RESUMO
B7-2(-/-) non-obese diabetic (NOD) mice develop a spontaneous autoimmune polyneuropathy (SAP) that mimics the progressive form of chronic inflammatory demyelinating polyradiculoneuropathy (CIDP). In this study, we focused on the role of regulatory T cells (Tregs ) and regulatory B cells (Bregs ) in SAP. We found that deletion of B7-2 in female NOD mice led to a lower frequency and number of Tregs and Bregs in spleens and lymph nodes. Tregs but not Bregs suppressed antigen-stimulated splenocyte proliferation, whereas Bregs inhibited the T helper type 1 (Th1) cytokine response. Both Tregs and Bregs induced an increase in CD4(+) interleukin (IL)-10(+) cells, although less effectively in the absence of B7-2. Adoptive transfer studies revealed that Tregs , but not Bregs , suppressed SAP, while Bregs attenuated disease severity when given prior to symptom onset. B cell deficiency in B cell-deficient (muMT)/B7-2(-/-) NOD mice prevented the development of SAP, which would indicate that the pathogenic role of B cells predominates over its regulatory role in this model. We conclude that Bregs and Tregs control the immunopathogenesis and progression of SAP in a non-redundant fashion, and that therapies aimed at expansion of Bregs and Tregs may be an effective approach in autoimmune neuropathies.
Assuntos
Linfócitos B Reguladores/imunologia , Antígeno B7-2/imunologia , Linfonodos/imunologia , Polirradiculoneuropatia Desmielinizante Inflamatória Crônica/imunologia , Baço/imunologia , Linfócitos T Reguladores/imunologia , Transferência Adotiva , Animais , Autoimunidade , Linfócitos B Reguladores/patologia , Linfócitos B Reguladores/transplante , Antígeno B7-2/deficiência , Antígeno B7-2/genética , Proliferação de Células , Modelos Animais de Doenças , Feminino , Expressão Gênica , Humanos , Interleucina-10/genética , Interleucina-10/imunologia , Linfonodos/patologia , Contagem de Linfócitos , Camundongos , Camundongos Endogâmicos NOD , Polirradiculoneuropatia Desmielinizante Inflamatória Crônica/genética , Polirradiculoneuropatia Desmielinizante Inflamatória Crônica/patologia , Polirradiculoneuropatia Desmielinizante Inflamatória Crônica/terapia , Baço/patologia , Linfócitos T Reguladores/patologia , Linfócitos T Reguladores/transplante , Células Th1/imunologia , Células Th1/patologia , Equilíbrio Th1-Th2RESUMO
Spontaneous autoimmune polyneuropathy (SAP) in B7-2 knock-out non-obese diabetic (NOD) mice is mediated by myelin protein zero (P0)-reactive T helper type 1 (Th1) cells. In this study, we investigated the role of B cells in SAP, focusing on CD19 as a potential therapeutic target. We found that P0-specific plasmablasts and B cells were increased in spleens of SAP mice compared to wild-type NOD mice. Depletion of B cells and plasmablasts with anti-CD19 monoclonal antibody (mAb) led to attenuation of disease severity when administered at 5 months of age. This was accompanied by decreased serum immunoglobulin (Ig)G and IgM levels, depletion of P0-specific plasmablasts and B cells, down-regulation/internalization of surface CD19 and increased frequency of CD4(+) regulatory T cells in spleens. We conclude that B cells are crucial to the pathogenesis of SAP, and that CD19 is a promising B cell target for the development of disease-modifying agents in autoimmune neuropathies.
Assuntos
Antígenos CD19/imunologia , Linfócitos B/imunologia , Neurite Autoimune Experimental/imunologia , Neurite Autoimune Experimental/terapia , Polineuropatias/imunologia , Polineuropatias/terapia , Animais , Anticorpos Monoclonais/imunologia , Anticorpos Monoclonais/uso terapêutico , Citotoxicidade Celular Dependente de Anticorpos/imunologia , Antígenos CD19/metabolismo , Proliferação de Células , Regulação para Baixo , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Camundongos , Camundongos Endogâmicos NOD , Camundongos Knockout , Proteína P0 da Mielina/imunologia , Plasmócitos/imunologia , Células Th1/imunologiaRESUMO
Fingolimod (FTY720) and its phosphorylated form FTY720P are modulators of sphingosine-1-phosphate (S1P) receptors, which are G-protein coupled receptors linked to cell migration and vascular maturation. The efficacy of FTY720 in autoimmune diseases such as multiple sclerosis and its animal models has been attributed to its inhibition of lymphocyte trafficking to target organs. In this study, we examined the role of S1P receptors in cultured rat oligodendrocytes (OLGs) and OLG progenitor cells (OPCs) using the active phosphorylated form of FTY720. We found that (1) FTY720P improves the survival of neonatal rat OLGs during serum withdrawal, which is associated with the phosphorylation of extracellular signal regulated kinases (ERK1/2) and Akt; (2) FTY720P regulates OPC differentiation into OLGs in a concentration-dependent manner; and (3) S1P receptors are differentially modulated by platelet-derived growth factor (PDGF) resulting in downregulation of S1P5 and upregulation of S1P1 in OPCs. In addition, siRNA studies revealed that S1P1 participates in PDGF-induced OPC mitogenesis. We conclude that S1P1 and S1P5 serve different functions during oligodendroglial development, and possibly during remyelination.
Assuntos
Oligodendroglia/fisiologia , Receptores de Lisoesfingolipídeo/metabolismo , Células-Tronco/fisiologia , Animais , Diferenciação Celular/efeitos dos fármacos , Linhagem da Célula , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Meios de Cultura Livres de Soro/farmacologia , Relação Dose-Resposta a Droga , Regulação para Baixo , Mitógenos/farmacologia , Mitose/fisiologia , Oligodendroglia/metabolismo , Organofosfatos/administração & dosagem , Organofosfatos/farmacologia , Concentração Osmolar , Fator de Crescimento Derivado de Plaquetas/farmacologia , Ratos , Esfingosina/administração & dosagem , Esfingosina/análogos & derivados , Esfingosina/farmacologia , Células-Tronco/citologia , Células-Tronco/efeitos dos fármacos , Células-Tronco/metabolismo , Regulação para CimaRESUMO
Glial cell line-derived neurotrophic factor (GDNF), a known survival factor for neurons, has recently been shown to stimulate the migration of Schwann cells (SCs) and to enhance myelination. GDNF exerts its biological effects by activating the Ret tyrosine kinase in the presence of glycosylphosphatidylinositol-linked receptor, GDNF family receptor (GFR) alpha1. In Ret-negative cells, the alternative transmembrane coreceptor is the 140-kDa isoform of neural cell adhesion molecule (NCAM) associated with a non-receptor tyrosine kinase Fyn. We confirmed that GDNF, GFRalpha1 and NCAM are expressed in neonatal rat SCs. We found that GDNF induces an increase in the partitioning of NCAM and heparan sulfate proteoglycan agrin into lipid rafts and that heparinase inhibits GDNF-signaling in SCs. In addition to activation of extracellular signal-regulated kinases, and phosphorylation of cAMP response element binding protein, we found that cAMP-dependent protein kinase A and protein kinase C are involved in GDNF-mediated signaling in SCs. Although GDNF did not promote the differentiation of purified SCs into the myelinating phenotype, it enhanced myelination in neuron-SC cocultures. We conclude that GDNF utilizes NCAM signaling pathways to regulate SC function prior to myelination and at early stages of myelin formation.
Assuntos
Comunicação Celular/fisiologia , Bainha de Mielina/metabolismo , Fatores de Crescimento Neural/metabolismo , Células de Schwann/metabolismo , Transdução de Sinais/fisiologia , Agrina/metabolismo , Animais , Animais Recém-Nascidos , Axônios/metabolismo , Linhagem Celular Transformada , Células Cultivadas , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Gânglios Espinais/metabolismo , Fator Neurotrófico Derivado de Linhagem de Célula Glial , Receptores de Fator Neurotrófico Derivado de Linhagem de Célula Glial , Microdomínios da Membrana/metabolismo , Regeneração Nervosa/fisiologia , Moléculas de Adesão de Célula Nervosa/metabolismo , Neurônios Aferentes/metabolismo , Proteína Quinase C/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-ret , Ratos , Receptores Proteína Tirosina Quinases/metabolismo , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/fisiologiaRESUMO
This report illustrates a case of chronic inflammatory demyelinating polyradiculoneuropathy (CIDP) masquerading as neurofibromatosis due to multifocal enlargements of spinal nerve roots. The patient initially complained of intermittent numbness of the hands and leg weakness at age 62. Nerve conduction velocities were reported to be abnormally slow, suggesting a diagnosis of demyelinating neuropathy. A complaint of progressive lower back pain 4 years later prompted a lumbar CT myelogram, which demonstrated bilateral nerve root enlargements. A biopsy of an enlarged lumbar root obtained during decompressive laminectomy was interpreted as consistent with a plexiform neurofibroma. He suffered recurrent paraparesis, at times with a sensory level indicating spinal cord compression, which responded to corticosteroid therapy. An autopsy 15 years after the onset of symptoms revealed hypertrophic radiculopathy and peripheral neuropathy due to CIDP with no evidence of neurofibromatosis. This case illustrates how the hypertrophic neuropathy accompanying CIDP can be mistaken for neurofibromatosis.
Assuntos
Neurofibromatoses/patologia , Polirradiculoneuropatia Desmielinizante Inflamatória Crônica/patologia , Raízes Nervosas Espinhais/patologia , Idoso , Diagnóstico Diferencial , Humanos , Hipertrofia/diagnóstico por imagem , Hipertrofia/etiologia , Hipertrofia/patologia , Imageamento por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Condução Nervosa/fisiologia , Paraparesia/etiologia , Paraparesia/patologia , Polirradiculoneuropatia Desmielinizante Inflamatória Crônica/complicações , Polirradiculoneuropatia Desmielinizante Inflamatória Crônica/diagnóstico por imagem , Polirradiculoneuropatia Desmielinizante Inflamatória Crônica/fisiopatologia , Radiculopatia/diagnóstico por imagem , Radiculopatia/etiologia , Radiculopatia/patologia , Radiografia , Raízes Nervosas Espinhais/diagnóstico por imagemRESUMO
An increasing number of studies have documented the central role of T cell costimulation in autoimmunity. Here we show that the autoimmune diabetes-prone nonobese diabetic (NOD) mouse strain, deficient in B7-2 costimulation, is protected from diabetes but develops a spontaneous autoimmune peripheral polyneuropathy. All the female and one third of the male mice exhibited limb paralysis with histologic and electrophysiologic evidence of severe demyelination in the peripheral nerves beginning at 20 wk of age. No central nervous system lesions were apparent. The peripheral nerve tissue was infiltrated with dendritic cells, CD4(+), and CD8(+) T cells. Finally, CD4(+) T cells isolated from affected animals induced the disease in NOD.SCID mice. Thus, the B7-2-deficient NOD mouse constitutes the first model of a spontaneous autoimmune disease of the peripheral nervous system, which has many similarities to the human disease, chronic inflammatory demyelinating polyneuropathy (CIDP). This model demonstrates that NOD mice have "cryptic" autoimmune defects that can polarize toward the nervous tissue after the selective disruption of CD28/B7-2 costimulatory pathway.
Assuntos
Antígenos CD/imunologia , Glicoproteínas de Membrana/imunologia , Doença Autoimune do Sistema Nervoso Experimental/imunologia , Doenças do Sistema Nervoso Periférico/imunologia , Linfócitos T/imunologia , Envelhecimento , Animais , Antígenos CD/genética , Antígeno B7-2 , Encéfalo/imunologia , Encéfalo/patologia , Cruzamentos Genéticos , Gânglios Espinais/imunologia , Gânglios Espinais/patologia , Inflamação , Glicoproteínas de Membrana/deficiência , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos NOD , Camundongos Knockout , Doença Autoimune do Sistema Nervoso Experimental/genética , Doença Autoimune do Sistema Nervoso Experimental/patologia , Doenças do Sistema Nervoso Periférico/genética , Doenças do Sistema Nervoso Periférico/patologia , Nós Neurofibrosos/imunologia , Nós Neurofibrosos/patologia , Nervo Isquiático/imunologia , Nervo Isquiático/patologiaRESUMO
Tumor necrosis factor-alpha and interferon-gamma are pleiotropic cytokines that regulate Schwann cell responses during injury and inflammatory demyelination. We have previously shown that cyclic AMP (cAMP)-elevating agents decrease the demyelination and Wallerian degeneration in experimental allergic neuritis. In this study, we examined the role of cAMP in cytokine-mediated signaling in a spontaneously immortal Schwann cell clone (iSC). We found that tumor necrosis factor-alpha and interferon-gamma exert synergistic inhibitory action on Schwann cell viability via the production of nitric oxide (NO) and ceramide (cer). Furthermore, we found that: (i) NO synthase inhibitors attenuate the cytokine-induced cer accumulation and cell death indicating that NO acts upstream of cer; and (ii) cytokine-induced cell death is decreased in iSCs pretreated continuously for 48-72 h with forskolin, an activator of adenylate cyclase. Although forskolin modulates the phosphorylation of ERKs and Akt, it decreases the susceptibility of iSC to cytokines via a separate mechanism operating after NO induction and before cer accumulation. We propose that the protective effect of cAMP-elevating agents in experimental allergic neuritis may be mediated in part via modulation of Schwann cell responses to cytokines.
Assuntos
Apoptose/imunologia , AMP Cíclico/metabolismo , Interferon gama/farmacologia , Óxido Nítrico/metabolismo , Células de Schwann/citologia , Fator de Necrose Tumoral alfa/farmacologia , Animais , Apoptose/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/imunologia , Linhagem Celular Transformada , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/imunologia , Ceramidas/metabolismo , Colforsina/farmacologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Doenças Desmielinizantes/enzimologia , Doenças Desmielinizantes/imunologia , Inibidores Enzimáticos/farmacologia , Síndrome de Guillain-Barré/imunologia , Síndrome de Guillain-Barré/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , NG-Nitroarginina Metil Éster/farmacologia , Neuroimunomodulação/fisiologia , Óxido Nítrico Sintase/metabolismo , Óxido Nítrico Sintase Tipo II , Fosfatidilinositol 3-Quinases/metabolismo , Ratos , Células de Schwann/enzimologia , Células de Schwann/imunologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/imunologiaRESUMO
Intracellular Ca2+ is the key signal that regulates the efficacy of neurotransmitter release and synaptic plasticity in neurons but is also an important second messenger involved in the signal transduction and modulation of gene expression in both excitable and non-excitable cells. Glial cells, including cells of oligodendroglial (OLG) lineage, are capable of responding to extracellular stimuli via changes in the intracellular Ca2+. This review article focuses on the mechanisms of Ca2+ signalling in cells of OLG lineage with the goal of providing the basis for understanding the relevance of receptor- and non-receptor-mediated signalling to oligodendroglial development, myelination, and demyelination. Conclusions to date indicate that cells of OLG lineage exhibit remarkable plasticity with regard to the expression of ion channels and receptors linked to Ca2+ signalling and that perturbation of [Ca2](i) homeostasis contributes to the pathogenesis of demyelinating diseases.
Assuntos
Sinalização do Cálcio/fisiologia , Oligodendroglia/fisiologia , Transdução de Sinais/fisiologia , Animais , CamundongosRESUMO
Proliferation and apoptosis are two dynamic, interrelated processes that are regulated by growth factors and cytokines. We investigated the effects of tumor necrosis factor-alpha (TNF-alpha) and transforming growth factor-beta (TGF-beta) on apoptosis and regulation of cell cycle proteins in OLG lineage cells. We found that: (1) both cytokines enhanced apoptosis in neonatal pre-OLGs but only TNFalpha-mediated apoptosis persisted in the presence of a mitogen, fibroblast growth factor (FGF); (2) cell cycle proteins such as p21(waf1/cip1), p27(kip1), cyclin D1 and PCNA were differentially regulated by TNF-alpha and TGF-beta. We conclude that differential modulation of cell cycle proteins by TNF-alpha and TGF-beta contributes to the diversity of their biological effects in OLG lineage cells.
Assuntos
Quinases relacionadas a CDC2 e CDC28 , Proteínas de Ciclo Celular/metabolismo , Linfotoxina-alfa/farmacologia , Oligodendroglia/efeitos dos fármacos , Oligodendroglia/metabolismo , Fator de Crescimento Transformador alfa/farmacologia , Proteínas Supressoras de Tumor , Envelhecimento , Animais , Apoptose/efeitos dos fármacos , Linhagem da Célula , Células Cultivadas , Ciclina D1/metabolismo , Quinase 2 Dependente de Ciclina , Inibidor de Quinase Dependente de Ciclina p21 , Inibidor de Quinase Dependente de Ciclina p27 , Quinases Ciclina-Dependentes/metabolismo , Ciclinas/metabolismo , DNA/biossíntese , DNA/genética , Fatores de Crescimento de Fibroblastos/farmacologia , Marcação In Situ das Extremidades Cortadas , Proteínas Associadas aos Microtúbulos/metabolismo , Mitógenos/farmacologia , Oligodendroglia/citologia , Antígeno Nuclear de Célula em Proliferação/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Ratos , Ratos Sprague-DawleyRESUMO
Sphingolipid products such as ceramide (cer), sphingosine (sph), and sphingosine-1-phosphate (SPP) are implicated in the regulation of cell growth and apoptosis. We have recently shown that cer, sph, and SPP differentially modulate ionic events in cultured oligodendrocytes (OLGs). Cer but not sph or SPP inhibits the inward rectifier (I(Kir)) in OLGs. To further investigate the role of sphingolipid products in OLGs, we studied the effect of cer, sph, and SPP on OLG survival and on the regulation of mitogen-activated protein kinases (MAPKs). We found that cer, sph, and SPP differentially modulate OLG survival and activation of MAPK members. Cer causes OLG apoptosis, sph causes OLG lysis, and SPP does not affect OLG survival. Cer induces a preferential activation of p38alpha, whereas sph and SPP induce a preferential activation of extracellular signal-regulated kinase 2 (ERK2) in OLGs. In addition, the effect of cer on p38alpha activity is mimicked by the inhibition of I(Kir) with Ba(2+). In contrast, exposure to cer results in increased activity of ERK2 but not of p38alpha in astrocytes. Cer-induced OLG apoptosis is attenuated by a p38 inhibitor, SB203580, and by expression of a p38alpha dominant negative mutant. We conclude that p38alpha is the mediator in cer-induced OLG apoptosis and that cer-induced I(Kir) inhibition may contribute to the sustained activation of p38alpha in OLGs.
Assuntos
Proteínas Quinases Dependentes de Cálcio-Calmodulina/metabolismo , Proteínas Quinases Ativadas por Mitógeno , Oligodendroglia/fisiologia , Esfingolipídeos/fisiologia , Animais , Animais Recém-Nascidos , Apoptose , Astrócitos/citologia , Astrócitos/efeitos dos fármacos , Astrócitos/fisiologia , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Ceramidas/farmacologia , Ceramidas/fisiologia , Ativação Enzimática/fisiologia , Proteínas Quinases JNK Ativadas por Mitógeno , Proteína Quinase 1 Ativada por Mitógeno , Modelos Neurológicos , Oligodendroglia/citologia , Oligodendroglia/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Proteínas Recombinantes/metabolismo , Esfingolipídeos/farmacologia , Esfingosina/farmacologia , Esfingosina/fisiologia , Transfecção , Proteínas Quinases p38 Ativadas por MitógenoRESUMO
Ceramide is a lipid mediator implicated in apoptosis induced by proinflammatory cytokines in many cell types, including oligodendrocytes (OLGs). To determine whether ceramide modulates transmembrane signaling events in OLGs, we studied its effect on intracellular Ca2+ (Cai), resting membrane potential and inwardly rectifying K+ currents (IKir) in cultured neonatal rat OLGs. We report here that (1) exposure to C2-ceramide (cer) rarely increases OLG Cai, whereas sphingosine elicits sustained increase in Cai; (2) cer causes OLG depolarization, an effect mimicked by sphingosine-1-phosphate but not by sphingosine; and (3) cer, but not its inactive analog dihydroceramide, inhibits OLG IKir. The cer effect is attenuated by Ras antibody Y13-259, by protein kinase C inhibitory peptide (19-36), and by suppression of c-Raf-1 expression with antisense raf-1 oligonucleotides. We conclude that cer-induced OLG depolarization is mediated via inhibition of IKir by a Ras- and raf-1-dependent pathway, which results in the phosphorylation of the inward rectifier K+ channel protein.
Assuntos
Ceramidas/farmacologia , Lisofosfolipídeos , Oligodendroglia/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização , Canais de Potássio/metabolismo , Proteínas Proto-Oncogênicas c-raf/fisiologia , Proteínas ras/fisiologia , Animais , Animais Recém-Nascidos , Cálcio/metabolismo , Células Cultivadas , Técnicas In Vitro , Potenciais da Membrana/efeitos dos fármacos , Oligodendroglia/efeitos dos fármacos , Técnicas de Patch-Clamp , Canais de Potássio/efeitos dos fármacos , Proteínas Tirosina Quinases/antagonistas & inibidores , Ratos , Esfingosina/análogos & derivados , Esfingosina/farmacologiaRESUMO
Protein phosphorylation is a well-known mechanism by which extracellular molecules or factors transduce their signals into intracellular effects. In the context of myelin assembly, phosphorylation of major myelin proteins affects the electrostatic repulsion between adjacent proteins within myelin structure and therefore constitutes one of the mechanisms by which myelin stability is regulated. We report here that arachidonic acid (AA) decreases the phosphorylation of myelin basic protein (MBP) both in the absence and in the presence of phorbol esters in cultured rat oligodendrocytes (OLGs). The effect of AA on MBP phosphorylation is not mediated by cyclooxygenase products, though the possibility that leukotrienes or other epoxides may have a role cannot be excluded. AA did not act by inactivation of protein kinase C. Based on our findings from gadolinium and low K+ experiments, we conclude that inhibition of MBP phosphorylation is not dependent on AA-induced increases in OLG Ca(i), but rather on its depolarizing action. We have thus demonstrated that a brief exposure to AA, which either acts as a diffusible paracrine signal to OLGs or as a signal transducer, can trigger changes in protein phosphorylation in OLGs/myelin via ionic signaling events at the plasma membrane.
Assuntos
Ácido Araquidônico/farmacologia , Proteína Básica da Mielina/metabolismo , Oligodendroglia/metabolismo , Animais , Cálcio/metabolismo , Membrana Celular/fisiologia , Células Cultivadas , Proteína Básica da Mielina/antagonistas & inibidores , Oligodendroglia/citologia , Oligodendroglia/efeitos dos fármacos , Fosforilação , Proteína Quinase C/metabolismo , RatosRESUMO
We examined the molecular identity of K+ channel genes underlying the delayed rectifier (IK) in differentiated cultured oligodendrocytes (OLGs) and oligodendrocyte progenitor (OP) cells. Using reverse transcription-PCR cloning, we found that OP cells and OLGs expressed multiple Kv transcripts, namely Kv1.2, Kv1.4, Kv.1.5, and Kv1.6. Immunocytochemical and Western blot analyses revealed that Kv1.5 and Kv1.6 as well as Kv1.2 and Kv1.4 channel proteins could be detected in these cells, but definitive evidence for functional K+ channel expression was obtained only for the Kv1.5 channel. In addition, mRNA and immunoreactive protein levels of both Kv1.5 and Kv1.6 channels were significantly lower in differentiated OLGs when compared with levels in OP cells. Proliferation of OP cells was inhibited by K+ channel blockers, but not by incubation with either Kv1.5 or Kv1.6 antisense oligonucleotides. We conclude that (1) IK in OP cells and OLGs is encoded partly by Kv1.5 subunits, possibly forming heteromultimeric channels with Kv1.6 or other Kv subunits; and (2) inhibition of Kv1.5 or Kv1.6 channel expression alone does not prevent mitogenesis. Concomitant inhibition of other Kv channels underlying IK may be necessary for OP cells to exit from cell cycle.
Assuntos
Proteínas do Tecido Nervoso/metabolismo , Oligodendroglia/metabolismo , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Canais de Potássio/metabolismo , Potássio/metabolismo , Células-Tronco/metabolismo , 4-Aminopiridina/farmacologia , Animais , Diferenciação Celular , Linhagem Celular , Canais de Potássio de Retificação Tardia , Humanos , Hibridização In Situ , Transporte de Íons/efeitos dos fármacos , Rim , Canal de Potássio Kv1.2 , Canal de Potássio Kv1.4 , Canal de Potássio Kv1.5 , Substâncias Macromoleculares , Família Multigênica , Proteínas do Tecido Nervoso/classificação , Proteínas do Tecido Nervoso/genética , Oligonucleotídeos Antissenso/farmacologia , Técnicas de Patch-Clamp , Reação em Cadeia da Polimerase , Bloqueadores dos Canais de Potássio , Canais de Potássio/classificação , Canais de Potássio/genética , Compostos de Amônio Quaternário/farmacologia , Quinidina/farmacologia , Quinina/farmacologia , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Ratos , Proteínas Recombinantes de Fusão/metabolismo , Tetraetilamônio/farmacologia , TransfecçãoRESUMO
Suramin, a promising chemotherapeutic agent, causes a dose-limiting sensorimotor polyneuropathy. We undertook a phase 1 study of suramin that included serial neurologic and electrophysiologic examinations as part of the safety evaluation. We found that 6 of 41 (15%) patients developed suramin-induced demyelinating neuropathy which resembled Guillain-Barre syndrome clinically. There was 1 asymptomatic patient with electrophysiologic abnormalities suggestive of a demyelinating neuropathy. In addition, 1 patient with mild axonal neuropathy at baseline had deterioration of his symptoms during suramin treatment. Four asymptomatic patients developed electrophysiologic findings suggestive of a mild axonal neuropathy. We conclude that: (1) serial electrophysiologic monitoring is helpful for early detection of suramin-induced neuropathy; and (2) fixed dosing schedule of suramin without adaptive control does not lead to an increased incidence of demyelinating neuropathy when compared to adaptively controlled dosing schedules.
Assuntos
Doenças Desmielinizantes/induzido quimicamente , Doença dos Neurônios Motores/induzido quimicamente , Condução Nervosa/fisiologia , Neoplasias da Próstata/tratamento farmacológico , Suramina/efeitos adversos , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Suramina/uso terapêuticoAssuntos
Antineoplásicos/administração & dosagem , Neoplasias/tratamento farmacológico , Suramina/administração & dosagem , Adulto , Idoso , Idoso de 80 Anos ou mais , Antineoplásicos/efeitos adversos , Ensaios Clínicos Fase I como Assunto , Humanos , Infusões Intravenosas , Pessoa de Meia-Idade , Suramina/efeitos adversosRESUMO
PURPOSE: Suramin is a promising agent for the treatment of hormone-refractory metastatic prostate cancer. However, questions about the relationship of severe neurotoxicity to sustained peak plasma concentrations greater than 300 micrograms/mL raised concerns that this drug could not be safely administered without adaptive control. To test the adaptive-control hypothesis, we designed a phase I study that relied on clinical end points, using a fixed dosing scheme that did not rely on adaptive control. PATIENTS AND METHODS: In a phase I dose-escalation study using fixed dosing without adaptive control, gradually decreasing doses of suramin were administered to 63 patients on days 1 (loading dose), 2, 8, and 9 of a 28-day cycle. Fifty-four patients with hormone-refractory metastatic prostate cancer and nine patients with other solid tumors have been treated. RESULTS: Doses of 400 mg/m2 to 2,080 mg/m2 on the first day have been administered. The mean peak plasma concentration following the loading dose at a dose level of 1,730 mg/m2 was 933 micrograms/mL (26% coefficient of variation), and the mean trough concentration was 139 micrograms/mL (40% CV) on day 1 of cycle 2 [corrected]. At 1,730 mg/m2, five of 13 patients experienced dose-limiting toxicity (DLT), including malaise, neurotoxicity, pericardial effusion, and coagulopathy. At 2,080 mg/m2, three of five patients experienced DLT. Two patients treated at this dose level died while on study. One of these patients died of a subdural hematoma sustained after a fall and had a prolonged prothrombin time at the time of his death. One patient developed classic suramin neurotoxicity, which led to respiratory failure, for which the patient refused intubation. No significant associations were noted between peak or trough concentrations during either cycles 1 or 2 and the occurrence of neurotoxicity. CONCLUSION: (1) Suramin can be safely administered without adaptive control, (2) suramin on this schedule may exhibit significant activity against hormone-refractory metastatic prostate cancer, and (3) based strictly on toxicity considerations, we recommended that a day-1 dose of 1,440 mg/m2 be used in subsequent clinical trials, with a maximum of three cycles. Further studies to establish the optimal empiric dosing regimen are needed.
Assuntos
Adaptação Fisiológica , Neoplasias/tratamento farmacológico , Suramina/administração & dosagem , Adulto , Idoso , Estudos de Coortes , Esquema de Medicação , Monitoramento de Medicamentos , Feminino , Humanos , Infusões Intravenosas , Tábuas de Vida , Modelos Logísticos , Masculino , Pessoa de Meia-Idade , Neoplasias/sangue , Neoplasias/mortalidade , Doenças do Sistema Nervoso/induzido quimicamente , Neoplasias da Próstata/sangue , Neoplasias da Próstata/tratamento farmacológico , Indução de Remissão , Suramina/efeitos adversos , Suramina/farmacocinética , Taxa de SobrevidaRESUMO
Arachidonic acid (AA) and its metabolites play a dual role as intracellular second messengers and as transcellular mediators of neural activity. We have previously shown that AA increases cytosolic Ca2+ in oligodendrocytes. In this work, we studied the effects of AA and other fatty acids on whole cell K+ currents of cultured rat oligodendrocytes using the patch-clamp technique. We found that 1) AA decreased the current amplitudes of both the inwardly rectifying K+ current (IKir) and the outward K+ currents (IKo) resulting in membrane depolarization; 2) AA also induced IKo current inactivation/blocked state; 3) AA appeared to act directly on K+ channels and not indirectly via its metabolic products, activation of protein kinase C, or by generation of oxygen free radicals. We have thus demonstrated an additional mechanism for AA-induced signaling in oligodendrocytes, i.e., via modulation of K+ conductances leading to membrane depolarization. The latter has been shown to influence protein phosphorylation and perhaps other important functional output of oligodendrocytes.
Assuntos
Ácido Araquidônico/farmacologia , Oligodendroglia/efeitos dos fármacos , Oligodendroglia/fisiologia , Bloqueadores dos Canais de Potássio , Canais de Potássio/fisiologia , Animais , Células Cultivadas , Condutividade Elétrica , Técnicas de Patch-Clamp , Ratos , Ratos EndogâmicosRESUMO
The syntax of neuronal-glial or axonal-glial interaction is frequently communicated through transient changes in internal calcium (Cai). We examined mechanisms for Cai signaling and intercellular propagation of Cai responses in cultured oligodendrocytes (OLGs) derived from adult spinal cord (SC), postnatal day 21 (P21) SC, and P21 brain. We found that (1) cultured OLGs exhibited a heterogeneous response to norepinephrine, carbachol, ATP, histamine, and glutamate; (2) receptor-mediated Cai increases were derived from both Ca2+ influx and intracellular Ca2+ release; (3) the percentage of responders to neuroligands varied as a function of cell origin; (4) cultured OLGs exhibited a thapsigargin-sensitive, but not a caffeine-sensitive, intracellular Ca2+ pool; and (5) gap junctional contacts between OLGs permitted limited intercellular propagation of mechanically stimulated Cai responses. Receptor-mediated Cai signaling appears to occur not only in cultured OLGs but also in acutely dissociated OLGs. The heterogeneity in Cai responses as a function of cell origin may reflect the existence of OLG subsets or differences in the maturation stage of OLGs.
Assuntos
Cálcio/fisiologia , Oligodendroglia/fisiologia , Transdução de Sinais/fisiologia , Trifosfato de Adenosina/farmacologia , Animais , Carbacol/farmacologia , Células Cultivadas/fisiologia , Ácido Glutâmico/farmacologia , Histamina/farmacologia , Ativação do Canal Iônico/fisiologia , Norepinefrina/farmacologia , Oligodendroglia/citologia , Oligodendroglia/efeitos dos fármacos , Ratos , Ratos EndogâmicosRESUMO
We have used a combination of electrophysiological and biochemical approaches to investigate the effects and the mechanisms of action of tumor necrosis factor-alpha (TNF-alpha) on cultured oligodendrocytes (OLGs). Our studies have led to the following conclusions: (1) prolonged exposure of mature ovine OLGs to TNF-alpha leads to inhibition of process extension, membrane depolarization and a decrease in the amplitudes of both inwardly rectifying and outward K+ currents; (2) brief exposure of OLGs to TNF-alpha does not elicit membrane depolarization or consistent changes in cytosolic Ca2+ levels; (3) incubation of OLGs with TNF-alpha for 1 hr results in inhibition of phosphorylation of myelin basic protein and 2',3'-cyclic nucleotide phosphohydrolase. Ceramides, which have been shown to be effectors of TNF-alpha, are ineffective in inhibiting phosphorylation, whereas sphingomyelinase mimics TNF-alpha in this action. These observations suggest that other products of sphingomyelin hydrolysis may be the mediator(s) of TNF-alpha effect on protein phosphorylation. We have thus demonstrated that TNF-alpha can perturb the functions of OLGs via modulation of ion channels and of protein phosphorylation without necessarily inducing cell death. It is conceivable that modulation of ion channels and protein phosphorylation constitutes effective mechanisms for the participation of cytokines in signal transduction during myelination, demyelination and remyelination.
Assuntos
Oligodendroglia/fisiologia , Transdução de Sinais/fisiologia , Fator de Necrose Tumoral alfa/fisiologia , Animais , Adesão Celular/fisiologia , HumanosRESUMO
It is recognized that tumor necrosis factor-alpha (TNF-alpha), a pleiotropic cytokine, influences hormone secretion and transmitter release from central neurons. To examine the role of TNF-alpha as a modulator of autonomic function of the PNS, we measured [3H]norepinephrine ([3H]NE) secretion evoked by 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP), a nicotinic agonist, in cultures from neonatal rat superior cervical ganglia (SCG). We found that (1) DMPP-evoked [3H]NE secretion was enhanced in SCG mixed cultures treated for 48 h with recombinant human TNF-alpha (rhTNF-alpha) plus rat interferon-gamma (IFN-gamma) but not in cultures treated with either cytokine alone; (2) an increase in [3H]NE secretion was also observed in mixed cultures treated with recombinant murine TNF-alpha (rmTNF-alpha) alone; and (3) the presence of nonneuronal cells or soluble factors released by them was required for the effect of these cytokines on secretion. Electrophysiologic experiments revealed an increase in nicotinic receptor current density in neurons from mixed cultures treated with rhTNF-alpha plus IFN-gamma or with rmTNF-alpha when compared with control cultures. We conclude that prolonged exposure to rhTNF-alpha plus IFN-gamma or rmTNF-alpha regulates nicotinic responses in SCG cultures via a soluble factor or factors secreted by nonneuronal cells.