaly-inducing retrovirus Friend virus complex (FVC) infection, we find that while CD169 promoted draining lymph node infection, it limited systemic spread to the spleen. At the spleen, CD169-expressing macrophages captured incoming blood-borne retroviruses and limited their spread to the erythroblasts in the red pulp where FVC manifests its pathogenesis. CD169-mediated retroviral capture activated conventional dendritic cells 1 (cDC1s) and promoted cytotoxic CD8+ T cell responses, resulting in efficient clearing of FVC-infected cells. Accordingly, CD169 blockade led to higher viral loads and accelerated death in susceptible mouse strains. Thus, CD169 plays a protective role during FVC pathogenesis by reducing viral dissemination to erythroblasts and eliciting an effective cytotoxic T lymphocyte response via cDC1s.
Assuntos
Lectinas/farmacologia , Substâncias Protetoras/farmacologia , Infecções por Retroviridae/tratamento farmacológico , Retroviridae/efeitos dos fármacos , Retroviridae/patogenicidade , Lectina 1 Semelhante a Ig de Ligação ao Ácido Siálico/farmacologia , Animais , Linfócitos T CD8-Positivos , Proliferação de Células , Células Dendríticas/virologia , Modelos Animais de Doenças , Eritroblastos/virologia , Feminino , Interferon Tipo I/metabolismo , Linfonodos/virologia , Macrófagos/efeitos dos fármacos , Macrófagos/virologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Baço , Linfócitos T Citotóxicos , Carga ViralAssuntos
Anemia Aplástica/diagnóstico , Anemia Falciforme/diagnóstico , Eritroblastos/patologia , Isquemia Miocárdica/diagnóstico , Infecções por Parvoviridae/diagnóstico , Anemia Aplástica/complicações , Anemia Aplástica/fisiopatologia , Anemia Aplástica/virologia , Anemia Falciforme/complicações , Anemia Falciforme/fisiopatologia , Anemia Falciforme/virologia , Criança , Eritroblastos/virologia , Hematopoese , Humanos , Unidades de Terapia Intensiva , Masculino , Isquemia Miocárdica/complicações , Isquemia Miocárdica/fisiopatologia , Isquemia Miocárdica/virologia , Infecções por Parvoviridae/complicações , Infecções por Parvoviridae/fisiopatologia , Infecções por Parvoviridae/virologia , Parvovirus B19 Humano/isolamento & purificaçãoAssuntos
Transplante de Coração/efeitos adversos , Terapia de Imunossupressão/métodos , Infecções por Parvoviridae/diagnóstico por imagem , Infecções por Parvoviridae/etiologia , Idoso , Exame de Medula Óssea , Transformação Celular Viral , Eritroblastos/patologia , Eritroblastos/virologia , Humanos , Hospedeiro Imunocomprometido , Corpos de Inclusão Intranuclear/patologia , Masculino , Parvovirus B19 Humano , Aplasia Pura de Série Vermelha/virologia , Vacúolos/patologia , Ativação ViralAssuntos
Anemia Hemolítica Congênita/patologia , Anemia Hemolítica Congênita/virologia , Eritroblastos/patologia , Herpesvirus Humano 6/isolamento & purificação , Infecções por Roseolovirus/complicações , Anemia Hemolítica Congênita/sangue , Anemia Hemolítica Congênita/terapia , Transfusão de Sangue , Medula Óssea/patologia , Medula Óssea/virologia , Eritroblastos/virologia , Feminino , Humanos , Lactente , Infecções por Roseolovirus/sangue , Infecções por Roseolovirus/patologia , Infecções por Roseolovirus/virologiaRESUMO
This protocol describes the efficient isolation of peripheral blood mononuclear cells from circulating blood via density gradient centrifugation and subsequent generation of integration-free human induced pluripotent stem cells. Peripheral blood mononuclear cells are cultured for 9 days to allow expansion of the erythroblast population. The erythroblasts are then used to derive human induced pluripotent stem cells using Sendai viral vectors, each expressing one of the four reprogramming factors Oct4, Sox2, Klf4, and c-Myc.
Assuntos
Técnicas de Cultura de Células/métodos , Técnicas de Reprogramação Celular/métodos , Reprogramação Celular , Vetores Genéticos/genética , Células-Tronco Pluripotentes Induzidas/citologia , Leucócitos Mononucleares/citologia , Vírus Sendai/genética , Animais , Separação Celular/métodos , Células Cultivadas , Meios de Cultura , Citocinas/farmacologia , Eritroblastos/citologia , Eritroblastos/efeitos dos fármacos , Eritroblastos/virologia , Fibroblastos/citologia , Humanos , Fator 4 Semelhante a Kruppel , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/fisiologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/virologia , Camundongos , Fator 3 de Transcrição de Octâmero/genética , Fator 3 de Transcrição de Octâmero/fisiologia , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/fisiologia , Proteínas Recombinantes/genética , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/fisiologiaRESUMO
Erythrocytes are highly abundant circulating cells in the vertebrates, which, with the notable exception of mammals, remain nucleated throughout the entire life cycle. The major function associated with these cells is respiratory gas exchange however other functions including interaction with the immune system have been attributed to these cells. Many viral, prokaryotic and eukaryotic pathogens directly target this cell type and across the vertebrate group a significant number of related pathologies have been reported. Across the primary literature mechanisms of interaction, invasion and replication between viruses and erythrocytes have been well described however the functional response of the erythrocyte has been poorly studied. A fragmented series of reports spanning the vertebrates suggests that these cells are capable of functional responses to viral infection. In contrast, in-depth proteomic studies using human erythrocytes have strongly progressed throughout the past decade providing a rich source of information related to protein expression and potential function. Furthermore information at the gene expression level is becoming available. Here we provide a review of erythrocyte-pathogen interactions, erythrocyte functions in immunity and propose in light of recent -omics research that the nucleated erythrocytes may have a direct role in the immune response.
Assuntos
Eritrócitos/imunologia , Vertebrados/imunologia , Animais , Eritroblastos/imunologia , Eritroblastos/microbiologia , Eritroblastos/parasitologia , Eritroblastos/virologia , Eritrócitos/microbiologia , Eritrócitos/parasitologia , Eritrócitos/virologia , Interações Hospedeiro-Patógeno , Proteômica , Vertebrados/microbiologia , Vertebrados/parasitologia , Vertebrados/virologiaRESUMO
Aggresome formation, a cellular response to misfolded protein aggregates, is linked to cancer and neurodegenerative disorders. Previously we showed that Gag-v-ErbA (v-ErbA), a retroviral variant of the thyroid hormone receptor (TRα1), accumulates in and sequesters TRα1 into cytoplasmic foci. Here, we show that foci represent v-ErbA targeting to aggresomes. v-ErbA colocalizes with aggresomal markers, proteasomes, hsp70, HDAC6, and mitochondria. Foci have hallmark characteristics of aggresomes: formation is microtubule-dependent, accelerated by proteasome inhibitors, and they disrupt intermediate filaments. Proteasome-mediated degradation is critical for clearance of v-ErbA and T(3)-dependent TRα1 clearance. Our studies highlight v-ErbA's complex mode of action: the oncoprotein is highly mobile and trafficks between the nucleus, cytoplasm, and aggresome, carrying out distinct activities within each compartment. Dynamic trafficking to aggresomes contributes to the dominant negative activity of v-ErbA and may be enhanced by the viral Gag sequence. These studies provide insight into novel modes of oncogenesis across multiple cellular compartments.
Assuntos
Corpos de Inclusão/metabolismo , Proteínas Oncogênicas v-erbA/metabolismo , Alpharetrovirus/genética , Alpharetrovirus/metabolismo , Transporte Biológico , Biomarcadores/metabolismo , Dineínas/metabolismo , Eritroblastos/citologia , Eritroblastos/metabolismo , Eritroblastos/virologia , Produtos do Gene gag/genética , Produtos do Gene gag/metabolismo , Proteínas de Choque Térmico HSP70/metabolismo , Células HeLa , Desacetilase 6 de Histona , Histona Desacetilases/genética , Histona Desacetilases/metabolismo , Humanos , Filamentos Intermediários/metabolismo , Microtúbulos/metabolismo , Mitocôndrias/metabolismo , Proteínas Oncogênicas v-erbA/genética , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Vimentina/metabolismoRESUMO
During chronic viral infection, persistent exposure to viral Ags leads to the overexpression of multiple inhibitory cell-surface receptors that cause CD8(+) T cell exhaustion. The severity of exhaustion correlates directly with the level of infection and the number and intensity of inhibitory receptors expressed, and it correlates inversely with the ability to respond to the blockade of inhibitory pathways. Friend virus (FV) is a murine retrovirus complex that induces acute high-level viremia, followed by persistent infection and leukemia development, when inoculated into immunocompetent adult mice. In this article, we provide conclusive evidence that FV infection results in the generation of virus-specific effector CD8(+) T cells that are terminally exhausted. Acute FV-induced disease is characterized by a rapid increase in the number of virus-infected erythroblasts, leading to massive splenomegaly. Most of the expanded erythroblasts strongly express programmed death ligand-1 and MHC class I, thereby creating a highly tolerogenic environment. Consequently, FV-specific effector CD8(+) T cells uniformly express multiple inhibitory receptors, such as programmed cell death 1 (PD-1), T cell Ig domain and mucin domain 3 (Tim-3), lymphocyte activation gene-3, and CTLA-4, rapidly become nonresponsive to restimulation and are no longer reinvigorated by combined in vivo blockade of PD-1 and Tim-3 during the memory phase. However, combined blockade of PD-1 and Tim-3 during the priming/differentiation phase rescued FV-specific CD8(+) T cells from becoming terminally exhausted, resulting in improved CD8(+) T cell functionality and virus control. These results highlight FV's unique ability to evade virus-specific CD8(+) T cell responses and the importance of an early prophylactic approach for preventing terminal exhaustion of CD8(+) T cells.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Vírus da Leucemia Murina de Friend/imunologia , Ativação Linfocitária/imunologia , Receptores KIR/biossíntese , Animais , Antígeno B7-1/fisiologia , Antígeno B7-H1 , Linfócitos T CD8-Positivos/virologia , Células Cultivadas , Epitopos de Linfócito T/imunologia , Eritroblastos/imunologia , Eritroblastos/patologia , Eritroblastos/virologia , Feminino , Receptor Celular 1 do Vírus da Hepatite A , Receptor Celular 2 do Vírus da Hepatite A , Evasão da Resposta Imune/imunologia , Masculino , Glicoproteínas de Membrana/antagonistas & inibidores , Glicoproteínas de Membrana/fisiologia , Proteínas de Membrana/antagonistas & inibidores , Proteínas de Membrana/fisiologia , Camundongos , Camundongos Endogâmicos A , Camundongos Endogâmicos C57BL , Peptídeos/antagonistas & inibidores , Peptídeos/fisiologia , Receptores KIR/fisiologia , Receptores Virais/antagonistas & inibidores , Receptores Virais/fisiologia , Infecções por Retroviridae/imunologia , Infecções por Retroviridae/patologia , Infecções por Retroviridae/virologia , Infecções Tumorais por Vírus/imunologia , Infecções Tumorais por Vírus/patologia , Infecções Tumorais por Vírus/virologiaRESUMO
Erythropoiesis is the process by which nucleated erythroid progenitors proliferate and differentiate to generate, every second, millions of nonnucleated red cells with their unique discoid shape and membrane material properties. Here we examined the time course of appearance of individual membrane protein components during murine erythropoiesis to throw new light on our understanding of the evolution of the unique features of the red cell membrane. We found that the accumulation of all of the major transmembrane and all skeletal proteins of the mature red blood cell, except actin, accrued progressively during terminal erythroid differentiation. At the same time, and in marked contrast, accumulation of various adhesion molecules decreased. In particular, the adhesion molecule, CD44 exhibited a progressive and dramatic decrease from proerythroblast to reticulocyte; this enabled us to devise a new strategy for distinguishing unambiguously between erythroblasts at successive developmental stages. These findings provide unique insights into the genesis of red cell membrane function during erythroblast differentiation and also offer a means of defining stage-specific defects in erythroid maturation in inherited and acquired red cell disorders and in bone marrow failure syndromes.
Assuntos
Proteínas Sanguíneas/genética , Células Precursoras Eritroides/citologia , Eritropoese/fisiologia , Proteínas de Membrana/genética , Animais , Proteína 1 de Troca de Ânion do Eritrócito/genética , Antígeno CD47/genética , Moléculas de Adesão Celular/genética , Eritroblastos/citologia , Eritroblastos/fisiologia , Eritroblastos/virologia , Membrana Eritrocítica/fisiologia , Membrana Eritrocítica/ultraestrutura , Células Precursoras Eritroides/fisiologia , Citometria de Fluxo , Vírus da Leucemia Murina de Friend , Regulação da Expressão Gênica , Humanos , Receptores de Hialuronatos/fisiologia , Proteínas de Membrana/sangue , CamundongosRESUMO
Infection of mice with Friend spleen focus-forming virus (SFFV) results in a multistage erythroleukemia. In the first stage, the SFFV envelope glycoprotein interacts with the erythropoietin receptor and a short form of the receptor tyrosine kinase sf-Stk, resulting in constitutive activation of signal transducing molecules and the development of erythropoietin (Epo)-independent erythroid hyperplasia and polycythemia. The second stage results from the outgrowth of a rare virus-infected erythroid cell that expresses nonphysiological levels of the myeloid transcription factor PU.1. These cells exhibit a differentiation block and can be grown as murine erythroleukemia (MEL) cell lines. In this study, we examined SFFV MEL cells to determine whether their transformed phenotype was associated with a block in the activation of any Epo signal-transducing molecules. Our studies indicate that Epo- or SFFV-induced activation of STAT1/3 DNA binding activity is blocked in SFFV MEL cells. The block is at the level of tyrosine phosphorylation of STAT1, although Jak2 phosphorylation is not blocked in these cells. In contrast to Epo, alpha interferon can induce STAT1 phosphorylation and DNA binding in SFFV MEL cells. The SFFV-transformed cells were shown to express elevated levels of the hematopoietic phosphatase SHP-1, and treatment of the cells with a phosphatase inhibitor restored STAT1 tyrosine phosphorylation. MEL cells derived from Friend murine leukemia virus (MuLV) or ME26 MuLV-infected mice, which do not express PU.1, express lower levels of SHP-1 and are not blocked in STAT1/3 DNA-binding activity. Our studies suggest that SFFV-infected erythroid cells become transformed when differentiation signals activated by STAT1/3 are blocked due to high SHP-1 levels induced by inappropriate expression of the PU.1 protein.
Assuntos
Transformação Celular Viral , DNA/metabolismo , Eritroblastos/virologia , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas Tirosina Fosfatases/genética , Fator de Transcrição STAT1/metabolismo , Vírus Formadores de Foco no Baço/patogenicidade , Animais , Diferenciação Celular , Eritropoetina/fisiologia , Regulação da Expressão Gênica , Camundongos , Fosforilação , Proteína Tirosina Fosfatase não Receptora Tipo 6 , Proteínas Proto-Oncogênicas/genética , Transativadores/genéticaRESUMO
Regulatory factors other than erythropoietin (Epo) dependence, that control mammalian erythroid terminal differentiation, are currently uncertain. Here we report the existence of erythroid differentiation factors in erythroid cytoplasm. Purification of these factors from cultured Friend virus anaemia (FVA)-infected mouse splenic erythroblasts was carried out using isoelectrophoresis and high performance of liquid chromatography techniques. We have identified intracellular erythroid differentiation denucleation factors (EDDFs) that were able to mediate the events of post-Epo-dependent erythroblast terminal differentiation. Purified EDDF proteins bound specifically to the enhancer HS2 sequence of the globin gene activated the expression of haemoglobin in mouse erythroleukaemia and K562 erythroleukaemic cells and promoted them to differentiate into mature erythrocytes. EDDF proteins began to emerge at the pro-early erythroblast stages upon exposure to Epo in culture, and increased dramatically in early erythroblast stage. The dynamic of EDDF expression and its action on the key events of erythroblast differentiation and denucleation appeared to be closely consistent with its spatiotemporal distribution. These results suggest that EDDFs are the critical intracellular regulatory factors that may act as the successive regulators to Epo, responsible for the final stages of erythroid terminal differentiation.
Assuntos
Ativinas/metabolismo , Eritrócitos/citologia , Células Precursoras Eritroides/citologia , Eritropoese/fisiologia , Eritropoetina/fisiologia , Subunidades beta de Inibinas/metabolismo , Ativinas/isolamento & purificação , Animais , Diferenciação Celular , Fusão Celular , Linhagem Celular Transformada , Células Cultivadas , Citoplasma/metabolismo , Elementos Facilitadores Genéticos , Eritroblastos/citologia , Eritroblastos/metabolismo , Eritroblastos/virologia , Feminino , Vírus da Leucemia Murina de Friend , Globinas/genética , Globinas/metabolismo , Humanos , Subunidades beta de Inibinas/isolamento & purificação , Camundongos , Camundongos Endogâmicos BALB C , Baço/citologiaRESUMO
Friend virus is an acutely oncogenic retrovirus that causes erythroblastosis and polycythemia in mice. Previous studies suggested that the Friend virus oncoprotein, gp55, constitutively activates the erythropoietin receptor (EPOR), causing uncontrolled erythroid proliferation. Those studies showed that gp55 confers growth factor independence on an interleukin-3 (IL-3)-dependent cell line (Ba/F3) when the EPOR is coexpressed. Subsequently, we showed that a truncated form of the stem-cell kinase receptor (sf-STK) is required for susceptibility to Friend disease. Given the requirement for sf-STK, we sought to establish the in vivo significance of gp55-mediated activation of the EPOR. We found that the cytoplasmic tyrosine residues of the EPOR, and signal transducer and activator of transcription-5 (STAT5), which acts through these sites, are not required for Friend virus-induced erythroblastosis. The EPOR itself was required for the development of erythroblastosis but not for gp55-mediated erythroid proliferation. Interestingly, the murine EPOR, which is required for gp55-mediated Ba/F3-cell proliferation, was dispensable for erythroblastosis in vivo. Finally, gp55-mediated activation of the EPOR and STAT5 are required for Friend virus-induced polycythemia. These results suggest that Friend virus activates both sf-STK and the EPOR to cause deregulated erythroid proliferation and differentiation.
Assuntos
Vírus da Leucemia Murina de Friend/patogenicidade , Leucemia Experimental/virologia , Policitemia/virologia , Receptores da Eritropoetina/fisiologia , Proteínas do Envelope Viral/fisiologia , Animais , Diferenciação Celular , Proliferação de Células , Transformação Celular Viral , Eritroblastos/patologia , Eritroblastos/virologia , Leucemia Experimental/etiologia , Leucemia Experimental/patologia , Camundongos , Camundongos Knockout , Receptores da Eritropoetina/metabolismo , Infecções por Retroviridae , Fator de Transcrição STAT5/metabolismo , Fator de Transcrição STAT5/fisiologia , Transdução de Sinais/fisiologia , Infecções Tumorais por VírusRESUMO
The etiology of transient erythroblastopenia of childhood (TEC) remains unknown, although an association with viral infections has been proposed. The authors describe a 3.5-year-old girl with classic TEC concomitantly with human parvovirus B19 (HPV) infection. The infection was evident by detection of HPV genome in the blood and the bone marrow by polymerase chain reaction. Viral genome was no longer detected when the TEC resolved clinically. The patient was immunocompetent and the anemia has not recurred. To the authors' knowledge, this is one of the few documented cases of classic TEC attributable to HPV infection.
Assuntos
Eritroblastos/virologia , Infecções por Parvoviridae/sangue , Infecções por Parvoviridae/complicações , Parvovirus/isolamento & purificação , Aplasia Pura de Série Vermelha/virologia , Sangue/virologia , Células da Medula Óssea/virologia , Núcleo Celular/virologia , Pré-Escolar , Feminino , Genoma Viral , Hemoglobinas/análise , Humanos , Parvovirus/genética , Aplasia Pura de Série Vermelha/sangue , Contagem de ReticulócitosRESUMO
In primary chicken erythroblasts (stem cell factor [SCF] erythroblasts), transferrin receptor (TfR) messenger RNA (mRNA) and protein were hyperexpressed as compared to nonerythroid chicken cell types. This erythroid-specific hyperexpression was abolished in transformed erythroblasts (HD3E22 cells) expressing the v-ErbA and v-ErbB oncogenes of avian erythroblastosis virus. TfR expression in HD3E22 cells could be modulated by changes in exogenous iron supply, whereas expression in SCF erythroblasts was not subject to iron regulation. Measurements of TfR mRNA half-life indicated that hyperexpression in SCF erythroblasts was due to a massive stabilization of transcripts even in the presence of high iron levels. Changes in mRNA binding activity of iron regulatory protein 1 (IRP1), the primary regulator of TfR mRNA stability in these cells, correlated well with TfR mRNA expression; IRP1 activity in HD3E22 cells and other nonerythroid cell types tested was iron dependent, whereas IRP1 activity in primary SCF erythroblasts could not be modulated by iron administration. Analysis of avian erythroblasts expressing v-ErbA alone indicated that v-ErbA was responsible for these transformation-specific alterations in the regulation of iron metabolism. In SCF erythroblasts high amounts of TfR were detected on the plasma membrane, but a large fraction was also located in early and late endosomal compartments, potentially concealing temporary iron stores from the IRP regulatory system. In contrast, TfR was almost exclusively located to the plasma membrane in HD3E22 cells. In summary, stabilization of TfR mRNA and redistribution of Fe-Tf/TfR complexes to late endosomal compartments may contribute to TfR hyperexpression in primary erythroblasts, effects that are lost on leukemic transformation.
Assuntos
Alpharetrovirus , Transformação Celular Viral/fisiologia , Eritroblastos/metabolismo , Eritroblastos/virologia , Receptores da Transferrina/metabolismo , Animais , Compartimento Celular , Galinhas , Homeostase , Ferro/metabolismo , Ferro/farmacocinética , Ferro/farmacologia , Proteína 1 Reguladora do Ferro , Proteínas Reguladoras de Ferro , Proteínas Ferro-Enxofre/metabolismo , Transporte Proteico , Estabilidade de RNA , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/metabolismo , Receptores da Transferrina/efeitos dos fármacos , Receptores da Transferrina/genéticaRESUMO
During the initial stage of Friend virus-induced erythroleukemia in mice, interaction of the viral protein gp55 with the erythropoietin receptor, and other host factors, drives the expansion of erythroid precursor cells. Recently, we demonstrated that the Friend virus susceptibility locus, Fv2, which is required for the expansion of infected cells, encodes a naturally occurring, N-terminally truncated form of the Stk receptor tyrosine kinase (Sf-Stk). Here we show that in vitro expression of Sf-Stk confers Friend virus sensitivity to erythroid progenitor cells from Fv2(rr) mice. Furthermore, our data reveal that Sf-Stk kinase activity and Y436, but not Y429, are required for Epo-independent colony formation following Friend virus infection. Introduction of a mutation that results in failure to bind Grb2 abrogates the ability of Sf-Stk to induce colonies in response to Friend virus, while the Grb2 binding site from EGFR restores this response. Consistent with the ability of Grb2 to recruit SOS and Gab1, the Ras/MAPK and PI3K pathways are activated by Sf-Stk, and both of these pathways are required for gp55-mediated erythroblast proliferation. These data clearly demonstrate a requirement for signaling through Sf-Stk in the Epo-independent expansion of Friend virus-infected cells, and suggest a pivotal role for Grb2 in this response.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Eritroblastos/virologia , Vírus da Leucemia Murina de Friend/patogenicidade , Proteínas/metabolismo , Receptores Proteína Tirosina Quinases/química , Receptores Proteína Tirosina Quinases/metabolismo , Receptores de Superfície Celular/química , Receptores de Superfície Celular/metabolismo , Sequência de Aminoácidos , Animais , Sítios de Ligação , Divisão Celular , Linhagem Celular , Células Cultivadas , Eritroblastos/metabolismo , Eritropoetina/fisiologia , Proteína Adaptadora GRB2 , Leucemia Eritroblástica Aguda/virologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Dados de Sequência Molecular , Mutação , Receptores Proteína Tirosina Quinases/genética , Receptores de Superfície Celular/genética , Homologia de Sequência de Aminoácidos , Transdução de Sinais , Tirosina/genética , Tirosina/fisiologia , Proteínas do Envelope Viral/genéticaRESUMO
PURPOSE: It has been proposed that human parvovirus B19 or human herpesvirus 6 (HHV-6) causes transient erythroblastopenia of childhood (TEC). This study was performed prospectively and accurately in a sufficiently large population to confirm or reject these suggestions, or possibly to find another causative agent for TEC. PATIENTS AND METHODS: The authors studied prospectively the causative aspects of 10 consecutive children with TEC presenting at five Swedish pediatric clinics from 1994 to 1998 using serologic assays and polymerase chain reaction assays for B19, HHV-6, cytomegalovirus, and Epstein-Barr virus (EBV). The authors also performed isolation investigations of virus from bone marrow, stools, and aspirate from the nasopharynx. RESULTS: The authors found acute EBV infection in one child. There were no other instances of IgM positivity for any viruses at presentation, but a few children did show IgG positivity to HHV-6 (n = 2), EBV (n = 1), and cytomegalovirus (n = 1). No children were positive with respect to IgG for B19. However, it could not be determined when these children had been infected, and thus, a connection to TEC cannot be proved or excluded. No child showed B19 or HHV-6 DNA in serum or bone marrow collected at presentation. Isolation of virus from bone marrow, stools, and aspirate from nasopharynx culture was negative in all children but one, who had rotavirus in stool. CONCLUSIONS: HHV-6, B19, EBV, and cytomegalovirus are not common causative agents of TEC. The authors found no proof of any single agent causing TEC.