RESUMO
Alymphoplasia (aly) mice, which carry a point mutation in the nuclear factor kappaB-inducing kinase (NIK) gene, are characterized by the systemic absence of lymph nodes and Peyer's patches, disorganized splenic and thymic architectures, and immunodeficiency. Another unique feature of aly/aly mice is that their peritoneal cavity contains more B1 cells than normal and aly/+ mice. Transfer experiments of peritoneal lymphocytes from aly/aly mice into recombination activating gene (RAG)-2(-/-) mice revealed that B and T cells fail to migrate to other lymphoid tissues, particularly to the gut-associated lymphatic tissue system. In vivo homing defects of aly/aly peritoneal cells correlated with reduction of their in vitro chemotactic responses to secondary lymphoid tissue chemokine (SLC) and B lymphocyte chemoattractant (BLC). The migration defect of aly/aly lymphocytes was not due to a lack of expression of chemokines and their receptors, but rather to impaired signal transduction downstream of the receptors for SLC, indicating that NIK is involved in the chemokine signaling pathway known to couple only with G proteins. The results showed that the reduced serum levels of immunoglobulins (Igs) and the absence of class switch to IgA in aly/aly mice are due, at least in part, to a migration defect of lymphocytes to the proper microenvironment where B cells proliferate and differentiate into Ig-producing cells.
Assuntos
Quimiotaxia , Intestino Delgado/imunologia , Tecido Linfoide/anormalidades , Peritônio/imunologia , Proteínas Serina-Treonina Quinases/genética , Receptores de Quimiocinas/metabolismo , Animais , Quimiocinas/metabolismo , Proteínas de Ligação a DNA , Isotipos de Imunoglobulinas/biossíntese , Linfonodos/anormalidades , Camundongos , Camundongos Mutantes , Peritônio/citologia , Nódulos Linfáticos Agregados/anormalidades , Plasmócitos , Transdução de Sinais , Quinase Induzida por NF-kappaBRESUMO
Notch1 has been shown to induce glia in the peripheral nervous system. However, it has not been known whether Notch can direct commitment to glia from multipotent progenitors of the central nervous system. Here we present evidence that activated Notch1 and Notch3 promotes the differentiation of astroglia from the rat adult hippocampus-derived multipotent progenitors (AHPs). Quantitative clonal analysis indicates that the action of Notch is likely to be instructive. Transient activation of Notch can direct commitment of AHPs irreversibly to astroglia. Astroglial induction by Notch signaling was shown to be independent of STAT3, which is a key regulatory transcriptional factor when ciliary neurotrophic factor (CNTF) induces astroglia. These data suggest that Notch provides a CNTF-independent instructive signal of astroglia differentiation in CNS multipotent progenitor cells.
Assuntos
Astrócitos/metabolismo , Fator 2 de Crescimento de Fibroblastos/metabolismo , Proteínas de Membrana/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Receptores de Superfície Celular , Células-Tronco/metabolismo , Fatores de Transcrição , Animais , Diferenciação Celular/efeitos dos fármacos , Linhagem da Célula/efeitos dos fármacos , Células Cultivadas , Fator Neurotrófico Ciliar/metabolismo , Fator Neurotrófico Ciliar/farmacologia , Células Clonais/efeitos dos fármacos , Proteínas de Ligação a DNA/metabolismo , Fator 2 de Crescimento de Fibroblastos/farmacologia , Hipocampo/citologia , Hipocampo/metabolismo , Proteínas de Membrana/farmacologia , Neurônios/citologia , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Proteínas Proto-Oncogênicas/farmacologia , Ratos , Receptor Notch1 , Fator de Transcrição STAT3 , Transdução de Sinais/efeitos dos fármacos , Células-Tronco/citologia , Células-Tronco/efeitos dos fármacos , Transativadores/metabolismoRESUMO
BACKGROUND: 3-Hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors have been demonstrated to suppress glomerular injuries in various renal diseases. These agents inhibit in vitro proliferation of several cell types, including mesangial cells. This effect indicates the ability to ameliorate mesangioproliferative lesions, independent of the improvement of hypercholesterolemia. On the other hand, it is not clear whether HMG-CoA reductase inhibitors directly regulate extracellular matrix (ECM) accumulation from mesangial cells. METHODS: In this study, to examine the in vitro effects of simvastatin, an HMG-CoA reductase inhibitor, on mRNA expressions of matrix proteins, growth factors, and matrix turnover proteins, we incubated cultured murine mesangial cells stimulated by fetal calf serum (FCS) with or without simvastatin for 24 hours, and Northern analysis was performed. RESULTS: Simvastatin showed a slightly suppressive effect on mRNA expression of type IV collagen and fibronectin and a slightly up-regulative effect on that of type I collagen, whereas mRNA expression of type III collagen was markedly up-regulated. mRNA expression of platelet-derived growth factor (PDGF)-B chain and PDGF receptor beta-subunit was suppressed, whereas that of transforming growth factor-beta (TGF-beta) was not affected by simvastatin. Concerning matrix turnover proteins, simvastatin markedly reduced mRNA expression of plsminogen activator inhibitor-1 (PAI-1) without affecting the expression of tissue-type plasminogen activator (tPA). CONCLUSION: These results suggest type-specific modulation of matrix protein production independent of TGF-beta and the suppressive effects of autocrine PDGF by administration of HMG-CoA reductase inhibitors in mesangial cells. Moreover, the beneficial effects of these agents on matrix protein accumulation may be through promoting ECM degradation derived from PAI-1 suppression.
Assuntos
Proteínas da Matriz Extracelular/efeitos dos fármacos , Mesângio Glomerular/efeitos dos fármacos , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Sinvastatina/farmacologia , Animais , Células Cultivadas , Colágeno/genética , Proteínas da Matriz Extracelular/genética , Proteínas da Matriz Extracelular/metabolismo , Fibronectinas/genética , Expressão Gênica/efeitos dos fármacos , Mesângio Glomerular/citologia , Mesângio Glomerular/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Inibidor 1 de Ativador de Plasminogênio/genética , Fator de Crescimento Derivado de Plaquetas/genética , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptor beta de Fator de Crescimento Derivado de Plaquetas , Receptores do Fator de Crescimento Derivado de Plaquetas/genética , Ativador de Plasminogênio Tipo Uroquinase/genéticaRESUMO
A 50-year-old man with WPW syndrome and chronic atrial fibrillation controlled by digoxin exhibited nephrotic syndrome due to membranous nephropathy in 1994. For massive edema resistant to loop diuretics, administration of 2 mg/day of Temocapril, an ACE inhibitor with biliary excretion, was started. It provided marked natriuresis and rapid improvement of massive anasarca. Moreover, 6 months later at the time of writing this report, nephrotic proteinuria has decreased and abnormally depressed Ccr level has elevated. For massive edema resistant to loop diuretics in nephrotic syndrome, Temocapril might bring about a beneficial effect possibly through the induction of natriuresis, which has been suppressed by the over-expression of angiotensin II in tubulointerstitium.
Assuntos
Inibidores da Enzima Conversora de Angiotensina/uso terapêutico , Diuréticos/farmacologia , Edema/tratamento farmacológico , Glomerulonefrite Membranosa/complicações , Natriurese/efeitos dos fármacos , Proteinúria/tratamento farmacológico , Tiazepinas/uso terapêutico , Resistência a Medicamentos , Edema/fisiopatologia , Glomerulonefrite Membranosa/patologia , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
A 47-year-old woman was admitted on August 4th, 1995, because of edema of the lower extremities. She had been suffering from RA for about 20 years and underwent total knee-replacements 5 years previously. On admission, nephrotic syndrome and rapidly progressive glomerulonephritis had developed in association with ileus, melena, diarrhea, dyspnea and hemoptysis. She showed a high titer of serum rheumatoid factor (357.0 IU/ml) and amyloid A protein (83.9 micrograms/ml) with positive antinuclear antibodies (homogeneous and speckled patterns). However, anti-neutrophil cytoplasmic autoantibody (ELISA), immune complexes and anti-glomerular basement membrane antibody (ELISA) were negative. Renal biopsy showed microscopic PN overlapping A-type positive amyloidosis. Although the maintenance of hemodialysis was necessary, aggressive immunosuppressive therapy with steroid pulse therapy and frequent plasma exchange provided a rapid improvement of systemic symptoms possibly due to vasculitis. We suggested that in this case, massive necrotizing crescentic glomerulonephritis with systemic arteritis developed on the basis of secondary amyloidosis due to rheumatoid arthritis. In such a case, even if various serum autoantibodies and immune complexes were negative, plasma exchange was suggested to be effective to remove not only pathogenic autoantibodies but also various serum inflammatory cytokines which may be related with severe vasculitis and glomerulitis, in addition to aggressive steroid therapy which may suppress the invasion of inflammatory cells producing these cytokines.
Assuntos
Amiloidose/etiologia , Arterite/etiologia , Artrite Reumatoide/complicações , Glomerulonefrite/etiologia , Anti-Inflamatórios/administração & dosagem , Feminino , Glomerulonefrite/terapia , Humanos , Glomérulos Renais/patologia , Metilprednisolona/administração & dosagem , Pessoa de Meia-Idade , Necrose , Troca PlasmáticaAssuntos
Subpopulações de Linfócitos B/citologia , Nódulos Linfáticos Agregados/imunologia , Proteínas Serina-Treonina Quinases/deficiência , Animais , Diferenciação Celular , Movimento Celular , Quimiocinas/genética , Quimiocinas/fisiologia , Citocinas/fisiologia , Proteínas de Ligação a DNA/deficiência , Proteínas de Ligação a DNA/genética , Regulação da Expressão Gênica , Deficiência de IgA/imunologia , Imunoglobulina A/imunologia , Síndromes de Imunodeficiência/genética , Síndromes de Imunodeficiência/imunologia , Síndromes de Imunodeficiência/patologia , Mucosa Intestinal/citologia , Mucosa Intestinal/imunologia , Tecido Linfoide/anormalidades , Camundongos , Camundongos Knockout , Camundongos Mutantes , NF-kappa B/metabolismo , Nódulos Linfáticos Agregados/citologia , Plasmócitos/patologia , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/fisiologia , RNA Mensageiro/biossíntese , Receptores de Quimiocinas/genética , Receptores de Quimiocinas/fisiologia , Transdução de Sinais , Baço/patologia , Quinase Induzida por NF-kappaBRESUMO
BACKGROUND: In a previous clinicopathological study, we observed mesangial factor V expression accompanied by the intact form of cross-linked fibrin deposition in the active type of IgA nephropathy. The conversion of prothrombin to thrombin by factor Xa is potently accelerated more than 104-fold by the presence of factor V, which is a membrane-bound cofactor. Another membrane-bound cofactor, tissue factor, is known to play an initiating role in the coagulation cascade and to be synthesized in mesangial cells (MCs) by the stimulation of tumor necrosis factor-alpha (TNF-alpha). However, the synthesis of factor V, which plays on the terminating stage of prothrombin activation, has not been reported previously in MCs by in vitro study. Our current study tested the coagulation process via expression of factor V by the stimulation of proinflammatory cytokine, TNF-alpha, in cultured human MCs. METHODS: To evaluate factor V protein expression, immunoperoxidase staining with densitometric evaluation and Western blot analysis were conducted after stimulation of TNF-alpha. To test factor V activity, stimulated MCs were incubated in combination with factor Xa, prothrombin, fibrinogen and factor XIII, and fibrin production on MCs was assessed after immunoperoxidase staining on the cell surface. In a blocking test using an antibody against factor V, suppression of fibrin production was evaluated to clarify the role of factor V activity. For the evaluation of factor V mRNA expression in cultured human MCs, in situ hybridization and Northern blot analysis were performed. RESULTS: Factor V protein expression in MCs after TNF-alpha stimulation increased both time- and dose-dependently. As a marker of factor V activity with exogenous factor Xa, fibrin production on TNF-alpha-stimulated MCs was increased in a time-dependent manner and was inhibited by the addition of anti-factor V antibody. Factor V mRNA was identified in MCs by in situ hybridization and showed an increase after stimulation with TNF-alpha on Northern blot analysis. CONCLUSIONS: Our data suggest that the coagulation process proceeds on MCs as the result of increased expression of endogenous factor V activity on its cell surface in cooperation with exogenous factor Xa.
Assuntos
Fator V/biossíntese , Mesângio Glomerular/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Anticorpos/imunologia , Células Cultivadas , Fator V/genética , Fator V/imunologia , Fator Xa/farmacologia , Fibrina/metabolismo , Mesângio Glomerular/efeitos dos fármacos , Humanos , Imuno-Histoquímica , Modelos Químicos , RNA Mensageiro/análise , RNA Mensageiro/biossínteseRESUMO
Because abnormalities of mucosal immunity have been suggested in human IgA nephropathy, we examined the involvement of mucosal immunity in IgA deposition to the kidney in hyper IgA (HIGA) mice, which was established as a mouse model for human IgA nephropathy with hyperserum IgA. The number of surface IgA+B220- lymphocytes in the intestinal lamina propria (LP) of HIGA mice increased 2.7-fold at 30 wk of age as compared with those at 10 wk of age, whereas normal mice did not show such increase. The surface IgA+B220- LP lymphocytes spontaneously secreted IgA in culture. Morphological studies showed that the surface IgA+B220- lymphocytes of murine intestinal LP are identical with plasma cells (PCs). About 20% of IgA+B220- PC in LP expressed both Mac-1 and CD19, suggesting that they may derive from peritoneal B-1 cells. Cell cycle study on intestinal IgA-PCs using bromodeoxyuridine revealed no difference between HIGA mice and normal mice, suggesting that the high frequency of IgA-producing PCs in HIGA mice is not due to enhanced proliferation or prolonged survival of IgA-producing PCs in LP. In addition, IgA secretion into the gut lumen of HIGA mice decreased drastically (to one forth) with aging. These data suggest that the increased number of intestinal IgA-producing PCs and the down-regulation of IgA excretion into the intestinal lumen might synergistically contribute to the hyperserum IgA in HIGA mice and resultant IgA deposition to the kidney.
Assuntos
Glomerulonefrite por IGA/imunologia , Imunoglobulina A/biossíntese , Mucosa Intestinal/citologia , Mucosa Intestinal/imunologia , Plasmócitos/metabolismo , Receptores de Antígenos de Linfócitos B/biossíntese , Animais , Ciclo Celular/imunologia , Fezes/química , Feminino , Glomerulonefrite por IGA/sangue , Glomerulonefrite por IGA/metabolismo , Imunoglobulina A/sangue , Imunoglobulina A/metabolismo , Mucosa Intestinal/metabolismo , Antígenos Comuns de Leucócito/biossíntese , Contagem de Linfócitos , Subpopulações de Linfócitos/imunologia , Subpopulações de Linfócitos/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Plasmócitos/imunologia , RNA Mensageiro/biossíntese , Receptores de Antígenos de Linfócitos B/sangue , Receptores de Imunoglobulina Polimérica/biossíntese , Receptores de Imunoglobulina Polimérica/genética , Especificidade da EspécieRESUMO
BACKGROUND/AIMS: Recently, we established a high serum IgA-prone inbred (HIGA) mouse strain as a murine model of spontaneous IgA nephropathy by selective mating of high serum IgA ddY mice, and found that they showed enhanced production of glomerular extracellular matrix components with increased expression of TGF-beta mRNA and protein in the kidneys. In this study, we examined the roles of lymphocytes in the development of high serum IgA in this strain. METHODS: We performed flow cytometric analyses of T and B cells in splenic mononuclear cells (SMNCs) from these mice using BALB/c mice as normal controls. We also compared serum TGF-beta1 concentrations and TGF-beta mRNA expression levels in the B-cell-depleted (T-cell-rich) fraction of SMNCs in these mice. RESULTS: HIGA mice showed significantly fewer CD3-positive cells compared with BALB/c mice when young, but not when aged. The CD4/CD8 ratio of HIGA mice was lower than that of BALB/c mice, but this difference was not significant. Although the number of B220-positive cells did not vary significantly, the ratio of surface IgA-positive B cells was significantly increased in both young and adult HIGA mice. The B-cell-depleted SMNCs from HIGA mice exhibited higher levels of expression of TGF-beta and TGF-beta1 mRNA than controls from a young age, which were maintained throughout life, but there were no differences in PDGF, MCP-1 or bFGF expression between these two strains. In contrast to local mRNA expression, serum TGF-beta1 concentration was decreased in HIGA mice compared with BALB/c controls. CONCLUSION: These findings suggest that the mating procedure performed to establish HIGA mice selected for a unique phenotype of local up-regulation of TGF-beta production in the kidneys, as well as T cells that may contribute to both the early and consistently high serum IgA expression and enhanced production of renal extracellular matrix components in HIGA mice. Additionally, TGF-beta1 may act locally, not systemically, in a paracrine or autocrine manner.
Assuntos
Glomerulonefrite por IGA/complicações , Glomerulosclerose Segmentar e Focal/complicações , Baço/imunologia , Linfócitos T/imunologia , Fator de Crescimento Transformador beta/biossíntese , Animais , Subpopulações de Linfócitos B , Linfócitos B/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Modelos Animais de Doenças , Citometria de Fluxo , Glomerulonefrite por IGA/sangue , Glomerulonefrite por IGA/metabolismo , Imunoglobulina A/sangue , Imunoglobulina G/sangue , Glomérulos Renais/imunologia , Contagem de Linfócitos , Camundongos , Camundongos Endogâmicos , RNA Mensageiro/biossíntese , RNA Mensageiro/sangue , Baço/metabolismo , Linfócitos T/metabolismo , Fator de Crescimento Transformador beta/sangue , Regulação para CimaRESUMO
BACKGROUND: Factor V in its active form (Va) plays a key role at the termination of the intrinsic coagulation pathway, serving as a membrane-bound cofactor for the conversion of prothrombin to thrombin by factor Xa. Cross-linked fibrin (XFb) is often observed in mesangial areas in active types of human glomerulonephritis. In this study, to clarify contribution of factor V in intramesangial coagulation, mesangial factor V expression and its relationship to mesangial proliferation and fibrin deposition in IgA nephropathy (IgAN) were investigated. METHODS: Twenty-two patients with IgAN were studied. XFb was detected in renal biopsy specimens using anti-d-dimer antibody combined with plasmin exposure, and factor V was detected with rabbit antibody against human factor V. Double-labeling immunohistochemistry was used to investigate the relationship of the glomerular distribution of factor V to XFb. The relationship of factor V staining to the activity index or XFb deposition was evaluated. The expression of factor V mRNA was assessed by in situ hybridization in relationship to the antigen staining of alpha-smooth muscle actin (alpha-SMA). The ultrastructural distribution of factor V in glomeruli was studied by immunoelectron microscopy. RESULTS: XFb and factor V were observed in the mesangium and along capillary loops in seven and nine specimens, respectively. Factor V had intense, frequent expression in the proliferating and necrotizing areas, showing a significant relationship to XFb (P < 0.05). Furthermore, XFb deposition and factor V expression were markedly correlated with disease activity (P = 0.005 and P = 0.008, respectively). By double-labeling experiments, XFb and factor V were often seen colocalized in mesangial areas of the glomeruli, which showed necrotizing lesions and/or intense cellular proliferation. By in situ hybridization, factor V mRNA was detected mainly in the mesangial cells, which were positive for alpha-SMA, and partly in the endothelial cells. By immunoelectron microscopy, factor V presence was confirmed in the mesangium and endothelium. CONCLUSION: The present findings suggest that factor V is strongly expressed in mesangial cells in active IgAN accompanied with mesangial proliferation and may exert procoagulant activity, leading to intramesangial coagulation.
Assuntos
Fator V/análise , Fibrina/análise , Mesângio Glomerular/química , Glomerulonefrite por IGA/metabolismo , Actinas/análise , Adulto , Biópsia , Reagentes de Ligações Cruzadas/análise , Fator V/metabolismo , Feminino , Mesângio Glomerular/patologia , Mesângio Glomerular/ultraestrutura , Glomerulonefrite por IGA/imunologia , Glomerulonefrite por IGA/patologia , Humanos , Técnicas Imunoenzimáticas , Masculino , Microscopia Imunoeletrônica , Pessoa de Meia-IdadeRESUMO
BACKGROUND: Our recently established high immunoglobulin (Ig)A inbred strain (HIGA) of ddY mice showed constantly high serum IgA levels, progressive mesangial sclerosis accompanied by IgA deposits, and elevated renal expression of transforming growth factor (TGF)-beta, mimicking IgA nephropathy. In the present study, we assessed the role of the immune system, especially of T cells, in this strain. METHODS: The in vitro production of interferon (IFN)-gamma, interleukin (IL)-4 and TGF-beta1 by splenic CD4+ T cells was assessed in HIGA mice at 14 and 28 weeks of age by comparison with age-matched C57BL/6 and BALB/c mice, T-helper (Th) 1, and Th2 prone controls respectively. Moreover, recombinant murine IL-12 was administered intraperitoneally to HIGA mice and serum IgA and renal lesions were analysed. RESULTS: The production of IFN-gamma by splenic CD4+ T cells was markedly upregulated in HIGA mice at both ages as compared with age-matched C57BL/6 and BALB/c mice. Although splenic CD4+ T cells from HIGA mice produced less IL-4 than those from BALB/c mice at both ages, the former produced significantly more IL-4 with age, which contrasted with the age-associated decrease in the latter. Moreover, TGF-beta1 production of these cells in HIGA mice was equal to or greater than that in the two groups of control mice at both ages. Daily intraperitoneal administration of IL-12 for 1 week significantly enhanced crescent formation with glomerular macrophage accumulation and interstitial cell infiltration, whereas it reduced the serum IgA level. CONCLUSIONS: In HIGA mice, Th1 is markedly upregulated from a young age and there is an age-associated Th2 increase with TGF-beta1 upregulation in helper T cells. The former may be related to the exacerbation of inflammatory renal lesions on IL-12 administration, while the latter may contribute to increased IgA production, leading to glomerular IgA deposition and progressive glomerulosclerosis in HIGA mice. The pathogenic role of T cell function and fluctuation of these subsets, especially the Th1/Th2 balance, is crucial to the immunopathological phenotype of the renal lesions in HIGA mice.