RESUMO
OBJECTIVES: Ameloblastoma is an odontogenic neoplasm of the mandible and maxilla with various histological types and subtypes. It has been reported that some ameloblastomas could arise from dentigerous cyst walls; thus, the development of ameloblastoma from dentigerous cysts may be due to differential protein expression. Our aim was to identify a membrane protein that is differentially expressed in ameloblastomas with respect to dentigerous cysts. METHODS: We analyzed the SDS-PAGE profiles of membrane proteins from ameloblastomas and dentigerous cysts. The protein in a band present in the ameloblastoma sample, but apparently absent in the dentigerous cyst sample was identified via mass spectrometry as the chaperonin Hsp60. We used western blotting and immunohistochemistry to analyze its overexpression and localization in ameloblastoma. RESULTS: We found a differential band of 95 kDa in the membrane proteins of ameloblastoma. In this band, the chaperonin Hsp60 was identified, and its overexpression was corroborated using western blotting and immunohistochemistry. Hsp60 was localized in the plasma membrane of all ameloblastoma samples studied; in addition, it was found in the cell nucleus of the plexiform subtype of conventional ameloblastoma. CONCLUSIONS: Our results suggest that Hsp60 may be involved in ameloblastoma development, and could therefore be a potential therapeutic target for ameloblastoma treatment.
Assuntos
Ameloblastoma , Chaperonina 60/genética , Cisto Dentígero , Proteínas Mitocondriais/genética , Tumores Odontogênicos , Ameloblastoma/genética , Chaperoninas , Humanos , Imuno-HistoquímicaRESUMO
The Bardet-Biedl syndrome is an autosomal recessive hereditary disorder with vast locus heterogeneity that belongs to the so-called ciliopathies, whose proteins are localized in the primary cilia and present functional deficiency. The multisystemic features of the disease include ocular, renal, cognitive, skeletal, as well as gonadal involvement and obesity, among others, with high inter- and intrafamilial variability. We describe the clinical case of an adolescent male patient with Bardet-Biedl syndrome, including the approach, the results from a 22-gene sequencing panel, and the analysis of updated scientific literature. We collected the clinical data of the patient and, after obtaining the informed consent, we conducted a multigenic sequencing panel oriented to known implicated genes. The patient was born to consanguineous parents and was the first affected member of the family. He presented with postaxial polydactyly, obesity, micropenis, retinitis pigmentosa, and learning disability. The multigenic panel allowed the identification of the homozygous pathogenic variant c.39_46del in the BBS10 gene and in other BBS genes variants associated with obesity. As the Bardet-Biedl syndrome is a rare disease, it is challenging to interpret its pleiotropism and gene/allelic heterogeneity. Its confirmation by molecular tests allows an adequate approach, follow-up, and genetic counseling of the patient and the family.
Assuntos
Síndrome de Bardet-Biedl/genética , Chaperoninas do Grupo II/genética , Adolescente , Chaperoninas , Consanguinidade , Análise Mutacional de DNA , Genes Recessivos , Homozigoto , Humanos , Masculino , Linhagem , Deleção de SequênciaRESUMO
Resumen El síndrome de Bardet-Biedl es una enfermedad hereditaria, autosómica recesiva, con gran heterogeneidad de locus, que pertenece a las denominadas ciliopatías, denominadas así por la deficiencia funcional presente y porque las proteínas afectadas se localizan en el cilio primario. El síndrome afecta múltiples sistemas, con compromiso visual, renal, cognitivo, esquelético y gonadal, y obesidad. Este síndrome presenta una gran variabilidad intrafamiliar e interfamiliar. Se presenta el caso clínico de un paciente adolescente con diagnóstico de síndrome de Bardet-Biedl, así como su manejo, los resultados de la secuenciación de 22 genes y el análisis actualizado de la literatura médica. Se recopiló la información clínica y, previo consentimiento informado, se hizo la prueba de panel de secuenciación multigénica de los genes implicados. El paciente es hijo de la unión de personas consanguíneas. Fue el primer afectado en la familia y presentaba polidactilia posaxial, obesidad, micropene, retinitis pigmentaria y dificultades de aprendizaje. En el panel multigénico, se identificó la variante patogénica homocigótica c.39_46del en el gen BBS10 y otras variantes de genes BBS asociadas con la obesidad. Dado que el síndrome de Bardet-Biedl es una enfermedad huérfana rara, interpretar el pleiotropismo y la heterogeneidad de locus y de alelos, constituye un reto. La confirmación molecular permite el manejo adecuado de los pacientes, así como el seguimiento y el asesoramiento genético apropiados.
Abstract The Bardet-Biedl syndrome is an autosomal recessive hereditary disorder with vast locus heterogeneity that belongs to the so-called ciliopathies, whose proteins are localized in the primary cilia and present functional deficiency. The multisystemic features of the disease include ocular, renal, cognitive, skeletal, as well as gonadal involvement and obesity, among others, with high inter- and intrafamilial variability. We describe the clinical case of an adolescent male patient with Bardet-Biedl syndrome, including the approach, the results from a 22-gene sequencing panel, and the analysis of updated scientific literature. We collected the clinical data of the patient and, after obtaining the informed consent, we conducted a multigenic sequencing panel oriented to known implicated genes. The patient was born to consanguineous parents and was the first affected member of the family. He presented with postaxial polydactyly, obesity, micropenis, retinitis pigmentosa, and learning disability. The multigenic panel allowed the identification of the homozygous pathogenic variant c.39_46del in the BBS10 gene and in other BBS genes variants associated with obesity. As the Bardet-Biedl syndrome is a rare disease, it is challenging to interpret its pleiotropism and gene/allelic heterogeneity. Its confirmation by molecular tests allows an adequate approach, follow-up, and genetic counseling of the patient and the family.
Assuntos
Adolescente , Humanos , Masculino , Síndrome de Bardet-Biedl/genética , Chaperoninas do Grupo II/genética , Linhagem , Análise Mutacional de DNA , Deleção de Sequência , Chaperoninas , Consanguinidade , Genes Recessivos , HomozigotoRESUMO
Anaplasmataceae family members include vector-borne bacteria of veterinary importance that may also affect humans. Ehrlichia canis and Anaplasma platys are the main members of this family detected in dogs worldwide. In Uruguay there are not many published studies on tick-borne pathogens affecting dogs, the only haemoparasite molecularly confirmed in dogs, is the piroplasm Rangelia vitalii. The aim of the present work was to detect the presence of A. platys and E. canis in dogs and dogs-associated ticks of two localities in Northwestern Uruguay. Blood samples from dogs with and without clinical signs associated with vector-borne diseases, and Rhipicephalus sanguineus obtained from these dogs were analyzed by PCR for Anaplasmataceae. Positive dogs were further analyzed by PCR for Ehrlichia spp. and A. platys. All the ticks were found negative. No dog was detected infected with E. canis, while eight dogs (4.2%) were found to be infected with A. platys. Phylogenetic analysis of groESL operon sequence for A. platys revealed no differences with sequences described for A. platys in neighbor countries and from other regions of the world. This is the first report of the presence of A. platys in Uruguay.
Assuntos
Anaplasma/isolamento & purificação , Anaplasmose/microbiologia , Doenças do Cão/microbiologia , Anaplasma/classificação , Anaplasmose/epidemiologia , Animais , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Chaperoninas/genética , Chaperoninas/metabolismo , Doenças do Cão/epidemiologia , Cães , Feminino , Regulação Bacteriana da Expressão Gênica , Masculino , Ninfa/microbiologia , Filogenia , Uruguai/epidemiologiaRESUMO
The interferon (IFN)-γ response to peptides can be a useful diagnostic marker of Mycobacterium tuberculosis (MTB) latent infection. We identified promiscuous and potentially protective CD4+ T-cell epitopes from the most conserved regions of MTB antigenic proteins by scanning the MTB antigenic proteins GroEL2, phosphate-binding protein 1 precursor and 19 kDa antigen with the TEPITOPE algorithm. Seven peptide sequences predicted to bind to multiple human leukocyte antigen (HLA)-DR molecules were synthesised and tested with IFN-γ enzyme-linked immunospot (ELISPOT) assays using peripheral blood mononuclear cells (PBMCs) from 16 Mantoux tuberculin skin test (TST)-positive and 16 TST-negative healthy donors. Eighty-eight percent of TST-positive donors responded to at least one of the peptides, compared to 25% of TST-negative donors. Each individual peptide induced IFN-γ production by PBMCs from at least 31% of the TST-positive donors. The magnitude of the response against all peptides was 182 ± 230 x 106 IFN-γ spot forming cells (SFC) among TST-positive donors and 36 ± 62 x 106 SFC among TST-negative donors (p = 0.007). The response to GroEL2 (463-477) was only observed in the TST-positive group. This combination of novel MTB CD4 T-cell epitopes should be tested in a larger cohort of individuals with latent tuberculosis (TB) to evaluate its potential to diagnose latent TB and it may be included in ELISPOT-based IFN-γ assays to identify individuals with this condition.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Epitopos/imunologia , Interferon gama/metabolismo , Tuberculose Latente/diagnóstico , Mycobacterium tuberculosis/imunologia , Teste Tuberculínico , Adulto , Algoritmos , Antígenos de Bactérias/análise , Proteínas de Bactérias/sangue , Biomarcadores/análise , Brasil , Linfócitos T CD4-Positivos/metabolismo , Chaperoninas/sangue , ELISPOT , Mapeamento de Epitopos , Antígenos HLA-DR/imunologia , Voluntários Saudáveis , Humanos , Tuberculose Latente/imunologia , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/metabolismo , Pessoa de Meia-Idade , Proteínas de Ligação a Fosfato/sangueRESUMO
The interferon (IFN)-γ response to peptides can be a useful diagnostic marker of Mycobacterium tuberculosis (MTB) latent infection. We identified promiscuous and potentially protective CD4+ T-cell epitopes from the most conserved regions of MTB antigenic proteins by scanning the MTB antigenic proteins GroEL2, phosphate-binding protein 1 precursor and 19 kDa antigen with the TEPITOPE algorithm. Seven peptide sequences predicted to bind to multiple human leukocyte antigen (HLA)-DR molecules were synthesised and tested with IFN-γ enzyme-linked immunospot (ELISPOT) assays using peripheral blood mononuclear cells (PBMCs) from 16 Mantoux tuberculin skin test (TST)-positive and 16 TST-negative healthy donors. Eighty-eight percent of TST-positive donors responded to at least one of the peptides, compared to 25% of TST-negative donors. Each individual peptide induced IFN-γ production by PBMCs from at least 31% of the TST-positive donors. The magnitude of the response against all peptides was 182 ± 230 x 106 IFN-γ spot forming cells (SFC) among TST-positive donors and 36 ± 62 x 106 SFC among TST-negative donors (p = 0.007). The response to GroEL2 (463-477) was only observed in the TST-positive group. This combination of novel MTB CD4 T-cell epitopes should be tested in a larger cohort of individuals with latent tuberculosis (TB) to evaluate its potential to diagnose latent TB and it may be included in ELISPOT-based IFN-γ assays to identify individuals with this condition.
Assuntos
Adulto , Humanos , Pessoa de Meia-Idade , /imunologia , Epitopos/imunologia , Interferon gama/metabolismo , Tuberculose Latente/diagnóstico , Mycobacterium tuberculosis/imunologia , Teste Tuberculínico , Algoritmos , Antígenos de Bactérias/análise , Brasil , Proteínas de Bactérias/sangue , Biomarcadores/análise , /metabolismo , Chaperoninas/sangue , ELISPOT , Mapeamento de Epitopos , Voluntários Saudáveis , Antígenos HLA-DR/imunologia , Tuberculose Latente/imunologia , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/metabolismo , Proteínas de Ligação a Fosfato/sangueRESUMO
Mitochondria are critical for cell survival and normal development, as they provide energy to the cell, buffer intracellular calcium, and regulate apoptosis. They are also major targets of oxidative stress, which causes bioenergetics failure in astrocytes through the activation of different mechanisms and production of oxidative molecules. This review provides an insightful overview of the recent discoveries and strategies for mitochondrial protection in astrocytes. We also discuss the importance of rotenone as an experimental approach for assessing oxidative stress in the brain and delineate some molecular strategies that enhance mitochondrial function in astrocytes as a promising strategy against brain damage.
Assuntos
Astrócitos/fisiologia , Mitocôndrias/fisiologia , Rotenona/toxicidade , Doenças dos Trabalhadores Agrícolas/induzido quimicamente , Doenças dos Trabalhadores Agrícolas/epidemiologia , Animais , Apoptose/efeitos dos fármacos , Apoptose/fisiologia , Astrócitos/efeitos dos fármacos , Astrócitos/ultraestrutura , Encéfalo/metabolismo , Chaperoninas/fisiologia , Transporte de Elétrons/efeitos dos fármacos , Regulação da Expressão Gênica , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Peroxidação de Lipídeos , Mitocôndrias/efeitos dos fármacos , Membranas Mitocondriais/efeitos dos fármacos , Membranas Mitocondriais/fisiologia , NF-kappa B/fisiologia , Proteínas do Tecido Nervoso/fisiologia , Doenças Neurodegenerativas/metabolismo , Doenças Neurodegenerativas/patologia , Estresse Oxidativo , Transtornos Parkinsonianos/induzido quimicamente , Praguicidas/toxicidade , Superóxido Dismutase/fisiologia , Fatores de Transcrição/fisiologiaAssuntos
Proteínas de Homeodomínio/metabolismo , Meristema/citologia , Proteínas de Plantas/metabolismo , Plasmodesmos/metabolismo , Dobramento de Proteína , RNA Mensageiro/metabolismo , RNA de Plantas/metabolismo , Zea mays/metabolismo , Arabidopsis/genética , Chaperoninas/metabolismo , Flores , Proteínas de Homeodomínio/química , Meristema/metabolismo , Folhas de Planta/citologia , Folhas de Planta/metabolismo , Proteínas de Plantas/química , Transporte Proteico , Zea mays/genéticaRESUMO
OBJECTIVES: The aim of the present work was to standardise a PCR-Restriction Fragment Length Polymorphism analysis (PRA) as a tool to detect the mycobacteriologic composition of lepromas from leprosy patients used in the production of lepromin to improve the quality of the Mitsuda test. DESIGN: PCR-Restriction Fragment Length Polymorphism analysis using hsp65 and rpoB genes were applied to 11 reference strains of mycobacteria, including M. leprae, and the obtained PRA profiles were compared to mycobacteria in clinical specimens. RESULTS: Out of the biopsies studied, 522% had DNA fragment amplified for both genes (hsp65 and rpoB) for M. leprae. However, other Mycobacterium species were observed in samples of lepromatous leprosy patients. Here we discussed the importance of mycobacteria identification in the antigen of Mitsuda production to be used in the evaluation of leprosy. CONCLUSIONS: Our results suggest that the use of the molecular approach for sample selection can contribute to an improvement in the quality of produced lepromin.
Assuntos
Impressões Digitais de DNA/métodos , Antígeno de Mitsuda/isolamento & purificação , Hanseníase/microbiologia , Mycobacterium leprae/classificação , Reação em Cadeia da Polimerase/métodos , Polimorfismo de Fragmento de Restrição , Proteínas de Bactérias/genética , Técnicas de Tipagem Bacteriana/métodos , Técnicas de Tipagem Bacteriana/normas , Chaperonina 60 , Chaperoninas/genética , Análise por Conglomerados , Impressões Digitais de DNA/normas , RNA Polimerases Dirigidas por DNA/genética , Genótipo , Humanos , Hanseníase/patologia , Mycobacterium leprae/genética , Mycobacterium leprae/isolamento & purificação , Reação em Cadeia da Polimerase/normasRESUMO
An epidemic of infections after video-assisted surgery (1,051 possible cases) caused by rapidly growing mycobacteria (RGM) and involving 63 hospitals in the state of Rio de Janeiro, Brazil, occurred between August 2006 and July 2007. One hundred ninety-seven cases were confirmed by positive acid-fast staining and/or culture techniques. Thirty-eight hospitals had cases confirmed by mycobacterial culture, with a total of 148 available isolates recovered from 146 patients. Most (n = 144; 97.2%) isolates presented a PRA-hsp65 restriction pattern suggestive of Mycobacterium bolletii or Mycobacterium massiliense. Seventy-four of these isolates were further identified by hsp65 or rpoB partial sequencing, confirming the species identification as M. massiliense. Epidemic isolates showed susceptibility to amikacin (MIC at which 90% of the tested isolates are inhibited [MIC(90)], 8 microg/ml) and clarithromycin (MIC(90), 0.25 microg/ml) but resistance to ciprofloxacin (MIC(90), >or=32 microg/ml), cefoxitin (MIC(90), 128 microg/ml), and doxycycline (MIC(90), >or=64 microg/ml). Representative epidemic M. massiliense isolates that were randomly selected, including at least one isolate from each hospital where confirmed cases were detected, belonged to a single clone, as indicated by the analysis of pulsed-field gel electrophoresis (PFGE) patterns. They also had the same PFGE pattern as that previously observed in two outbreaks that occurred in other Brazilian cities; we designated this clone BRA100. All five BRA100 M. massiliense isolates tested presented consistent tolerance to 2% glutaraldehyde. This is the largest epidemic of postsurgical infections caused by RGM reported in the literature to date in Brazil.