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2.
Arthritis Rheumatol ; 68(11): 2740-2751, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27159593

RESUMO

OBJECTIVE: In lupus nephritis, tubulointerstitial inflammation (TII) is associated with in situ adaptive immune cell networks that amplify local tissue damage. Since conventional therapy appears ineffective for severe TII, and these patients often progress to renal failure, understanding in situ mechanisms might reveal new therapeutic targets. This study was undertaken to assess whether dysregulated apoptotic regulators maintain local adaptive immunity and drive inflammation in TII. METHODS: This study utilized novel computational approaches that, when applied to multicolor confocal images, quantified apoptotic regulator protein expression in selected lymphocyte subsets. This approach was validated using laser-capture microdissection (LCM) coupled to quantitative polymerase chain reaction (qPCR). Furthermore, the consequences of dysregulated apoptotic mediator expression were explored in a murine model of lupus nephritis. RESULTS: Analyses of renal biopsy tissue from patients with lupus nephritis and those with mixed cellular renal allograft rejection revealed that the B cell lymphoma 2 protein (Bcl-2) was frequently expressed in infiltrating lymphocytes, whereas expression of myeloid cell leukemia 1 was low. In contrast, the reciprocal pattern of expression was observed in tonsil germinal centers. These results were consistent with RNA expression data obtained using LCM and qPCR. Bcl-2 was also highly expressed in tubulointerstitial infiltrates in (NZB × NZW)F1 (NZB/NZW) mice. Furthermore, treatment of NZB/NZW mice with ABT-199, a selective oral inhibitor of Bcl-2, prolonged survival and prevented proteinuria and development of TII in a lupus prevention model. Interestingly, glomerular immune complexes were partially ameliorated by ABT-199 treatment, and serum anti-double-stranded DNA antibody titers were unaffected. CONCLUSION: These data demonstrate that Bcl-2 is an attractive therapeutic target in patients with lupus nephritis who manifest TII.


Assuntos
Apoptose , Nefrite Lúpica/metabolismo , Linfócitos/metabolismo , Nefrite Intersticial/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Imunidade Adaptativa/imunologia , Animais , Complexo Antígeno-Anticorpo/efeitos dos fármacos , Complexo Antígeno-Anticorpo/imunologia , Complexo Antígeno-Anticorpo/metabolismo , Proteína 11 Semelhante a Bcl-2/genética , Proteína 11 Semelhante a Bcl-2/metabolismo , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Feminino , Centro Germinativo/metabolismo , Rejeição de Enxerto/imunologia , Rejeição de Enxerto/metabolismo , Humanos , Processamento de Imagem Assistida por Computador , Imuno-Histoquímica , Inflamação , Rim/efeitos dos fármacos , Rim/imunologia , Rim/metabolismo , Glomérulos Renais/efeitos dos fármacos , Glomérulos Renais/imunologia , Glomérulos Renais/metabolismo , Transplante de Rim , Microdissecção e Captura a Laser , Nefrite Lúpica/imunologia , Linfócitos/imunologia , Camundongos , Camundongos Endogâmicos NZB , Microscopia Confocal , Microscopia de Fluorescência , Proteína de Sequência 1 de Leucemia de Células Mieloides/genética , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Nefrite Intersticial/imunologia , Tonsila Palatina , Reação em Cadeia da Polimerase , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-bcl-2/genética , Sulfonamidas/farmacologia
3.
J Autoimmun ; 60: 51-58, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25921064

RESUMO

Systemic lupus erythematosus (SLE) is a complex autoimmune disease of uncertain etiology. Patients from different ancestral backgrounds demonstrate differences in clinical manifestations and autoantibody profiles. We examined genome-wide transcriptional patterns in major immune cell subsets across different ancestral backgrounds. Peripheral blood was collected from African-American (AA) and European-American (EA) SLE patients and controls. CD4 T-cells, CD8 T-cells, monocytes, and B cells were purified by flow sorting, and each cell subset from each subject was run on a genome-wide expression array. Cases were compared to controls of the same ancestral background. The overlap in differentially expressed gene (DEG) lists between different cell types from the same ancestral background was modest (<10%), and only 5-8% overlap in DEG lists was observed when comparing the same cell type between different ancestral backgrounds. IFN-stimulated gene (ISG) expression was not up-regulated synchronously in all cell types from a given patient, for example a given subject could have high ISG expression in T and B cells, but not in monocytes. AA subjects demonstrated more concordance in ISG expression between cell types from the same individual, and AA patients demonstrated significant down-regulation of metabolic gene expression which was not observed in EA patients. ISG expression was significantly decreased in B cells in patients taking immunosuppressants, while ISGs in other cell types did not differ with medication use. In conclusion, gene expression was strikingly different between immune cell subsets and between ancestral backgrounds in SLE patients. These findings emphasize the critical importance of studying multiple ancestral backgrounds and multiple cell types in gene expression studies. Ancestral backgrounds which are not studied will not benefit from personalized medicine strategies in SLE.


Assuntos
Afro-Americanos/genética , Grupo com Ancestrais do Continente Europeu/genética , Interferons/biossíntese , Lúpus Eritematoso Sistêmico/genética , Linfócitos B/classificação , Linfócitos B/imunologia , Linfócitos T CD4-Positivos/classificação , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/classificação , Linfócitos T CD8-Positivos/imunologia , Feminino , Expressão Gênica/genética , Estudo de Associação Genômica Ampla , Humanos , Interferons/genética , Monócitos/classificação , Monócitos/imunologia
4.
Arthritis Rheumatol ; 66(12): 3359-70, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-25306868

RESUMO

OBJECTIVE: In lupus nephritis (LN), severe tubulointerstitial inflammation (TII) predicts progression to renal failure. Severe TII is associated with tertiary lymphoid neogenesis and in situ antigen-driven clonal B cell selection. The autoantigen(s) driving in situ B cell selection in TII are not known. This study was undertaken to identify the dominant driving autoantigen(s). METHODS: Single CD38+ or Ki-67+ B cells were laser captured from 7 biopsy specimens that were diagnostic for LN. Eighteen clonally expanded immunoglobulin heavy- and light-chain variable region pairs were cloned and expressed as monoclonal antibodies. Seven more antibodies were cloned from flow-sorted CD38+ cells from an eighth biopsy specimen. Antigen characterization was performed using a combination of confocal microscopy, enzyme-linked immunosorbent assay, screening protoarrays, immunoprecipitation, and mass spectrometry. Serum IgG titers to the dominant antigen in 48 LN and 35 non-nephritic lupus samples were determined using purified antigen-coated arrays. Autoantigen expression on normal and LN kidney was localized by immunohistochemistry and immunofluorescence. RESULTS: Eleven of 25 antibodies reacted with cytoplasmic structures, 4 reacted with nuclei, and none reacted with double-stranded DNA. Vimentin was the only autoantigen identified by both mass spectrometry and protoarray. Ten of the 11 anticytoplasmic TII antibodies directly bound vimentin. Vimentin was highly expressed by tubulointerstitial inflammatory cells, and the TII antibodies tested preferentially bound inflamed tubulointerstitium. Finally, high titers of serum antivimentin antibodies were associated with severe TII (P = 0.0001). CONCLUSION: Vimentin, an antigenic feature of inflammation, is a dominant autoantigen targeted in situ in LN TII. This adaptive autoimmune response likely feeds forward to worsen TII and renal damage.


Assuntos
Autoanticorpos/imunologia , Autoantígenos/imunologia , Rim/imunologia , Nefrite Lúpica/imunologia , Nefrite Intersticial/imunologia , Vimentina/imunologia , Adolescente , Adulto , Biópsia , Estudos de Casos e Controles , Ensaio de Imunoadsorção Enzimática , Humanos , Imunidade Humoral , Imuno-Histoquímica , Imunoprecipitação , Inflamação , Rim/patologia , Nefrite Lúpica/patologia , Espectrometria de Massas , Microscopia Confocal , Nefrite Intersticial/patologia , Índice de Gravidade de Doença , Adulto Jovem
5.
Curr Opin Nephrol Hypertens ; 23(3): 204-10, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24685591

RESUMO

PURPOSE OF REVIEW: To focus on the latest data that elucidate the role of the NLRP3 inflammasome in kidney diseases. RECENT FINDINGS: The NLRP3 inflammasome is not limited by the traditional microbial stimuli of innate immunity and its connection with autophagy, apoptosis, fibrosis, and pro-inflammatory cytokines has broader implications for a variety of kidney diseases. In a wide spectrum of glomerular and tubulointerstitial diseases, the NLRP3 inflammasome is upregulated in both classical immune cells such as infiltrating macrophages and resident dendritic cells as well as in renal tubular epithelial cells, and even podocytes. Inhibition of the NLRP3 inflammasome ameliorates renal injury in a variety of animal models. Interestingly, this extends to models of proteinuria, which suggests that the deleterious effect of albuminuria on the proximal tubular epithelium and podocytes is, in part, mediated by inflammasome activation. SUMMARY: Recent studies in animal models, and limited studies in humans, suggest a broad role for inflammasome activation in renal disease. Surprisingly, individual components of the inflammasome, independent of inflammasome activation, may also contribute to progressive renal injury. Additional, studies are needed to define the relative importance of the inflammasome in specific diseases and the therapeutic opportunities afforded by targeting the inflammasome.


Assuntos
Proteínas de Transporte/metabolismo , Inflamassomos/metabolismo , Nefropatias/metabolismo , Rim/metabolismo , Transdução de Sinais , Animais , Anti-Inflamatórios/uso terapêutico , Proteínas de Transporte/antagonistas & inibidores , Humanos , Inflamassomos/antagonistas & inibidores , Inflamassomos/imunologia , Mediadores da Inflamação/metabolismo , Rim/efeitos dos fármacos , Rim/imunologia , Rim/fisiopatologia , Nefropatias/tratamento farmacológico , Nefropatias/imunologia , Nefropatias/fisiopatologia , Proteína 3 que Contém Domínio de Pirina da Família NLR , Transdução de Sinais/efeitos dos fármacos
6.
Front Immunol ; 4: 309, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24101921

RESUMO

BACKGROUND: In systemic lupus erythematosus (SLE), antibodies directed at RNA-binding proteins (anti-RBP) are associated with high serum type I interferon (IFN), which plays an important role in SLE pathogenesis. African-Americans (AA) are more likely to develop SLE, and SLE is also more severe in this population. We hypothesized that peripheral blood gene expression patterns would differ between AA and European-American (EA) SLE patients, and between those with anti-RBP antibodies and those who lack these antibodies. METHODS: Whole blood RNA from 33 female SLE patients and 16 matched female controls from AA and EA ancestral backgrounds was analyzed on Affymetrix Gene 1.0 ST gene expression arrays. Ingenuity Pathway Analysis was used to compare the top differentially expressed canonical pathways amongst the sample groups. An independent cohort of 116 SLE patients was used to replicate findings using quantitative real-time PCR (qPCR). RESULTS: Both AA and EA patients with positive anti-RBP antibodies showed over-expression of similar IFN-related canonical pathways, such as IFN Signaling (P = 1.3 × 10(-7) and 6.3 × 10(-11) in AA vs. EA respectively), Antigen Presenting Pathway (P = 1.8 × 10(-5) and 2.5 × 10(-6)), and a number of pattern recognition receptor pathways. In anti-RBP negative (RBP-) patients, EA subjects demonstrated similar IFN-related pathway activation, whereas no IFN-related pathways were detected in RBP-AA patients. qPCR validation confirmed similar results. CONCLUSION: Our data show that IFN-induced gene expression is completely dependent on the presence of autoantibodies in AA SLE patients but not in EA patients. This molecular heterogeneity suggests differences in IFN-pathway activation between ancestral backgrounds in SLE. This heterogeneity may be clinically important, as therapeutics targeting this pathway are being developed.

7.
Clin Dev Immunol ; 2012: 780436, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-23251221

RESUMO

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by multiple genetic risk factors, high levels of interferon alpha (IFN-α), and the production of autoantibodies against components of the cell nucleus. Interferon regulatory factor 5 (IRF5) is a transcription factor which induces the transcription of IFN-α and other cytokines, and genetic variants of IRF5 have been strongly linked to SLE pathogenesis. IRF5 functions downstream of Toll-like receptors and other microbial pattern-recognition receptors, and immune complexes made up of SLE-associated autoantibodies seem to function as a chronic endogenous stimulus to this pathway. In this paper, we discuss the physiologic role of IRF5 in immune defense and the ways in which IRF5 variants may contribute to the pathogenesis of human SLE. Recent data regarding the role of IRF5 in both serologic autoimmunity and the overproduction of IFN-α in human SLE are summarized. These data support a model in which SLE-risk variants of IRF5 participate in a "feed-forward" mechanism, predisposing to SLE-associated autoantibody formation, and subsequently facilitating IFN-α production downstream of Toll-like receptors stimulated by immune complexes composed of these autoantibodies.


Assuntos
Doenças Autoimunes/imunologia , Fatores Reguladores de Interferon/imunologia , Lúpus Eritematoso Sistêmico/imunologia , Animais , Autoanticorpos/imunologia , Autoimunidade/imunologia , Humanos
8.
Clin Dev Immunol ; 2012: 682018, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22988468

RESUMO

Systemic lupus erythematosus (SLE) is a highly heterogeneous autoimmune disorder characterized by differences in autoantibody profiles, serum cytokines, and clinical manifestations. We have previously conducted a case-case genome-wide association study (GWAS) of SLE patients to detect associations with autoantibody profile and serum interferon alpha (IFN-α). In this study, we used public gene expression data sets to rationally select additional single nucleotide polymorphisms (SNPs) for validation. The top 200 GWAS SNPs were searched in a database which compares genome-wide expression data to genome-wide SNP genotype data in HapMap cell lines. SNPs were chosen for validation if they were associated with differential expression of 15 or more genes at a significance of P < 9 × 10(-5). This resulted in 11 SNPs which were genotyped in 453 SLE patients and 418 matched controls. Three SNPs were associated with SLE-associated autoantibodies, and one of these SNPs was also associated with serum IFN-α (P < 4.5 × 10(-3) for all). One additional SNP was associated exclusively with serum IFN-α. Case-control analysis was insensitive to these molecular subphenotype associations. This study illustrates the use of gene expression data to rationally select candidate loci in autoimmune disease, and the utility of stratification by molecular phenotypes in the discovery of additional genetic associations in SLE.


Assuntos
Autoanticorpos/genética , Perfilação da Expressão Gênica , Lúpus Eritematoso Sistêmico/genética , Lúpus Eritematoso Sistêmico/imunologia , Polimorfismo de Nucleotídeo Único , Autoanticorpos/imunologia , Linhagem Celular , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Humanos , Interferon-alfa/sangue , Fenótipo
9.
J Rheumatol ; 39(6): 1238-40, 2012 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-22505704

RESUMO

OBJECTIVE: To investigate and refine the relationships among systemic lupus erythematosus (SLE) and related autoantibodies, interferon-α (IFN-α), and various ancestral backgrounds. METHODS: We investigated quantitatively defined genetic ancestry through principal component analysis in place of self-reported ancestry. RESULTS: African ancestry was found to be associated with presence of anti-RNP antibody (p = 0.0026), and anti-RNP was correlated with high levels of IFN-α (p = 2.8 × 10(-5)). CONCLUSION: Our data support a model in which African ancestry increases the likelihood of SLE-associated autoantibody formation, which subsequently results in higher levels of serum IFN-α.


Assuntos
Grupo com Ancestrais do Continente Africano/genética , Anticorpos Antinucleares/sangue , Predisposição Genética para Doença , Interferon-alfa/sangue , Lúpus Eritematoso Sistêmico/genética , Grupo com Ancestrais do Continente Africano/etnologia , Feminino , Humanos , Lúpus Eritematoso Sistêmico/sangue , Lúpus Eritematoso Sistêmico/etnologia , Masculino , Pessoa de Meia-Idade , Ohio/epidemiologia , Análise de Componente Principal , Ribonucleoproteínas/sangue , Ribonucleoproteínas/imunologia
10.
Am Surg ; 69(3): 225-9; discussion 229-30, 2003 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-12678479

RESUMO

False negative (FN) results limit the efficacy of technetium-99m-sestamibi scanning for parathyroid localization. We determined the incidence of FN results and attempted to correlate it with clinical and operative findings. One hundred forty-six patients underwent parathyroidectomy; 89 had primary hyperparathyroidism (76 single adenoma and 13 multiglandular disease) and underwent sestamibi scanning. The false negative rate was 22 per cent with an overall sensitivity of 77 per cent and a positive predictive value of 99 per cent. Patients with single adenomas were more likely to have a true positive scan than those with multiglandular disease [83% vs 38%; odds ratio (OR) = 7.754, 95% confidence interval (CI) = 2.184-27.524; P < or = 0.0001]. Inferior adenomas (90% vs 59%; OR = 6.261, 95% CI = 2.037-19.243; P < or = 0.0001) and larger adenomas (1422.3 +/- 1576.2 vs 474.6 +/- 193.2 g; P < or = 0.0001) were more likely to be detected by sestamibi imaging. Patients with normal preoperative calcium levels were more likely to have an FN sestamibi scan. Sestamibi parathyroid imaging is limited by a 22 per cent FN rate and is less accurate for detecting abnormal parathyroid tissue in patients with small adenomas, multiglandular disease, superior adenomas, or preoperative normocalcemia.


Assuntos
Hiperparatireoidismo/diagnóstico por imagem , Glândulas Paratireoides/diagnóstico por imagem , Paratireoidectomia , Compostos Radiofarmacêuticos , Tecnécio Tc 99m Sestamibi , Adenoma/diagnóstico por imagem , Adulto , Idoso , Cálcio/sangue , Reações Falso-Negativas , Feminino , Humanos , Hiperparatireoidismo/sangue , Hiperparatireoidismo/cirurgia , Masculino , Pessoa de Meia-Idade , Neoplasias das Paratireoides/diagnóstico por imagem , Cintilografia , Sensibilidade e Especificidade
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