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1.
Nat Commun ; 15(1): 6971, 2024 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-39138218

RESUMO

Ligation of the B cell antigen receptor (BCR) initiates humoral immunity. However, BCR signaling without appropriate co-stimulation commits B cells to death rather than to differentiation into immune effector cells. How BCR activation depletes potentially autoreactive B cells while simultaneously primes for receiving rescue and differentiation signals from cognate T lymphocytes remains unknown. Here, we use a mass spectrometry-based proteomic approach to identify cytosolic/nuclear shuttling elements and uncover transcription factor EB (TFEB) as a central BCR-controlled rheostat that drives activation-induced apoptosis, and concurrently promotes the reception of co-stimulatory rescue signals by supporting B cell migration and antigen presentation. CD40 co-stimulation prevents TFEB-driven cell death, while enhancing and prolonging TFEB's nuclear residency, which hallmarks antigenic experience also of memory B cells. In mice, TFEB shapes the transcriptional landscape of germinal center B cells. Within the germinal center, TFEB facilitates the dark zone entry of light-zone-residing centrocytes through regulation of chemokine receptors and, by balancing the expression of Bcl-2/BH3-only family members, integrates antigen-induced apoptosis with T cell-provided CD40 survival signals. Thus, TFEB reprograms antigen-primed germinal center B cells for cell fate decisions.


Assuntos
Apoptose , Linfócitos B , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos , Antígenos CD40 , Centro Germinativo , Receptores de Antígenos de Linfócitos B , Animais , Centro Germinativo/imunologia , Centro Germinativo/citologia , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/metabolismo , Linfócitos B/imunologia , Linfócitos B/metabolismo , Camundongos , Antígenos CD40/metabolismo , Antígenos CD40/imunologia , Receptores de Antígenos de Linfócitos B/metabolismo , Receptores de Antígenos de Linfócitos B/imunologia , Camundongos Endogâmicos C57BL , Ativação Linfocitária/imunologia , Diferenciação Celular/imunologia , Transdução de Sinais , Apresentação de Antígeno/imunologia
2.
Nat Neurosci ; 25(7): 887-899, 2022 07.
Artigo em Inglês | MEDLINE | ID: mdl-35773544

RESUMO

The meninges, comprising the leptomeninges (pia and arachnoid layers) and the pachymeninx (dura layer), participate in central nervous system (CNS) autoimmunity, but their relative contributions remain unclear. Here we report on findings in animal models of CNS autoimmunity and in patients with multiple sclerosis, where, in acute and chronic disease, the leptomeninges were highly inflamed and showed structural changes, while the dura mater was only marginally affected. Although dural vessels were leakier than leptomeningeal vessels, effector T cells adhered more weakly to the dural endothelium. Furthermore, local antigen-presenting cells presented myelin and neuronal autoantigens less efficiently, and the activation of autoreactive T cells was lower in dural than leptomeningeal layers, preventing local inflammatory processes. Direct antigen application was required to evoke a local inflammatory response in the dura. Together, our data demonstrate an uneven involvement of the meningeal layers in CNS autoimmunity, in which effector T cell trafficking and activation are functionally confined to the leptomeninges, while the dura remains largely excluded from CNS autoimmune processes.


Assuntos
Autoimunidade , Meninges , Esclerose Múltipla , Animais , Aracnoide-Máter , Sistema Nervoso Central , Dura-Máter , Humanos , Meninges/fisiologia
3.
J Clin Invest ; 131(5)2021 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-33645550

RESUMO

Approximately 80% of neuromyelitis optica spectrum disorder (NMOSD) patients harbor serum anti-aquaporin-4 autoantibodies targeting astrocytes in the CNS. Crucial for NMOSD lesion initiation is disruption of the blood-brain barrier (BBB), which allows the entrance of Abs and serum complement into the CNS and which is a target for new NMOSD therapies. Astrocytes have important functions in BBB maintenance; however, the influence of their loss and the role of immune cell infiltration on BBB permeability in NMOSD have not yet been investigated. Using an experimental model of targeted NMOSD lesions in rats, we demonstrate that astrocyte destruction coincides with a transient disruption of the BBB and a selective loss of occludin from tight junctions. It is noteworthy that BBB integrity is reestablished before astrocytes repopulate. Rather than persistent astrocyte loss, polymorphonuclear leukocytes (PMNs) are the main mediators of BBB disruption, and their depletion preserves BBB integrity and prevents astrocyte loss. Inhibition of PMN chemoattraction, activation, and proteolytic function reduces lesion size. In summary, our data support a crucial role for PMNs in BBB disruption and NMOSD lesion development, rendering their recruitment and activation promising therapeutic targets.


Assuntos
Astrócitos/imunologia , Barreira Hematoencefálica/imunologia , Leucócitos Mononucleares/imunologia , Neuromielite Óptica/imunologia , Animais , Astrócitos/patologia , Barreira Hematoencefálica/patologia , Modelos Animais de Doenças , Feminino , Humanos , Leucócitos Mononucleares/patologia , Neuromielite Óptica/patologia , Ratos , Ratos Endogâmicos Lew
4.
Environ Pollut ; 274: 115900, 2021 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-33246767

RESUMO

During March 2020, most European countries implemented lockdowns to restrict the transmission of SARS-CoV-2, the virus which causes COVID-19 through their populations. These restrictions had positive impacts for air quality due to a dramatic reduction of economic activity and atmospheric emissions. In this work, a machine learning approach was designed and implemented to analyze local air quality improvements during the COVID-19 lockdown in Graz, Austria. The machine learning approach was used as a robust alternative to simple, historical measurement comparisons for various individual pollutants. Concentrations of NO2 (nitrogen dioxide), PM10 (particulate matter), O3 (ozone) and Ox (total oxidant) were selected from five measurement sites in Graz and were set as target variables for random forest regression models to predict their expected values during the city's lockdown period. The true vs. expected difference is presented here as an indicator of true pollution during the lockdown. The machine learning models showed a high level of generalization for predicting the concentrations. Therefore, the approach was suitable for analyzing reductions in pollution concentrations. The analysis indicated that the city's average concentration reductions for the lockdown period were: -36.9 to -41.6%, and -6.6 to -14.2% for NO2 and PM10, respectively. However, an increase of 11.6-33.8% for O3 was estimated. The reduction in pollutant concentration, especially NO2 can be explained by significant drops in traffic-flows during the lockdown period (-51.6 to -43.9%). The results presented give a real-world example of what pollutant concentration reductions can be achieved by reducing traffic-flows and other economic activities.


Assuntos
Poluentes Atmosféricos , Poluição do Ar , COVID-19 , Poluentes Atmosféricos/análise , Poluição do Ar/análise , Áustria , Controle de Doenças Transmissíveis , Monitoramento Ambiental , Europa (Continente) , Humanos , Aprendizado de Máquina , Material Particulado/análise , SARS-CoV-2
5.
Nature ; 567(7749): E15, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30867589

RESUMO

In this Article, owing to an error during the production process, the y-axis label of Fig. 2c should read "Number of Tß-syn cells" rather than "Number of T1ß-syn cells" and the left and right panels of Fig. 4 should be labelled 'a' and 'b', respectively. These errors have been corrected online.

6.
Nature ; 566(7745): 503-508, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30787438

RESUMO

The grey matter is a central target of pathological processes in neurodegenerative disorders such as Parkinson's and Alzheimer's diseases. The grey matter is often also affected in multiple sclerosis, an autoimmune disease of the central nervous system. The mechanisms that underlie grey matter inflammation and degeneration in multiple sclerosis are not well understood. Here we show that, in Lewis rats, T cells directed against the neuronal protein ß-synuclein specifically invade the grey matter and that this is accompanied by the presentation of multifaceted clinical disease. The expression pattern of ß-synuclein induces the local activation of these T cells and, therefore, determined inflammatory priming of the tissue and targeted recruitment of immune cells. The resulting inflammation led to significant changes in the grey matter, which ranged from gliosis and neuronal destruction to brain atrophy. In humans, ß-synuclein-specific T cells were enriched in patients with chronic-progressive multiple sclerosis. These findings reveal a previously unrecognized role of ß-synuclein in provoking T-cell-mediated pathology of the central nervous system.


Assuntos
Substância Cinzenta/imunologia , Substância Cinzenta/patologia , Esclerose Múltipla Crônica Progressiva/imunologia , Esclerose Múltipla Crônica Progressiva/patologia , Linfócitos T/imunologia , beta-Sinucleína/imunologia , Animais , Encéfalo/patologia , Movimento Celular/imunologia , Feminino , Regulação da Expressão Gênica , Gliose/patologia , Humanos , Inflamação/imunologia , Inflamação/patologia , Ativação Linfocitária , Contagem de Linfócitos , Masculino , Esclerose Múltipla Crônica Progressiva/sangue , Doenças Neurodegenerativas/imunologia , Doenças Neurodegenerativas/patologia , Neurônios/patologia , Ratos , Ratos Endogâmicos Lew , Linfócitos T/metabolismo , Linfócitos T/patologia , beta-Sinucleína/análise , beta-Sinucleína/genética , beta-Sinucleína/metabolismo
7.
Hum Vaccin Immunother ; 14(9): 2263-2273, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29771574

RESUMO

Pertussis or whooping cough, a highly infectious respiratory infection, causes significant morbidity and mortality in infants. In adolescents and adults, pertussis presents with atypical symptoms often resulting in under-diagnosis and under-reporting, increasing the risk of transmission to more vulnerable groups. Maternal vaccination against pertussis protects mothers and newborns. This evaluation assessed the cost-effectiveness of adding maternal dTpa (reduced antigen diphtheria, Tetanus, acellular pertussis) vaccination to the 2016 nationally-funded pertussis program (DTPa [Diphtheria, Tetanus, acellular Pertussis] at 2, 4, 6, 18 months, 4 years and dTpa at 12-13 years) in Australia. A static cross-sectional population model was developed using a one-year period at steady-state. The model considered the total Australian population, stratified by age. Vaccine effectiveness against pertussis infection was assumed to be 92% in mothers and 91% in newborns, based on observational and case-control studies. The model included conservative assumptions around unreported cases. With 70% coverage, adding maternal vaccination to the existing pertussis program would prevent 8,847 pertussis cases, 422 outpatient cases, 146 hospitalizations and 0.54 deaths per year at the population level. With a 5% discount rate, 138.5 quality-adjusted life-years (QALYs) would be gained at an extra cost of AUS$ 4.44 million and an incremental cost-effectiveness ratio of AUS$ 32,065 per QALY gained. Sensitivity and scenario analyses demonstrated that outcomes were most sensitive to assumptions around vaccine effectiveness, duration of protection in mothers, and disutility of unreported cases. In conclusion, dTpa vaccination in the third trimester of pregnancy is likely to be cost-effective from a healthcare payer perspective in Australia.


Assuntos
Análise Custo-Benefício , Doenças do Recém-Nascido/prevenção & controle , Vacina contra Coqueluche/administração & dosagem , Complicações Infecciosas na Gravidez/prevenção & controle , Cuidado Pré-Natal/métodos , Coqueluche/prevenção & controle , Adolescente , Adulto , Austrália , Criança , Pré-Escolar , Estudos Transversais , Transmissão de Doença Infecciosa/prevenção & controle , Feminino , Humanos , Lactente , Recém-Nascido , Doenças do Recém-Nascido/economia , Masculino , Vacina contra Coqueluche/economia , Gravidez , Complicações Infecciosas na Gravidez/economia , Cuidado Pré-Natal/economia , Coqueluche/economia , Adulto Jovem
8.
Proc Natl Acad Sci U S A ; 113(12): 3323-8, 2016 Mar 22.
Artigo em Inglês | MEDLINE | ID: mdl-26957602

RESUMO

Multiple sclerosis (MS) is caused by T cells that are reactive for brain antigens. In experimental autoimmune encephalomyelitis, the animal model for MS, myelin-reactive T cells initiate the autoimmune process when entering the nervous tissue and become reactivated upon local encounter of their cognate CNS antigen. Thereby, the strength of the T-cellular reactivation process within the CNS tissue is crucial for the manifestation and the severity of the clinical disease. Recently, B cells were found to participate in the pathogenesis of CNS autoimmunity, with several diverse underlying mechanisms being under discussion. We here report that B cells play an important role in promoting the initiation process of CNS autoimmunity. Myelin-specific antibodies produced by autoreactive B cells after activation in the periphery diffused into the CNS together with the first invading pathogenic T cells. The antibodies accumulated in resident antigen-presenting phagocytes and significantly enhanced the activation of the incoming effector T cells. The ensuing strong blood-brain barrier disruption and immune cell recruitment resulted in rapid manifestation of clinical disease. Therefore, myelin oligodendrocyte glycoprotein (MOG)-specific autoantibodies can initiate disease bouts by cooperating with the autoreactive T cells in helping them to recognize their autoantigen and become efficiently reactivated within the immune-deprived nervous tissue.


Assuntos
Autoanticorpos/imunologia , Doenças Autoimunes/imunologia , Doenças do Sistema Nervoso Central/imunologia , Ativação Linfocitária/imunologia , Linfócitos T/imunologia , Diferenciação Celular , Humanos , Linfócitos T/patologia
9.
Nature ; 530(7590): 349-53, 2016 Feb 18.
Artigo em Inglês | MEDLINE | ID: mdl-26863192

RESUMO

In multiple sclerosis, brain-reactive T cells invade the central nervous system (CNS) and induce a self-destructive inflammatory process. T-cell infiltrates are not only found within the parenchyma and the meninges, but also in the cerebrospinal fluid (CSF) that bathes the entire CNS tissue. How the T cells reach the CSF, their functionality, and whether they traffic between the CSF and other CNS compartments remains hypothetical. Here we show that effector T cells enter the CSF from the leptomeninges during Lewis rat experimental autoimmune encephalomyelitis (EAE), a model of multiple sclerosis. While moving through the three-dimensional leptomeningeal network of collagen fibres in a random Brownian walk, T cells were flushed from the surface by the flow of the CSF. The detached cells displayed significantly lower activation levels compared to T cells from the leptomeninges and CNS parenchyma. However, they did not represent a specialized non-pathogenic cellular sub-fraction, as their gene expression profile strongly resembled that of tissue-derived T cells and they fully retained their encephalitogenic potential. T-cell detachment from the leptomeninges was counteracted by integrins VLA-4 and LFA-1 binding to their respective ligands produced by resident macrophages. Chemokine signalling via CCR5/CXCR3 and antigenic stimulation of T cells in contact with the leptomeningeal macrophages enforced their adhesiveness. T cells floating in the CSF were able to reattach to the leptomeninges through steps reminiscent of vascular adhesion in CNS blood vessels, and invade the parenchyma. The molecular/cellular conditions for T-cell reattachment were the same as the requirements for detachment from the leptomeningeal milieu. Our data indicate that the leptomeninges represent a checkpoint at which activated T cells are licensed to enter the CNS parenchyma and non-activated T cells are preferentially released into the CSF, from where they can reach areas of antigen availability and tissue damage.


Assuntos
Movimento Celular , Líquido Cefalorraquidiano/citologia , Encefalomielite Autoimune Experimental/patologia , Meninges/patologia , Esclerose Múltipla/patologia , Linfócitos T/patologia , Transferência Adotiva , Animais , Adesão Celular , Líquido Cefalorraquidiano/imunologia , Quimiocinas/metabolismo , Plexo Corióideo , Colágeno/metabolismo , Modelos Animais de Doenças , Encefalomielite Autoimune Experimental/imunologia , Feminino , Integrina alfa4beta1/metabolismo , Ativação Linfocitária , Antígeno-1 Associado à Função Linfocitária/metabolismo , Macrófagos/imunologia , Macrófagos/metabolismo , Masculino , Meninges/imunologia , Esclerose Múltipla/imunologia , Ratos , Ratos Endogâmicos Lew , Receptores CCR5/metabolismo , Receptores CXCR3/metabolismo , Linfócitos T/imunologia
10.
Nat Med ; 19(6): 784-90, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23624600

RESUMO

Multiple sclerosis is an autoimmune disease of the central nervous system (CNS) that is initiated when self-reactive T cells enter the brain and become locally activated after encountering their specific nervous antigens. When and where the disease-relevant antigen encounters occur is unclear. Here we combined fluorescently labeled nuclear factor of activated T cells (NFAT) with histone protein H2B to create a broadly applicable molecular sensor for intravital imaging of T cell activation. In experimental autoimmune encephalomyelitis, an animal model for multiple sclerosis, we report that effector T cells entering the CNS become activated after short contacts with leptomeningeal phagocytes. During established disease, the activation process is extended to the depth of the CNS parenchyma, where the cells form contacts with microglia and recruited phagocytes. We show that it is the activation processes during the preclinical phase rather than during established disease that are essential for the intensity and duration of the disease bout.


Assuntos
Autoimunidade , Encéfalo/imunologia , Histonas/fisiologia , Fatores de Transcrição NFATC/fisiologia , Linfócitos T/imunologia , Animais , Técnicas Biossensoriais , Encefalomielite Autoimune Experimental/imunologia , Fluorescência , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Transporte Proteico , Ratos , Ratos Endogâmicos Lew , Transdução de Sinais
11.
Drug Metab Dispos ; 34(9): 1582-99, 2006 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-16763017

RESUMO

Biliary excretion of bile salts and other bile constituents from hepatocytes is mediated by the apical (canalicular) transporters P-glycoprotein 3 (MDR3, ABCB4) and the bile salt export pump (ABCB11). Mutations in ABCB4 and ABCB11 contribute to cholestatic diseases [e.g., progressive familial intrahepatic cholestasis 2 (PFIC2), PFIC3, and intrahepatic cholestasis of pregnancy], and our objective was to establish genetic variability and haplotype structures of ABCB4 and ABCB11 in healthy populations of different ethnic backgrounds. All coding exons, 5 of 6 noncoding exons, 50 to 300 base pairs of the flanking intronic regions, and 2.5 to 2.8 kilobase pairs of the promoter regions of ABCB4 and ABCB11 were sequenced in 159 and 196 DNA samples of Caucasian, African-American, Japanese, and Korean origin. In total, 76 and 86 polymorphisms were identified in ABCB4 and ABCB11, respectively; among them, 14 and 28 exonic polymorphisms, and 8 and 10 protein-altering variants, of which 4 were predicted to have functional consequences. Both genes showed substantial ethnic differences with respect to allele number, frequency of common and population-specific sites, and patterns of linkage disequilibrium. Population genetic analysis suggested some selective pressure against changes in the protein, supporting the important endogenous role of these transporters. Haplotype variability was greater in ABCB11 than in ABCB4. An ABCB11 promoter haplotype was associated with significant decrease of activity compared with wild type. Our results contribute to a better understanding of the molecular basis and of ethnic differences in drug response, and provide a valuable tool for future research on the heredity of cholestatic liver injury.


Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Transportadores de Cassetes de Ligação de ATP/genética , Povo Asiático/genética , Negro ou Afro-Americano/genética , Haplótipos , Polimorfismo de Nucleotídeo Único , População Branca/genética , Região 5'-Flanqueadora/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/química , Subfamília B de Transportador de Cassetes de Ligação de ATP/metabolismo , Membro 11 da Subfamília B de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/química , Transportadores de Cassetes de Ligação de ATP/metabolismo , Sequência de Aminoácidos , Ácidos e Sais Biliares/metabolismo , Linhagem Celular Tumoral , Colestase/etnologia , Colestase/genética , Colestase/metabolismo , Frequência do Gene , Genes Reporter , Testes Genéticos , Humanos , Desequilíbrio de Ligação , Fígado/metabolismo , Luciferases , Modelos Genéticos , Dados de Sequência Molecular , Regiões Promotoras Genéticas/genética , Análise de Sequência de DNA/métodos , Transfecção
12.
Pharmacogenet Genomics ; 16(1): 59-71, 2006 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-16344723

RESUMO

Cytochrome P450 3A enzymes (CYP3A) play a major role in the metabolism of steroid hormones, drugs and other chemicals, including many carcinogens. The individually variable CYP3A expression, which remains mostly unexplained, has been suggested to affect clinical phenotypes. We investigated the CYP3A locus in five ethnic groups. The degree of linkage disequilibrium (LD) differed among ethnic groups, but the most common alleles of the conserved LD regions were remarkably similar. Non-African haplotypes are few; for example, only four haplotypes account for 80% of common European Caucasian alleles. Large LD blocks of high frequencies were suggestive of selection. Accordingly, European Caucasian and Asian cohorts each contained a block of single nucleotide polymorphism (SNPs) with very high P excess values. The overlap between these blocks in these two groups contained only two of the investigated 26 SNPs and one of them was the CYP3A4*1B allele. The region centromeric of CYP3A4*1B exhibited high haplotype homozygosity in European Caucasians as opposed to African-Americans. CYP3A4*1B showed a moderate effect on CYP3A4 mRNA and protein expression, as well as on CYP3A activity assessed as Vmax of testosterone 6beta-hydroxylation in a liver bank. Our data are consistent with a functional relevance of CYP3A4*1B and with selection against this allele in non-African populations. The elimination of CYP3A4*1B involved different parts of the CYP3A locus in European Caucasians and Asians. Because CYP3A4 is involved in the vitamin D metabolism, rickets may have been the underlying selecting factor.


Assuntos
Povo Asiático/genética , Sistema Enzimático do Citocromo P-450/genética , Sistema Enzimático do Citocromo P-450/metabolismo , Seleção Genética , Alelos , População Negra/genética , Citocromo P-450 CYP3A , Variação Genética/genética , Haplótipos/genética , Humanos , Desequilíbrio de Ligação/genética , Fígado/enzimologia , População Branca/genética
13.
Pharmacogenet Genomics ; 15(9): 609-24, 2005 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-16041240

RESUMO

The human cytochrome P450 2A6 (CYP2A6) enzyme metabolizes several xenobiotic compounds of clinical or toxicological importance. We aimed to identify genetic variants and major CYP2A6 haplotypes associated with CYP2A6 phenotypic variation. CYP2A6 mRNA level, protein level, activity and haplotypes were determined in Caucasian liver samples via real-time polymerase chain reaction, Western blot, coumarin 7-hydroxylation, DNA sequencing and genotyping, respectively. Phenotypes were then analyzed for associations with haplotypes. CYP2A6 transcript, protein and activity levels were correlated among each other. In 45 African-American, 156 Caucasian, 47 Chinese, 50 Japanese and 47 Korean DNA samples, we detected 95 different polymorphisms in the CYP2A6 gene, 49 of which had not been described previously. Caucasian variants formed 33 haplotypes which built four clades. Allele *9B and the CYP2A7/2A6 partial deletion allele CYP2A6*12B were both associated with decreased expression. The latter haplotype extends at least over 147 kb up into the CYP2B6 gene. A haplotype almost identical to allele *1A was associated with decreased expression and activity of CYP2A6 compared to all other haplotypes. In summary A CYP2A6*1A-like allele, *9B and *12B are major genetic determinants of CYP2A6 phenotype variation in Caucasians.


Assuntos
Hidrocarboneto de Aril Hidroxilases/genética , Hidrocarboneto de Aril Hidroxilases/metabolismo , Oxigenases de Função Mista/genética , Oxigenases de Função Mista/metabolismo , Alelos , Apolipoproteínas/química , Sequência de Bases , Western Blotting , Clonagem Molecular , Cumarínicos/farmacologia , Citocromo P-450 CYP2A6 , DNA/metabolismo , Primers do DNA/química , DNA Complementar/metabolismo , Éxons , Deleção de Genes , Variação Genética , Genótipo , Haplótipos , Humanos , Fígado/metabolismo , Modelos Genéticos , Dados de Sequência Molecular , Fenótipo , Polimorfismo Genético , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Sequência de DNA , Homologia de Sequência do Ácido Nucleico , População Branca , Xenobióticos
14.
J Hepatol ; 43(3): 536-43, 2005 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-16039748

RESUMO

BACKGROUND/AIMS: Inherited dysfunction of the bile salt export pump BSEP (ABCB11) causes a progressive and a benign form of familial intrahepatic cholestasis, denominated as PFIC2 and BRIC2, respectively. We functionally characterized novel ABCB11 mutations encountered in two patients with a PFIC2 and a BRIC2 phenotype, respectively. METHODS: BSEP expression was determined in liver biopsies by immunohistochemistry. ABCB11 mutations were functionally characterized by taurocholate transport in SF9 cells transfected with human ABCB11. RESULTS: The PFIC2 patient was compound heterozygous for a splicing mutation in intron 4 ((+3)A > C) combined with an early stop codon at position 930 (R930X), while the BRIC2 patient was compound heterozygous for two nonsynonymous mutations in exon 9 (E297G) and exon 12 (R432T), respectively. Hepatic BSEP expression was absent in PFIC2 and preserved in BRIC2. In BRIC2, taurocholate transport was decreased to 13% and 20% of reference levels for R432T and E297G, respectively. CONCLUSIONS: The intron 4 (+3)A > C, R930X and R432T represent previously undescribed mutations of the ABCB11 gene that confer a PFIC2 and a BRIC2 phenotype, respectively. By combining functional in-vitro characterization with immunohistochemical detection of variant BSEP we provide direct evidence for the role of ABCB11 mutations in the pathogenesis of different forms of intrahepatic cholestasis.


Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Colestase Intra-Hepática/genética , Regulação da Expressão Gênica , Mutação , Membro 11 da Subfamília B de Transportadores de Cassetes de Ligação de ATP , Transportadores de Cassetes de Ligação de ATP/química , Transportadores de Cassetes de Ligação de ATP/metabolismo , Sequência de Aminoácidos , Sequência de Bases , Ácidos e Sais Biliares/metabolismo , Transporte Biológico , Primers do DNA , Feminino , Humanos , Masculino , Modelos Moleculares , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Linhagem , Conformação Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo
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