RESUMO
Sporadic Creutzfeldt-Jakob disease (CJD) is a rare neurodegenerative spongiform encephalopathy that causes neuronal derangement secondary to prion protein. Its initial diagnosis is often complex and challenging due to non-specific clinical presentation, lack of awareness, and low clinical suspicion. This disease is invariably fatal, and most patients die within 12 months of presentation. Definite diagnosis of prion disease requires neuropathological analysis, usually done at autopsy. Here, we present the autopsy findings of a 57-year-old male patient, illustrating the complexity of diagnosing this disease early in the clinical course and the need for a broad differential diagnosis at the onset.
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Since prion diseases result from infection and neurodegeneration of the central nervous system (CNS), experimental characterizations of prion strain properties customarily rely on the outcomes of intracerebral challenges. However, natural transmission of certain prions, including those causing chronic wasting disease (CWD) in elk and deer, depends on propagation in peripheral host compartments prior to CNS infection. Using gene-targeted GtE and GtQ mice, which accurately control cellular elk or deer PrP expression, we assessed the impact that peripheral or intracerebral exposures play on CWD prion strain propagation and resulting CNS abnormalities. Whereas oral and intraperitoneal transmissions produced identical neuropathological outcomes in GtE and GtQ mice and preserved the naturally convergent conformations of elk and deer CWD prions, intracerebral transmissions generated CNS prion strains with divergent biochemical properties in GtE and GtQ mice that were changed compared to their native counterparts. While CWD replication kinetics remained constant during iterative peripheral transmissions and brain titers reflected those found in native hosts, serial intracerebral transmissions produced 10-fold higher prion titers and accelerated incubation times. Our demonstration that peripherally and intracerebrally challenged Gt mice develop dissimilar CNS diseases which result from the propagation of distinct CWD prion strains points to the involvement of tissue-specific cofactors during strain selection in different host compartments. Since peripheral transmissions preserved the natural features of elk and deer prions, whereas intracerebral propagation produced divergent strains, our findings illustrate the importance of experimental characterizations using hosts that not only abrogate species barriers but also accurately recapitulate natural transmission routes of native strains.
Assuntos
Encéfalo , Cervos , Príons , Doença de Emaciação Crônica , Animais , Doença de Emaciação Crônica/transmissão , Camundongos , Encéfalo/metabolismo , Encéfalo/patologia , Príons/metabolismo , Príons/genética , Príons/patogenicidade , Camundongos TransgênicosRESUMO
ABSTRACT Sporadic Creutzfeldt-Jakob disease (CJD) is a rare neurodegenerative spongiform encephalopathy that causes neuronal derangement secondary to prion protein. Its initial diagnosis is often complex and challenging due to non-specific clinical presentation, lack of awareness, and low clinical suspicion. This disease is invariably fatal, and most patients die within 12 months of presentation. Definite diagnosis of prion disease requires neuropathological analysis, usually done at autopsy. Here, we present the autopsy findings of a 57-year-old male patient, illustrating the complexity of diagnosing this disease early in the clinical course and the need for a broad differential diagnosis at the onset.
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Resumo A doença de Creutzfeldt-Jakob é uma condição rara causada por príons. Embora a forma mais notória da doença seja a infecciosa, a forma mais comum é a chamada esporádica, onde ocorre a transformação de proteínas citoplasmáticas das células gliais em príons. Caracteriza-se por uma demência rapidamente progressiva cujo diagnóstico pode ser feito com grande precisão por meio de sinais clínicos, alterações típicas na ressonância magnética de crânio e o exame Real-Time Quaking-Induced Conversion (Rt-QuIC) no líquido cefalorraquidiano. Relatamos um caso da doença sem distúrbios cognitivos, mas com outros sinais clínicos comuns como anormalidades comportamentais, ataxia, reações extrapiramidais e mioclonia; observamos ainda alterações típicas na ressonância magnética do crânio (alterações de sinal afetando áreas dos lobos parietal e temporal) e um Rt-QuIC fortemente positivo. Entendemos que o relato do caso possa servir de alerta para que outros profissionais de saúde possam reconhecer a doença, aumentando as possibilidades de um diagnóstico mais preciso em casos semelhantes.
Abstract Creutzfeldt-Jakob disease is a rare condition caused by prions. Although the infectious form of the disease is the most well-known, the most common form is the so-called sporadic type, where the transformation of cytoplasmic proteins from glial cells into prions occurs. The disease is characterized by rapidly progressive dementia whose diagnosis can be made with great accuracy based on clinical signs, typical changes on magnetic resonance imaging and real-time quaking-induced conversion (Rt-QuIC) testing in cerebrospinal fluid. We report a case of the disease without cognitive disorders in the initial phase, but with other common clinical signs including behavioral abnormalities, ataxia, extrapyramidal features, and myoclonus; typical changes on magnetic resonance imaging of the skull (signal alterations affecting parietal and temporal lobes areas) and strongly positive Rt-QuIC test. This case report can serve to alert other health professionals on recognizing the disease and contribute to a more accurate diagnosis in similar cases.
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Prions are proteins that cause fatal neurodegenerative diseases. The misfolded conformation adopted by prions can be transmitted to other normally folded proteins. Therapeutics to stop prion proliferation have been studied experimentally; however, it is not clear how the combination of different types of treatments can decrease the growth rate of prions in the brain. In this article, we combine the implementation of pharmacological chaperones and interferons to develop a novel model using a non-linear system of ordinary differential equations and study the quantitative effects of these two treatments on the growth rate of prions. This study aims to identify how the two treatments affect prion proliferation, both individually and in tandem. We analyze the model, and qualitative global results on the disease-free and disease equilibria are proved analytically. Numerical simulations, using parameter values from in vivo experiments that provide a pharmaceutically important demonstration of the effects of these two treatments, are presented here. This mathematical model can be used to identify and optimize the best combination of the treatments within their safe ranges.
Assuntos
Doenças Priônicas , Príons , Proliferação de Células , Humanos , Interferons , Doenças Priônicas/tratamento farmacológicoRESUMO
Structural conversion of cellular prion protein (PrPC) into scrapie PrP (PrPSc) and subsequent aggregation are key events associated with the onset of transmissible spongiform encephalopathies (TSEs). Experimental evidence supports the role of nucleic acids (NAs) in assisting this conversion. Here, we asked whether PrP undergoes liquid-liquid phase separation (LLPS) and if this process is modulated by NAs. To this end, two 25-mer DNA aptamers, A1 and A2, were selected against the globular domain of recombinant murine PrP (rPrP90-231) using SELEX methodology. Multiparametric structural analysis of these aptamers revealed that A1 adopts a hairpin conformation. Aptamer binding caused partial unfolding of rPrP90-231 and modulated its ability to undergo LLPS and fibrillate. In fact, although free rPrP90-231 phase separated into large droplets, aptamer binding increased the number of droplets but noticeably reduced their size. Strikingly, a modified A1 aptamer that does not adopt a hairpin structure induced formation of amyloid fibrils on the surface of the droplets. We show here that PrP undergoes LLPS, and that the PrP interaction with NAs modulates phase separation and promotes PrP fibrillation in a NA structure and concentration-dependent manner. These results shed new light on the roles of NAs in PrP misfolding and TSEs.
Assuntos
Amiloide/metabolismo , Aptâmeros de Nucleotídeos/química , Aptâmeros de Nucleotídeos/metabolismo , Extração Líquido-Líquido/métodos , Doenças Priônicas/patologia , Proteínas Priônicas/química , Proteínas Priônicas/metabolismo , Animais , Camundongos , Conformação de Ácido Nucleico , Doenças Priônicas/metabolismo , Proteínas Priônicas/isolamento & purificação , Ligação Proteica , Conformação Proteica , Dobramento de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Técnica de Seleção de AptâmerosRESUMO
The infectious protein or prion (PrPSC) is a transmissible and replicable polypeptide, which arises from an abnormal folding of the PrP protein, by unknown mechanisms and without changes in the primary sequence of its amino acids. Its new spatial disposition arises from the substitution of its alpha helices by beta bands, which increase its structural stability, avoiding its complete proteolysis, resulting in a residual accumulation of prions. These prions induce the misfolding of normal PrP protein, generating their exponential increase, leading to a disturbance of neuronal homeostasis which results in the development of the fatal spongiform encephalopathy of the Creutzfeldt-Jakob disease (CJD). This is the most prevalent human prion disease, and 90% of cases are sporadic, suggesting the endogenous genesis of prions. There are different types of prions, identified based on the genetic variance of codon 129 amino acids of the prion protein. Meteonin (M) and Valine (V)), associated with the result of their enzymatic proteolysis, define prions type 1 (21 kDa) and type 2 (19 kDa). The Classical form of CJD produced by MM1 prion occurs in 70% of the cases. The Cerebellar form originated by the VV2 prion occurs in 15% of cases, the form with Kuru plates, associated with the prion MV2 occurs in 5%, and the Vacuolar, related to the MM2 prion occurs in 4%. CJD is always characterized by behavioral, motor, cognitive, and vision alterations and by findings in magnetic resonance imaging, electroencephalogram and cerebrospinal fluid that define each clinical and neuropathological form.
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Humanos , Príons , Síndrome de Creutzfeldt-Jakob/genética , Doenças PriônicasRESUMO
The search for antiprion compounds has been encouraged by the fact that transmissible spongiform encephalopathies (TSEs) share molecular mechanisms with more prevalent neurodegenerative pathologies, such as Parkinson's and Alzheimer's diseases. Cellular prion protein (PrPC) conversion into protease-resistant forms (protease-resistant PrP [PrPRes] or the scrapie form of PrP [PrPSc]) is a critical step in the development of TSEs and is thus one of the main targets in the screening for antiprion compounds. In this work, three trimethoxychalcones (compounds J1, J8, and J20) and one oxadiazole (compound Y17), previously identified in vitro to be potential antiprion compounds, were evaluated through different approaches in order to gain inferences about their mechanisms of action. None of them changed PrPC mRNA levels in N2a cells, as shown by reverse transcription-quantitative real-time PCR. Among them, J8 and Y17 were effective in real-time quaking-induced conversion reactions using rodent recombinant PrP (rPrP) from residues 23 to 231 (rPrP23-231) as the substrate and PrPSc seeds from hamster and human brain. However, when rPrP from residues 90 to 231 (rPrP90-231), which lacks the N-terminal domain, was used as the substrate, only J8 remained effective, indicating that this region is important for Y17 activity, while J8 seems to interact with the PrPC globular domain. J8 also reduced the fibrillation of mouse rPrP23-231 seeded with in vitro-produced fibrils. Furthermore, most of the compounds decreased the amount of PrPC on the N2a cell surface by trapping this protein in the endoplasmic reticulum. On the basis of these results, we hypothesize that J8, a nontoxic compound previously shown to be a promising antiprion agent, may act by different mechanisms, since its efficacy is attributable not only to PrP conversion inhibition but also to a reduction of the PrPC content on the cell surface.
Assuntos
Chalconas/farmacologia , Drogas em Investigação/farmacologia , Neurônios/efeitos dos fármacos , Oxidiazóis/farmacologia , Proteínas Priônicas/antagonistas & inibidores , Animais , Sítios de Ligação , Linhagem Celular Tumoral , Chalconas/síntese química , Clonagem Molecular , Drogas em Investigação/síntese química , Retículo Endoplasmático/efeitos dos fármacos , Retículo Endoplasmático/metabolismo , Retículo Endoplasmático/ultraestrutura , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Cinética , Camundongos , Simulação de Acoplamento Molecular , Neurônios/metabolismo , Neurônios/patologia , Oxidiazóis/síntese química , Proteínas Priônicas/química , Proteínas Priônicas/genética , Proteínas Priônicas/metabolismo , Ligação Proteica , Conformação Proteica em alfa-Hélice , Domínios e Motivos de Interação entre Proteínas , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Relação Estrutura-Atividade , TermodinâmicaRESUMO
ABSTRACT Gerstmann-Sträussler-Scheinker is a genetic prion disease and the most common mutation is p.Pro102Leu. We report clinical, molecular and neuropathological data of seven individuals, belonging to two unrelated Brazilian kindreds, carrying the p.Pro102Leu. Marked differences among patients were observed regarding age at onset, disease duration and clinical presentation. In the first kindred, two patients had rapidly progressive dementia and three exhibited predominantly ataxic phenotypes with variable ages of onset and disease duration. In this family, age at disease onset in the mother and daughter differed by 39 years. In the second kindred, different phenotypes were also reported and earlier ages of onset were associated with 129 heterozygosis. No differences were associated with apoE genotype. In these kindreds, the codon 129 polymorphism could not explain the clinical variability and 129 heterozygosis was associated with earlier disease onset. Neuropathological examination in two patients confirmed the presence of typical plaques and PrPsc immunopositivity.
RESUMO A doença de Gerstmann-Sträussler-Scheinker é uma doença priônica genética, cuja mutação mais frequente é p.Pro102Leu. Descrevem-se dados clínicos, moleculares e neuropatológicos de sete indivíduos em duas famílias não relacionadas com p.Pro102Leu. Diferenças notáveis entre os pacientes em relação à idade de início, duração da doença e apresentação clínica foram encontradas. Na primeira família, dois pacientes apresentaram demência rapidamente progressiva e três apresentaram fenótipo de ataxia com idade variáveis de início e duração da doença. Nesta família, a idade de início entre mãe e filha diferiu em 39 anos. Na segunda família, fenótipos diferentes foram observados e idades precoces de início dos sintomas foram associadas à heterozigose no códon 129. Não houve diferença em relação ao genótipo do gene da apoE. O genótipo do códon 129 não foi responsável pela variabilidade clínica; heterozigose no códon 129 esteve associada ao início precoce da doença. O exame neuropatológico em dois pacientes confirmou presença de placas típicas e imunohistoquímica para PrPsc.
Assuntos
Humanos , Masculino , Feminino , Adulto , Pessoa de Meia-Idade , Idoso , Adulto Jovem , Príons/genética , DNA , Doença de Gerstmann-Straussler-Scheinker/genética , Mutação , Linhagem , Fenótipo , Polimorfismo Genético , Encéfalo/patologia , Doença de Gerstmann-Straussler-Scheinker/patologiaRESUMO
Prion diseases are rare neurodegenerative disorders occurring worldwide and affecting both humans and animals. Herein, we present the case of a patient diagnosed with definite sporadic Creutzfeldt-Jakob disease in Cali, Colombia. Besides neurological examination, 14-3-3 and tau proteins were valuable tools supporting the diagnosis. We also present a brief perspective of the prion diseases reported in Colombia to date. Although the incidence of prion diseases is unknown in Colombia, our literature review revealed that one case of scrapie in 1981 and 29 human sporadic cases of Creutzfeldt-Jakob disease have been documented and published in our country.
Assuntos
Síndrome de Creutzfeldt-Jakob/genética , Doenças Priônicas/genética , Proteínas tau/genética , Proteínas 14-3-3/metabolismo , Animais , Western Blotting , Colômbia , Síndrome de Creutzfeldt-Jakob/metabolismo , Humanos , Incidência , Doenças Priônicas/metabolismo , Proteínas tau/metabolismoRESUMO
Prion diseases are rare neurodegenerative disorders occurring worldwide and affecting both humans and animals. Herein, we present the case of a patient diagnosed with definite sporadic Creutzfeldt-Jakob disease in Cali, Colombia. Besides neurological examination, 14-3-3 and tau proteins were valuable tools supporting the diagnosis. We also present a brief perspective of the prion diseases reported in Colombia to date. Although the incidence of prion diseases is unknown in Colombia, our literature review revealed that one case of scrapie in 1981 and 29 human sporadic cases of Creutzfeldt-Jakob disease have been documented and published in our country.
Las enfermedades priónicas son alteraciones neurodegenerativas raras que ocurren en todo el mundo y afectan tanto a humanos como a animales. En el presente artículo, se reporta un caso con diagnóstico confirmado de enfermedad esporádica de Creutzfeldt-Jakob. Además del examen neuropatológico, las proteínas 14-3-3 y tau fueron herramientas valiosas que ayudaron en el diagnóstico. También, se presenta una breve reseña de las enfermedades priónicas reportadas en Colombia hasta la fecha. Aunque en el país se desconoce la incidencia de las enfermedades priónicas, nuestra búsqueda en la literatura científica reveló informes publicados sobre un caso de tembladera de las ovejas ( scrapie o encefalopatía espongiforme ovina) en 1981 y 29 casos esporádicos de Creutzfeldt-Jakob en el país.
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Síndrome de Creutzfeldt-Jakob , Relatos de Casos , PríonsRESUMO
La enfermedad de Creutzfeldt-Jakob es la encefalopatía espongiforme más común en el ser humano y prototipode las patologías causadas por priones. Se caracteriza histológicamente por astrogliosis y degeneración dela sustancia gris. Típicamente inicia con síntomas prodrómicos no específicos progresando a demencia conmioclonias y ataxia. Presentamos dos casos de mujeres en edad media con deterioro cognitivo progresivo,dificultades motrices, alteraciones del lenguaje y mioclonias que conducen a la muerte. En electroencefalogramasde ondas trifásicas lentas periódicas así como elevación de proteínas tau y 14-3-3 en LCR por apoyodel The National Prion Disease Pathology Surveillance Center - Cleveland, todos estos hallazgos definen lascondiciones para el diagnóstico clínico de enfermedad por priones. El diagnóstico diferencial en el contextode demencia rápidamente progresiva es amplio, incluyendo infecciones, intoxicaciones, trastornos metabólicos,autoinmunidad, vasculopatías y neoplasias que podrían explicar un posible subregistro en las estadísticasregionales. Existe una posible asociación de riesgo entre enfermedad por priones y médicos patólogos que,aunque discutida, podría limitar el estudio de los especímenes histológicos que son la clave del diagnósticodefinitivo. A pesar de la importancia en salud pública de estas condiciones, el actual modelo de salud limita elmanejo integral de los pacientes.
Creutzfeldt-Jakob is the most common spongiform encephalopathy in humans and the prototype of prions diseases. Astrogliosis and degeneration of the gray matter are the histological features. Typically starts with nonspecific prodromal symptoms that progressing to dementia with myoclonus and ataxia. We present two cases of women in middle age with progressive cognitive impairment, motor difficulties, language disorders and myoclonus that lead to death. EEG slow periodic triphasic waves and elevated protein tau and CSF14-3-3 support for The National Prion Disease Pathology Surveillance Center - Cleveland, all these findings define the conditions for the clinical diagnosis of prion disease. The differential diagnosis in the context of rapidly progressive dementia is broad including infections, poisoning, metabolic disorders, autoimmunity, vascular disease and neoplasms that could explain a possible underreporting in regional statistics. There is a possible risk association between disease and Medical Pathologists that although discussed could limit the study of histological specimens that are key to definitive diagnosis. Despite the public health importance of these conditions the current model of health limits the comprehensive management of patients.
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Humanos , Síndrome de Creutzfeldt-Jakob , Demência , Mioclonia , PríonsRESUMO
A emergência e a reemergência das doenças infecciosas oferece desafios à saúde pública, gerando demandas para os governos e para a comunidade científica; o que leva à priorização de ações em saúde, estabelecimento de políticas, aprimoramento da vigilância, além da manutenção de uma boa infraestrutura laboratorial e do cumprimento das medidas de biossegurança. As encefalopatias espongiformes transmissíveis são doenças neurodegenerativas, causadas por um agente infeccioso desprovido de material genético, composto por elementos proteicos, altamente estáveis e resistentes aos processos de descontaminação utilizados rotineiramente nos serviços de saúde e assim representam riscos à saúde pública. Este estudo objetivou identificar os óbitos registrados no Brasil causados por príons, para estabelecer medidas de biossegurança relativas aos riscos oferecidos aos profissionais de saúde, no sentido de prevenir doenças ocupacionais. Foram levantados os óbitos por doenças priônicas no Brasil, no período de janeiro de 2005 a dezembro de 2010, utilizando como fonte de obtenção de dados o Sistema de Informações sobre Mortalidade (SIM), do Ministério da Saúde. Foi identificado 1 caso de Kuru e 132 casos de doença de Creutzfeldt-Jakob, do total de 171.223 óbitos causados por doenças infecciosas e parasitárias. Os príons foram classificados quanto ao risco e a seguir foram identificadas as medidas de biossegurança.
The emergence and re-emergence of infectious diseases poses challenges to public health, demanding priority health actions to governments and the scientific community, while improving health surveillance, good laboratory infrastructure and compliance with biosafety measures. Transmissible spongiform encephalopaties are a group of neuro-degenerative infections caused by a "naked" infectious agent, with protein elements in its structure, but without genetic material. They are highly stable agents, resistant to most decontamination processes used routinely on health services, therefore representing a public health hazard. The present study aims at analyzing deaths registered in Brazil caused by prions, and establishing biosafety measures related to risks for the health professionals, in order to prevent occupational diseases. A survey of deaths from prion disease was made in Brazil from January 2005 to December 2010. The Mortality Information System (SIM) of the Brazilian Ministry of Health was used to obtain these data. We identified 1 case of Kuru and 132 cases of Creutzfeldt-Jakob disease of the total 171,223 deaths caused by infectious and parasitic diseases. Prions were classified according to risk and the biosafety measures were identified.
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La enfermedad de Creutzfeldt-Jakob (ECJ) es una enfermedad neurológica fatal producida por la isoforma patológica de la proteína priónica humana. Se reporta las características clínicas de seis casos de la forma esporádica de ECJ con diagnóstico definitivo por histopatología, y cinco casos con diagnóstico probable, en pacientes atendidos en el Instituto Nacional de Ciencias Neurológicas del Perú. La edad de inicio en los casos definitivos fue de 55,8 años y, en los probables, de 59,6 años, con predominio del sexo masculino. El tiempo de enfermedad fue de 8,8 meses. Se encontró un EEG típico en 50% de los casos definitivos y 80% de los probables. La proteína 14-3-3 en líquido cefalorraquídeo fue positiva en un caso probable y los hallazgos típicos en resonancia magnética se observaron en dos casos probables. Todos los casos cursaron con una evolución clínica típica de la enfermedad, y se considera el primer reporte de ECJ en el Perú.
Creutzfeldt-Jakob disease (CJD) is a fatal neurological disease caused by pathological isoform of the human prion protein. Clinical features of six cases of the sporadic form of CJD with definitive diagnosis by histopathology, and five cases with probable diagnosis were reported in patients treated at the Peruvian National Institute of Neurological Sciences. The average age of onset in definite cases was 55.8 years and in probable cases was 59.6, mostly males. The average disease duration was 8.8 months. A typical EEG was found in 50% of definite cases and in 80% of probable. The 14-3-3 protein in cerebrospinal fluid was positive in a probable case, and typical MRI findings were observed in two probable cases. All cases studied had a typical clinical course of the disease, and it is considered as the first report of CJD in Peru.
Assuntos
Humanos , Masculino , Feminino , Adulto , Pessoa de Meia-Idade , Idoso , Síndrome de Creutzfeldt-Jakob/diagnóstico , PeruRESUMO
Se presenta el caso de una paciente de 67 años que consultó por un cuadro progresivo de alteraciones neurológicas y deterioro cognitivo, las cuales, el curso de 6 meses, condujeron a su fallecimiento. Con base en los hallazgos de la resonancia magnética cerebral se hizo un diagnóstico presuntivo de enfermedad de Creutzfeldt-Jakob, confirmado por la presencia de proteína 14-3-3 elevada en el líquido cefalorraquídeo. Se explica brevemente la fisiopatología y la clínica de los pacientes con enfermedad de Creutzfeldt-Jakob y se revisan con detalle los cambios que se observan en las imágenes por resonancia magnética.
We present the case of a 67-year-old female patient with progressive neurologic symptoms and cognitive decline, which caused her demise in 6 months. Based on brain MRI findings, a presumptive diagnosis of Creutzfeldt-Jakob disease was established, confirmed by the presence of elevated 14-3-3 protein in spinal fluid. We briefly explain the physiopathology and clinical changes in patients with Creutzfeldt-Jakob disease, and review in detail the imaging findings on brain MRI.
Assuntos
Humanos , Síndrome de Creutzfeldt-Jakob , Príons , Imageamento por Ressonância Magnética , DiagnósticoRESUMO
Las enfermedades ocasionadas por priones son también conocidas como encefalitis espongiformes transmisibles o demencias de tipo infeccioso. En humanos las presentaciones clínicas más reconocidas son la enfermedad de Creutzfeldt-Jakob, el síndrome de Gerstmann-Strãussler-Scheinker y el insomnio fatal familiar. Son consideradas patologías poco comunes, pero los descubrimientos en biología molecular de los últimos años muestran que los mecanismos patológicos que llevan a su desarrollo pueden ser comunes a varias enfermedades neurodegenerativas, lo cual puede ampliar el espectro patológico, convirtiéndolas en alteraciones no tan infrecuentes en neurología. Esta revisión pretende dar herramientas al clínico para reconocer estas enfermedades discutiendo las presentaciones clínicas en seres humanos con sus variantes: esporádicas, infecciosas y familiares, comentando además el uso de laboratorios, criterios diagnósticos y aproximación terapéutica.
Illnesses caused by prions are also known as Transmissible Spongiform Encephalopathies or infectious dementias. The most recognized clinic presentations in humans are the Creutzfeldt-Jakob disease, the Gerstmann-Sträussler-Scheinker syndrome and the Fatal Familial Insomnia. They are considered uncommon pathologies, but discoveriesin molecular biology in recent years, reveal that pathological mechanisms that allow its development could becommon to several neurodegenerative diseases in which the pathologic spectrum can be amplified, becomingnot infrequent neurological diseases. This review gives tools to the clinical attendant in order to recognize thesedisorders, discussing the clinical presentations: sporadic, infectious and familiar, diagnostic work up, diagnosticcriteria and therapeutic approach.
Assuntos
Humanos , Biologia Molecular , Encefalite , Príons , NeurologiaRESUMO
Doenças por prions são encefalopatias neurodegenerativas de origem infecciosa, esporádica ou hereditária que atingem tanto animais quanto o homem. Desde a identificação da isoforma celular da proteína prion e sequenciamento do gene que a codifica, diversas mutações têm sido descritas no gene PRNP, sendo que mais de 55 destas segregam com enfermidades. A partir da década de 90 foram relatadas diversas funções fisiológicas de PrPC e a hipótese de que as doenças de príon estão relacionadas à perda de função da proteína normal, e não apenas à neurotoxicidade da isoforma infecciosa, PrPSc, vem ganhando força desde então. Com o intuito de verificar a influência destas mutações na função fisiológica de PrPC, a interação da proteína com seus ligantes e sua distribuição celular, foram geradas moléculas de PrPC com as mutações V180I, T183A e E200K fusionadas a proteína fluorescente verde (GFPPrPC). Optamos por estudar estes mutantes uma vez que se localizam dentro sítio de interação entre PrPC e um de seus ligantes, a laminina, descrito por nosso grupo. Esta interação é fundamental para vários processos biológicos de células do sistema nervoso, como adesão e diferenciação celular. Dada a influência do polimorfismo no códon 129 no fenótipo de algumas doenças priônicas de origem genética, a alteração de aminoácido neste códon foi associada a cada mutação, gerando haplótipos 129M/180I, 129V/180I, 129M/183A, 129V/183A, 129M/200K e 129V/200K, além do tipo selvagem associado a 129M ou 129V. Através de microscopia confocal, mostramos que a GFP-PrPV180I e GFP-PrPE200K, assim como a proteína do tipo selvagem, estão distribuídas corretamente na célula Zanith da S. P. Cook ii (membrana plasmática e região perinuclear). Ao contrário, a mutação T183A afeta a distribuição celular da molécula de PrPC, independente do aminoácido presente no códon 129, resultando no acúmulo da proteína na região perinuclear, sugerindo retenção no retículo endoplasmático. Ainda, através da indução de internação da molécula de PrPC presente na superfície celular por adição de íons cobre ao meio de cultura, mostramos que as construções GFP-PrPV180I e GFP-PrPE200K são funcionais.Ensaios de migração celular utilizando laminina como substrato foram realizados a fim de avaliar funcionalmente os mutantes de PrPC. Para estes ensaios, as contruções foram transfectadas em células N2a, das quais selecionamos clones para obtenção de populações com níveis de expressão homogêneos. Não foi possível detectar diferença significativa na taxa de migração das células mutantes em relação às que expressam a proteína tipo-selvagem. No entanto, apenas com esta observação não podemos afirmar que as mutações não afetam a função normal da proteína. Assim, outras abordagens devem ser realizadas a fim de melhor avaliar o impacto destas mutações na função normal de PrPC.
Prion diseases are neurodegenerative encephalopathies that can have infectious, sporadic or inherited origin and affect both animals and humans. Since the identification of the cellular isoform of the prion protein, followed by the sequencing of its corresponding gene, several mutations have been described on the PRNP gene and more than 55 of those being related to pathologies. Over the years, many physiological functions of PrPC have been described, and the PrPC loss-of-function hypothesis, for the prion diseases, is getting more strength. In order to verify the influence of those mutations on the physiological functions of PrPC, we generated PrPC molecules containing the mutations V180I, T183A and E200K fused to green fluorescence protein (GFP). Those mutants were chosen because they are localized in the lamminin-PrPC binding site, which was previously described by our group. This interaction is required for various biological processes on the nervous system, including cell adhesion and differentiation. Interestingly, the polymorphism at codon 129 affects the phenotype of some prion diseases when associated with mutations, and need to be considered. Thus, PrPC molecules presenting a combination of the 129 polimorphism and the described mutations were generated resulting in the haplotypes: 129M/180I, 129V/180I, 129M/183A, 129V/183A, 129M/200K and 129V/200K, as well as wild-type with methionine or valine on codon 129. Using confocal microscopy, we showed that GFP-PrPV180I and GFP-PrPE200K, as well as the wild-type molecules, were correctly localized on the plasma membrane and perinuclear region. On the other hand, the mutation T183A altered the subcellular distribution of PrPC, independently of the amino acid present at Zanith da S. P. Cook iv codon 129, resulting in the protein accumulation at the perinuclear region, wich suggests PrPC retention in the endoplasmatic reticulum. Also, we showed that the GFP-PrPV180I and GFP-PrPE200K molecules are functional since they are internalized after the addition of copper ions to the culture medium. To evaluate the effect of the mutations on the PrPC function, we next performed migration assays using lamminin as the substrate. For that, the constructs were stably transfected in N2a cells, from which clones were selected to obtain homogeneous expression levels. No significative differences were detected on the migration rates of the mutants when comparing with their normal counterparts. However, with these preliminary observations is not possible to conclude that PrPC mutations do not affect the normal function of the protein. Further work, with different approaches, should be addressed for a better evaluation of the impact of these mutations on the normal functions of t he cellular prion protein.
Assuntos
Movimento Celular , Laminina , Proteínas PrPCRESUMO
A proteína prion celular, PrPC, tem sido relacionada com doenças que atingem animais e o homem conhecidas como encefalopatias espongiformes transmissíveis (TSEs), ou doenças por prions. PrPC é uma glicoproteína de superfície que está ligada à membrana plasmática por uma âncora de glicosilfosfatidilinositol (GPI). Ela é abundantemente expressa em neurônios e células da glia e é extremamente conservada entre as espécies. Mutações no gene de PrPC humano, PRNP, causam doenças por prions hereditárias como insônia familial fatal (FFI 129M/178N), e a doença de Creutzfeldt-Jakob (CJD 129V/178N) que estão associadas a mutação no codon 178 e ao polimorfismo no codon 129 de PRNP. Da mesma forma, o polimorfismo no codon 171 de PRNP foi associado a uma forma atípica de esquizofrenia familial. No intuito de esclarecer o papel biológico de PrPC, um crescente número de trabalhos vem sendo publicado associando-o a fenômenos celulares relevantes, como proteção contra estresse oxidativo, adesão neuronal e diferenciação e sobrevivência celulares. Nesse sentido, nosso grupo tem estudado e caracterizado a interação de PrPC com importantes proteínas de matriz extracelular, laminina (Ln) e vitronectina (Vn). Essa interação está associada ao crescimento, formação e manutenção de neuritos. Interessantemente, a mutação D178N e o polimorfismo N171S estão dentro ou próximos ao domínio de ligação de PrPC à Ln (173 a 182, em PrPC murino). Assim, para investigar se essas modificações poderiam levar a perda de função relacionadas à interação de PrPC-Ln, estudamos o polimorfismo N171S e a mutação D178N, associadas ou não ao polimorfismo Val. ou Met. no codon 129. Construções do gene humano de PrPC 129M (tipo selvagem), PrPC 129V, PrPC 129M/171S, PrPC 129V/171S e PrPC 129M/178N fusionadas a proteína fluorescente verde (GFP) foram transfectadas nas linhagens celulares de SN56, HEK e N2a, para análise de localização subcelular, padrão de glicosilação e migração celular mediada por laminina, respectivamente. Observamos um padrão de localização e glicosilação semelhantes ao da proteína tipo selvagem em todas as quimeras, mas houve uma importante inibição no estímulo de migração celular estimulado por Ln nas células transfectadas com as construções GFP- PrPC 129M/171S, GFP- PrPC 129V/171S, GFP- PrPC 129M/178N. Esses resultados sugerem que modificações em PrPC em seu sítio de interação com a Ln podem afetar a migração celular, um fenômeno importante em várias atividade biológicas.
The cellular prion protein, PrPC, has been implicated in a variety of human and animal illness referred to as transmissible spongiform encephalopathies, TSEs, or prion diseases. PrPC is a glycosylated glycosylphosphatidyl inositol (GPI) anchored protein mostly expressed on the surface of neurons and glial cells. Mutations in the PrPC gene, PRNP, have been reported to be associated with inherited forms of human prion diseases. A mutation at codon 178 associated to Met (methionine) at codon 129 is linked to fatal familial insomnia (FFI 129M/178N) whereas one type of hereditary CreutzfeldtJakob disease present the same mutation at codon 178 but Val (valine) at codon 129 (CJD 129V/178N). Additionally, a PRNP polymorphism at codon 171 has been associated with familial form of atypical schizophrenia. Recent studies have provided evidence that PrPC might have physiological functions such as protection against oxidative stress, neuronal adhesion, cellular differentiation and survival. In this direction, our group have studied and characterized the interaction of PrPC with vitronectin (Vn) and laminin (Ln), two important proteins of extracellular matrix. These interactions mediate neurite formation, outgrowth, and maintenance. Thus, alterations on the PrPC functions may result in its loss-of-function. Interestingly, the mutation D178N and polymorphism N171S are inside or closer to the Ln binding site at the PrPC molecule (173 to 182, in murine PrPC). In attempt to investigate if PrPC mutations or polymorphisms associated to inherited diseases (prion or psychiatric) interfere with its binding to Ln and consequently with biological function mediated by this ligand, we constructed PrPC mutants (PrPC 171S, PrPC 129V/171S and PrPC 178N) fused to green fluorescence protein. To address that question, we analyzed the glicosylation pattern and sub-cellular localization of PrPC in HEK and SN56 cell lines, respectively, and cell migration mediated by Ln in N2a cells. Transient transfection in SN56 and HEK cells demonstrated that all PrPC mutant molecules were located at the cell surface and at peri-nuclear region similarly to the wild-type counterpart. We also showed the same glicosylation patterns of the PrPC mutated molecules when compared to the wild-type protein. Clones of permanent transfected N2a cells expressing the human wild-type or the mutant/polymorphic (PrPC 171S, PrPC 129V/171S and PrPC 178N) molecules were generated to evaluate cell migration mediated by laminin. Cells expressing PrPC mutants showed an impaired migration to laminin when compared to those expressing wild-type molecules. Taken together, these data suggest that PrPC mutation/polymorphism at the Ln binding domain cause an impairment of important cellular functions.