RESUMEN
Immunoglobulin A (IgA) glycosylation, recognized as an important pathogenic factor in IgA nephropathy (IgAN), is apparently controlled by the polarity of T helper (Th) cytokine responses. To examine the role of cytokine polarity in IgAN, inbred mice were immunized by intraperitoneal priming with inactivated Sendai virus (SeV) emulsified in either complete Freund's adjuvant (CFA) or incomplete Freund's adjuvant (IFA), which promote Th1- or Th2-immune response, respectively, and then boosted identically twice orally with aqueous suspensions of inactivated virus. Next, some mice were challenged intranasally with infectious SeV. Mice primed with CFA or IFA had equal reductions in nasal viral titre relative to non-immune controls, and equally increased serum levels of SeV-specific IgA antibody. Mice primed with CFA showed higher SeV-specific IgG than those with IFA. Splenocytes from mice primed with IFA produced copious amounts of interleukin (IL)-4 and IL-5, but little interferon-gamma and IL-2; those primed with CFA had reciprocal cytokine recall responses. Total serum IgA and especially SeV-specific IgA from mice primed with IFA showed a selective defect in sialylation and galactosylation. Although the frequency and intensity of glomerular deposits and haematuria did not differ, glomerulonephritis in mice primed with IFA and challenged with infectious virus was more severe than in those given CFA, as judged by serum creatinine level. We conclude that the polarity of T cell cytokines controls the pattern of IgA glycosylation and exerts direct or indirect effects on functional glomerular responses to immune complex deposition.
Asunto(s)
Citocinas/inmunología , Glomerulonefritis por IGA/inmunología , Inmunoglobulina A/inmunología , Virus Sendai/inmunología , Células TH1/inmunología , Células Th2/inmunología , Adyuvantes Inmunológicos , Animales , Citocinas/metabolismo , Modelos Animales de Enfermedad , Ensayo de Inmunoadsorción Enzimática , Femenino , Adyuvante de Freund , Glicosilación , Inmunoglobulina A/sangre , Inmunoglobulina G/sangre , Inmunoglobulina G/inmunología , Lípidos , Ratones , Ratones Endogámicos BALB C , Líquido del Lavado Nasal/inmunología , Células TH1/metabolismo , Células Th2/metabolismoRESUMEN
Mucosal antigenic exposure is implicated in pathogenesis of IgA nephropathy. Although IgG and/or IgM codeposits may promote disease, protracted mucosal antigenic exposure reduces IgG and IgM antibody, a process termed mucosal tolerance. We immunized mice intranasally with infectious or inactivated Sendai virus for 6 or 14 weeks. Anti-virus IgG remained high in mice given infectious virus for 14 weeks, but decreased after 6 weeks in mice given inactivated virus; IgA antibody remained high in both groups. Upon viral challenge, glomerular IgG and complement deposits and the frequency of hematuria, all equal after 6 weeks of immunization, were lower in mice immunized with inactivated virus for 14 weeks but remained high in mice given infectious virus; glomerular IgA increased over time in both immunized groups. Viremia in a non-tolerized immune host can promote glomerulonephritis with IgG and complement codeposits and glomerular dysfunction. These preliminary experiments may guide future, more mechanistic, investigation.
Asunto(s)
Inmunoglobulina A/inmunología , Enfermedades Renales/inmunología , Infecciones por Respirovirus/inmunología , Virus Sendai/inmunología , Animales , Técnica del Anticuerpo Fluorescente , Hematuria/inmunología , Enfermedades del Complejo Inmune/inmunología , Inmunidad Mucosa , Inmunoglobulina A/sangre , Riñón/inmunología , Ratones , Proteinuria/inmunologíaRESUMEN
Hyperfunction of Th2 cells and aberrant glycosylation of IgA have been proposed independently as factors in the pathogenesis of IgA nephropathy (IgAN), the most common form of glomerulonephritis. To investigate the relationship between Th2 cytokines and IgA glycosylation in the genesis of IgAN, we induced IgAN in C3HeB and BALB/c mice by oral immunization and intranasal challenge with Sendai virus. Although both strains of mice developed microhaematuria and glomerular IgA immune deposits to similar degrees, only BALB/c mice developed significant renal insufficiency. More profound reductions of terminal galactosylation and sialylation occurred in Sendai virus-specific IgA from BALB/c versus C3HeB mice, and splenocytes from immunized BALB/c mice produced more Th2 and less Th1 cytokines compared to C3HeB mice when stimulated with antigen in vitro. Furthermore, the decreased glycosylation of IgA elicited by Th2 cytokines in vitro was blunted by the addition of IFN-gamma. We conclude that increased production of Th2 cytokines can lead to abnormalities in IgA glycosylation, which in turn promote heightened phlogistic responses to IgA immune complexes lodging in the glomerulus. We suggest that a relative or absolute increase in Th2 cytokine production in response to mucosal infection is a significant pathogenic factor in human IgAN.
Asunto(s)
Citocinas/biosíntesis , Glomerulonefritis por IGA/inmunología , Inmunoglobulina A/química , Linfocitos T/inmunología , Animales , Anticuerpos Antivirales/sangre , Complejo Antígeno-Anticuerpo/metabolismo , Femenino , Glicosilación , Inmunoglobulina A/sangre , Interferón gamma/biosíntesis , Interleucina-2/biosíntesis , Interleucina-4/biosíntesis , Interleucina-5/biosíntesis , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C3H , Virus Sendai/inmunología , Células TH1/inmunología , Células Th2/inmunologíaRESUMEN
Decay-accelerating factor (DAF) is a membrane regulator of C3 activation that protects self cells from autologous complement attack. In humans, DAF is uniformly expressed as a glycosylphosphatidylinositol (GPI)-anchored molecule. In mice, both GPI-anchored and transmembrane-anchored DAF proteins are produced, each of which can be derived from two different genes (Daf1 and Daf2). In this report, we describe a Daf1 gene knock-out mouse arising as the first product of a strategy for targeting one or both Daf genes. As part of the work, we characterize recently described monoclonal antibodies against murine DAF protein using deletion mutants synthesized in yeast, and then employ the monoclonal antibodies in conjunction with wild-type and the Daf1 knock-out mice to determine the tissue distribution of the mouse Daf1 and Daf2 gene products. To enhance the immunohistochemical detection of murine DAF protein, we utilized the sensitive tyramide fluorescence method. In wild-type mice, we found strong DAF labelling of glomeruli, airway and gut epithelium, the spleen, vascular endothelium throughout all tissues, and seminiferous tubules of the testis. In Daf1 knock-out mice, DAF labelling was ablated in most tissues, but strong labelling of the testis and splenic dendritic cells remained. In both sites, reverse transcription-polymerase chain reaction analyses identified both GPI and transmembrane forms of Daf2 gene-derived protein. The results have relevance for studies of in vivo murine DAF function and of murine DAF structure.
Asunto(s)
Antígenos CD55/metabolismo , Proteínas de Saccharomyces cerevisiae , Animales , Anticuerpos Monoclonales/inmunología , Sitios de Unión/inmunología , Antígenos CD55/genética , Antígenos CD55/inmunología , Proteínas de Caenorhabditis elegans , Complemento C3/análisis , Ciclinas/genética , Proteínas Fúngicas/genética , Técnicas para Inmunoenzimas , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Receptor de Insulina/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Distribución TisularRESUMEN
OBJECTIVE: To investigate the efficacy of a novel therapy (proteases) in an animal model of rheumatoid arthritis, and to investigate the mechanisms of arthritogenesis. METHODS: We induced progressive arthritis in male DBA/1 mice by immunization and boosting with Type II collagen; groups of mice were treated orally twice daily with either ibuprofen or proteases, or were left untreated. After 2 weeks, joints were scored for clinical, radiographic, and histologic changes. In addition, we measured serum levels of IgG anti-collagen II, the glycosylation of circulating total and anti-collagen II IgG, and cytokine production by lymphocytes isolated from lymph nodes. RESULTS: Amelioration of joint inflammation, and accentuation of a prototypical Th2 cytokine (interleukin 5) were similar in the ibuprofen and protease treatment groups. However, protease treatment protects and preserves articular cartilage, normalizes the sialylation of IgG and anti-collagen antibody, and fully restores Th1 (interferon-gamma) synthesis, distinct from ibuprofen. CONCLUSION: Protease therapy has antiinflammatory efficacy in the early (inflammatory) phase of collagen induced arthritis, similar to ibuprofen. The immunomodulatory effects of proteases, not seen with ibuprofen, may underlie a correction of aberrant IgG glycosylation and/or contribute to the increased capacity of protease to delay or forestall erosive and destructive arthritis or ankylosis. Similar effects may apply to spontaneous RA in humans.
Asunto(s)
Anticuerpos Antiidiotipos/análisis , Artritis Reumatoide/tratamiento farmacológico , Artritis Reumatoide/patología , Ibuprofeno/farmacología , Inmunoglobulina G/análisis , Péptido Hidrolasas/farmacología , Animales , Artritis Reumatoide/diagnóstico por imagen , Artritis Reumatoide/inmunología , Colágeno , Modelos Animales de Enfermedad , Inmunohistoquímica , Masculino , Ratones , Ratones Endogámicos DBA , Radiografía , Valores de Referencia , Sensibilidad y Especificidad , Resultado del TratamientoRESUMEN
The capacity of mucosal IgA Abs to serve as an excretory immune system in vivo was investigated. Mice expressing a transgenic TCR were immunized intragastrically with the cognate Ag to elicit a vigorous mucosal IgA Ab response. Soon after i.v. challenge, Ag was detected within the epithelial cells of the small intestinal crypts and to a lesser degree within the epithelial cells higher up the villi, paralleling the gradient in expression of the polymeric Ig receptor and the transport of its ligand, oligomeric IgA. Uptake of Ag into the epithelial cells occurred only from the basolateral aspect and only when Ag complexed to IgA Ab could be present in the lamina propria. The results support the concept that local IgA Abs can excrete Ags from the body by transporting them directly through mucosal epithelial cells, using the same mechanism that transports free IgA into the mucosal secretions.
Asunto(s)
Inmunoglobulina A/fisiología , Mucosa Intestinal/inmunología , Mucosa Intestinal/metabolismo , Animales , Especificidad de Anticuerpos , Complejo Antígeno-Anticuerpo/metabolismo , Biotinilación , Pollos , Femenino , Inmunidad Mucosa , Inmunización Secundaria , Inmunoglobulina A/biosíntesis , Inmunoglobulina A/metabolismo , Inyecciones Intravenosas , Mucosa Intestinal/citología , Intubación Gastrointestinal , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Transgénicos , Ovalbúmina/administración & dosificación , Ovalbúmina/inmunología , Ovalbúmina/metabolismoRESUMEN
IgA nephropathy (IgAN) is a form of immune complex glomerulonephritis that occurs spontaneously in humans. This unit describes the induction of active disease in inbred mice, utilizing inert proteins or a common viral pathogen as the inciting antigen. An alternate protocol is offered for the induction of disease in rats by noninfectious protein antigens. Support protocols are presented for the evaluation of the extent of disease, for preparation of infectious and inactivated suspensions of viral stock, and for quantification of the virus.
Asunto(s)
Modelos Animales de Enfermedad , Glomerulonefritis por IGA/inmunología , Administración Oral , Animales , Apoferritinas/administración & dosificación , Bovinos , Pollos , Femenino , Glomerulonefritis por IGA/virología , Caballos , Humanos , Inyecciones , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C3H , Ovalbúmina/administración & dosificación , Ratas , Ratas Endogámicas Lew , Ratas Wistar , Virus Sendai/inmunología , Albúmina Sérica/administración & dosificación , gammaglobulinas/administración & dosificaciónRESUMEN
BACKGROUND: Signal transduction by mesangial cell (MC) integrins regulates cell growth and survival, extracellular matrix production, and organization. The aim of the study was to investigate human MC integrin modulation by differently glycosylated IgA and macromolecular IgA, which are thought to play a pathogenetic role in IgA nephropathy (IgAN). METHODS: MCs were incubated with purified human polymeric IgA, heat-aggregated IgA, IgA glycoforms generated by enzymatic hydrolysis of saccharide residues and serum fractions from IgAN patients, and controls isolated by lectin affinity and containing IgA with peculiar glycan patterns. Integrins were quantitated by flow cytometry. RESULTS: Cultured MCs highly expressed alphavbeta3 and some alpha3beta1; alphavbeta3 was up-regulated by matrix components (P < 0.02). In vitro desialylated and degalactosylated polymeric human IgA enhanced alphavbeta3 expression on cultured MCs (P < 0.001). Serum IgA glycoforms isolated from IgAN patients with high exposure of internal sugars, GalNAc, Neu5Ac2,6GalNAc, and Man enhanced alphav expression on cultured MCs more than healthy controls. CONCLUSIONS.: These data support the hypothesis that IgA glycation plays a role in modulating the cell-matrix interaction, and that this mechanism can be operating in IgAN.
Asunto(s)
Glomerulonefritis por IGA/inmunología , Glomerulonefritis por IGA/metabolismo , Inmunoglobulina A/metabolismo , Inmunoglobulina A/farmacología , Receptores de Vitronectina/biosíntesis , Anticuerpos Monoclonales , Apoptosis , Células Cultivadas , Matriz Extracelular/metabolismo , Citometría de Flujo , Galactosa/metabolismo , Glicosilación , Humanos , Técnicas para Inmunoenzimas , Técnicas In Vitro , Integrina alfa3beta1 , Integrinas/análisis , Integrinas/biosíntesis , Integrinas/inmunología , Glomérulos Renales/citología , Glomérulos Renales/inmunología , Glomérulos Renales/metabolismo , Sustancias Macromoleculares , Ácido N-Acetilneuramínico/metabolismo , Receptores de Vitronectina/análisis , Receptores de Vitronectina/inmunologíaRESUMEN
We sought to determine whether selected cytokines, known to stimulate profoundly B-cell activation and differentiation, also have as yet unrecognized effects upon the glycosylation of secreted Ig and/or membrane-associated proteins. The glycosylation of both secreted IgM and membrane-bound MHC Class-I synthesized by CH12LX cells was detected by enzyme-lectin conjugates in immunoabsorption assays. Stimulation of B cells with IL-4 plus IL-5 significantly decreases the terminal glycosylation of secreted IgM, whereas LPS has a minor effect, despite the fact that both stimuli are equipotent for IgM secretion. Neither LPS nor IL-4 plus IL-5 affect MHC Class-I expression. However, IL-4 plus IL-5 substantially increases the terminal glycosylation of MHC Class-I produced from both mIgM(+)and mIgA(+)CH12LX cells. LPS has no or a modest effect on the terminal glycosylation of MHC Class-I produced from CH12LX cells. These results suggest that Th(2)-derived cytokines differentially influence the glycosylation of secreted and membrane-associated glycoproteins of B cells. In turn, this might elucidate the basis of aberrant glycosylation reported in conditions such as IgA nephropathy, cancer and rheumatoid arthritis.
Asunto(s)
Linfocitos B/inmunología , Antígenos de Histocompatibilidad Clase I/metabolismo , Inmunoglobulina M/metabolismo , Interleucina-4/inmunología , Interleucina-5/inmunología , Animales , Glicosilación , Antígenos de Histocompatibilidad Clase I/biosíntesis , Hibridomas , Inmunoglobulina M/biosíntesis , Ratones , Células Tumorales CultivadasRESUMEN
BACKGROUND: The clinical use of cyclosporine (CsA) is limited by its nephrotoxicity. Apoptosis, perhaps instigated by increased nitric oxide synthase (NOS) activity, may play a role in such toxicity. METHODS: Human mesangial cells, human tubular cells, human umbilical vein endothelial cells, or murine endothelial cells were cultured with CsA at final concentrations of 0 to 1000 ng/mL for 4 to 24 hours. As inhibitors of apoptosis, 0.01 mol/L L-nitromethylarginine (L-NAME) or 1 microg/mL cycloheximide (CHX) was added, whereas 0.01 mol/L sodium nitroprusside (as a nitric oxide donor) was used as a positive control. Apoptosis was assessed by using TUNEL method and by DNA fragmentation by electrophoresis. In addition, NOS enzymatic activity, Northern blots for inducible NOS (iNOS) mRNA, and immunohistochemically demonstrable iNOS protein were evaluated. RESULTS: Within 12 to 24 hours, CsA significantly increased the fraction (8 to 35%) of apoptotic cells in each cell line, according to the dose. Fragmentation of DNA confirmed apoptosis. L-NAME and CHX inhibited the phenomenon, whereas sodium nitroprusside enhanced it. Each cell line significantly increased NOS activity in response to CsA, an effect blunted by L-NAME and CHX. Neither inhibitor modified the increased iNOS mRNA expression elicited by CsA. Positive staining for both iNOS and p53 proteins was observed in all cell lines incubated with CsA that were inhibited by CHX; L-NAME inhibited only p53 staining. CONCLUSIONS: CsA induces apoptosis in various renal cell lines, and this effect is mediated by the induction of iNOS via p53. These effects may contribute to the acellular fibrosis characteristic of late CsA nephrotoxicity.
Asunto(s)
Apoptosis/fisiología , Ciclosporina/farmacología , Riñón/efectos de los fármacos , Riñón/fisiología , Óxido Nítrico/fisiología , Células Cultivadas , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/farmacología , Humanos , Inmunohistoquímica , Riñón/citología , Riñón/enzimología , NG-Nitroarginina Metil Éster/farmacología , Óxido Nítrico Sintasa/genética , Óxido Nítrico Sintasa/metabolismo , ARN Mensajero/metabolismoRESUMEN
Immunoglobulin (Ig)A provides the initial immune barrier to viruses at mucosal surfaces. Specific IgA interrupts viral replication in polarized epithelium during receptor-mediated transport, probably by binding to newly synthesized viral proteins. Here, we demonstrate by immunoelectron microscopy that specific IgA monoclonal antibodies (mAbs) accumulate within Sendai virus-infected polarized cell monolayers and colocalize with the hemagglutinin- neuraminidase (HN) viral protein in a novel intracellular structure. Neither IgG specific for HN nor irrelevant IgA mAbs colocalize with viral protein. Treatment of cultures with viral-specific IgA but not with viral-specific IgG or irrelevant IgA decreases viral titers. These observations provide definitive ultrastructural evidence of a subcellular compartment in which specific IgA and viral envelope proteins interact, further strengthening our hypothesis of intracellular neutralization of virus by specific IgA antibodies. Our results have important implications for intracellular protein trafficking, viral replication, and viral vaccine development.
Asunto(s)
Proteína HN/análisis , Inmunoglobulina A/análisis , Respirovirus/enzimología , Animales , Línea Celular , Polaridad Celular , Perros , Epitelio/metabolismo , Epitelio/virología , Ratones , Ratones Endogámicos BALB C , Respirovirus/fisiologíaRESUMEN
BACKGROUND: Chronic renal allograft rejection is characterized histologically by transplantation-associated arteriosclerosis and glomerulosclerosis (Tx-AA and Tx-AGS). Recent studies in animal models implicate the mitogenic and pressor actions of endothelin-1 (ET-1) in Tx-AA. In humans, however, a link between elevated ET-1 secretion and Tx-AA or Tx-AGS remains unclear. In this study we analyzed expression of ET-1 in the vasculature of renal transplant patients with chronic or acute rejection and in normal controls. METHODS: Renal vascular and glomerular ET-1 was assessed by immunohistochemistry in 12 patients with clinically and histologically defined chronic rejection, in 11 patients with acute rejection, and in 5 normal kidneys. ET-1 staining was also correlated with various clinical parameters and with a morphometric index of neointima formation. ET-1 secretion was measured by ELISA in cultured human vascular cell types treated with T cell- and macrophage-associated cytokines. RESULTS: We found that renal allografts with chronic rejection and Tx-AA expressed 6.1-fold more ET-1 in the vasculature relative to allografts with acute rejection or to normal kidneys (P < 0.01). In Tx-AA, ET-1 was detected predominantly in the neointima, which contained mostly endothelial cells and smooth muscle cells. A strong positive correlation (r = 0.82, P < 0.01) was observed between vascular ET-1 peptide expression and hypertension in patients with chronic rejection. We also showed that macrophage-associated cytokines, but not T cell-associated cytokines, stimulated ET-1 secretion in human endothelial cells, vascular smooth muscle and mesangial cells. CONCLUSIONS: These results demonstrate that elevated ET-1 in the neointima is associated with Tx-AA and chronic rejection. In addition, these results point to an important role for endothelial dysfunction in chronic renal allograft rejection.
Asunto(s)
Arteriosclerosis/sangre , Endotelina-1/sangre , Rechazo de Injerto , Trasplante de Riñón/inmunología , Músculo Liso Vascular/química , Adolescente , Adulto , Anciano , Arteriosclerosis/etiología , Células Cultivadas , Citocinas/farmacología , Endotelina-1/metabolismo , Femenino , Mesangio Glomerular/patología , Humanos , Masculino , Persona de Mediana Edad , Trasplante HomólogoRESUMEN
To understand better the function of endothelin-1 (ET-1) in renal physiology, we examined vascular and glomerular expression of ET-1 in normal human kidney and in lupus nephritis. Immunohistochemical analysis revealed that renal endothelium of glomeruli, arteries, veins, and capillaries expressed ET-1. Endothelial cells were the principal source of glomerular ET-1; positive immunostaining was detected only rarely in mesangial cells and vascular smooth muscle cells from normal kidney. However, mesangial staining for ET-1 was elevated in patients with lupus nephritis, suggesting that under certain conditions mesangial cells elaborate ET-1. Indeed cultured human mesangial cells from normal subjects secreted ET-1 peptide. ET-1 secretion was augmented by the protein kinase C activator phorbol ester and by transforming growth factor-beta1 (TGF-beta1), a cytokine implicated in the development of glomerulosclerosis. Transient transfection of cultured mesangial cells with a preproET-1 reporter construct showed that the preproET-1 promoter is transcriptionally active in mesangial cells and is stimulated by TGF-beta1, phorbol ester, or ectopic expression of protein kinase beta1. Cultured human mesangial cells have both ETA and ETB receptors that contribute to ET-1-stimulated mitogenesis. Taken together, these results demonstrate that ET-1 is expressed at sites where paracrine or autocrine signaling by ET-1 might control renal vasoconstriction, glomerular filtration rate, and remodeling of the glomerulus in renal disease.
Asunto(s)
Endotelina-1/biosíntesis , Endotelio Vascular/metabolismo , Glomérulos Renales/metabolismo , Nefritis Lúpica/metabolismo , Circulación Renal/fisiología , Células Cultivadas , Regulación de la Expresión Génica/efectos de los fármacos , Mesangio Glomerular/metabolismo , Humanos , Músculo Liso Vascular/metabolismo , Reacción en Cadena de la Polimerasa , Proteína Quinasa C/metabolismo , ARN Mensajero/biosíntesis , Proteínas Recombinantes/biosíntesis , Acetato de Tetradecanoilforbol/farmacología , Transcripción Genética , Transfección , Factor de Crecimiento Transformador beta/farmacologíaRESUMEN
Despite progress in techniques, early or late deep infection develops in 1 to 5% of patients with prosthetic joint replacements. We report a retrospective review of 64 original frozen sections (FSs) compared with permanent sections, the preoperative clinical and intraoperative findings, and the subsequent culture results. A histologic section, frozen or permanent, was considered positive for infection if there were more than five polymorphonuclear leukocytes (PMNs) per high power field, excluding surface fibrin and inflammatory exudate, in at least five separate microscopic fields. The sensitivity of an FS as a diagnostic test to detect prosthetic infection when present was 43%. The specificity of an FS to correctly identify the absence of infection was 97%. These data support the conclusion that the FS is reasonably specific but not a sensitive diagnostic modality. If the preoperative evaluation is completely negative and the FS does not reveal PMNs, the reimplantation can be performed with minimal concern regarding the possibility of infection. If the preoperative and intraoperative evaluations are completely negative but PMNs are observed in the FS, then the surgeon should consider proceeding with caution; the decision to proceed with reimplantation might be delayed, depending on the surgeon's clinical assessment.
Asunto(s)
Artroplastia de Reemplazo de Cadera , Artroplastia de Reemplazo de Rodilla , Infecciones Bacterianas/patología , Articulación de la Cadera/patología , Articulación de la Rodilla/patología , Neutrófilos/patología , Infecciones Bacterianas/microbiología , Biopsia/métodos , Reacciones Falso Negativas , Secciones por Congelación , Articulación de la Cadera/microbiología , Humanos , Articulación de la Rodilla/microbiología , Recuento de Leucocitos , Reoperación , Estudios Retrospectivos , Sensibilidad y EspecificidadRESUMEN
We sought to determine whether selected cytokines, known to profoundly increase proliferation and/or production of Ig by B cells and their progeny, also have as yet unrecognized effects upon IgA glycosylation. For these studies, we selected CH12LX mouse B lymphoma cells, a widely used model of B cell differentiation. Glycosylation was assessed by detection with enzyme-lectin conjugates in an immunoabsorption assay and verified by profiling and sequencing of the N-linked oligosaccharides. Stimulation of B cells with IL-4 plus IL-5 significantly alters the terminal glycosylation of secreted IgA, whereas LPS has a minor effect, despite the fact that both stimuli are equipotent at inducing Ig class switching and Ig secretion. Moreover, the alteration in terminal glycosylation was more profound on IgA secreted from surface IgM+ than from surface IgA+ CH12LX cells. These results suggest that the increased production of IL-4 and IL-5 by peripheral blood lymphocytes from IgA nephropathy patients might result in the production of abnormally glycosylated IgA. In turn, this abnormally glycosylated IgA may promote deposition of IgA in glomeruli in this disease.
Asunto(s)
Linfocitos B/metabolismo , Inmunoglobulina A/metabolismo , Interleucina-4/farmacología , Interleucina-5/farmacología , Lectinas de Plantas , Células Th2/metabolismo , Animales , Conformación de Carbohidratos , Secuencia de Carbohidratos , Cromatografía de Afinidad , Glicosilación/efectos de los fármacos , Lectinas/metabolismo , Lipopolisacáridos/farmacología , Linfoma de Células B/patología , Ratones , Datos de Secuencia Molecular , Monosacáridos/análisis , Procesamiento Proteico-Postraduccional/efectos de los fármacos , Proteínas Recombinantes/farmacología , Proteínas Inactivadoras de Ribosomas , Células Tumorales CultivadasRESUMEN
Renal disease in patients infected with human immunodeficiency virus (HIV) often presents with significant proteinuria and progressive renal failure; focal glomerulosclerosis is the most common renal pathology identified. To our knowledge, we report the first case of nephrotic-range proteinuria and preserved renal function in an HIV-infected patient in association with disseminated histoplasmosis. The initial level of proteinuria was 12.5 g/24 h. The patient developed a concomitant lesion on his neck, which was biopsied and identified as Histoplasma capsulatum by fungal stains and culture. The serum CF titer of antibody against yeast antigens of H. capsulatum was 1:8. The level of serum albumin decreased to 2.0 g/dL, and the level of serum cholesterol increased to 284 mg/dL. Immunohistochemical staining of renal biopsy tissue demonstrated immune complexes within the mesangium; H. capsulatum antigen was also demonstrated in the mesangium. Therapy with oral itraconazole resulted in marked clinical improvement. The findings in this case emphasize the need to rule out treatable causes of the nephrotic syndrome in AIDS, especially in cases of immune-complex glomerulonephritis.
Asunto(s)
Infecciones Oportunistas Relacionadas con el SIDA/microbiología , Glomerulonefritis Membranoproliferativa/diagnóstico , Histoplasmosis/complicaciones , Proteinuria/diagnóstico , Administración Oral , Anticuerpos Antifúngicos/análisis , Anticuerpos Antifúngicos/sangre , Antifúngicos/administración & dosificación , Antifúngicos/uso terapéutico , Complejo Antígeno-Anticuerpo/análisis , Biopsia , Colesterol/análisis , Colesterol/sangre , Diagnóstico Diferencial , Glomerulonefritis Membranoproliferativa/microbiología , Histoplasmosis/tratamiento farmacológico , Histoplasmosis/inmunología , Humanos , Inmunohistoquímica , Itraconazol/administración & dosificación , Itraconazol/uso terapéutico , Riñón/inmunología , Riñón/patología , Masculino , Persona de Mediana Edad , Proteinuria/microbiología , Albúmina Sérica/análisisRESUMEN
Heavy alcohol intake and/or lipotrope-deficient diet induced hepatocellular injury and mesangial deposition of IgA and often IgG in Lewis rats. The experimental animals showing more severe urinary abnormalities and histologic damage in the glomeruli had increased levels of IgA antibodies to dietary antigens and altered intestinal permeability. Based on human studies, the prolonged circulation of IgA-containing complexes associated with the liver disease could be envisaged as important for the development of mesangial IgA deposits. In order to verify this hypothesis, four groups (G) of Lewis rats were studied: G1 received thrice a weak an intragastric infusion of 1.5 ml/100 g body wt of whiskey; G2 rats were nourished with lipotrope-deficient diet; G3 rats were given both whiskey and LD diet; G4 rats were nourished with regular chow. After 12 weeks, heat-aggregated rat monomeric IgA was labeled with 133I and intravenously injected. Three control subgroups of rats, one given whiskey, one nourished with LD diet, and one with regular chow, were injected with radiolabeled heat-aggregated rat IgG. A large field-of-view digital gamma camera, equipped with an ultra-high-resolution collimator and interfaced to a dedicated computer, was used to analyze tracer kinetics and fate. The liver was the main organ involved in clearance of both test probes. The hepatic mean transit (MTT) was 11.4 +/- 11 min in G1 (proteinuria of 6.9 +/- 1.41 mg/day and hematuria +/+2), 221 +/- 19 min in G2 (proteinuria 9.1 +/- 0.64 mg/day and hematuria +2/+3), and 230 +/- 15 min in G3 (proteinuria 9.5 +/- 0.58 mg/day and hematuria +2/+3). In each case MTT value was found to be significantly prolonged compared to G4 (85 +/- 4 min). The multiple regression analysis showed that MTT values, proteinuria, and hematuria were significantly correlated (P < 0.01). Controls had trace amount proteinuria (0.82 +/- 0.17 mg/day, significantly lower than for each study group, P < 0.08) and undetectable hematuria. Similar results were obtained in control rats injected with aggregated IgG; i.e., MTT values were more prolonged in rats given whiskey or LD diet than normally nourished rats (P < 0.01). The lipotrope-deficient diet and the chronic alcohol abuse per se seem to lead to critical changes in hepatic uptake and catabolism of both an IgA and an IgG aggregate, which could account in turn for the reported appearance of renal immunoglobulin deposits in this experimental model. Due to the comparable delay in removal of IgA and IgG probes in equally nourished animals, additional factors are likely to be involved in the prominent deposition of IgA.
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Inmunoglobulina A/metabolismo , Inmunoglobulina A/fisiología , Hepatopatías/inmunología , Hepatopatías/metabolismo , Hígado/metabolismo , Animales , Complejo Antígeno-Anticuerpo/sangre , Enfermedad Crónica , Modelos Animales de Enfermedad , Técnica del Anticuerpo Fluorescente Directa , Glomerulonefritis por IGA/sangre , Humanos , Inmunoglobulina A/sangre , Radioisótopos de Yodo , Hígado/diagnóstico por imagen , Masculino , Cintigrafía , Ratas , Ratas Endogámicas LewRESUMEN
BACKGROUND & AIMS: Microvascular endothelial cells mediate leukocyte homing, angiogenesis, and inflammation and healing and show tissue-specific adhesion molecules and functions. The activation of human intestinal mucosal microvascular endothelial cells (HIMECs) was studied in vitro to uncover possible abnormalities associated with inflammatory bowel disease. METHODS: HIMECs were isolated from normal and inflammatory bowel disease mucosa and assessed for phenotypic and morphological features, proliferative response, leukocyte binding capacity, and adhesion molecule expression. RESULTS: Basal proliferation by HIMECs was less than that of human umbilical vein endothelial cells (HUVECs) but increased proportionally more in response to vascular endothelial growth factor. Proinflammatory stimuli induced an activated, spindle-shaped morphology in HIMEC monolayers. Compared with HUVECs, unstimulated HIMECs showed less adhesiveness for U937 and MOLT4 cells and neutrophils, but cytokines and lipopolysaccharide substantially increased the binding capacity of HIMECs. HIMECs derived from inflammatory bowel disease mucosa showed a markedly greater leukocyte-binding capacity than normal mucosal HIMECs. Patterns of intercellular adhesion molecule 1, vascular cell adhesion molecule 1 and E-selectin messenger RNA expression were distinct in HIMECs, HUVECs, and mucosal mesenchymal cells. CONCLUSIONS: HIMECs represent differentiated endothelial cells with unique functional properties. Their dramatically enhanced capacity to bind leukocytes in inflammatory bowel disease suggests that HIMECs play an important role in initiating or maintaining inflammation.
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Endotelio/metabolismo , Enfermedades Inflamatorias del Intestino/metabolismo , Mucosa Intestinal/metabolismo , Leucocitos/metabolismo , Humanos , InmunohistoquímicaAsunto(s)
Bases de Datos Factuales/normas , Glomerulonefritis por IGA , Biopsia , Supervivencia sin Enfermedad , Genotipo , Glomerulonefritis por IGA/genética , Glomerulonefritis por IGA/inmunología , Glomerulonefritis por IGA/mortalidad , Glomerulonefritis por IGA/patología , Humanos , Pronóstico , Índice de Severidad de la Enfermedad , Análisis de SupervivenciaRESUMEN
IgA nephropathy (IgAN) is defined by the predominant deposition of IgA immune complexes (IC) in the glomerular mesangium. Interaction between IgA immune complexes and mesangial cells (MC) could be a linchpin for the genesis of IgAN. We studied the modulation of MC expression of IgA receptors (Fc alphaR) by selected cytokines. Binding of 125I-IgA to quiescent human MC showed 2.55 x 10(5) sites/cell with an affinity (Ka) of 3.2 x 10(7) M(-1). Addition of selected recombinant cytokines had no significant influence on Ka, but increased the number of sites/cell relative to unstimulated cells. Northern hybridization using the pHuFc alphaR cDNA probe showed time-dependent increases in mRNA expression in stimulated versus control cells. IL-6 and tumour necrosis factor-alpha (TNF-alpha) had a biphasic effect on the Fc alphaR mRNA level; at 48 h, IL-6 increased steady state mRNA levels about six-fold relative to control, TNF-alpha increased mRNA four-fold, and interferon-gamma (IFN-gamma) induced Fc alphaR mRNA two-fold. By reverse transcriptase-polymerase chain reaction (RT-PCR), the Fc alphaR expressed on human MC appears highly homologous to that expressed by U937 cells. Altered Fc alphaR expression in response to cytokines may influence the pathogenesis of IgAN by affecting deposition and/or clearance of IgA-IC in the mesangium.