RESUMO
Agammaglobulinemia induced in chickens by surgical bursectomy and irradiation at hatching is transmissible at 4 mo of age to sublethally irradiated normal chickens by bone marrow transplantation. The immunodeficiency which develops in the recipients persists for life and is characterized by inability to produce antibody and by progressive loss of detectable serum IgM and IgG.
Assuntos
Agamaglobulinemia/transmissão , Células da Medula Óssea , Transplante de Medula Óssea , Agamaglobulinemia/etiologia , Agamaglobulinemia/imunologia , Animais , Formação de Anticorpos , Medula Óssea/imunologia , Bolsa de Fabricius/imunologia , Galinhas , Hemocianinas/imunologia , Imunidade Materno-Adquirida , Imunização Passiva , Imunoglobulina G/análise , Imunoglobulina M/análise , Terapia de Imunossupressão , Moluscos/imunologia , Quimera por Radiação , Transplante HomólogoRESUMO
The direct effects of steroid hormones on the production of immunoglobulins and DNA synthesis by human T and B lymphocytes was evaluated in cultures of peripheral blood mononuclear cells. As detected by a reverse hemolytic plaque assay, the addition of 0.1 mM to 10 nM hydrocortisone to lymphocytes in culture in the absence of other stimulants or mitogens, resulted in the dramatic induction of immunoglobulin production with responses comparable to those seen in similar cultures stimulated with pokeweed mitogen. Steroid-stimulated immunoglobulin production was first seen after 48 h and peaked at 8-10 d of culture. The production of IgG, IgA, and IgM was induced following incubation with steroid. Glucocorticoids, but not estrogens or androgens, were capable of mediating this effect, and only compounds with affinity for the glucocorticoid receptor were active. The induction of immunoglobulin production was dependent on both T cells and monocytes; cultures depleted of either cell type did not produce immunoglobulin when stimulated with glucocorticoid hormones. Proliferation of B cells or T cells could not be detected by [3H]thymidine incorporation or total cell recovery from steroid-stimulated cultures, even though such cultures demonstrated marked increases in immunoglobulin production. The mechanism responsible for this functional maturation of B cells to become high rate immunoglobulin producing cells is as yet undefined, although it appears to involve more than merely steroid mediated inactivation of suppressor T cells.
Assuntos
Linfócitos B/efeitos dos fármacos , Glucocorticoides/farmacologia , Imunoglobulinas/biossíntese , Linfócitos T/efeitos dos fármacos , Adulto , Linfócitos B/metabolismo , Células Cultivadas , Técnica de Placa Hemolítica , Humanos , Hidrocortisona/farmacologia , Cinética , Cooperação Linfocítica , Mitógenos/farmacologia , Linfócitos T/efeitos da radiação , Fatores de TempoRESUMO
The functional maturity of T and B lymphocyte populations from human newborns was evaluated using a reverse hemolytic plaque assay to detect immunoglobulin-secreting cells generated in in vitro cultures stimulated with pokeweed mitogen (PWM), a T cell-dependent polyclonal activator, and the Epstein-Barr virus (EBV), a T cell-independent B cell activator. Cord blood lymphocytes failed to produce immunoglobulin in response to PWM, but did respond with immunoglobulin synthesis to stimulation with EBV. Co-culture experiments demonstrated that cord blood T cells would inhibit immunoglobulin production by adult cells stimulated with PWM, but not with EBV. Cord blood T cells did suppress immunoglobulin production by cord blood B cells when stimulated with a mixture of EBV and PWM, indicating that cord blood, in contrast to adult blood, contains a population of suppressor T cell precursors that are easily activated by PWM. Irradiation of the cord blood T cells with 2,000 rad eliminated the suppressor activity and revealed normal helper function for immunoglobulin (Ig) G, A, and M when these T cells were co-cultured with adult B cells. Cord blood B cells co-cultured with adult T cells or irradiated cord blood T cells did produce immunoglobulin in response to PWM, but the response was significantly lower than that of adult B cells, and only IgM was produced in these cultures. These studies demonstrate that both the T and B cells of the human newborn have significant functional differences compared with the functions of T and B lymphocyte populations in adults.
Assuntos
Formação de Anticorpos , Linfócitos B/imunologia , Sangue Fetal/imunologia , Linfócitos T/imunologia , Herpesvirus Humano 4/imunologia , Humanos , Tolerância Imunológica , Imunoglobulina A/biossíntese , Imunoglobulina G/biossíntese , Imunoglobulina M/biossíntese , Cooperação Linfocítica , Mitógenos , Linfócitos T Reguladores/imunologiaRESUMO
Vitronectin (VN) is an extracellular matrix (ECM) protein, the synthesis of which in vivo by glioma cells correlates with tumor grade. Although the role of VN as a permissive substrate for glioma migration has been well characterized, its role in conferring a survival advantage for tumor cells has not been addressed previously. By using an in vitro assay of DNA fragmentation as a quantitative measure of apoptotic cell death, we sought to determine whether the sensitivity of two human glioma cell lines (D54 and U251) to drug-induced apoptosis could be inhibited by VN. As well, the extent to which apoptosis could be inhibited was correlated with the levels of the Bcl-2 family of proteins that are known to modulate apoptosis and chemoresistance. Results of the study were: (a) VN coatings, in a dose-dependent manner, inhibited topoisomerase (Topo)-induced apoptosis by up to 50% (optimal coating density, 500 ng/cm2); in contrast, fibronectin (FN), an ECM protein present in abundance in the brain, demonstrated no protection; (b) in a dose-response study, VN clearly conferred a survival advantage (LD50 of Topo: on VN, 120 ng/ml; on FN, 35 ng/ml); (c) the protective effect of VN was not due to enhanced cell adhesion or alterations in the cell cycle distribution; (d) both of the classic integrin receptors that bind VN (alpha(v)beta3, alpha(v)beta5) were capable of mediating this protective effect, because ligation of either of the two classic integrins conferred chemoresistance to Topo; and (e) chemoresistance observed with VN was associated with an increase in expression of two antiapoptotic proteins, Bcl-2 and Bcl-X(L), with a consequent increase in the ratios for Bcl-2:Bax and Bcl-X(L):Bax. VN, an ECM protein preferentially expressed at the tumor-brain interface in vivo, may confer a survival advantage to glioma cells at the advancing tumor margin and may thus, in part, underlie the high level of tumor recurrence at this interface.
Assuntos
Apoptose , Glioma/patologia , Vitronectina/fisiologia , Anticorpos Monoclonais/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Fibronectinas/farmacologia , Humanos , Marcação In Situ das Extremidades Cortadas , Integrinas/antagonistas & inibidores , Integrinas/imunologia , Integrinas/fisiologia , Proteínas Proto-Oncogênicas c-bcl-2/biossíntese , Receptores de Fibronectina/antagonistas & inibidores , Receptores de Fibronectina/imunologia , Receptores de Fibronectina/fisiologia , Receptores de Vitronectina/antagonistas & inibidores , Receptores de Vitronectina/imunologia , Receptores de Vitronectina/fisiologia , Topotecan/farmacologia , Células Tumorais Cultivadas , Proteína bcl-XRESUMO
A hallmark of invasive tumors is their ability to effect degradation of the surrounding extracellular matrix (ECM) by the local production of proteolytic enzymes, such as the matrix metalloproteases (MMPs). In this paper, we demonstrate that the invasion of human gliomas is mediated by a 72 kDa MMP, referred to as MMP-2, and provide further evidence that the activity of MMP-2 is regulated by protein kinase C (PKC). The invasiveness of five human glioma cell lines (A172, U87, U118, U251, U563) was assessed in an in vitro invasion assay and was found to correlate with the level of MMP-2 activity (r2 = 0.95); in contrast, the activity of this 72 kDa metalloprotease was barely detectable in non-invasive control glial cells (non-transformed human astrocytes and oligodendrocytes). Treatment with 1,10-phenanthroline, a metalloprotease inhibitor, or with a synthetic dipeptide, containing a blocking sequence (ala-phe) specific for MMPs, resulted in a > 90% reduction in glioma invasion. Furthermore, this MMP-2 activity could be inhibited by the treatment of tumor cells with calphostin C, a specific inhibitor of PKC. Glioma cell lines treated with calphostin C demonstrated a dose-dependent decrease (IC50 = 30 nM) in tumor invasiveness with a concomitant reduction in the activity of the MMP-2. Conversely, treatment of non-invasive control astrocytes with a PKC activator (phorbol ester) led to a corresponding increase in their invasiveness and metalloprotease activity. These findings support the postulate that MMP-2 activity constitutes an important effector of human glioma invasion and that the regulation of this proteolytic activity can be modulated by PKC.
Assuntos
Gelatinases/fisiologia , Glioma/patologia , Metaloendopeptidases/fisiologia , Invasividade Neoplásica , Proteína Quinase C/fisiologia , Divisão Celular , Colágeno/metabolismo , Colagenases/metabolismo , Combinação de Medicamentos , Inibidores Enzimáticos/farmacologia , Proteínas da Matriz Extracelular/metabolismo , Gelatinases/antagonistas & inibidores , Glioma/enzimologia , Humanos , Laminina/metabolismo , Metaloproteinase 2 da Matriz , Metaloproteinase 9 da Matriz , Metaloendopeptidases/antagonistas & inibidores , Naftalenos/farmacologia , Proteína Quinase C/antagonistas & inibidores , Proteoglicanas/metabolismo , Células Tumorais CultivadasRESUMO
Recent studies have implicated protein kinase C (PKC)-mediated signaling in the proliferation of gliomas. In this study, we have investigated the role of PKC mediated signaling in the proliferation of medulloblastoma cell lines DAOY, D283-Med and D341-Med. By Western blot analyses, conventional PKC (cPKC) alpha was detectable in DAOY only, while atypical PKC (aPKC) zeta was present in all three cell lines. cPKC beta1, beta11, gamma novel PKC (nPKC) delta, and epsilon were not detectable in any of the cell lines. Antisense oligonucleotides to PKC alpha , Calphostin C (a specific PKC inhibitor) and prolonged treatment with phorbol 12-myristate 13-acetate (PMA) with down regulation of cPKCalpha caused a decrease in proliferation in DAOY and no effect on D283-Med. Furthermore, PMA treatment was also associated with upregulation of p21cip1 in DAOY. Since cPKCalpha is the only PMA responsive isoform in DAOY, this observation implicates the cPKCalpha isoform in the proliferation of DAOY but not in D283-Med. A comparison of DAOY and D283-Med showed a higher proliferation index in DAOY. In contrast, multiprobe riboquant ribonuclease protection assay revealed higher levels of p27kip1 and p21cip1 mRNA in D283-Med. These transcripts were barely detectable in untreated DAOY. These observations indicate possible significant molecular heterogeneity among medulloblastomas with implications for differing biology among medulloblastoma cell lines and tumors.
Assuntos
Meduloblastoma/enzimologia , Proteína Quinase C/fisiologia , Divisão Celular , Humanos , Isoenzimas/análise , Meduloblastoma/patologia , Proteína Quinase C/análise , Proteína Quinase C/genética , RNA Mensageiro/análise , Acetato de Tetradecanoilforbol/farmacologia , Células Tumorais CultivadasRESUMO
Microglia, a population of central nervous system (CNS) macrophages, have been demonstrated to support immune accessory and effector functions in the CNS. Numerous studies support the role of microglia in CNS development and pathology, where activation of microglia is consistently noted. The current study investigated microglial immune functions under basal and activation conditions and assessed the ability of interleukin-10 (IL-10), added exogenously or produced by microglia, to down-regulate microglial functions. This report demonstrates that microglia from the adult human brain produce IL-10 following interferon-gamma/lipopolysaccharide activation. Functionally, recombinant human IL-10 down-regulated basal HLA-DR expression by microglia and inhibited, in a dose-dependent response, the ability of microglia to stimulate CD4+ T-cells in antigen presentation assays. These data, together with recent observations of the inhibition of experimental allergic encephalomyelitis (EAE) following IL-10 administration and reduced CNS infection by Listeria monocytogenes after anti-IL-10 treatment, suggest that IL-10 production by microglia may have important immune-regulatory functions in CNS disease and disease models.
Assuntos
Interleucina-10/biossíntese , Microglia/metabolismo , Adulto , Células Apresentadoras de Antígenos/efeitos dos fármacos , Astrócitos/metabolismo , Encéfalo/citologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Células Cultivadas , DNA Complementar/biossíntese , Regulação para Baixo/efeitos dos fármacos , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Humanos , Imunidade Celular/imunologia , Imunidade Celular/fisiologia , Imuno-Histoquímica , Interleucina-10/farmacologia , Microglia/efeitos dos fármacos , Reação em Cadeia da Polimerase , RNA Mensageiro/biossínteseRESUMO
The protein kinase C (PKC) activity of human glioma cells correlates with their rate of proliferation. We report here that the down-regulation of the predominant PKC isoform of glioma cells, alpha, by antisense phosphorothioate oligonucleotides (AS PTO) significantly reduced the rate of proliferation of three human glioma cell lines. This reduction in growth rate was attributed to apoptosis, as assessed by terminal deoxynucleotidyl transferase (TdT) assay. Unexpectedly, when low concentrations of the protein synthesis inhibitor cycloheximide (CHX) were administered to A172 cells immediately prior to AS PTO treatment, a marked enhancement in the number of apoptotic cells was observed. These findings suggest that PKC alpha plays a pivotal role in the ability of gliomas to avoid apoptotic cell death.
Assuntos
Apoptose/efeitos dos fármacos , Cicloeximida/farmacologia , Isoenzimas/metabolismo , Oligonucleotídeos Antissenso/farmacologia , Proteína Quinase C/metabolismo , Inibidores da Síntese de Proteínas/farmacologia , Divisão Celular/efeitos dos fármacos , Regulação para Baixo , Humanos , Proteína Quinase C-alfa , Células Tumorais CultivadasRESUMO
One of the most lethal properties of high grade gliomas is their ability to invade the surrounding normal brain tissue, as infiltrated cells often escape surgical resection and inevitably lead to tumour recurrence. The consequent poor prognosis and survival rate underscore the need to further understand and target the cellular mechanisms that underly tumour invasiveness. Proteases which degrade the surrounding stromal cells and extracellular matrix proteins have been demonstrated to be critical effectors of invasion for tumours of both central and peripheral origin. Within the nervous system, the role of metalloproteinases as well as other classes of proteases in mediating the invasive phenotype of high grade gliomas has been an intense area of research. We present in this article a review of this literature and address the possibility that these proteases and the biochemical pathways that regulate their expression, such as protein kinase C, may represent potential targets in the therapy of high grade gliomas.
Assuntos
Neoplasias Encefálicas/enzimologia , Neoplasias Encefálicas/patologia , Matriz Extracelular/enzimologia , Glioma/enzimologia , Glioma/patologia , Metaloendopeptidases/metabolismo , Animais , HumanosRESUMO
This article reviews the role of the signal transduction enzyme protein kinase C in the regulation of growth of malignant gliomas, and describes how targetting this enzyme clinically can provide a novel approach to glioma therapy.
Assuntos
Neoplasias Encefálicas/metabolismo , Glioma/metabolismo , Proteína Quinase C/metabolismo , Animais , HumanosRESUMO
INTRODUCTION: Pain that has lasted for 2 months is considered to be persistent, as defined by The International Association for the Study of Pain (IASP). To date, there is no data on the incidence of persistent pain after vaginal delivery available for Ireland. The aim of this pilot study is to determine the incidence of persistent pain in women post-vaginal delivery at 8 weeks postpartum in the Coombe Women's & Infants University Hospital, Dublin (CWIUH). METHODS: 50 women post-vaginal delivery at the CWIUH were recruited over a 2-month period. At 8 weeks after delivery, the patients were contacted by telephone interview to assess the presence of pain, if any, using a modified painDETECT Questionnaire. RESULTS: The incidence of persistent pain at 8 weeks from our pilot study is 28%.
Assuntos
Dor Crônica/epidemiologia , Parto Obstétrico , Complicações do Trabalho de Parto/epidemiologia , Adolescente , Adulto , Feminino , Humanos , Incidência , Irlanda , Medição da Dor , Projetos Piloto , Período Pós-Parto , Gravidez , Prevalência , Inquéritos e Questionários , Adulto JovemAssuntos
Agamaglobulinemia/imunologia , Formação de Anticorpos , Terapia de Imunossupressão , Linfócitos T/imunologia , Animais , Linfócitos B/imunologia , Bolsa de Fabricius/imunologia , Galinhas , Relação Dose-Resposta Imunológica , Imunoglobulina A/metabolismo , Imunoglobulina G/metabolismo , Imunoglobulina M/metabolismoRESUMO
The extension of cellular processes from the oligodendrocyte soma is an early and critical event in myelin formation. Previous reports from this laboratory have implicated a role for protein kinase C (PKC) as an important intracellular mediator of this critical step in myelinogenesis. In the current study, the regrowth of fibers by adult human oligodendrocytes was examined and was found to be significantly enhanced by the PKC stimulator, 4 beta-phorbol-12,13-didecanoate (PDB); this was accompanied by a 400-500% increase in oligodendroglial PKC activity. In contrast to other cell types, the increased PKC activity in oligodendrocytes was not followed by subsequent down-regulation of the enzyme. The role of PKC in oligodendroglial process formation was further demonstrated by the ability of inhibitors of PKC to block the basal- or PDB-enhanced fiber outgrowth. As well, studies employing isoform-specific agonists implicated PKC alpha as the major determinant of fiber outgrowth by oligodendrocytes. The potential significance of PKC in myelin formation was further underscored by the observation that the synthesis of myelin basic protein, a prerequisite component for myelinogenesis, was increased by 2-fold in PDB-treated oligodendrocytes. Collectively, these observations suggest that PKC, in particular the alpha isoform, constitutes an important mediator in the initiation of myelin formation.
Assuntos
Isoenzimas/fisiologia , Proteínas do Tecido Nervoso/fisiologia , Oligodendroglia/enzimologia , Proteína Quinase C/fisiologia , Adulto , Alcaloides/farmacologia , Animais , Sequência de Bases , Benzofenantridinas , Encéfalo/citologia , Células Cultivadas , Diterpenos/farmacologia , Feminino , Humanos , Isoenzimas/antagonistas & inibidores , Dados de Sequência Molecular , Bainha de Mielina/fisiologia , Proteínas do Tecido Nervoso/antagonistas & inibidores , Oligodendroglia/efeitos dos fármacos , Oligodendroglia/ultraestrutura , Fenantridinas/farmacologia , Ésteres de Forbol/farmacologia , Proteína Quinase C/antagonistas & inibidores , Ratos , Ratos Wistar , EstaurosporinaRESUMO
The expression of beta-amyloid precursor protein (BAPP) and its mRNAs was studied in fibroblasts obtained from patients afflicted with Alzheimer's disease (AD) and age-matched controls. Using reverse transcriptase-polymerase chain reaction (RT-PCR), transcripts corresponding to 770, 751, 714, and 695 amino acids were detected in both AD and control fibroblasts. Antibody 22C11 against BAPP (Boehringer Mannheim) labeled an intracellular protein, specifically localized to the intermediate filament network. In addition to bands of the predicted molecular weights for BAPP (120-135 kDa), Western blotting revealed a 57 kDa band which was not evident in samples of human brain. As cytoskeletal elements are vital in maintaining cellular architecture and various cell interactions, localization of BAPP or a related molecule to the cytoskeleton suggests a possible structural role for this protein within the cell.
Assuntos
Doença de Alzheimer/metabolismo , Precursor de Proteína beta-Amiloide/imunologia , Fibroblastos/metabolismo , Proteínas de Filamentos Intermediários/imunologia , Doença de Alzheimer/imunologia , Especificidade de Anticorpos , Western Blotting , Citoesqueleto/imunologia , Citoesqueleto/metabolismo , Fibroblastos/imunologia , Humanos , Imuno-Histoquímica , Microinjeções , Reação em Cadeia da Polimerase , RNA Mensageiro/biossíntese , RNA Mensageiro/metabolismo , Pele/imunologia , Pele/metabolismo , Vimentina/metabolismoRESUMO
We have previously demonstrated that the protein kinase C (PKC) activity of human glioma cell lines was significantly elevated (by 3 orders of magnitude) when compared to non-malignant adult human glia, and that the proliferation rate of several established human glioma cell lines correlated with the measured protein kinase C activity. The purpose of this study was to determine whether 1) elevated PKC activity was also a characteristic of fast growing cell lines of non-glial origin, 2) the proliferation rate of non-glioma cell lines correlated with their PKC activity and 3) the proliferation of non-glioma cell lines could be inhibited by staurosporine, a relatively selective PKC inhibitor, which significantly attenuates the growth of glioma cells. Eight established human non-glioma cell lines (bladder, colorectal, rhabdomyosarcoma-oligodendrocyte hybrid, melanoma, cervix, and fibroblast) were compared to the highly proliferative A172 glioma cell line. PKC activity was significantly higher in the glioma cell lines even though 3 of 8 of the non-glioma lines had higher proliferation rates than A172. In non-glioma cell lines, no correlation was found between the PKC activity and proliferation rates. Staurosporine was more effective in decreasing the proliferation of three glioma cell lines compared to the non-glioma cell lines. We conclude that PKC activity is differentially increased in glioma cell lines and that their proliferation rate is more sensitive to inhibition by staurosporine. Targetting the PKC system may prove to be of therapeutic benefit to patients with malignant gliomas.
Assuntos
Alcaloides/farmacologia , Glioma/tratamento farmacológico , Proteína Quinase C/antagonistas & inibidores , Divisão Celular/efeitos dos fármacos , Células Cultivadas , Glioma/enzimologia , Humanos , Proteína Quinase C/análise , Estaurosporina , Células Tumorais CultivadasRESUMO
It was previously reported that a monoclonal antibody to beta-amyloid precursor protein (mab22C11; Boehringer Mannheim, Indianapolis, IN) labels an intermediate filament-associated protein (beta APP-IFAP) in cultured human skin fibroblasts (Dooley et al.: J Neurosci Res 33:60-67, 1992). The time course of its expression and association with different classes of intermediate filaments has been assessed in neurons, Schwann cells, and astrocytes in dissociated cultures of murine brain and spinal cord-dorsal root ganglia; in primary cultures of human muscle; and in the epithelial cell line PtK1. beta APP-IFAP was expressed in all non-neuronal cell types examined. Mab22C11 immunoreactivity was minimal or absent following dissociation or subculture, but gradually increased with time. In fibroblasts, myoblasts, and epithelial cells, the distribution eventually resembled that of vimentin. With the exception of glial fibrillary acidic protein (GFAP), beta APP-IFAP was not associated with the intermediate filament proteins characteristically found in differentiated cells, i.e., desmin, the cytokeratins, and neurofilament proteins. No labeling of neurons by mab22C11 was observed at any stage of in vitro maturation. In sections of Alzheimer's brain, the antibody labeled a subpopulation of reactive astrocytes. It is suggested that beta APP-IFAP may be the product of a member of the beta APP multigene family expressed developmentally in non-neuronal cells.
Assuntos
Precursor de Proteína beta-Amiloide/metabolismo , Sistema Nervoso Central/embriologia , Embrião de Mamíferos/metabolismo , Proteínas de Filamentos Intermediários/metabolismo , Animais , Células Cultivadas , Embrião de Mamíferos/citologia , Embrião de Mamíferos/fisiologia , Desenvolvimento Embrionário e Fetal , Células Epiteliais , Epitélio/metabolismo , Fibroblastos/metabolismo , Humanos , Rim/citologia , Rim/metabolismo , Marsupiais , Camundongos/embriologia , Músculos/citologia , Músculos/metabolismo , Neuroglia/metabolismo , Neurônios/metabolismo , Vimentina/metabolismoRESUMO
The localized hemolysis in gel (LHG) assay for antibody-secreting cells was used to evaluate antibody production in heterologous erythrocyte-immunized cultures of human peripheral blood lymphocytes supplemented with 10% human plasma. Antigen-specific plaques were detected in such cultures and the number of plaques varied with different lymphocyte donors and were inhibited by heat killing the cultures before assay. However, plaque number varied directly with the number of immunizing erythrocytes added to the cultures and inversely with the number of lymphocytes added. Maximum plaque production occurred within 10 min of initiation of the cultures and occurred in the absence of any lymphoid cells. Absorption of the plasma used as culture supplement with the immunizing erythrocyte resulted in complete abrogation of subsequent specific plaque formation. Mice previously passively immunized with high titered anti-erythrocyte antibody had large numbers of plaques detected 30 min after i.v. immunization with the appropriate erythrocyte. These plaques detected by LHG are the result of carry-over of aggregates of antibody-coated erythrocytes and subsequent release of this antibody in the LHG assay and are not the result of active antibody synthesis. This "pseudoplaque" production may lead to the false interpretation of plaque formation as indicating active synthesis of antibody in vivo and in vitro where such experiments are carried out in immune animals or with antibody containing serum culture supplements.
Assuntos
Especificidade de Anticorpos , Técnica de Placa Hemolítica , Imunização Passiva , Animais , Formação de Anticorpos , Reações Falso-Positivas , Humanos , CamundongosRESUMO
Matrix metalloproteinases (MMPs), the key effectors of extracellular matrix remodeling, have been demonstrated to regulate the extension of neurites from neuronal cell bodies. In this report we have addressed the hypothesis that oligodendrocytes (OLs) may utilize a similar mechanism in extending their processes during the initial phase of myelination. Furthermore, given our previous findings linking protein kinase C (PKC) to the OL process outgrowth, we tested the postulate that this signal transduction pathway may regulate MMPs and thus the process outgrowth phenotype. We demonstrate that in response to pharmacologic activators of PKC, cultured human OLs augment their process extension with a concomitant increase in the activity of an MMP, MMP-9, as measured by gelatin zymography. Similarly, the phorbol ester-enhanced process extension and increased MMP-9 activity were both inhibited by calphostin C, a selective PKC inhibitor. Also, MMP inhibitors such as 1,10-phenanthroline and synthetic dipeptides that inactivate the MMP catalytic site negated the 4beta-phorbol-12,13-dibutyrate (PDB)-mediated process extension, further supporting the key role of MMPs in process extension in vitro. Finally, the elevation of MMP-9 protein expression in the mouse corpus callosum, a tissue rich in OL and myelin, coincided with the previously documented temporal increase in myelination that occurs postnatally. Taken together, these data suggest that MMP-9 constitutes an important mediator of OL process outgrowth, and that this protease in turn can be regulated by PKC. The results are relevant not only to the initial steps of myelination during development, but also to the attempted remyelination that has been shown to occur in pathologic conditions such as MS.
Assuntos
Astrócitos/ultraestrutura , Colagenases/fisiologia , Matriz Extracelular/enzimologia , Matriz Extracelular/ultraestrutura , Oligodendroglia/enzimologia , Oligodendroglia/ultraestrutura , Animais , Western Blotting , Encéfalo/citologia , Comunicação Celular/fisiologia , Células Cultivadas , Corpo Caloso/citologia , Humanos , Imuno-Histoquímica , Radioisótopos do Iodo , Metaloproteinase 9 da Matriz , Camundongos , Bainha de Mielina/metabolismo , Testes de Precipitina , Proteína Quinase C/metabolismoRESUMO
Infectious mononucleosis is caused by the Epstein-Barr virus (EBV), an unusual human pathogen because it preferentially infects B lymphocytes and consequently activates them to produce immunoglobulins. When cultures of lymphocytes from patients with infectious mononucleosis were stimulated with polyclonal activators, unseparated cells failed to produce immunoglobulins, whereas purified B cells responded normally. Cocultures demonstrated profound suppressor T-cell activity in blood from patients with infectious mononucleosis. Early in this disease, circulating immunoglobulin-secreting cells were elevated, but during the second week their number was strikingly depressed. These data indicate that during infectious mononucleosis, EBV causes polyclonal activation of B cells, reflected by hypergammaglobulinemia and increased circulating immunoglobulin-secreting cells. Next, suppressor T cells become activated and inhibit further B-cell activation. Thus, activation of suppressor T cells in infectious mononucleosis provides a unique additional mechanism of host defense because these T cells inhibit the activation and proliferation of an important target of the causative virus.