RESUMO
In most forms of prion disease, infectivity is present primarily in the central nervous system or immune system organs such as spleen and lymph node. However, a transgenic mouse model of prion disease has demonstrated that prion infectivity can also be present as amyloid deposits in heart tissue. Deposition of infectious prions as amyloid in human heart tissue would be a significant public health concern. Although abnormal disease-associated prion protein (PrP(Sc)) has not been detected in heart tissue from several amyloid heart disease patients, it has been observed in the heart tissue of a patient with sporadic Creutzfeldt-Jakob Disease (sCJD), the most common form of human prion disease. In order to determine whether prion infectivity can be found in heart tissue, we have inoculated formaldehyde fixed brain and heart tissue from two sCJD patients, as well as prion protein positive fixed heart tissue from two amyloid heart disease patients, into transgenic mice overexpressing the human prion protein. Although the sCJD brain samples led to clinical or subclinical prion infection and deposition of PrP(Sc) in the brain, none of the inoculated heart samples resulted in disease or the accumulation of PrP(Sc). Thus, our results suggest that prion infectivity is not likely present in cardiac tissue from sCJD or amyloid heart disease patients.
Assuntos
Amiloidose/metabolismo , Amiloidose/patologia , Cardiomiopatias/metabolismo , Cardiomiopatias/patologia , Síndrome de Creutzfeldt-Jakob/metabolismo , Síndrome de Creutzfeldt-Jakob/patologia , Miocárdio/metabolismo , Miocárdio/patologia , Proteínas PrPSc/metabolismo , Proteínas PrPSc/patogenicidade , Animais , Encéfalo/metabolismo , Encéfalo/patologia , Síndrome de Creutzfeldt-Jakob/transmissão , Cricetinae , Modelos Animais de Doenças , Humanos , Imuno-Histoquímica , Camundongos , Camundongos TransgênicosRESUMO
Persistent viral infections are a major health concern. One obstacle inhibiting the clearance of persistent infections is functional inactivation of antiviral T cells. Although such immunosuppression occurs rapidly after infection, the mechanisms that induce the loss of T-cell activity and promote viral persistence are unknown. Herein we document that persistent viral infection in mice results in a significant upregulation of interleukin (IL)-10 by antigen-presenting cells, leading to impaired T-cell responses. Genetic removal of Il10 resulted in the maintenance of robust effector T-cell responses, the rapid elimination of virus and the development of antiviral memory T-cell responses. Therapeutic administration of an antibody that blocks the IL-10 receptor restored T-cell function and eliminated viral infection. Thus, we identify a single molecule that directly induces immunosuppression leading to viral persistence and demonstrate that a therapy to neutralize IL-10 results in T-cell recovery and the prevention of viral persistence.
Assuntos
HIV/fisiologia , Hepacivirus/fisiologia , Interleucina-10/fisiologia , Vírus da Coriomeningite Linfocítica/fisiologia , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Citometria de Fluxo , Memória Imunológica , Camundongos , Receptores de Interleucina-10/antagonistas & inibidores , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
After infection with RML murine scrapie agent, transgenic (tg) mice expressing prion protein (PrP) without its glycophosphatidylinositol (GPI) membrane anchor (GPI(-/-) PrP tg mice) continue to make abundant amounts of the abnormally folded disease-associated PrPres but have a normal life span. In contrast, all age-, sex-, and genetically matched mice with a GPI-anchored PrP become moribund and die due to a chronic progressive neurodegenerative disease by 160 days after RML scrapie agent infection. We report here that infected GPI(-/-) PrP tg mice, although free from progressive neurodegenerative disease of the cerebellum and extrapyramidal and pyramidal systems, nevertheless suffer defects in learning and memory, long-term potentiation, and neuronal excitability. Such dysfunction increases over time and is associated with an increase in gamma aminobutyric acid (GABA) inhibition but not loss of excitatory glutamate/N-methyl-d-aspartic acid. Enhanced deposition of abnormally folded infectious PrP (PrPsc or PrPres) in the central nervous system (CNS) localizes with GABAA receptors. This occurs with minimal evidence of CNS spongiosis or apoptosis of neurons. The use of monoclonal antibodies reveals an association of PrPres with GABAA receptors. Thus, the clinical defects of learning and memory loss in vivo in GPI(-/-) PrP tg mice infected with scrapie agent may likely involve the GABAergic pathway.
Assuntos
Deficiências da Aprendizagem/fisiopatologia , Transtornos da Memória/fisiopatologia , Camundongos Transgênicos , Príons/metabolismo , Scrapie/metabolismo , Ácido gama-Aminobutírico/metabolismo , Animais , Comportamento Animal/fisiologia , Encéfalo/anatomia & histologia , Encéfalo/metabolismo , Encéfalo/patologia , Eletrofisiologia , Glicosilfosfatidilinositóis/genética , Glicosilfosfatidilinositóis/metabolismo , Humanos , Deficiências da Aprendizagem/patologia , Transtornos da Memória/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Testes Neuropsicológicos , Príons/genética , Distribuição Aleatória , Receptores de GABA-A/genética , Receptores de GABA-A/metabolismo , Scrapie/genética , Scrapie/patologia , Scrapie/fisiopatologiaRESUMO
Earlier studies indicated that transgenic (tg) mice engineered to express prion protein (PrP) lacking the glycophosphatidylinositol (GPIâ»/â») membrane anchor formed abnormal proteinase-resistant prion (PrPsc) amyloid deposits in their brains and hearts when infected with the RML strain of murine scrapie. In contrast, RML scrapie infection of normal mice with a GPI-anchored PrP did not deposit amyloid with PrPsc in the brain or the heart. Here we report that scrapie-infected GPIâ»/â» PrP tg mice also deposit PrP and transmissible infectious material in the gut, kidneys, and islets of Langerhans. Similar to previously reported amyloid deposits in the brain and heart, amyloid deposits were found in the gut; however, no amyloid deposited in the islets. By high-resolution electron microscopy, we show PrP is located primarily in α cells and also ß cells. Islets contain abundant insulin and there is no abnormality in glucose metabolism in infected GPIâ»/â» PrP tg mice.
Assuntos
Glicosilfosfatidilinositóis/deficiência , Doenças Priônicas/veterinária , Príons/metabolismo , Doenças dos Roedores/patologia , Amiloide/análise , Estruturas Animais/patologia , Animais , Feminino , Masculino , Camundongos , Camundongos Transgênicos , Doenças Priônicas/patologia , Proteínas PriônicasRESUMO
To study the pathogenesis of chronic wasting disease (CWD) in deer and elk, transgenic (tg) mice were generated that expressed the prion protein (PrP) of deer containing a glycine at amino acid (aa) 96 and a serine at aa 225 under transcriptional control of the murine PrP promoter. This construct was introduced into murine PrP-deficient mice. As anticipated, neither non-tg mice nor PrP ko mice were susceptible when inoculated intracerebrally (i.c.) or orally with CWD brain material (scrapie pool from six mule deer) and followed for 600+ days (dpi). Deer PrP tg mice were not susceptible to i.c. inoculation with murine scrapie. In contrast, a fatal neurologic disease occurred accompanied by conversion of deer PrPsen to PrPres by western blot and immunohistochemistry after either i.c. inoculation with CWD brain into two lines of tg mice studied (312+32 dpi [mean+2 standard errors] for the heterozygous tg line 33, 275+46 dpi for the heterozygous tg line 39 and 210 dpi for the homozygous tg line 33) or after oral inoculation (381+55 dpi for the homozygous tg line 33 and 370+26 dpi for the homozygous tg line 39). Kinetically, following oral inoculation of CWD brain, PrPres was observed by day 200 when mice were clinically healthy in the posterior surface of the dorsum of the tongue primarily in serous and mucous glands, in the intestines, in large cells at the splenic marginal zone that anatomically resembled follicular dendritic cells and macrophages and in the olfactory bulb and brain stem but did not occur in the cerebellum, cerebral cortex or hippocampus or in hearts, lungs and livers of infected mice. After 350 days when mice become clinically ill the cerebellum, cerebral cortex and hippocampus became positive for PrPres and displayed massive spongiosis, neuronal drop out, gliosis and florid plaques.
Assuntos
Administração Oral , Encéfalo/patologia , Transmissão de Doença Infecciosa , Doenças Priônicas/transmissão , Príons/metabolismo , Doença de Emaciação Crônica/transmissão , Animais , Bovinos , Cervos , Modelos Animais de Doenças , Camundongos , Camundongos Transgênicos , Doença de Emaciação Crônica/etiologia , Doença de Emaciação Crônica/patologiaRESUMO
Prion protein (PrP) is a required factor for susceptibility to transmissible spongiform encephalopathy or prion diseases. In transgenic mice, expression of prion protein (PrP) from another species often confers susceptibility to prion disease from that donor species. For example, expression of deer or elk PrP in transgenic mice has induced susceptibility to chronic wasting disease (CWD), the prion disease of cervids. In the current experiments, transgenic mice expressing two naturally occurring allelic variants of deer PrP with either glycine (G) or serine (S) at residue 96 were found to differ in susceptibility to CWD infection. G96 mice were highly susceptible to infection, and disease appeared starting as early as 160 days postinfection. In contrast, S96 mice showed no evidence of disease or generation of disease-associated protease-resistant PrP (PrPres) over a 600-day period. At the time of clinical disease, G96 mice showed typical vacuolar pathology and deposition of PrPres in many brain regions, and in some individuals, extensive neuronal loss and apoptosis were noted in the hippocampus and cerebellum. Extraneural accumulation of PrPres was also noted in spleen and intestinal tissue of clinically ill G96 mice. These results demonstrate the importance of deer PrP polymorphisms in susceptibility to CWD infection. Furthermore, this deer PrP transgenic model is the first to demonstrate extraneural accumulation of PrPres in spleen and intestinal tissue and thus may prove useful in studies of CWD pathogenesis and transmission by oral or other natural routes of infection.
Assuntos
Imunidade Inata/genética , Polimorfismo Genético , Príons/metabolismo , Doença de Emaciação Crônica/genética , Animais , Sequência de Bases , Encéfalo/patologia , Cervos/metabolismo , Immunoblotting , Camundongos , Camundongos Transgênicos , Dados de Sequência Molecular , Oligonucleotídeos/genética , Príons/genética , Doença de Emaciação Crônica/patologiaRESUMO
We investigated extraneural manifestations in scrapie-infected transgenic mice expressing prion protein lacking the glycophosphatydylinositol membrane anchor. In the brain, blood, and heart, both abnormal protease-resistant prion protein (PrPres) and prion infectivity were readily detected by immunoblot and by inoculation into nontransgenic recipients. The titer of infectious scrapie in blood plasma exceeded 10(7) 50% infectious doses per milliliter. The hearts of these transgenic mice contained PrPres-positive amyloid deposits that led to myocardial stiffness and cardiac disease.
Assuntos
Amiloide/análise , Amiloidose/patologia , Cardiopatias/patologia , Miocárdio/química , Miocárdio/patologia , Proteínas PrPSc/análise , Scrapie/patologia , Amiloidose/sangue , Amiloidose/etiologia , Amiloidose/fisiopatologia , Animais , Western Blotting , Cateterismo Cardíaco , Vasos Coronários/química , Vasos Coronários/patologia , Modelos Animais de Doenças , Glicosilfosfatidilinositóis , Cardiopatias/sangue , Cardiopatias/etiologia , Cardiopatias/fisiopatologia , Testes de Função Cardíaca , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Microcirculação/química , Microcirculação/patologia , Contração Miocárdica , Proteínas PrPC/química , Proteínas PrPSc/sangue , Scrapie/sangue , Scrapie/fisiopatologia , Coloração e Rotulagem , Fatores de TempoRESUMO
Understanding, treating, and preventing diseases caused by immunosuppression and/or persistent infections remain both a major challenge in biomedical research and an important health goal. For a virus or any infectious agent to persist, it must utilize strategies to suppress or evade the host's immune response. Here, we report that two dissimilar viruses employ a common maneuver to cause a profound immunosuppression. Measles virus (MV) and lymphocytic choriomeningitis virus (LCMV) interfere with dendritic cell (DC) development and expansion in vivo and in vitro. The underlying mechanism for this is through the generation of type I interferon (IFN) that acts via a signal transducer and activator of a transcription (STAT)2-dependent, but STAT1-independent, pathway. Thus, viruses subvert the known antiviral effect of type I IFN through STAT2-specific signaling to benefit their survival. These observations have implications for understanding and developing therapies to treat diseases caused by immunosuppression and/or persistent infections.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Interferon Tipo I/metabolismo , Vírus da Coriomeningite Linfocítica/fisiologia , Vírus do Sarampo/fisiologia , Transdução de Sinais , Transativadores/metabolismo , Animais , Infecções por Arenaviridae/imunologia , Infecções por Arenaviridae/metabolismo , Infecções por Arenaviridae/virologia , Células da Medula Óssea/citologia , Células da Medula Óssea/imunologia , Diferenciação Celular , Células Cultivadas , Proteínas de Ligação a DNA/imunologia , Células Dendríticas/citologia , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Interferon Tipo I/imunologia , Vírus da Coriomeningite Linfocítica/imunologia , Vírus do Sarampo/imunologia , Camundongos , Infecções por Paramyxoviridae/imunologia , Infecções por Paramyxoviridae/metabolismo , Infecções por Paramyxoviridae/virologia , Fator de Transcrição STAT1 , Fator de Transcrição STAT2 , Transativadores/imunologiaRESUMO
In prion and Alzheimer's diseases, the roles played by amyloid versus nonamyloid deposits in brain damage remain unresolved. In scrapie-infected transgenic mice expressing prion protein (PrP) lacking the glycosylphosphatidylinositol (GPI) membrane anchor, abnormal protease-resistant PrPres was deposited as amyloid plaques, rather than the usual nonamyloid form of PrPres. Although PrPres amyloid plaques induced brain damage reminiscent of Alzheimer's disease, clinical manifestations were minimal. In contrast, combined expression of anchorless and wild-type PrP produced accelerated clinical scrapie. Thus, the PrP GPI anchor may play a role in the pathogenesis of prion diseases.
Assuntos
Glicosilfosfatidilinositóis/metabolismo , Príons/metabolismo , Scrapie/etiologia , Animais , Encéfalo/metabolismo , Encéfalo/patologia , Encéfalo/ultraestrutura , Glicosilfosfatidilinositóis/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Placa Amiloide/metabolismo , Placa Amiloide/patologia , Proteínas PrPSc/química , Proteínas PrPSc/metabolismo , Doenças Priônicas/etiologia , Doenças Priônicas/metabolismo , Doenças Priônicas/patologia , Príons/biossíntese , Príons/química , Príons/genética , Scrapie/metabolismo , Scrapie/patologiaRESUMO
The role of CC chemokine ligand 3 (CCL3) in activation of dendritic cells (DCs) following mouse hepatitis virus (MHV) infection of the central nervous system (CNS) was examined. The results indicate that CCL3 participates in an effective host response to MHV infection by contributing to CD11c+CD11b+CD8alpha- DC maturation, activation, and migration to cervical lymph nodes (CLN). Diminished CD8alpha- DC activation correlated with reduced IFN-gamma expression by virus-specific T cells accompanied by increased IL-10 production suggesting that CCL3 contributes to an effective host response to viral infection by enhancing the T cell activation potential of DC.
Assuntos
Viroses do Sistema Nervoso Central/imunologia , Infecções por Coronavirus/imunologia , Células Dendríticas/citologia , Imunidade Inata , Proteínas Inflamatórias de Macrófagos/metabolismo , Vírus da Hepatite Murina/patogenicidade , Animais , Encéfalo/citologia , Encéfalo/imunologia , Encéfalo/virologia , Antígeno CD11b/metabolismo , Antígeno CD11c/metabolismo , Antígenos CD8/metabolismo , Diferenciação Celular , Movimento Celular , Viroses do Sistema Nervoso Central/virologia , Quimiocina CCL3 , Quimiocina CCL4 , Infecções por Coronavirus/virologia , Células Dendríticas/imunologia , Linfonodos/citologia , Linfonodos/imunologia , Linfonodos/virologia , Ativação Linfocitária , Camundongos , Vírus da Hepatite Murina/genética , Linfócitos T/imunologiaRESUMO
In the present study, the authors evaluated the contributions of the CXC chemokine ligand (CXCL)10 to leukocyte recruitment into the central nervous system (CNS) and disease development. Instillation of a replication-deficient adenovirus that expresses CXCL10 (AdCXCL10) into the CNS of C57BL/6 mice resulted in a rapid (day 3) and prolonged (day 21) infiltration of both CD4(+) and CD8(+) T cells as compared to mice infected with an adenovirus vector containing beta-galactosidase (Adbetagal). Despite increased T-cell infiltration into the CNS of AdCXCL10-infected mice, production of proinflammatory chemokines normally associated with the recruitment of activated T cells into the CNS was muted and mice developed limited neuropathology. Therefore, these results indicate that T-cell infiltration in the absence of appropriate activation is not sufficient to induce pathology within the CNS and that additional signals other than CXCL10 are required for induction of an immune-mediated neurologic disease.
Assuntos
Adenoviridae/genética , Quimiocinas CXC/fisiologia , Quimiotaxia de Leucócito , Vírus Defeituosos/genética , Encefalite/imunologia , Vetores Genéticos/genética , Meningite/imunologia , Subpopulações de Linfócitos T/imunologia , Animais , Encéfalo , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Quimiocina CXCL10 , Quimiocinas CXC/biossíntese , Quimiocinas CXC/genética , Doenças Desmielinizantes/etiologia , Encefalite/patologia , Injeções , Óperon Lac , Masculino , Meningite/patologia , Camundongos , Camundongos Endogâmicos C57BL , Degeneração Neural/etiologia , Transcrição GênicaRESUMO
Chemokines induce the directional migration of targeted populations of leukocytes during periods of inflammation. Moreover, these molecules also regulate T-cell activation and differentiation following antigenic stimulation. In the present study, the contributions of the CC chemokine ligand 3 (CCL3) to the differentiation and migration of effector T cells in response to viral infection of the central nervous system (CNS) were analyzed. CCL3(-/-) mice infected with mouse hepatitis virus exhibited a significant reduction of virus-specific CD8(+) T cells within the CNS, correlating with delayed viral clearance. Decreased infiltration of CD8(+) T cells into infected CCL3(-/-) mice was associated with enhanced accumulation of primed CD8(+) T cells in cervical lymph nodes. Although virus-specific CD8(+) T cells from CCL3(-/-) mice were CD44(high), they remained CD62L(high) and CD25(low), retained CCR7 expression, and contained limited transcripts of the proinflammatory chemokine receptors CCR5 and CXCR3 compared with virus-specific CD8(+) T cells from CCL3(+/+) mice. Furthermore, the absence of CCL3 impaired the cytokine production and cytolytic activity of CD8(+) T cells. In addition, macrophage accumulation within the CNS was significantly decreased in infected CCL3(-/-) mice, correlating with reduced demyelination. These results suggest that CCL3 not only mediates macrophage chemotaxis but also significantly enhances differentiation of primed CD8(+) T cells into effector cells and their release into circulation, thus potentiating effective migration to the site of infection.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Quimiocinas CC/fisiologia , Infecções por Coronavirus/imunologia , Hepatite Viral Animal/imunologia , Proteínas Inflamatórias de Macrófagos/fisiologia , Vírus da Hepatite Murina/imunologia , Transferência Adotiva , Animais , Linfócitos T CD8-Positivos/fisiologia , Movimento Celular , Doenças do Sistema Nervoso Central/imunologia , Doenças do Sistema Nervoso Central/virologia , Quimiocina CCL3 , Quimiocina CCL4 , Quimiocinas CC/deficiência , Quimiocinas CC/genética , Infecções por Coronavirus/virologia , Doenças Desmielinizantes/etiologia , Doenças Desmielinizantes/imunologia , Doenças Desmielinizantes/virologia , Hepatite Viral Animal/virologia , Ligantes , Proteínas Inflamatórias de Macrófagos/deficiência , Proteínas Inflamatórias de Macrófagos/genética , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Vírus da Hepatite Murina/patogenicidade , Fenótipo , Transdução de SinaisRESUMO
Phosphoinositide 3-kinase activation is important for lymphocyte proliferation and survival. Disrupting the gene that encodes the major phosphoinositide 3-kinase regulatory isoform p85alpha impairs B cell development and proliferation. However, T cell functions are intact in the absence of p85alpha. In this study, we test the hypothesis that the related isoform p85beta is an essential regulatory subunit for T cell signaling. Unexpectedly, T cells lacking p85beta showed a marked increase in proliferation and decreased death when stimulated with anti-CD3 plus IL-2. Both CD4(+) and CD8(+) T cells completed more cell divisions. Transcriptional profiling revealed reduced levels of caspase-6 mRNA in p85beta-deficient T cells, which was paralleled by reduced caspase-6 enzyme activity. Increased T cell accumulation was also observed in vivo following infection of p85beta-deficient mice with mouse hepatitis virus. Together, these results suggest a unique role for p85beta in limiting T cell expansion.
Assuntos
Ativação Linfocitária , Fosfatidilinositol 3-Quinases/fisiologia , Linfócitos T/imunologia , Animais , Linfócitos B/fisiologia , Cálcio/metabolismo , Caspase 6 , Caspases/fisiologia , Interleucina-2/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Fosfatidilinositol 3-Quinases/química , Fosfolipase C gama , Subunidades Proteicas , Linfócitos T/fisiologia , Fosfolipases Tipo C/fisiologiaRESUMO
How chemokines shape the immune response to viral infection of the central nervous system (CNS) has largely been considered within the context of recruitment and activation of antigen-specific lymphocytes. However, chemokines are expressed early following viral infection, suggesting an important role in coordinating innate immune responses. Herein, we evaluated the contributions of CXC chemokine ligand 10 (CXCL10) in promoting innate defense mechanisms following coronavirus infection of the CNS. Intracerebral infection of RAG1(-/-) mice with a recombinant CXCL10-expressing murine coronavirus (mouse hepatitis virus) resulted in protection from disease and increased survival that correlated with a significant increase in recruitment and activation of natural killer (NK) cells within the CNS. Accumulation of NK cells resulted in a reduction in viral titers that was dependent on gamma interferon secretion. These results indicate that CXCL10 expression plays a pivotal role in defense following coronavirus infection of the CNS by enhancing innate immune responses.