RESUMO
OBJECTIVE: Historical clinical studies suggest the potential for insect-borne transmission of human hepatitis viruses. Studies of hepatitis B virus (HBV) persistence in insects were performed before the advent of molecular techniques, and studies to assess possible insect-borne transmission of hepatitis viruses have not yet been performed. The aim of this study was to determine, using molecular techniques, whether HBV and hepatitis C virus (HCV) persist in and are excreted in the feces of the bedbug Cimex lectularius L. and kissing bug Rodnius prolixus after an infectious meal. METHODS: Blood-feeding insects from the insect order Hemiptera (Cimex lectularius L. and Rhodnius prolixus) were fed on blood from infected patients with high titers of HBV, HCV, and control uninfected patients. Insects and insect excrement were collected at weekly intervals and tested for HBV DNA and HCV RNA using the polymerase chain reaction. RESULTS: HBV DNA was detected in bedbugs and excrement up to 6 wk after feeding on an infectious meal. HBV DNA was also detected in most kissing bugs and excrement up to 2 wk after feeding. HCV RNA was not detected in bedbugs at any time after feeding. CONCLUSIONS: We did not detect HCV RNA in bedbugs after feeding on an infectious meal. Our data provide molecular evidence to suggest that HBV may persist in Hemiptera. Additional studies are ongoing to determine whether this viral persistence is capable of infection.
Assuntos
Percevejos-de-Cama/virologia , Hepacivirus/isolamento & purificação , Vírus da Hepatite B/isolamento & purificação , Hepatite Viral Humana/transmissão , Insetos Vetores/virologia , Rhodnius/virologia , Animais , DNA Viral/análise , Fezes/virologia , Hepacivirus/genética , Hepatite B/transmissão , Vírus da Hepatite B/genética , Hepatite C/transmissão , Humanos , RNA Viral/análiseRESUMO
We examined the proliferative responses of peripheral blood mononuclear cells obtained from 60 untreated patients who were seropositive by enzyme immunoassay, but negative for hepatitis C virus (HCV) RNA by reverse transcriptase-polymerase chain reaction (RT-PCR). We used second- and third-generation recombinant immunoblot assay (RIBA) for further serological characterization. In vitro HCV-specific proliferative responses of mononuclear cells were compared to those of both untreated chronic HCV patients and patients who showed sustained virological response to interferon-alpha monotherapy, in order to assess the relative contribution of the immune response to the eradication of HCV. We found that frequency of responses to nonstructural proteins showed statistically significant differences, which were attributable to vigorous, polyspecific responses by cells from the RIBA-positive patients. In this group, core-specific proliferation was significantly associated with intravenous drug use as route of acquisition. Both other patient groups and the RIBA-indeterminate patients showed indistinguishable frequencies of proliferative responses. No association was detected between residual humoral responses, as determined from the RIBA results, and elapsed time since infection. The frequency of antibodies to NS5 differs between spontaneous cure and chronically infected patients. Cell-mediated and humoral immunity appear to be maintained in this population of patients.
Assuntos
Hepacivirus/imunologia , Anticorpos Anti-Hepatite C/sangue , Hepatite C/imunologia , Ativação Linfocitária , Adolescente , Adulto , Criança , Feminino , Hepacivirus/isolamento & purificação , Hepatite C/tratamento farmacológico , Hepatite C/virologia , Hepatite C Crônica/imunologia , Humanos , Immunoblotting , Técnicas Imunoenzimáticas , Interferon-alfa/uso terapêutico , Fígado/virologia , Masculino , RNA Viral/análise , RNA Viral/sangue , Proteínas Recombinantes/imunologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Risco , Abuso de Substâncias por Via Intravenosa/complicações , Fatores de Tempo , Proteínas não Estruturais Virais/imunologia , Proteínas Virais/imunologiaRESUMO
We previously reported that recovery of Lewis rats from experimental autoimmune encephalomyelitis (EAE) is associated with the appearance of suppressor T cells (Ts). These Ts secrete TGF-beta which down-regulates the production of inflammatory cytokines by the effector T cells that mediate this disease. In the present study, we immunized Lewis rats with myelin basic protein (MBP)+CFA, and evaluated purified T cells and MBP-activated spleen cells (SpC) during the paralytic phase (day 12) and after recovery (days 30-33) for TGF-beta and interferon (IFN)-gamma mRNA. We used reverse transcriptase-polymerase chain reaction (RT-PCR), quantitated on the basis of beta-actin mRNA. Abundant IFN-gamma mRNA was present in MBP-activated SpC obtained on day 12. In contrast, only trace IFN-gamma mRNA was detected in day 30 activated SpC, and no IFN-gamma mRNA was present in purified, nonactivated T cells obtained at either time. The level of IFN-gamma mRNA correlated with secretion of IFN-gamma as determined by ELISA on SpC culture supernatants, and with severity of adoptively transferred EAE by the activated SpC. Thus, it appears that IFN-gamma mRNA is both transcribed and translated in response to antigen activation, resulting in secretion of IFN-gamma by the disease-inducing Te. In contrast, when we used RT-PCR to investigate the expression of TGF-beta mRNA, we found the transcript present in isolated T cells and MBP-activated SpC obtained from rats at both days 12 and 30. The presence of TGF-beta mRNA at time points corresponding to both clinical EAE and recovery suggests post-transcriptional regulation of the production of this immunoregulatory cytokine.
Assuntos
Encefalomielite Autoimune Experimental/genética , Regulação da Expressão Gênica , Interferon gama/biossíntese , Linfócitos T Reguladores/metabolismo , Fator de Crescimento Transformador beta/biossíntese , Doença Aguda , Animais , Células Cultivadas , Convalescença , Feminino , Interferon gama/genética , Ativação Linfocitária , Reação em Cadeia da Polimerase , RNA Mensageiro/biossíntese , Ratos , Ratos Endogâmicos Lew , Baço/imunologia , Fator de Crescimento Transformador beta/genéticaRESUMO
A modified splenic fragment assay was used to assess the role of antigen-specific helper T cells in B cell isotype expression. Limiting numbers of carrier-specific helper T cells from lines or clones were injected along with a source of B cells into lethally irradiated unprimed recipients. The incidence of lodging of the T cell lines in recipient spleens at 18 h was determined by autoradiography to be 1.5 to 4.3% of the injected cells. These T cells were necessary and sufficient for the generation of T-dependent B cell responses within splenic fragments cultured in vitro with specific antigen. A comparison of isotypic responses from splenic and Peyer's patch B cells generated with the same T cell population revealed that a high proportion of the response from Peyer's patch B cells consisted of IgA antibody exclusively (46-57%) while the percentage of such responses from splenic B cells was much lower (7-10%). Thus, the isotype pattern of the response reflected the B cell source. Experiments in which cloned hemocyanin-specific T cells provided help to T-depleted spleen cells within splenic fragments from athymic recipients indicated that a single specificity of helper T cell is both necessary and sufficient to support the generation of antibody responses consisting of multiple isotypes. Isotype-specific T cells do not appear to be required in this system.
Assuntos
Linfócitos B/imunologia , Isotipos de Imunoglobulinas/biossíntese , Cooperação Linfocítica , Linfócitos T Auxiliares-Indutores/imunologia , Animais , Linhagem Celular , Células Cultivadas , Quimiotaxia de Leucócito , Epitopos/imunologia , Hemocianinas/imunologia , Imunização Passiva , Camundongos , Camundongos Endogâmicos BALB C/imunologia , Camundongos Nus/imunologia , Nódulos Linfáticos Agregados/citologia , Nódulos Linfáticos Agregados/imunologia , Quimera por Radiação , Baço/citologia , Baço/imunologia , Baço/transplante , Linfócitos T Auxiliares-Indutores/transplanteAssuntos
Leucócitos Mononucleares/imunologia , Glândulas Mamárias Animais/imunologia , Animais , Antígenos de Diferenciação de Linfócitos T/análise , Antígenos Ly/análise , Antígenos de Superfície/análise , Ativação Linfocitária , Glândulas Mamárias Animais/citologia , Camundongos , Baço/citologia , Antígenos Thy-1 , Timo/citologiaRESUMO
The ontogeny of IL 2-responsive cells in the thymus of CBA/J mice was examined in neonatal animals and in fetuses at 14, 16, and 18 to 20 days gestation. The thymocytes were tested for responsiveness to 2 micrograms/ml Con A, TCGF, IL 2, and co-stimulation by Con A plus TCGF or IL 2. These responses were compared with those of thymocytes of 6- to 8-wk-old CBA/J. Thymocytes (1 X 10(5)) were cultured, and the reaction was measured at maximum response (96 hr). Neonatal animals gave an unusually high response to TCGF or partially purified IL 2 alone, approximately five times greater than the adult. A low but significantly enhanced proliferation, stimulated by partially purified IL 2 alone, was observed with 14-day fetal thymocytes, even though cultures of these cells in medium alone had higher background proliferation than any other age tested. In the co-stimulator reaction, proliferation significantly above background was measured at 16 days of gestation with Con A plus TCGF. The magnitude of the co-stimulator reaction increased with age, especially between the 16th and 18th day of gestation and immediately after birth.
Assuntos
Interleucina-2/fisiologia , Timo/embriologia , Animais , Divisão Celular/efeitos dos fármacos , Concanavalina A/farmacologia , Feminino , Camundongos , Camundongos Endogâmicos CBA , Gravidez , Timo/citologiaRESUMO
Previous work from our laboratory has indicated that the transplantation of pancreatic islets is a feasible approach to the problem of diabetes. A major obstacle to transplantation is presented by passenger leucocytes, which contaminate the preparations and can lead to the prompt rejection of fresh islets. We have extended our previous studies on the rejection of islet allografts by challenging transplanted animals with enriched lymphoid cell populations prepared from animals both syngeneic to the transplanted islets and third party. Rapid and complete rejection was observed when the challenge peritoneal exudate cell population was syngeneic with the transplanted islets; rejection was determined by both functional and histologic criteria. Peritoneal exudate cells from a third-party rat strain induced delayed and variable effects upon the function of the transplant. In contrast, splenic T-cells were capable of inducing rejection, regardless of the strain of origin, though the time course of T-cell-induced rejection was slower than that observed by syngeneic peritoneal exudate cells. Finally, splenic B-cells completely failed to induce rejection. Our data indicate that at least two mechanisms exist by which the rejection of islet allografts may be triggered. The first is a haplotype-specific mechanism initiated by a cell type present at high frequency in peritoneal exudate cells; these are probably macrophages. The second mechanism is initiated by immunocompetent T-cells; this mechanism shows no haplotype specificity. We suggest that both macrophages and T-cells must be considered when devising protocols for the removal of passenger leucocytes from allografts.
Assuntos
Rejeição de Enxerto , Transplante das Ilhotas Pancreáticas , Macrófagos/imunologia , Linfócitos T/imunologia , Animais , Líquido Ascítico/citologia , Linfócitos B/imunologia , Diabetes Mellitus Experimental/terapia , Ilhotas Pancreáticas/patologia , Fígado/patologia , Masculino , Ratos , Baço/citologia , Transplante HomólogoRESUMO
We describe the identification of a monoclonal antibody that recognizes a determinant on the delta chain of mice of the Iga, allotype groups. The monoclonal Ig in soluble form induces allotype-specific proliferation by splenic B lymphocytes from normal animals of these haplotypes. Spleen cells from mice bearing the X-linked defect of CBA/N mice fail to respond, although they bear the determinant. Proliferation is independent of T lymphocytes. The data indicate a direct triggering function for sIgD.
Assuntos
Linfócitos B/imunologia , Alótipos de Imunoglobulina/imunologia , Receptores de Antígenos de Linfócitos B/imunologia , Animais , Imunoglobulina D/imunologia , Imunoglobulina G/imunologia , Cadeias Pesadas de Imunoglobulinas/imunologia , Cadeias delta de Imunoglobulina/imunologia , Camundongos , Camundongos EndogâmicosRESUMO
Spleen cell cultures from young adult mice of a variety of strains were stimulated to incorporate tritiated thymidine ([3H]TdR) by a goat anti-mouse IgM antiserum and by purified anti-mu antibodies prepared from this serum. This stimulation was shown to depend upon the anti-mu activity of the antiserum. In addition, ultracentrifuged anti-mu and F(ab')2 fragments of anti-mu were shown to be stimulatory. The anti-mu preparation lacked detectable endotoxin contamination and was also shown to stimulate response by two strains (C57BL/10ScCr and C3H/HeJ) which are unresponsive to the mitogenic effects of endotoxin, while it failed to stimulate a response by cells from a mouse strain (CBA/N) which responds to endotoxin. In addition purified goat anti-mouse gamma, kappa antibodies and rabbit anti-mouse kappa-antib odies stimulated uptake of [3H]TdR by mouse spleen cells, although to a lesser degree than the anti-mu preparation. The cell density, culture requirements, and kinetics of the response are presented.
Assuntos
Anticorpos Anti-Idiotípicos , Cadeias Pesadas de Imunoglobulinas , Cadeias mu de Imunoglobulina , Ativação Linfocitária , Receptores de Antígenos de Linfócitos B , Animais , Anticorpos Anti-Idiotípicos/isolamento & purificação , Cromatografia de Afinidade , Relação Dose-Resposta Imunológica , Fragmentos Fab das Imunoglobulinas , Cadeias kappa de Imunoglobulina , Cinética , Camundongos , Polissacarídeos Bacterianos/farmacologia , Relação Estrutura-AtividadeRESUMO
An alloanti-delta antibody was prepared by immunizing C57BL/Ka mice with BALB/c spleen cells. Its specificity for delta-chain was demonstrated by immunoprecipitation and SDS-PAGE of 125I-labeled membrane proteins from BC8 spleen cells. BC8 mice possess C57BL/Ka "background" genes and BALB/c IgH genes. The anti-delta reagent without complement inhibited the primary in vitro anti-TNP antibody response to TNP-AECM-Ficoll by BC8 spleen cells, although it had no effect of the anti-TNP response of congenic C57BL/Ka spleen cells, which lack the delta-allotype identified by this antibody. On the other hand, the anti-delta antibody had no effect on the anti-TNP response of BC8 spleen cells to TNP-BA, except at limiting antigen concentrations. Both TNP-AECM-Ficoll and TNP-BA are T-I antigens, but they differ in that TNP-AECM-Ficoll fails to stimulate in vitro responses by immunologically defective CBA/N and neonatal spleen cells whereas TNP-BA can cause responses from both these animals. These results suggest that the IgD receptor is critical to T-I antibody responses initiated by TNP-AECM-Ficoll but that it is not required for T-I responses stimulated by TNP-BA.
Assuntos
Linfócitos B/imunologia , Receptores de Antígenos de Linfócitos B/fisiologia , Animais , Formação de Anticorpos , Antígenos , Epitopos , Imunoglobulina D/fisiologia , Camundongos , Camundongos Endogâmicos C57BLAssuntos
Linfócitos B/imunologia , Imunoglobulina D , Receptores de Antígenos de Linfócitos B , Animais , Antígenos , Membrana Celular/imunologia , Imunoglobulina D/análise , Terapia de Imunossupressão , Isoantígenos , Camundongos , Camundongos Endogâmicos CBA , Fenótipo , Receptores de Antígenos de Linfócitos B/análiseRESUMO
The operation of clonal selection for cells of the B-lymphocyte line is discussed with regard to: 1) The clonal repertoire determined by antigen binding to B lymphocytes, which is much larger than that determined by limiting dilution cloning assays. This quantitative difference is interpreted in terms of the multiple shared specificities of each antibody molecule. 2) Multiclonal responses and initial selection by antigen of particular clones (preferential primary selection). 3) Clonal dominance. During an immune response one clone (or a small number of clones) of B cells is preferentially selected and proliferated, apparently at random, from a heterogeneous population of cells capable of responding to the given antigen. Co-dominance of two or more clones simultaneously can be obtained by mixing selected clones. Secreted antibody is seen as playing a role in the establishment of clonal dominance. A model for clonal expansion is presented. The model attempts to explain the generation of memory and antibody secreting cells within each clonal expansion in terms of the ratio of two signals, one for proliferation and one for differentiation. The delivery of these signals is proposed to involve the receptor antibody-antigen interaction for proliferation and a self-recognition site interaction for differentiation.