RESUMO
Tubercidin (TUB) is an adenosine analog with potent antiparasite action, unfortunately associated with severe host toxicity. Prevention of TUB toxicity can be reached associating nitrobenzylthioinosine (NBMPR), an inhibitor of the purine nucleoside transport, specifically target to the mammal cells. It was demonstrated that this nucleoside transport inhibitor has no significant effect in the in vitro uptake of TUB by Schistosoma mansoni and Trypanosoma gambiense. Seeking to evaluate if the association of these compounds is also effective against leishmania, we analyzed the TUB-NBMPR combined treatment in in vitro cultures of promastigote forms of Leishmania (L.) amazonensis, Leishmania (L.) chagasi, Leishmania (L.) major, and Leishmania (V.) braziliensis as well as in cultures of amastigote forms of L. (L.) amazonensis, mice macrophages infected with L. (L.) amazonensis, and in vivo tests in BALB/c mice infected with L. (L.) amazonensis. We demonstrated that TUB-NBMPR combined treatment can be effective against leishmania cells protecting mammalian cells from TUB toxicity.
Assuntos
Antiparasitários/uso terapêutico , Inibidores Enzimáticos/uso terapêutico , Leishmania/efeitos dos fármacos , Leishmaniose/tratamento farmacológico , Tioinosina/análogos & derivados , Tionucleotídeos/uso terapêutico , Tubercidina/uso terapêutico , Animais , Antiparasitários/farmacologia , Antiparasitários/toxicidade , Células Cultivadas , Quimioterapia Combinada , Inibidores Enzimáticos/farmacologia , Macrófagos/efeitos dos fármacos , Macrófagos/parasitologia , Camundongos , Camundongos Endogâmicos BALB C , Schistosoma mansoni/efeitos dos fármacos , Tioinosina/farmacologia , Tioinosina/uso terapêutico , Tionucleotídeos/farmacologia , Trypanosoma brucei gambiense/efeitos dos fármacos , Tubercidina/farmacologia , Tubercidina/toxicidadeRESUMO
BACKGROUND: Cyclosporin A (CsA) is a widely used immunosuppressant that causes significant side effects including gingival overgrowth. The pathogenesis of this condition is not fully understood; however, recent studies show that CsA regulates the transcription of several cytokines including transforming growth factor-beta 1 (TGF-beta1). In this study, we evaluated the effects of CsA and TGF-beta1 on human normal gingival (NG) fibroblast proliferation, and explored a possible autocrine stimulation of TGF-beta1 as a cellular regulator of proliferation induced by CsA in NG fibroblasts. METHODS: NG fibroblast cell lines were incubated with increasing concentrations of CsA or TGF-beta1 and the proliferation index determined by automatic cell counting, BrdU incorporation, PCNA expression, and mitotic potential. To determine the effect of TGF-beta1 on the proliferation rate of NG fibroblasts under CsA treatment, NG fibroblast cultures were simultaneously treated with CsA and antisense oligonucleotides against the translation-start site of the TGF-beta1 mRNA. RESULTS: Treatment of NG fibroblasts with CsA or TGF-beta1 significantly stimulated the cell proliferation in a dose-dependent manner. Furthermore, neutralization of TGF-beta1 production in CsA-treated NG fibroblasts inhibited CsA's effect on NG fibroblast proliferation, demonstrating an autocrine stimulatory effect of TGF-beta1 in CsA-treated NG fibroblast proliferation. CONCLUSION: The results presented here suggest that CsA stimulatory induction of NG fibroblast proliferation is mediated via TGF-beta1 in an autocrine fashion.
Assuntos
Ciclosporina/farmacologia , Fibroblastos/efeitos dos fármacos , Gengiva/efeitos dos fármacos , Imunossupressores/farmacologia , Fator de Crescimento Transformador beta/efeitos dos fármacos , Análise de Variância , Antimetabólitos , Comunicação Autócrina/efeitos dos fármacos , Bromodesoxiuridina , Contagem de Células , Divisão Celular/efeitos dos fármacos , Linhagem Celular , Relação Dose-Resposta a Droga , Gengiva/citologia , Humanos , Índice Mitótico , Oligonucleotídeos Antissenso , Antígeno Nuclear de Célula em Proliferação/análise , Biossíntese de Proteínas/efeitos dos fármacos , Fator de Crescimento Transformador beta/antagonistas & inibidores , Fator de Crescimento Transformador beta/farmacologia , Fator de Crescimento Transformador beta1RESUMO
BACKGROUND: Increased collagen and extracellular matrix deposition within the gingiva is the main characteristic feature of hereditary gingival fibromatosis (HGF). To date, it is not well established if these events are a consequence of alterations in the collagen and other extracellular matrix molecules synthesis or disturbances in the homeostatic equilibrium between synthesis and degradation of extracellular matrix molecules. Cytokines are important regulators of expression of the profibrogenic genes, including type I collagen and its molecular chaperone heat shock protein (Hsp)47 and proteolytic enzymes degrading extracellular matrix such as matrix metalloproteinases-1 and -2 (MMP-1 and MMP-2). METHODS: In this study, we analyzed the expression and production of type I collagen, Hsp47, MMP-1, and MMP-2 in normal gingiva (NG) and HGF fibroblasts, and investigated the effects of transforming growth factor-beta1 (TGF-beta1), interleukin-6 (IL-6) and interferon-gamma (IFN-gamma) on the expression of these genes by NG and HGF fibroblasts. RESULTS: Our results obtained from semi-quantitative reverse transcription-polymerase chain reactions (RT-PCR), Western blots, enzyme-linked immunosorbent assays (ELISA), and enzymographies clearly demonstrated that the expression and production of type I collagen and Hsp47 were significantly higher in fibroblasts from HGF than from NG, whereas MMP-1 and MMP-2 expression and production were lower in fibroblasts from HGF patients. Addition of TGF-beta1 and IL-6, which are produced in greater amounts by HGF fibroblasts, promoted an increase in type I collagen and Hsp47 and a decrease in MMP-1 and MMP-2 expression. IFN-gamma reduced both type I collagen and Hsp47 expression, whereas it had a slight effect on the expression of MMP-1 and MMP-2. CONCLUSION: These patterns of expression and production suggest that enhanced TGF-beta1 and IL-6 production simultaneously increase the synthesis and reduce the proteolytic activities of fibroblasts from patients with HGF, which may favor the accumulation of extracellular matrix observed in patients with this condition.
Assuntos
Colágeno Tipo I/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Fibromatose Gengival/genética , Gengiva/efeitos dos fármacos , Proteínas de Choque Térmico/efeitos dos fármacos , Interferon gama/farmacologia , Interleucina-6/farmacologia , Metaloproteinase 1 da Matriz/efeitos dos fármacos , Metaloproteinase 2 da Matriz/efeitos dos fármacos , Fator de Crescimento Transformador beta/farmacologia , Adulto , Análise de Variância , Técnicas de Cultura de Células , Feminino , Fibromatose Gengival/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Proteínas de Choque Térmico HSP47 , Humanos , Masculino , Estatísticas não ParamétricasRESUMO
BACKGROUND: Gingival overgrowth is a common side effect following the administration of cyclosporin A (CsA). The pathogenesis of this condition is not fully understood; however, recent studies show that CsA regulates the transcription of several cytokines including transforming growth factor-beta1 (TGF-beta1). The aim of this study was to investigate the potential role of TGF-beta1 in the pathogenesis of CsA-induced gingival overgrowth, exploring a possible autocrine stimulation of TGF-beta1 as a cellular regulator of synthesis of matrix metalloproteinases (MMPs) and its tissue inhibitors (TIMPs). METHODS: Gingival fibroblasts from human normal gingiva were incubated with increasing concentrations of CsA, cultured for 24 hours, and the expression and production of TGF-beta1 determined by semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA), respectively. MMP and TIMP mRNA expression levels were also analyzed by RT-PCR. To determine the effect of TGF-beta1 on the expression of MMP and TIMP by human gingival fibroblasts under CsA treatment, human gingival fibroblast cultures were treated with sense oligonucleotides (SON) or antisense oligonucleotides (AON). RESULTS: CsA simultaneously stimulated TGF-beta1 expression and production and inhibited expression of MMP-1 and MMP-2 by human gingival fibroblasts, whereas CsA has a slight effect on TIMP-1 and TIMP-2 expression. AON reduced TGF-beta1 production as demonstrated by ELISA, whereas TGF-beta1 mRNA expression levels were not significantly modified. The inhibition of TGF-beta1 production by AON modulated MMP expression, demonstrating the autocrine inhibitory effect of TGF-beta1 in CsA-treated human gingival fibroblasts. CONCLUSIONS: The data presented here suggest that TGF-beta1 in an autocrine fashion may contribute to a reduction of proteolytic activity of human gingival fibroblasts in CsA-induced gingival overgrowth, which favors the accumulation of extracellular matrix.
Assuntos
Ciclosporina/farmacologia , Inibidores Enzimáticos/farmacologia , Gengiva/efeitos dos fármacos , Crescimento Excessivo da Gengiva/induzido quimicamente , Crescimento Excessivo da Gengiva/enzimologia , Metaloproteinases da Matriz/biossíntese , Fator de Crescimento Transformador beta/fisiologia , Análise de Variância , Células Cultivadas , Regulação para Baixo , Ensaio de Imunoadsorção Enzimática , Fibroblastos/efeitos dos fármacos , Fibroblastos/enzimologia , Gengiva/citologia , Gengiva/enzimologia , Humanos , Inibidores de Metaloproteinases de Matriz , Oligonucleotídeos Antissenso/farmacologia , Iniciação Traducional da Cadeia Peptídica/efeitos dos fármacos , RNA Mensageiro/análise , RNA Mensageiro/biossíntese , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Estatísticas não Paramétricas , Inibidores Teciduais de Metaloproteinases/biossíntese , Fator de Crescimento Transformador beta/biossíntese , Fator de Crescimento Transformador beta/efeitos dos fármacos , Fator de Crescimento Transformador beta1RESUMO
Background: Gingival overgrowth is a common side effect following the administration of cyclosporin A (CsA). The pathogenesis of this condition is not fully understood; however, recent studies show that CsA regulates the transcription of several cytokines including transforming growth factor-ß1 (TGF-ß1). The aim of this study was to investigate the potential role of TGF-ß1 in the pathogenesis of CsA-induced gingival overgrowth, exploring a possible autocrine stimulation of TGF-ß1 as a cellular regulator of synthesis of matrix metalloproteinases (MMPs) and its tissue inhibitors (TIMPs). Methods: Gingival fibroblasts from human normal gingiva were incubated with increasing concentrations of CsA, cultured for 24 hours, and the expression and production of TGF-ß1 determined by semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA), respectively. MMP and TIMP mRNA expression levels were also analyzed by RT-PCR. To determine the effect of TGF-ß1 on the expression of MMP and TIMP by human gingival fibroblasts under CsA treatment, human gingival fibroblast cultures were treated with sense oligonucleotides (SON) or anti-sense oligonucleotides (AON). Results: CsA simultaneously stimulated TGF-ß1 expression and production and inhibited expression of MMP-1 and MMP-2 by human gingival fibroblasts, whereas CsA has a slight effect on TIMP-1 and TIMP-2 expression. AON reduced TGF-ß1 production as demonstrated by ELISA, whereas TGF-ß1 mRNA expression levels were not significantly modified. The inhibition of TGF-ß1 production by AON modulated MMP expression, demonstrating the autocrine inhibitory effect of TGF-ß1 in CsA-treated human gingival fibroblasts. Conclusions: The data presented here suggest that TGF-ß1 in an autocrine fashion may contribute to a reduction of proteolytic activity of human gingival fibroblasts in CsA-induced gingival overgrowth, which favors the accumulation of extra-cellular matrix
Assuntos
Ciclosporina/efeitos adversos , Fibroblastos , Hiperplasia Gengival , Substâncias de Crescimento , Metaloproteinases da MatrizRESUMO
Basaloid squamous carcinoma (BSC) is an aggressive variant of squamous cell carcinoma (SCC) with a predilection for the upper aerodigestive tract. In the English literature, approximately 40 cases of BSC have been described in the oral cavity. BSC has frequently been confused with adenoid cystic carcinoma (ACC), basal cell adenocarcinoma, and undifferentiated SCC. The purpose of the investigation was to examine the histological features and immunohistochemical expression of differentiation-related substances, including cytokeratin (CK) subtypes, vimentin, S-100, chromogranin, laminin, and type IV collagen, for the characterization of biological features of these tumours. We studied three cases of BSC of the oral cavity, three cases of ACC, and one case of basal cell adenocarcinoma. Well-differentiated and undifferentiated SCCs were also studied for comparison. The BSCs showed many histopathologic similarities to cases previously reported. Among the CK subtypes analyzed, CK14 was the only subtype expressed by all basaloid cells of BSC. Potentially useful for the differential diagnosis was the finding of CKs 7 and 19 expression in the basaloid cells of ACC, and CKs 7 and 8 in basal cell adenocarcinoma. In BSCs, laminin and type IV collagen were found in the microcystic spaces between basaloid cells, but neither ACCs nor basal cell adenocarcinoma showed this feature. These data suggest that immunohistochemical findings are helpful in distinguishing BSC of the oral cavity from other histopathologically similar tumours.
Assuntos
Carcinoma de Células Escamosas/patologia , Carcinoma de Células de Transição/patologia , Neoplasias Bucais/patologia , Adenocarcinoma/química , Adenocarcinoma/diagnóstico , Adenocarcinoma/patologia , Carcinoma Adenoide Cístico/química , Carcinoma Adenoide Cístico/patologia , Carcinoma de Células Escamosas/química , Carcinoma de Células de Transição/química , Diagnóstico Diferencial , Humanos , Imuno-Histoquímica , Queratinas/análise , Neoplasias Bucais/químicaRESUMO
Basaloid squamous carcinoma (BSC) is an uncommon aggressive variant of squamous cell carcinoma (SCC) with a predilection for the head and neck. In the English literature, approximately 40 cases of BSC in the oral cavity have been described. In this study, the clinicopathologic features of 2 cases of BSC affecting the buccal mucosa are reported. In addition, we compare the proliferative and invasive potential of BSC cells with that of poorly differentiated SCC cells matched for age, sex, site, and TNM status. Proliferative activity was studied through use of the argyrophilic nuclear organizer region (AgNOR) method and immunohistochemical quantification of proliferating cell nuclear antigen (PCNA). The invasive potential was evaluated through use of the semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) for matrix metalloproteinases (MMPs). Alterations of p53 were also investigated through use of immunohistochemistry. The tumors showed many clinical and histopathologic similarities to tumors in cases previously reported. The AgNOR and PCNA indices were significantly higher in the 2 cases of BSC than in the cases of SCC. Immunostaining for p53 protein showed a higher percentage of positive cells and more intense staining in the BSC tissues than in the SCC tissues. RT-PCR studies clearly demonstrated that the expression of MMP-1, MMP-2, and MMP-9 was higher in cells from BSCs than in cells from SCCs. Taken together, the data described here are compatible with the concept that BSC has a more aggressive biologic behavior than conventional SCC.
Assuntos
Carcinoma Basoescamoso/patologia , Metaloproteinases da Matriz/análise , Neoplasias Bucais/patologia , Região Organizadora do Nucléolo/ultraestrutura , Antígeno Nuclear de Célula em Proliferação/análise , Proteína Supressora de Tumor p53/análise , Carcinoma Basoescamoso/enzimologia , Carcinoma de Células Escamosas/patologia , Estudos de Casos e Controles , Divisão Celular , Corantes , Feminino , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Imuno-Histoquímica , Masculino , Metaloproteinase 1 da Matriz/análise , Metaloproteinase 2 da Matriz/análise , Metaloproteinase 9 da Matriz/análise , Pessoa de Meia-Idade , Neoplasias Bucais/enzimologia , Invasividade Neoplásica , Estadiamento de Neoplasias , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Nitrato de PrataRESUMO
We tested a general method for the identification of drug resistance loci in the trypanosomatid protozoan parasite Leishmania major. Genomic libraries in a multicopy episomal cosmid vector were transfected into susceptible parasites, and drug selections of these transfectant libraries yielded parasites bearing cosmids mediating resistance. Tests with two antifolates led to the recovery of cosmids encoding DHFR-TS or PTR1, two known resistance genes. Overexpression/selection using the toxic nucleoside tubercidin similarly yielded the TOR (toxic nucleoside resistance) locus, as well as a new locus (TUB2) conferring collateral hypersensitivity to allopurinol. Leishmania synthesize ergosterol rather than cholesterol, making this pathway attractive as a chemotherapeutic target. Overexpression/selection using the sterol synthesis inhibitors terbinafine (TBF, targeting squalene epoxidase) and itraconazole (ITZ, targeting lanosterol C(14)-demethylase) yielded nine new resistance loci. Several conferred resistance to both drugs; several were drug-specific, and two TBF-resistant cosmids induced hypersensitivity to ITZ. One TBF-resistant cosmid encoded squalene synthase (SQS1), which is located upstream of the sites of TBF and ITZ action in the ergosterol biosynthetic pathway. This suggests that resistance to "downstream" inhibitors can be mediated by increased expression of ergosterol biosynthetic intermediates. Our studies establish the feasibility of overexpression/selection in parasites and suggest that many Leishmania drug resistance loci are amenable to identification in this manner.
Assuntos
Resistência a Medicamentos/genética , Ergosterol/antagonistas & inibidores , Leishmania major/efeitos dos fármacos , Seleção Genética , Tubercidina/antagonistas & inibidores , Sequência de Aminoácidos , Animais , Sequência de Bases , Primers do DNA , Ergosterol/metabolismo , Farnesil-Difosfato Farnesiltransferase/química , Farnesil-Difosfato Farnesiltransferase/genética , Humanos , Leishmania major/enzimologia , Leishmania major/genética , Dados de Sequência Molecular , Homologia de Sequência de Aminoácidos , Tubercidina/metabolismoRESUMO
We have studied the genomic organization and expression of the gene encoding a high molecular mass (300 kDa) repetitive antigen associated with the cytoskeleton of Trypanosoma cruzi. Protease digestion of the native protein, restriction analysis of genomic DNA and sequencing of genomic and cDNA clones indicated that most of the protein is built up by tandemly arranged, nearly identical repeats of 68 amino acids. The gene size was estimated to be approx. 9.4 kb based on the sizes of the transcript and the native protein. The nucleotide sequence conservation among the repeats indicates that selective sequence homogenization, presumably through gene conversion, maintained the amino-acid sequence conservation. Two duplicated allelic forms of this gene were mapped in fragments of about 20 kb. In some strains an additional allele was located in a fragment of 9.4 kb. Our results suggest that this repetitive antigen is a structural protein which could be involved in the attachment of the flagellum to the cell body.
Assuntos
Antígenos de Protozoários/genética , Citoesqueleto/química , Genes de Protozoários/genética , Proteínas de Protozoários/genética , Trypanosoma cruzi/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Clonagem Molecular , Sequência Conservada/genética , Proteínas do Citoesqueleto/genética , Regulação da Expressão Gênica , Dados de Sequência Molecular , Polimorfismo de Fragmento de Restrição , Proteínas de Protozoários/análise , RNA Mensageiro/análise , RNA de Protozoário/análise , Sequências Repetitivas de Ácido Nucleico/genética , Mapeamento por Restrição , Análise de Sequência de DNA , Transcrição Gênica , Trypanosoma cruzi/imunologiaRESUMO
A Trypanosoma cruzi DNA fragment encoding an immunodominant repetitive antigen (H49) was subcloned into a protein purification and expressed system. Purified H49 peptide reacted specifically in an enzyme-linked immunosorbent assay (ELISA) with sera from T. cruzi-infected patients, but not with sera from patients with other parasitic diseases such as leishmaniasis and T. rangeli-infection. The H49 recombinant ELISA was able to detect specific antibodies in 84% of chronic chagasic serum samples tested. One of the major advantage of the recombinant ELISA for serodiagnosis of chronic Chagas' disease resides in its high specificity (100%). Our data suggest that recombinant peptides could provide a practical basis for specific diagnosis tests for Chagas' disease.
Assuntos
Anticorpos Antiprotozoários/sangue , Doença de Chagas/imunologia , Epitopos Imunodominantes/imunologia , Proteínas Recombinantes/imunologia , Trypanosoma cruzi/imunologia , Animais , Antígenos de Protozoários/imunologia , Eletroforese em Gel de Poliacrilamida , Ensaio de Imunoadsorção Enzimática , Humanos , Epitopos Imunodominantes/genética , Infecções por Protozoários/imunologia , Testes Sorológicos , Trypanosoma cruzi/genéticaRESUMO
A monospecific antiserum raised to a Trypanosoma cruzi recombinant antigen, with tandem repeat of 68 amino acids, was used to screen urine samples of chagasic and nonchagasic patients. The antiserum detected a specific 150-160 kDa antigen in urine of 60% of chronic chagasic patients, but not in urine samples from nonchagasic patients and healthy control individuals. The reactivity to 150-160 kDa urinary antigen could be abolished by adsorption with the recombinant repetitive antigen. These results suggest that 150-160 kDa urinary antigen is a T. cruzi-derived antigen and specific for Chagas' disease.
Assuntos
Antígenos de Protozoários/urina , Doença de Chagas/urina , Trypanosoma cruzi/imunologia , Adulto , Animais , Doença de Chagas/diagnóstico , Doença de Chagas/imunologia , Doença de Chagas/parasitologia , Epitopos , Feminino , Humanos , Soros Imunes , Masculino , Pessoa de Meia-Idade , Peso Molecular , Proteínas Recombinantes de Fusão/imunologia , Trypanosoma cruzi/isolamento & purificaçãoAssuntos
Antígenos de Protozoários , Doença de Chagas/imunologia , Escherichia coli/imunologia , Trypanosoma cruzi/imunologia , Animais , Anticorpos Antiprotozoários/análise , Anticorpos Antiprotozoários/imunologia , Antígenos de Protozoários/imunologia , Humanos , Proteínas Recombinantes/imunologia , Sensibilidade e EspecificidadeRESUMO
Here we describe the characterization of a Trypanosoma cruzi DNA sequence (clone A13) that codes for a polypeptide recognized by IgM and IgG antibodies from sera of acute and congenital chagasic patients. Antibodies to A13 antigen are also detected in the sera of chronic patients with different clinical forms of Chagas' disease, but not in sera of patients with leishmaniasis or other parasitic diseases. The antigenic determinants encoded by clone A13 are found in amastigotes and trypomastigotes of several T. cruzi strains, but not in the noninfective epimastigotes. The DNA sequence of the recombinant clone reveals one open reading frame encoding 251 amino acids without tandemly repeated sequences. Our data suggest that the A13 antigen may be useful for the development of serodiagnostic procedures.
Assuntos
Antígenos de Protozoários/genética , Doença de Chagas/imunologia , DNA de Protozoário/genética , Trypanosoma cruzi/imunologia , Doença Aguda , Sequência de Aminoácidos , Animais , Antígenos de Protozoários/imunologia , Sequência de Bases , Doença de Chagas/diagnóstico , Doença Crônica , Clonagem Molecular , DNA de Protozoário/isolamento & purificação , Epitopos/genética , Epitopos/imunologia , Regulação da Expressão Gênica , Humanos , Imunoglobulina G/imunologia , Imunoglobulina M/imunologia , Dados de Sequência Molecular , Hibridização de Ácido Nucleico , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Trypanosoma cruzi/genéticaRESUMO
A genomic clone expressing a Trypanosoma cruzi antigen in Escherichia coli was identified using human chagasic sera. Chagasic antibodies affinity purified on extracts of this clone recognized a high-molecular-weight protein expressed in all developmental stages of the parasite life cycle, as well as in various T. cruzi strains. The antigen is associated with the cytoskeleton of the parasite and localizes along the attachment region between the flagellum and the cell body. Antibodies to the recombinant antigen were detected in the sera of 115 chagasic patients from different endemic regions, but not in sera of patients with leishmaniasis, T. rangeli infection, or other parasitic diseases. Our data suggest that the presence of antibodies to this antigen may be specifically associated with Chagas' disease.
Assuntos
Antígenos de Protozoários/genética , Doença de Chagas/imunologia , Trypanosoma cruzi/imunologia , Animais , Northern Blotting , Southern Blotting , Western Blotting , Clonagem Molecular , Citoesqueleto/imunologia , DNA/genética , Epitopos/imunologia , Escherichia coli/genética , Imunofluorescência , Expressão Gênica , Humanos , Soros Imunes/imunologia , Trypanosoma cruzi/genética , Trypanosoma cruzi/ultraestruturaRESUMO
We have isolated a clone of Trypanosoma cruzi genomic DNA, lambda 3b2-5, which contains sequences that are reiterated in the genome. Northern blot analysis showed that clone 3b2-5 hybridizes to 1,200-5,000 bases different mRNA species. The number of mRNAs species hybridized to clone 3b2-5 exceeds its coding capacity showing that this clone carries sequences that are common to several mRNAs species and conserved in the poly A(+) RNA. These sequences are not homologous to the T. cruzi spliced leader sequence, since clone 3b2-5 does not hybridize to a synthetic 20 nucleotide complementary to the spliced leader sequence. Clone 3b2-5 does not hybridize to DNA and RNA from several genera of Trypanosomatidae and other Trypanosoma species indicating that it carries T. cruzi species-specific sequences.
Assuntos
DNA/genética , RNA Mensageiro/genética , Sequências Repetitivas de Ácido Nucleico , Trypanosoma cruzi/genética , Animais , Northern Blotting , Southern Blotting , Biblioteca Genômica , Família Multigênica , Homologia de Sequência do Ácido Nucleico , Especificidade da EspécieRESUMO
Intact RNAs were isolated from epimastigote forms of different Trypanosoma cruzi strains. Translation of the mRNAs using rabbit reticulocyte lysate and analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed protein profiles comparable to those observed by labeling cells in vivo. No major interstrain differences were observed in the patterns of the polypeptides synthesized in vitro and in vivo, indicating that metabolic proteins are similar among distinct strains. Several T. cruzi polypeptides produced in the rabbit reticulocyte lysate system were immunoprecipitated by specific antisera. The patterns of antigenic polypeptides recognized by antisera raised against epimastigotes from different strains were also very similar.
Assuntos
Antígenos de Protozoários/isolamento & purificação , Trypanosoma cruzi/metabolismo , Animais , Eletroforese em Gel de Poliacrilamida , Peso Molecular , Biossíntese de Proteínas , RNA Mensageiro/isolamento & purificação , Coelhos , Reticulócitos/parasitologiaRESUMO
Um grupo de 1.288 indivíduos, residentes em Salvador, Bahia, Brasil, foi examinado para a determinaçäo de anti-HBs, utilizando-se a técnica do radioimunoensaio. Os resultados foram analisados de acordo com a idade, o sexo e o nível de renda. Uma prevalência de anti-HBs de 11,8% foi observada, variando de 6,7% nas crianças com idade inferior a três anos, a 26,1% para aqueles com idade superior a 30 anos. A prevalência de anti-HBs foi similar em indivíduos do sexo masculino e feminino. Aqueles com nível de renda mais alto tiveram prevalência de anti-HBs maior que o grupo de renda baixa. Dos resultados conclui-se: a alta prevalência de anti-HBs em crianças sugere o contato precoce com vírus da hepatite B, possivelmente relacionado com a transmissäo vertical e a disseminaçäo intrafamiliar do vírus; a freqüência de anti-HBs aumentou com a idade; a baixa prevalência de anti-HBs entre aqueles com baixo nível sócio-econômico sugere que neste grupo pode ser alta a prevalência de portadores do vírus B da hepatite