RESUMO
BACKGROUND: The Strengths and Difficulties Questionnaire (SDQ) is widely used to measure emotional and behavioural problems in typically developing young people, although there is some evidence that it may also be suitable for children with intellectual disability (ID). The Developmental Behaviour Checklist - Parent version (DBC-P) is a measure of emotional and behavioural problems that was specifically designed for children and adolescents with an ID. The DBC-P cut-off has high agreement with clinical diagnosis. The aim of this study was to estimate the relationship between DBC-P and SDQ scores in a sample of children with ID. METHOD: Parents of 83 young people with ID aged 4-17 years completed the parent versions of the SDQ and the DBC-P. We evaluated the concurrent validity of the SDQ and DBC-P total scores, and the agreement between the DBC-P cut-off and the SDQ cut-offs for 'borderline' and 'abnormal' behaviour. RESULTS: The SDQ total difficulties score correlated well with the DBC-P total behaviour problem score. Agreement between the SDQ borderline cut-off and the DBC-P cut-off for abnormality was high (83%), but was lower for the SDQ abnormal cut-off (75%). Positive agreement between the DBC-P and the SDQ borderline cut-off was also high, with the SDQ borderline cut-off identifying 86% of those who met the DBC-P criterion. Negative agreement was weaker, with the SDQ borderline cut-off identifying only 79% of the participants who did not meet the DBC-P cut-off. CONCLUSION: The SDQ borderline cut-off has some validity as a measure of overall levels of behavioural and emotional problems in young people with ID, and may be useful in epidemiological studies that include participants with and without ID. However, where it is important to focus on behavioural profiles in children with ID, a specialised ID instrument with established psychometric properties, such as the DBC-P, may provide more reliable and valid information.
Assuntos
Escala de Avaliação Comportamental/normas , Sintomas Comportamentais/diagnóstico , Transtornos do Comportamento Infantil/diagnóstico , Deficiência Intelectual/diagnóstico , Escalas de Graduação Psiquiátrica/normas , Adolescente , Sintomas Comportamentais/etiologia , Lista de Checagem , Criança , Transtornos do Comportamento Infantil/etiologia , Pré-Escolar , Feminino , Humanos , Deficiência Intelectual/complicações , Masculino , Comportamento Problema , Reprodutibilidade dos TestesRESUMO
Engagement of the B7 family of molecules on antigen-presenting cells with their T cell-associated ligands, CD28 and CD152 (cytotoxic T lymphocyte-associated antigen-4 [CTLA-4]), provides a pivotal costimulatory signal in T-cell activation. We investigated the role of the CD28/CD152 pathway in psoriasis in a 26-week, phase I, open-label dose-escalation study. The importance of this pathway in the generation of humoral immune responses to T cell-dependent neoantigens, bacteriophage phiX174 and keyhole limpet hemocyanin, was also evaluated. Forty-three patients with stable psoriasis vulgaris received 4 infusions of the soluble chimeric protein CTLA4Ig (BMS-188667). Forty-six percent of all study patients achieved a 50% or greater sustained improvement in clinical disease activity, with progressively greater effects observed in the highest-dosing cohorts. Improvement in these patients was associated with quantitative reduction in epidermal hyperplasia, which correlated with quantitative reduction in skin-infiltrating T cells. No markedly increased rate of intralesional T-cell apoptosis was identified, suggesting that the decreased number of lesional T cells was probably likely attributable to an inhibition of T-cell proliferation, T-cell recruitment, and/or apoptosis of antigen-specific T cells at extralesional sites. Altered antibody responses to T cell-dependent neoantigens were observed, but immunologic tolerance to these antigens was not demonstrated. This study illustrates the importance of the CD28/CD152 pathway in the pathogenesis of psoriasis and suggests a potential therapeutic use for this novel immunomodulatory approach in an array of T cell-mediated diseases.
Assuntos
Antígenos de Diferenciação/uso terapêutico , Imunoconjugados , Ativação Linfocitária , Psoríase/terapia , Linfócitos T/imunologia , Abatacepte , Adulto , Formação de Anticorpos , Antígenos CD , Antígenos de Diferenciação/sangue , Antígeno CTLA-4 , Estudos de Coortes , Relação Dose-Resposta Imunológica , Feminino , Humanos , Imuno-Histoquímica , Queratinócitos/metabolismo , Queratinócitos/patologia , Masculino , Pessoa de Meia-Idade , Psoríase/imunologia , Resultado do TratamentoRESUMO
Immunotoxins have shown promise as antitumor agents in clinical trials. However, they have not become part of standard cancer therapy because of factors that include their inherent immunogenicity, which limits the duration of therapy. To address this issue, we evaluated in preclinical models the concomitant use of the immunosuppressive agent CTLA4Ig and BR96 sFv-PE40, a single-chain immunotoxin that binds to carcinoma cells expressing Le(y). Cotreatment with CTLA4Ig, an inhibitor of the CD28/CTLA4-CD80/CD86 costimulation pathway, blocked the production of Abs against BR96 sFv-PE40 in immunocompetent rodents and dogs. It also blocked hypersensitivity reactions in rats carrying colon carcinoma allografts during a second course of BR96 sFv-PE40 therapy, and the cotreatment with CTLA4Ig resulted in enhanced antitumor activity. Cotreatment with CTLA4Ig also prevented hypersensitivity reactions induced by repeat dosing of BR96 sFv-PE40 (q3dx5) in dogs. The production of anti-BR96-sFv-PE40 Abs was decreased in CTLA4Ig-cotreated rodents and dogs resulting in increased plasma levels of BR96 sFv-PE40 relative to non-CTLA4Ig-cotreated animals. These data show that cotreatment of immunotoxins with CTLA4Ig, by inhibiting the production of anti-immunotoxin Abs, can extend the duration of BR96 sFv-PE40 therapy to give greater exposure, reduced toxicities, and increased efficacy.
Assuntos
Adjuvantes Imunológicos/administração & dosagem , Anticorpos Anti-Idiotípicos/biossíntese , Antígenos de Diferenciação/imunologia , Antineoplásicos/imunologia , Imunoconjugados , Fragmentos Fc das Imunoglobulinas/administração & dosagem , Imunossupressores/administração & dosagem , Imunotoxinas/imunologia , Abatacepte , Animais , Anticorpos Monoclonais , Antígenos CD , Antígenos de Diferenciação/administração & dosagem , Antineoplásicos/administração & dosagem , Antineoplásicos/farmacologia , Antígeno CTLA-4 , Carcinoma , Neoplasias do Colo , Cães , Feminino , Humanos , Imunotoxinas/administração & dosagem , Imunotoxinas/metabolismo , Imunotoxinas/farmacologia , Injeções Intraperitoneais , Injeções Intravenosas , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Ratos , Ratos Endogâmicos BN , Ratos Endogâmicos WF , Proteínas Recombinantes de Fusão/administração & dosagem , Proteínas Recombinantes de Fusão/imunologia , Proteínas Recombinantes de Fusão/metabolismo , Proteínas Recombinantes de Fusão/farmacologiaRESUMO
G28-5 sFv-PE40 is a single-chain immunotoxin targeted to CD40, which is highly expressed on human hematologic malignancies, including non-Hodgkin's lymphoma, B-lineage leukemias, multiple myeloma, and Hodgkin's disease, as well as certain carcinomas. In vitro analysis showed that this monovalent immunotoxin had a binding affinity of 3 nmol/L, within 15-fold of the bivalent parental monoclonal antibody. G28-5 sFv-PE40 was stable when incubated in mouse serum at 37 degrees C for 6 hours and cleared from the circulation of mice with a half-life of 16.7 minutes. This immunotoxin was effective in treating human Burkitt's lymphoma xenografted SCID mice with complete responses, defined by an asymptomatic phenotype for greater than 120 days, obtained at doses of 0.13 to 0.26 mg/kg. The efficacy of treatment was dependent on the schedule used, with every three days for five injections being the most effective tested. The toxicity of G28-5 sFv-PE40 was examined in SCID mice, rats, and monkeys, with the maximum tolerated dose being 0.48, 1.0, and 1.67 mg/kg, respectively. Comparative immunohistology showed that the G28-5 specificity was qualitatively similar between human and monkey tissue. In summary, G28-5 sFv-PE40 was effective at inducing complete antitumor responses in lymphoma xenografted mice at doses that were well tolerated in mice, rats, and monkeys.
Assuntos
Antígenos de Neoplasias/imunologia , Antineoplásicos/uso terapêutico , Linfoma de Burkitt/tratamento farmacológico , Antígenos CD40/imunologia , Imunotoxinas/uso terapêutico , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/farmacocinética , Anticorpos Monoclonais/uso terapêutico , Anticorpos Monoclonais/toxicidade , Antineoplásicos/imunologia , Antineoplásicos/farmacocinética , Antineoplásicos/toxicidade , Linfoma de Burkitt/imunologia , Esquema de Medicação , Exotoxinas , Feminino , Humanos , Imunotoxinas/imunologia , Imunotoxinas/farmacocinética , Imunotoxinas/toxicidade , Macaca fascicularis , Masculino , Camundongos , Camundongos SCID , Transplante de Neoplasias , Ratos , Ratos Sprague-Dawley , Especificidade da Espécie , Distribuição Tecidual , Transplante HeterólogoRESUMO
The toxicity of BMS-182248, an immunoglobulin (cBR96)-cytotoxic drug (doxorubicin) conjugate, was investigated in Sprague-Dawley rats at single intravenous doses of 508, 1,200, and 2,550 mg/m2 (conjugated doxorubicin doses of 14.7, 34.8, and 74 mg/m2, respectively) and compared to that obtained from administration of free doxorubicin at single doses of 33.6 and 72 mg/m2 (approximately equivalent to that contained in the 1,200- and 2,550-mg/m2 doses of BMS-182248, respectively). Necropsies were conducted on day 8, upon death/moribund sacrifice, or after an approximate 3-mo observation period following completion of treatment. Death/moribundity of all rats that received 72 mg/m2 and of 9 of 20 rats given 33.6 mg/m2 free doxorubicin were attributed primarily to delayed cardiotoxicity and glomerulonephropathy. With BMS-182248, death from glomerulonephropathy and cardiotoxicity occurred in only 4 of 20 rats given 2,550 mg/m2 (74 mg/m2 doxorubicin equivalent). No deaths or cardiotoxicity occurred in rats given 508 or 1,200 mg/m2 BMS-182248. Additional effects noted with either drug included testicular atrophy, axonal degeneration of sciatic nerve and nerve tracts of brain and spinal cord, teeth (incisor) abnormalities, thymic atrophy, bone marrow hypocellularity, splenic lymphoid and red-pulp depletion, and increased extramedullary hematopoiesis in the spleen and liver. Also noted were altered chief cells in the stomach, vacuolation of adrenal gland and corpora lutea in the ovary, uterine and seminal vesicle atrophy, ulceration and myocyte regeneration/degeneration in the tongue, increased osteoclasts and osteoblasts in bone, and lymphoid hyperplasia of mandibular lymph node. In general, these effects were more severe in doxorubicin-treated rats. All changes observed with BMS-182248 were considered primarily due to the effects of doxorubicin and were substantially less severe (most notably cardiotoxicity) compared to those produced by an equivalent amount of doxorubicin.
Assuntos
Anticorpos Monoclonais/toxicidade , Antineoplásicos/toxicidade , Doxorrubicina/toxicidade , Coração/efeitos dos fármacos , Imunotoxinas/toxicidade , Animais , Peso Corporal/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Feminino , Rim/efeitos dos fármacos , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
The purpose of this research was to determine whether prostaglandin E2 (PGE2), a major product of macrophages which can kill certain murine B cell lymphomas, induces death by a necrotic mechanism or by an alternate pathway called apoptosis. CH31 is a phenotypically "immature" B cell lymphoma which resembles immature neonatal B cells in its susceptibility to killing by reagents which cross-link surface immunoglobulin (sIg). In the present study we first show that PGE2, but not the closely related prostanoid, PGF2 alpha, kills CH31 lymphoma cells. In contrast, CH12, a phenotypically "mature" lymphoma which is not negatively affected by sIg cross-linking, is not induced to die after exposure to PGE2. Agarose gel electrophoresis demonstrated that the DNA of PGE2-treated CH31, but not CH12 cells, is cleaved into characteristic 200 base pair oligonucleosomal fragments indicative of an apoptotic mechanism of death. However, a necrotic form of death, indicated by random DNA cleavage which produces a smear following electrophoresis, could be induced by treatment of CH12 or CH31 with anti-class II MHC antibodies and complement. The apoptotic mechanism of CH31 cell killing by PGE2 was confirmed using scanning electron microscopy which demonstrated the unique membrane blebbing and bubbling pathognomonic of this form of death. Finally, using a recently devised flow cytometric method to study apoptosis in heterogeneous cell populations, we compared the ability of anti-IgM, PGE2, or PGF2 alpha to induce apoptosis in B lymphocytes from neonatal or adult mice. Anti-IgM, and to a lesser extent PGE2, but not PGF2 alpha, induces apoptosis in a fraction of neonatal B cells. None of these treatments induced cell death in B lymphocytes from mature mice. Overall, these observations suggest that PGE-secreting cells such as macrophages, which inhabit the B cell microenvironments of lymphoid organs, may eliminate a subset of immature B lymphocytes and may be important in controlling the spread of PGE-sensitive malignant B lymphoma cells.
Assuntos
Linfócitos B/citologia , Dinoprostona/farmacologia , Linfoma de Células B/patologia , Animais , Subpopulações de Linfócitos B/citologia , Morte Celular/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Dinoprosta/farmacologia , Humanos , Camundongos , Células Tumorais CultivadasAssuntos
Anticorpos , Antígenos CD/fisiologia , Divisão Celular , Imunoglobulina M/imunologia , Linfoma de Células B/imunologia , Fator de Crescimento Transformador beta/farmacologia , Animais , Antígenos CD/imunologia , Antígenos CD5 , Divisão Celular/efeitos dos fármacos , Cinética , Camundongos , Células Tumorais CultivadasRESUMO
Cross-linking of B-cell membrane immunoglobulin (Ig) receptors induces growth arrest at G1-S, leading to apoptosis and cell death in the immature lymphomas WEHI-231 and CH31, but not in the CH12 B-cell line. In this system, which has been used as a model for B-cell tolerance, we have established that these lymphomas produce active transforming growth factor beta (TGF-beta) when treated with anti-Ig and that their hierarchy of sensitivity to TGF-beta generally correlates with their growth inhibition by anti-Ig. TGF-beta, in turn, has been shown to interfere with the phosphorylation of the retinoblastoma gene product, pRB. Herein, we also demonstrate that in WEHI-231 B-lymphoma cells treated with anti-Ig for 24 h, the pRB protein is found to be predominantly in the underphosphorylated form, as previously reported for cells arrested by the exogenous addition of TGF-beta. However, neutralizing antibodies to TGF-beta failed to prevent growth inhibition by anti-Ig in WEHI-231 and CH31. When WEHI-231 lymphoma cells were selected for growth in TGF-beta, the majority of the TGF-beta-resistant clones remained sensitive to anti-Ig-mediated growth inhibition. In these clones, the retinoblastoma gene product was found to be in the underphosphorylated form after 24-h treatment with anti-Ig, but not with TGF-beta. These data show that anti-Ig treatment of murine B-cell lymphomas stimulates the production of active TGF-beta but that a TGF-beta-independent pathway may be responsible for the pRB underphosphorylation and cell cycle blockade.
Assuntos
Anticorpos Anti-Idiotípicos , Proteína do Retinoblastoma/metabolismo , Fator de Crescimento Transformador beta/fisiologia , Animais , Morte Celular/efeitos dos fármacos , Morte Celular/fisiologia , Divisão Celular/efeitos dos fármacos , Linfoma de Células B/tratamento farmacológico , Linfoma de Células B/fisiopatologia , Camundongos , Fosforilação/efeitos dos fármacos , Células Tumorais CultivadasRESUMO
Ly-1+ B cells have been reported to produce a number of autoantibodies, and to be involved in the selection and regulation of the conventional B cell repertoire. It is not known if these B cells, which are found in high numbers in the peritoneum of normal adult mice, themselves can be regulated. In this study, we evaluated the sensitivity of peritoneal B cells (PBCs) versus conventional splenic B cells to regulation in a model system for tolerance. Normal splenic (conventional) or PBCs (containing both CD5+ and CD5- 'sister' cells) were cultured overnight with either F(ab')2 or intact IgG anti-mouse Ig, washed, and then challenged with fluorescein(FL)-coupled to Brucella abortus (BA), trimethylammonium (TMA)-BA or lipopolysaccharide (LPS), and the IgM responses to the FL and TMA haptens measured. In contrast to spleen cells, which exhibited up to a 90% reduction in anti-FL responsiveness, pretreated PBCs were mostly resistant to this form of tolerance regardless of challenge. The anti-TMA response of PBCs, which reflects the skewed VH11 usage by peritoneal CD5 B cells, was also resistant to tolerance. However, splenic TMA-specific B cells appeared to be sensitive to unresponsiveness induced by anti-Ig. Signaling studies show that PBCs have a blunted initial Ca2+ response, suggesting that the consequence of anti-Ig crosslinking may be defective in these cells. Furthermore, phorbal myristate acetate and/or ionomycin treatment of both PB and splenic B cells led to hyporesponsiveness to LPS challenge. This suggests that PBCs may be defective in a signalling pathway, perhaps involving protein kinase C activation.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Líquido Ascítico/imunologia , Subpopulações de Linfócitos B/imunologia , Tolerância Imunológica , Animais , Líquido Ascítico/citologia , Subpopulações de Linfócitos B/efeitos dos fármacos , Subpopulações de Linfócitos B/metabolismo , Cálcio/metabolismo , Imunoglobulina M/biossíntese , Ionomicina/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos , Baço/citologia , Baço/imunologia , Acetato de Tetradecanoilforbol/farmacologiaRESUMO
Overnight exposure of adult splenic B cells to anti-Ig, a surrogate for antigen/tolerogen, can result in a hyporesponsive state in terms of antibody synthesis. Since B cells treated with either intact of F(ab')2 fragments of anti-Ig will exit the G0 phase of the cell cycle and enter G1 or S, respectively, we examined which steps in B-cell activation were required for this form of hyporesponsiveness. We found that B-cell hyporesponsiveness could be induced under conditions leading to either abortive or productive B-cell cycle progression, depending on the immunogenic challenge employed. Thus, PMA + ionomycin, concanavalin A, PMA alone, or ionomycin alone induced hyporesponsiveness. Each of these reagents is able to drive B-cell exit from G0 into G1 and cause class II hyperexpression. We next examined the effect of cyclosporin A (CSA), a reagent that blocks anti-Ig but not by PMA-induced class II hyperexpression. Interestingly, CSA only interfered with the induction of B-cell hyporesponsiveness with anti-Ig. These results suggest that upregulation of MHC class II may be coincident with a CSA-sensitive tolerance pathway in B cells stimulated by anti-Ig. Finally, IL-4 pretreatment was found to ablate hyporesponsiveness induced by either intact anti-Ig or PMA. These results parallel the Fc-dependent induction of hyporesponsiveness reported earlier (G. Warner and D. W. Scott, J. Immunol. 146, 2185, 1991). We propose that crosslinking of surface Ig, leading to cell cycle progression out of G0 as well as class II hyperexpression, in the absence of a cognate T cell signal, leads to B-cell hyporesponsiveness.
Assuntos
Linfócitos B/imunologia , Antígenos de Histocompatibilidade Classe II/análise , Tolerância Imunológica , Fase de Repouso do Ciclo Celular , Animais , Antígenos de Diferenciação/fisiologia , Concanavalina A , Ciclosporina/farmacologia , Interleucina-4/farmacologia , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos DBA , Modelos Biológicos , Proteína Quinase C/fisiologia , Receptores de Antígenos de Linfócitos B/fisiologia , Receptores Fc/fisiologia , Receptores de IgG , Acetato de Tetradecanoilforbol/farmacologia , Regulação para CimaRESUMO
We have developed a simple and adaptable, polyclonal model for B cell nonresponsiveness that is based on the inhibitory activity of anti-Ig as a surrogate for Ag. In our system the induction phase (treatment with anti-Ig) is separated from the challenge phase (Ag or mitogen), so that the critical events in each phase can be evaluated. Our results show that T cell-depleted B cells precultured for 18 to 24 h with rabbit anti-Ig reagents are rendered unresponsive to challenge with either Ag, fluorescein coupled to Brucella abortus (FL-BA), or mitogen (LPS). This state of nonresponsiveness (anergy) is reflected by an inhibition of a prototype response to the fluorescein hapten, as well as total Ig and IgG synthesis, but no reduction in proliferation to LPS. Interestingly, mitogen-induced polyclonal antibody formation was consistently reduced by 90% by treatment with either F(ab')2 or intact IgG anti-Ig. In contrast, the Ag-driven (FL-BA) response of pretreated B cells was inhibited by only 50 to 70%. Moreover, the latter effect usually required pretreatment with intact IgG anti-Ig, a result that suggests the importance of an Fc-dependent negative signal affecting the B cell's response to FL-BA. Furthermore, pretreatment and coculture of B cells with IL-4 blocked the Fc-dependent inhibition of the FL-BA responsiveness. These results, as well as kinetics experiments establishing a 4-h latent period, suggest that simple blocking of surface Ig receptor on target B cells is not responsible for the induction of anergy. Pretreated B cells displayed unique phenotypic changes after treatment with anti-Ig, including a diminution of Thy-1 expression in response to LPS + IL-4, as well as a reduction in membrane IgM and J11d expression (i.e., they were IgMlo, IgDmed, and J11dlo, as recently reported for anergic B cells in transgenic mice). These results suggest that B cell anergy can be induced in mature B cells by both Fc-dependent and Fc-independent processes that lead to unique phenotypic changes and may reflect egress from G0 in the absence of T cell help. The significance of these changes to tolerance mechanisms is discussed.
Assuntos
Subpopulações de Linfócitos B/imunologia , Tolerância Imunológica , Receptores de Antígenos de Linfócitos B/imunologia , Receptores Fc/imunologia , Animais , Formação de Anticorpos , Complexo Antígeno-Anticorpo/imunologia , Antígenos de Superfície/imunologia , Subpopulações de Linfócitos B/citologia , Interleucina-4/farmacologia , Lipopolissacarídeos/farmacologia , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos , Receptores de Antígenos de Linfócitos B/fisiologia , Baço/citologia , Antígenos Thy-1 , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
PG of the E series are generally known to suppress immune responses, however, we have found that PGE synergizes with IL-4 to induce IgE and IgG1 production in LPS-stimulated murine B lymphocytes. PGE2 and PGE1 (10(-6) to 10(-8) M) significantly increase IgE and IgG1 production (up to 26-fold) at all concentrations of IL-4 tested. In addition to its effects on IgE and IgG1, PGE also causes a significant decrease in IgM and IgG3 synthesis, suggesting that PGE may promote IL-4-induced class switching. The specificity of the E series PG effect is demonstrated by the fact that PGF2 alpha (10(-6) M) does not alter production of any of these isotypes. Because PGE can mediate its effects through cAMP in some cases, we investigated the importance of cAMP levels in regulation of isotype expression. Other agents that increase intracellular cAMP levels (cholera toxin and dibutyryl cAMP) were assessed for their ability to regulate isotype differentiation. Cholera toxin (100 pg/ml) and dibutyryl cAMP (100 microM) significantly enhanced IgE and IgG1 production and diminished IgM and IgG3 synthesis. We also show that PGE and cholera toxin elevate intracellular cAMP in B lymphocytes in a dose-dependent manner. In contrast, PGF2 alpha (10(-6) M) and the B subunit of cholera toxin (100 pg/ml) did not increase cAMP and did not regulate the isotype of Ig produced, reiterating the importance of cAMP in enhancing isotype differentiation. Although PGE is known to inhibit a number of immune responses, our data show that it is not always inhibitory. PGE may play a role in atopy in vivo where PGE-secreting cells such as macrophages, follicular dendritic cells, and fibroblasts can promote IgE synthesis. This research emphasizes the importance of PGE in regulation of the humoral immune response and adds a new stimulatory action to the repertoire of known PGE effects.
Assuntos
Linfócitos B/metabolismo , Dinoprostona/farmacologia , Imunoglobulina E/biossíntese , Imunoglobulina G/biossíntese , Interleucina-4/farmacologia , Alprostadil/farmacologia , Animais , Toxina da Cólera/farmacologia , AMP Cíclico/fisiologia , Dinoprosta/farmacologia , Isotipos de Imunoglobulinas/biossíntese , Imunoglobulina M/biossíntese , Ativação Linfocitária/efeitos dos fármacos , Masculino , Camundongos , Baço/citologiaRESUMO
ECH408-1 is a murine B cell lymphoma expressing idiotypically and allotypically distinguishable transfected and endogenous IgD. Previously, we demonstrated that this cell line was not growth inhibited by antibodies directed at membrane IgD, but could be inhibited by antibodies which crosslink membrane IgM. Herein, we demonstrate that both anti-mu and anti-delta will cause calcium mobilization in this transfected cell line; this is followed by a period during which antibodies against the alternative isotype are unable to induce significant increases in intracellular calcium concentrations. This phenomenon, called "desensitization," is short-lived, lasting 20 min. We further demonstrate that acute desensitization of these cells by anti-delta has no effect on immediate growth inhibition which is elicited by anti-mu. These data confirm our earlier proposal that the rapid, initial calcium response seen in these lymphomas is not required for the negative signal for growth. Moreover, we also demonstrate that pretreatment of these lymphoma cells with phorbol myristate acetate (PMA) also renders these lymphoma cells temporarily incapable of manifesting a significant calcium signal. Nonetheless, PMA-pretreated B lymphoma cells are not altered in their subsequent sensitivity to anti-mu growth inhibition, nor are they affected in their resistance to inhibition by anti-delta. Our data confirm the proposal that neither the calcium signal nor protein kinase-C activation is involved in the modulation of B lymphoma growth.
Assuntos
Linfócitos B/imunologia , Tolerância Imunológica , Ativação Linfocitária , Linfoma não Hodgkin/imunologia , Receptores de Antígenos de Linfócitos B/fisiologia , Animais , Anticorpos Anti-Idiotípicos , Cálcio/fisiologia , Regulação para Baixo , Imunoglobulina D/imunologia , Imunoglobulina M/imunologia , Ativação Linfocitária/efeitos dos fármacos , Camundongos , Agregação de Receptores , Receptores de Antígenos de Linfócitos B/imunologia , Transdução de Sinais , Acetato de Tetradecanoilforbol/farmacologia , TransfecçãoRESUMO
We have observed that a 2-h pretreatment of murine B cells with cholera toxin (CT) renders the B cell incapable of receiving an activation signal via surface Ig as measured by cell volume increase and entry into the S phase of the cell cycle. In contrast, CT pretreatment does not inhibit the delivery of a signal by IL-4, as measured by increase in cell volume. In fact, CT pretreated B cells are able to respond to anti-Ig in the presence of IL-4, as measured by both an increase in cell size and entry into S suggesting that IL-4 overcomes the effects of CT on normal B cell activation. Despite blocking the anti-Ig-mediated entry into the cell cycle, CT was not able to interfere with the induction of nonresponsiveness by anti-Ig in normal B cells or with the delivery of growth-inhibitory signal to the B cell lymphoma WEHI-231. These results suggest that there are two signaling pathways mediated by cross-linking of surface Ig: one pathway sensitive and the other insensitive to modulation by CT.
Assuntos
Linfócitos B/fisiologia , Toxina da Cólera/farmacologia , Receptores de Antígenos de Linfócitos B/fisiologia , Transdução de Sinais , Animais , Anticorpos Anti-Idiotípicos/fisiologia , Linfócitos B/imunologia , Linfócitos B/metabolismo , Cálcio/fisiologia , Linhagem Celular , Antígenos de Histocompatibilidade Classe II/análise , Fosfatos de Inositol/biossíntese , Interleucina-4 , Interleucinas/farmacologia , Cinética , Lipopolissacarídeos/farmacologia , Ativação Linfocitária , Linfoma/imunologia , CamundongosRESUMO
The ability of prostaglandins (PG) to inhibit the growth of B cell lymphomas was investigated. Macrophage-secreted PGE2 was previously shown to promote unresponsiveness to antigen in normal B lymphocytes. This observation suggested that B lymphomas might also be regulated by prostanoids. Five non-PG-secreting Ly-1+ B lymphomas (CH12, CH31, CH33, NBL and WEHI-231) were incubated for 24-72 h with PGE2, PGE1 or PGF2 alpha. The level of lymphoma growth at the end of culture was determined using a colorimetric assay which detects only viable cells. A marked heterogeneity was observed with respect to the sensitivity of these lymphomas to PGE2 and PGE1. CH31 was very sensitive, being growth inhibited by as little as 10(-8) M PGE. In contrast, CH12, a more mature lymphoma, was highly resistant, whereas CH33, NBL and WEHI-231 were of intermediate resistance. All five lymphomas demonstrated little or no growth inhibition when cultured with PGF2 alpha. Moreover, unlike PGE2, PGF2 alpha failed to elevate intracellular cAMP levels. It was previously shown that CH31, CH33 and WEHI-231 could be growth inhibited by anti-immunoglobulin antibodies which cross-link surface immunoglobulin. Interestingly, these three lymphomas were rendered more sensitive to this treatment if PGE2 was present. For example, 10(-8) M PGE2 alone had little effect on CH33, but significantly augmented growth inhibition induced by suboptimal quantities of anti-immunoglobulin antibody. Cholera toxin, another agent which was found to rapidly elevate intracellular cAMP levels, also synergized with suboptimal doses of anti-immunoglobulin to induce growth inhibition. Overall these data suggest that, in vivo, macrophage-secreted prostanoids may slow the growth of some lymphomas and that anti-immunoglobulin or anti-idiotype treatment may be more effective in the presence of agents which elevate cAMP such as E-series PG.
Assuntos
Inibidores do Crescimento , Leucemia Linfocítica Crônica de Células B/patologia , Prostaglandinas E/farmacologia , Animais , Linfócitos B , Divisão Celular/efeitos dos fármacos , Toxina da Cólera/farmacologia , AMP Cíclico/fisiologia , Dinoprosta/farmacologia , Esquema de Medicação , Sinergismo Farmacológico , Técnicas In Vitro , Camundongos , Receptores de Antígenos de Linfócitos B/fisiologia , Células Tumorais CultivadasRESUMO
The CH family of murine B-cell lymphomas includes several members that are sensitive to growth inhibition when their membrane IgM (mIgM) receptors are cross-linked by anti-mu chain, anti-kappa chain, or anti-idiotypic antibodies. These lymphomas are IgM+, Ia+, and IgD +/- and resemble neonatal B cells in terms of their exquisite sensitivity to anti-IgM-mediated negative signaling as a model for tolerance induction. In this report, we describe the properties of one of these lymphomas, CH33, which had been transfected with a construct containing an allotypically different delta chain constant region and the heavy chain variable region fragment from S107 (T15 idiotype positive). This transfected cell line allowed us to investigate the possibility that membrane IgD (mIgD) and mIgM can mediate different signals. Our results show that the transfected cells retained their exquisite sensitivity to anti-IgM-mediated growth inhibition; however, crosslinking of IgD with anti-delta chain antibody did not inhibit their growth. Furthermore, even prolonged pretreatment with anti-IgD antibodies did not affect cell growth nor did it modulate the inhibitory effects of anti-IgM antibody. Moreover, identical results were obtained with clones of CH33 that express significant amounts of endogenous IgD. Thus, the failure of mIgD to deliver a negative signal does not reflect a defect in the transfected IgD but appears to be a general property of IgD in these cells. The mIgD was shown to mediate transmembrane signals because anti-delta chain treatment resulted in Ca2+ mobilization in transfected CH33 cells and capping of those receptors. We conclude that mIgD can mediate qualitatively different signals than mIgM can and that mIgD expression per se is not sufficient to change the functional phenotype of immature B cells.
Assuntos
Imunoglobulina D/metabolismo , Imunoglobulina M/metabolismo , Animais , Linfócitos B , Linhagem Celular , Células Cultivadas , Citometria de Fluxo , Linfoma/imunologia , Camundongos , FenótipoRESUMO
WEHI-231, CH33, and CH31 are B-cell lymphomas that are inhibited in their growth by crosslinking of surface Ig receptors during early G1. This "negative signaling" process can be prevented or reversed under certain conditions. In the present paper, we use a cell synchronization procedure to demonstrate that activation of protein kinase C (PKC) is not involved in the negative signal for growth, but rather that PKC activation prevents growth inhibition when present early in the cell cycle. Indeed, the prevention of negative signaling is only accomplished by active phorbol esters. Moreover, phorbol esters and a calcium ionophore fail to deliver a negative signal under conditions in which anti-Ig can significantly prevent cell cycle progression into S phase, thereby ruling out synergy between PKC and calcium in growth inhibition. Whether phorbol esters reverse negative signaling by preventing internalization of the immune complex or phosphorylation of a critical intracellular protein is discussed.