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1.
BMJ Case Rep ; 12(10)2019 Oct 10.
Artigo em Inglês | MEDLINE | ID: mdl-31604714

RESUMO

We report the case of a woman with Cogan's syndrome concomitant with the wish to have children. After three major flares of the disease that led to unilateral deafness, immunosuppressive therapy with prednisolone and azathioprine was started. Because of the severe side effects, an off-label therapy with intravenous immunoglobulin (IVIG) was initiated, under which our patient has since given birth to three healthy children. To our knowledge this is the first report to describe Cogan's syndrome with multiple successful pregnancies under IVIG treatment.

2.
Brain ; 2018 Jul 09.
Artigo em Inglês | MEDLINE | ID: mdl-29985992

RESUMO

The transcription factor BCL11B is essential for development of the nervous and the immune system, and Bcl11b deficiency results in structural brain defects, reduced learning capacity, and impaired immune cell development in mice. However, the precise role of BCL11B in humans is largely unexplored, except for a single patient with a BCL11B missense mutation, affected by multisystem anomalies and profound immune deficiency. Using massively parallel sequencing we identified 13 patients bearing heterozygous germline alterations in BCL11B. Notably, all of them are affected by global developmental delay with speech impairment and intellectual disability; however, none displayed overt clinical signs of immune deficiency. Six frameshift mutations, two nonsense mutations, one missense mutation, and two chromosomal rearrangements resulting in diminished BCL11B expression, arose de novo. A further frameshift mutation was transmitted from a similarly affected mother. Interestingly, the most severely affected patient harbours a missense mutation within a zinc-finger domain of BCL11B, probably affecting the DNA-binding structural interface, similar to the recently published patient. Furthermore, the most C-terminally located premature termination codon mutation fails to rescue the progenitor cell proliferation defect in hippocampal slice cultures from Bcl11b-deficient mice. Concerning the role of BCL11B in the immune system, extensive immune phenotyping of our patients revealed alterations in the T cell compartment and lack of peripheral type 2 innate lymphoid cells (ILC2s), consistent with the findings described in Bcl11b-deficient mice. Unsupervised analysis of 102 T lymphocyte subpopulations showed that the patients clearly cluster apart from healthy children, further supporting the common aetiology of the disorder. Taken together, we show here that mutations leading either to BCL11B haploinsufficiency or to a truncated BCL11B protein clinically cause a non-syndromic neurodevelopmental delay. In addition, we suggest that missense mutations affecting specific sites within zinc-finger domains might result in distinct and more severe clinical outcomes.

3.
Front Immunol ; 8: 439, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28450870

RESUMO

Neutrophil extracellular traps (NETs) are chromatin filaments decorated with enzymes from neutrophil cytoplasmic granules. Anti-neutrophil cytoplasmic antibodies (ANCAs) bind to enzymes from neutrophil cytoplasmic granules and are biomarkers for the diagnosis of systemic vasculitides. ANCA diagnostics are based on indirect immunofluorescence (IIF) of ethanol-fixed neutrophils. IIF shows a cytoplasmic staining pattern (C-ANCA) due to autoantibodies against proteinase 3 (PR3) or a perinuclear staining pattern (P-ANCA) due to autoantibodies against myeloperoxidase (MPO). The distinct ANCA-staining patterns are an artifact of ethanol fixation. Here, we tested NETs as a substrate for the detection of ANCAs in human sera. We observed that P-ANCAs specifically stained NETs, while C-ANCAs targeted the cell bodies of netting neutrophils. The distinct ANCA-staining patterns were caused by the presence of MPO, but not PR3, in NETs. Using NETs as a substrate for IIF, we characterized ANCAs in sera of patients with ANCA-associated vasculitis (AAV). Furthermore, we inhibited serine proteases by diisopropylfluorophosphate to prevent chromatin unfolding and the release of NETs and thus generated neutrophils with MPO-positive nuclei and PR3-positive cytoplasm, which resembled the appearance of ethanol-fixed neutrophils. In conclusion, our data suggest that NETs are selectively loaded with antigens recognized by P-ANCAs, and netting neutrophils provide a physiological substrate for ANCA detection in patients with AAV.

4.
J Allergy Clin Immunol ; 135(4): 1031-43.e6, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25240785

RESUMO

BACKGROUND: Anaphylaxis is an acute, potentially lethal, multisystem syndrome resulting from the sudden release of mast cell-derived mediators into the circulation. OBJECTIVES AND METHODS: We report here that a plasma protease cascade, the factor XII-driven contact system, critically contributes to the pathogenesis of anaphylaxis in both murine models and human subjects. RESULTS: Deficiency in or pharmacologic inhibition of factor XII, plasma kallikrein, high-molecular-weight kininogen, or the bradykinin B2 receptor, but not the B1 receptor, largely attenuated allergen/IgE-mediated mast cell hyperresponsiveness in mice. Reconstitutions of factor XII null mice with human factor XII restored susceptibility for allergen/IgE-mediated hypotension. Activated mast cells systemically released heparin, which provided a negatively charged surface for factor XII autoactivation. Activated factor XII generates plasma kallikrein, which proteolyzes kininogen, leading to the liberation of bradykinin. We evaluated the contact system in patients with anaphylaxis. In all 10 plasma samples immunoblotting revealed activation of factor XII, plasma kallikrein, and kininogen during the acute phase of anaphylaxis but not at basal conditions or in healthy control subjects. The severity of anaphylaxis was associated with mast cell degranulation, increased plasma heparin levels, the intensity of contact system activation, and bradykinin formation. CONCLUSIONS: In summary, the data collectively show a role of the contact system in patients with anaphylaxis and support the hypothesis that targeting bradykinin generation and signaling provides a novel and alternative treatment strategy for anaphylactic attacks.


Assuntos
Anafilaxia/imunologia , Anafilaxia/metabolismo , Fator XII/metabolismo , Hipersensibilidade/imunologia , Hipersensibilidade/metabolismo , Mastócitos/imunologia , Adulto , Idoso , Anafilaxia/complicações , Anafilaxia/genética , Animais , Biomarcadores , Bradicinina/metabolismo , Modelos Animais de Doenças , Fator XII/antagonistas & inibidores , Fator XII/genética , Feminino , Humanos , Hipersensibilidade/complicações , Hipersensibilidade/genética , Hipotensão/etiologia , Cininogênios/metabolismo , Masculino , Camundongos Knockout , Pessoa de Meia-Idade , Receptor B2 da Bradicinina/genética , Receptor B2 da Bradicinina/metabolismo , Transdução de Sinais , Fatores de Tempo , Adulto Jovem
5.
Pediatr Transplant ; 17(8): 757-64, 2013 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-24164827

RESUMO

Individualization of immunosuppressive medications is an important objective in transplantation medicine. Reliable biomarkers to distinguish between patients dependent from intensive immunosuppressive therapy and those where therapy can be minimized among pediatric transplant recipients receiving immunosuppressive medications are still not established. We evaluated the potential of cross-sectional quantification of regulatory T cells, lymphocyte subsets, and cytokine concentrations as biomarkers in 60 pediatric liver transplant recipients with AR, CR, or normal graft function and in 11 non-transplanted patients. Transplant recipients presenting with AR had significantly higher CD8+ T-cell counts, significantly higher concentrations of IL-2, and increased levels of IFN-γ compared with asymptomatic patients or controls. Regulatory T-cell numbers did not differ between children with rejection and children with good graft function. A tendency toward increased concentrations of IL-4 and TGF-ß was detected in transplant recipients with good graft function. Cross-sectional parameters of peripheral regulatory T cells in pediatric liver transplant recipients do not seem to be valuable biomarkers for individualizing immunosuppressive therapy prior to the weaning process. Lymphocyte subsets, IL-2, IFN-γ, IL-4, and TGF-ß serum concentrations may be helpful to identify children in whom immunosuppression can be reduced or discontinued.


Assuntos
Biomarcadores/sangue , Transplante de Fígado , Linfócitos T Reguladores/citologia , Adolescente , Linfócitos T CD8-Positivos/citologia , Separação Celular , Criança , Pré-Escolar , Estudos Transversais , Feminino , Citometria de Fluxo , Rejeição de Enxerto , Sobrevivência de Enxerto , Humanos , Imunossupressão , Imunossupressores/uso terapêutico , Lactente , Interferon gama/sangue , Interleucina-2/sangue , Interleucina-4/sangue , Transplante de Rim , Masculino , Tacrolimo/uso terapêutico , Fator de Crescimento Transformador beta/sangue
6.
J Clin Endocrinol Metab ; 97(8): E1402-10, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-22659246

RESUMO

CONTEXT: Glucocorticoids (GC) are powerful endogenous and therapeutic modulators of inflammation and play a critical role for controlling autoimmunity. GC resistance can be seen in patients with cell-mediated autoimmune disorders, but it is unknown whether this represents a stable trait or a state. OBJECTIVE: The objective of the study was to determine whether GC resistance of T cell responses is dynamically regulated in experimental autoimmune encephalomyelitis (EAE) and multiple sclerosis (MS). DESIGN: This was a translational observational study. PATIENTS AND ANIMALS: EAE was induced in C57BL/6 mice. A cross-sectional sample of 25 patients with relapsing-remitting MS was included as well as four MS patients during pregnancy and postpartum. MAIN OUTCOME MEASURES: Outcome measures included GC sensitivity of T cell proliferation and GC-mediated apoptosis. RESULTS: GC resistance was seen in both autoantigen-specific and nonspecific responses of T cells obtained from mice with EAE. GC resistance preceded clinical symptoms and central nervous system infiltration of immune cells. T cells obtained during EAE were resistant to GC-induced apoptosis, and this was linked to down-regulation of GC receptor-α expression. GC resistance in T cells was also seen in MS patients with radiological evidence for ongoing inflammation. GC resistance was absent in the MS patients during pregnancy, when relapse risk is decreased, but recurred postpartum, a time of increased relapse risk. CONCLUSIONS: These data demonstrate that GC resistance during autoimmune neuroinflammation is dynamically regulated. This has implications for the timing of steroid treatments and provides a putative pathway to explain the observed association between psychological stress and exacerbation of autoimmune diseases.


Assuntos
Encefalomielite Autoimune Experimental/tratamento farmacológico , Glucocorticoides/uso terapêutico , Esclerose Múltipla/tratamento farmacológico , Animais , Apoptose/efeitos dos fármacos , Autoantígenos/imunologia , Estudos Transversais , Resistência a Medicamentos , Encefalomielite Autoimune Experimental/imunologia , Feminino , Glicoproteínas/imunologia , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Esclerose Múltipla/imunologia , Glicoproteína Mielina-Oligodendrócito , Fragmentos de Peptídeos/imunologia , Receptores de Glucocorticoides/genética , Linfócitos T/imunologia
7.
Psychoneuroendocrinology ; 37(10): 1712-8, 2012 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-22455832

RESUMO

Multiple sclerosis (MS) is an inflammatory, demyelinating disease of the CNS with a high prevalence of depression. Both MS and depression have been linked to elevated cortisol levels and inflammation, indicating disturbed endocrine-immune regulation. An imbalance in mineralocorticoid versus glucocorticoid signaling in the CNS has been proposed as a pathogenetic mechanism of depression. Intriguingly, both receptors are also expressed in lymphocytes, but their role for 'escape' of the immune system from endocrine control is unknown. Using steroid sensitivity of T cell function as a read-out system, we here investigate a potential role of mineralocorticoid receptor (MR) versus glucocorticoid receptor (GR) regulation in the immune system as a biological mechanism underlying MS-associated major depression. Twelve female MS patients meeting diagnostic criteria for current major depressive disorder (MDD) were compared to twelve carefully matched MS patients without depression. We performed lymphocyte phenotyping by flow cytometry. In addition, steroid sensitivity of T cell proliferation was tested using hydrocortisone as well as MR (aldosterone) and GR (dexamethasone) agonists. Sensitivity to hydrocortisone was decreased in T cells from depressed MS patients. Experiments with agonists suggested disturbed MR regulation, but intact GR function. Importantly, there were no differences in lymphocyte composition and frequency of T cell subsets, indicating that the differences in steroid sensitivity are unlikely to be secondary to shifts in the immune compartment. To our knowledge, this study provides first evidence for altered steroid sensitivity of T cells from MS patients with comorbid MDD possibly due to MR dysregulation.


Assuntos
Anti-Inflamatórios/farmacologia , Transtorno Depressivo Maior/tratamento farmacológico , Hidrocortisona/farmacologia , Esclerose Múltipla/tratamento farmacológico , Linfócitos T/efeitos dos fármacos , Adulto , Aldosterona/agonistas , Aldosterona/farmacologia , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Dexametasona/agonistas , Dexametasona/farmacologia , Relação Dose-Resposta a Droga , Feminino , Citometria de Fluxo , Humanos , Hidrocortisona/agonistas , Imunofenotipagem
8.
J Neurol Neurosurg Psychiatry ; 82(7): 814-8, 2011 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-21296901

RESUMO

OBJECTIVE: Depression and fatigue are among the most common symptoms of multiple sclerosis (MS). These symptoms frequently co-occur and partially overlap in MS but their underlying biological substrates are unclear. In this study, the relative role of cytokines and hypothalamic-pituitary-adrenal (HPA) axis activity in depression and fatigue were examined in patients with relapsing-remitting MS (RRMS). METHODS: HPA axis function and frequency of stimulated cytokine (interferon γ (IFNγ) and tumour necrosis factor α (TNFα)) producing T cells was measured cross sectionally in 44 female patients with RRMS. All subjects completed a neurological examination, the Structured Clinical Interview for DSM-IV Axis I Disorders (SCID-I) and self-report questionnaires. RESULTS: 10 patients met diagnostic criteria for major depressive disorder (MDD). MS patients with comorbid MDD showed normal morning but elevated evening salivary cortisol levels, resulting in a flattened slope. While a higher frequency of cytokine producing CD8+ T cells was also seen in MS patients with MDD, these markers were more closely associated with fatigue than depression. CONCLUSIONS: This study supports a role for HPA axis hyperactivity in major depression in MS. In addition, inflammatory and neuroendocrine factors may differentially mediate fatigue and depressive symptoms.


Assuntos
Transtorno Depressivo Maior/etiologia , Transtorno Depressivo Maior/fisiopatologia , Glândulas Endócrinas/fisiopatologia , Fadiga/etiologia , Sistema Imunitário/fisiopatologia , Esclerose Múltipla Recidivante-Remitente/complicações , Esclerose Múltipla Recidivante-Remitente/fisiopatologia , Adulto , Citocinas/metabolismo , Manual Diagnóstico e Estatístico de Transtornos Mentais , Feminino , Citometria de Fluxo , Humanos , Hidrocortisona/sangue , Sistema Hipotálamo-Hipofisário/fisiopatologia , Interferon gama/metabolismo , Contagem de Linfócitos , Pessoa de Meia-Idade , Esclerose Múltipla Recidivante-Remitente/psicologia , Análise de Regressão , Fatores Socioeconômicos , Inquéritos e Questionários , Linfócitos T/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Adulto Jovem
9.
Autoimmun Rev ; 9(11): 785-92, 2010 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-20647062

RESUMO

With all the incredible progress in scientific research over the past two decades, the trigger of the majority of autoimmune disorders remains largely elusive. Research on the biology of T helper type 17 (T(H)17) cells over the last decade not only clarified previous observations of immune regulations and disease manifestations, but also provided considerable information on the signaling pathways mediating the effects of this lineage and its seemingly dual role in fighting the invading pathogens on one hand, and in frightening the host by inducing chronic inflammation and autoimmunity on the other hand. In this context, recent reports have implicated T(H)17 cells in mediating host defense as well as a growing list of autoimmune diseases in genetically-susceptible individuals. Herein, we summarize the current knowledge on T(H)17 in autoimmunity with emphasis on its differentiation factors and some mechanisms involved in initiating pathological events of autoimmunity.


Assuntos
Doenças Autoimunes/imunologia , Autoimunidade , Interleucina-17/imunologia , Transdução de Sinais , Células Th17/imunologia , Células Th17/metabolismo , Animais , Linfócitos T CD4-Positivos/imunologia , Citocinas/metabolismo , Inflamação/imunologia , Inflamação/metabolismo , Camundongos , Receptores de Citocinas/metabolismo , Fator de Transcrição STAT3/metabolismo , Ativação Transcricional , Fatores de Crescimento Transformadores/metabolismo
10.
J Immunol ; 183(7): 4693-704, 2009 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-19734217

RESUMO

Infiltration of T cells into the kidney is a typical feature of human and experimental lupus nephritis that contributes to renal tissue injury. The chemokine receptor CXCR3 is highly expressed on Th1 cells and is supposed to be crucial for their trafficking into inflamed tissues. In this study, we explored the functional role of CXCR3 using the MRL/MpJ-Fas(lpr) (MRL/lpr) mouse model of systemic lupus erythematosus that closely resembles the human disease. CXCR3(-/-) mice were generated and backcrossed into the MRL/lpr background. Analysis of 20-wk-old CXCR3(-/-) MRL/lpr mice showed amelioration of nephritis with reduced glomerular tissue damage and decreased albuminuria and T cell recruitment. Most importantly, not only the numbers of renal IFN-gamma-producing Th1 cells, but also of IL-17-producing Th17 cells were significantly reduced. Unlike in inflamed kidneys, there was no reduction in the numbers of IFN-gamma- or IL-17-producing T cells in spleens, lymph nodes, or the small intestine of MRL/lpr CXCR3(-/-) mice. This observation suggests impaired trafficking of effector T cells to injured target organs, rather than the inability of CXCR3(-/-) mice to mount efficient Th1 and Th17 immune responses. These findings show a crucial role for CXCR3 in the development of experimental lupus nephritis by directing pathogenic effector T cells into the kidney. For the first time, we demonstrate a beneficial effect of CXCR3 deficiency through attenuation of both the Th1 and the newly defined Th17 immune response. Our data therefore identify the chemokine receptor CXCR3 as a promising therapeutic target in lupus nephritis.


Assuntos
Interleucina-17/fisiologia , Rim/imunologia , Rim/metabolismo , Nefrite Lúpica/imunologia , Nefrite Lúpica/metabolismo , Receptores CXCR3/fisiologia , Células Th1/imunologia , Células Th1/metabolismo , Animais , Movimento Celular/genética , Movimento Celular/imunologia , Modelos Animais de Doenças , Feminino , Regulação da Expressão Gênica/imunologia , Humanos , Imunoglobulina G/metabolismo , Rim/patologia , Lúpus Eritematoso Sistêmico/imunologia , Lúpus Eritematoso Sistêmico/patologia , Nefrite Lúpica/genética , Nefrite Lúpica/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos MRL lpr , Camundongos Knockout , Receptores CXCR3/biossíntese , Receptores CXCR3/deficiência , Receptores CXCR3/genética , Células Th1/patologia
11.
Ann N Y Acad Sci ; 1173: 166-73, 2009 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-19758146

RESUMO

The detection of autoantibodies is an important element in the diagnosis and monitoring of disease progression in patients with autoimmune diseases. In laboratory diagnostic tests for connective tissue and autoimmune liver diseases, indirect immunofluorescence on HEp-2 cells plays a central role in a multistage diagnostic process. Despite the high quality of diagnostics, findings at different laboratories can differ considerably due to a lack of standardization, as well as subjective factors. The present paper formulates recommendations for the standardized processing and interpretation of the HEp-2 cell test for the detection of non-organ-specific (especially antinuclear) antibodies. It provides requirements regarding the diagnostic tests used, instructions for laboratory procedure and evaluation, and recommendations for interpretation. For an optimal laboratory diagnostic process, it is useful to have an informative, tentative clinical diagnosis and an experienced laboratory diagnostician. In addition, the following key elements are recommended: initial screening using indirect immunofluorescence on carefully chosen HEp-2 cells beginning with a serum dilution of 1:80 and evaluation under a microscope with powerful illumination; results from a titer of 1:160 upwards being considered positive; internal laboratory quality control; and standardized interpretation. The aim is to improve diagnostic tests and care of patients with autoimmune diseases as a central concern of the European Autoimmunity Standardization Initiative (EASI).


Assuntos
Autoanticorpos/sangue , Técnica Indireta de Fluorescência para Anticorpo/métodos , Anticorpos Antinucleares/sangue , Linhagem Celular Tumoral , Técnica Indireta de Fluorescência para Anticorpo/normas , Humanos , Padrões de Referência , Reprodutibilidade dos Testes
12.
Autoimmun Rev ; 9(1): 17-22, 2009 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-19245860

RESUMO

Analysis of autoantibodies (AAB) by indirect immunofluorescence (IIF) remains the hallmark of diagnosing autoimmune diseases despite the introduction of multiplex techniques. Non-organ specific AAB are screened in routine diagnostics by IIF on HEp-2 cells. However, IIF results vary due to objective (e.g., cell fixation) and subjective factors (e.g., expert knowledge). Therefore, inter- and intralaboratory variance is relatively high. Standardisation of AAB testing by IIF remains a critical issue in and between routine laboratories and may be improved by automated interpretation systems. An overview of existing interpretation techniques will be given taking into account own data of the first fully automated reading system AKLIDES. The novel system provides fully automated reading of IIF images and software algorithms for the mathematical description of IIF AAB patterns. It can be used for screening and preclassification of non-organ specific AAB in routine diagnostics regarding systemic autoimmune and autoimmune liver diseases. Furthermore, this system paves the way for economic data processing of cell-based IIF assays and can contribute to the reduction of interlaboratory variance of AAB testing. More sophisticated pattern recognition algorithms and novel calibration systems will improve standardised quantifications of IIF image interpretation.


Assuntos
Autoanticorpos/sangue , Doenças Autoimunes/diagnóstico , Técnica Indireta de Fluorescência para Anticorpo/estatística & dados numéricos , Doenças Autoimunes/sangue , Doenças Autoimunes/epidemiologia , Doenças Autoimunes/imunologia , Linhagem Celular , Processamento Eletrônico de Dados , Técnica Indireta de Fluorescência para Anticorpo/normas , Humanos , Processamento de Imagem Assistida por Computador , Variações Dependentes do Observador
13.
Ann N Y Acad Sci ; 1109: 31-6, 2007 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-17785287

RESUMO

The German Regional Group of EASI was established during the annual Meeting of the German Society of Immunology in Kiel in September 2005. Since this initial informative meeting, an active core group of about a dozen rheumatologists, immunologists, and laboratory specialists has been generating starter projects. In general, these projects do focus on clinically associated diagnostic questions, and do integrate a variety of specialists with profound knowledge in several related subjects. The aims of the German EASI group are to contribute to the definition of standards and to improve patient care. Therefore, the group is establishing guidelines for the diagnosis of autoimmune diseases, to standardize and improve their quality, combining the experience of clinical and laboratory specialists. The diagnostic activities focus currently on systemic lupus erythematosus (SLE) and on rheumatoid arthritis. These activities include laboratory investigations and diagnosis through clinical manifestations. Standardized diagnostics cannot be based solely on vague symptoms and positive laboratory tests. In laboratory diagnostics, standardization and implementation of objective methods for the detection of autoantibodies has been identified as a central challenge. Here, immune fluorescence techniques and the evaluation of RibP are used as first parameters that could improve SLE diagnostics. Furthermore, guidelines and proposals from scientific medical organizations, and in particular from other national EASI groups will be adapted to the German health system. A cornerstone of implementation is the identification and logistic preparation of existing serum banks, the definition of gaps that should be bridged, and, particularly, the definition and collection of adequate control groups. Through these measures, the German EASI group will provide a standardized diagnostic model of autoimmune disorders throughout Europe starting in the field of rheumatology. Diagnostics may become more rational, efficient, faster, and cost-efficient. Patients with autoimmune rheumatic disorders will profit from receiving an earlier and more accurate diagnosis, which again will allow earlier therapeutic intervention and lead to a better long-term clinical outcome.


Assuntos
Doenças Autoimunes/diagnóstico , Técnicas e Procedimentos Diagnósticos/normas , Sociedades Médicas , Proteínas Adaptadoras de Transdução de Sinal/imunologia , Algoritmos , Anticorpos/imunologia , Doenças Autoimunes/epidemiologia , Bancos de Sangue , Doenças do Tecido Conjuntivo/diagnóstico , Doenças do Tecido Conjuntivo/imunologia , Alemanha/epidemiologia , Humanos
14.
BMC Biotechnol ; 7: 35, 2007 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-17594499

RESUMO

BACKGROUND: Interleukin-12 (IL-12) is well characterized to induce cellular antitumoral immunity by activation of NK-cells and T-lymphocytes. However, systemic administration of recombinant human IL-12 resulted in severe toxicity without perceptible therapeutic benefit. Even though intratumoral expression of IL-12 leads to tumor regression and long-term survival in a variety of animal models, clinical trials have not yet shown a significant therapeutic benefit. One major obstacle in the treatment with IL-12 is to overcome the relatively low expression of the therapeutic gene without compromising the safety of such an approach. Our objective was to generate an adenoviral vector system enabling the regulated expression of very high levels of bioactive, human IL-12. RESULTS: High gene expression was obtained utilizing the VP16 herpes simplex transactivator. Strong regulation of gene expression was realized by fusion of the VP16 to a tetracycline repressor with binding of the fusion protein to a flanking tetracycline operator and further enhanced by auto-regulated expression of its fusion gene within a bicistronic promoter construct. Infection of human colon cancer cells (HT29) at a multiplicity of infection (m.o.i.) of 10 resulted in the production of up to 8000 ng/106 cells in 48 h, thus exceeding any published vector system so far. Doxycycline concentrations as low as 30 ng/ml resulted in up to 5000-fold suppression, enabling significant reduction of gene expression in a possible clinical setting. Bioactivity of the human single-chain IL-12 was similar to purified human heterodimeric IL-12. Frozen sections of human colon cancer showed high expression of the coxsackie adenovirus receptor with significant production of human single chain IL-12 in colon cancer biopsies after infection with 3*107 p.f.u. Ad.3r-scIL12. Doxycycline mediated suppression of gene expression was up to 9000-fold in the infected colon cancer tissue. CONCLUSION: VP16 transactivator-mediated and doxycycline-regulated expression of the human interleukin-12 gene enables highly efficient and tightly controlled cytokine expression in human cancer. These data illustrate the potential of the described adenoviral vector system for the safe and superior expression of therapeutic genes in the treatment of colorectal cancer and other malignancies.


Assuntos
Adenoviridae/genética , Clonagem Molecular/métodos , Doxiciclina/administração & dosagem , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Interleucina-12/genética , Interleucina-12/metabolismo , Transdução Genética/métodos , Relação Dose-Resposta a Droga , Células HT29 , Humanos
16.
J Heart Lung Transplant ; 26(1): 70-7, 2007 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-17234520

RESUMO

BACKGROUND: This study examines the efficacy of FK778 regimens for prevention of different phases of xenograft rejection. METHODS: Antibody and complement tissue depositions were measured by immunofluorescence in a discordant ex vivo rat-to-human heart perfusion model of hyperacute rejection with immunosuppressant-enriched human blood. The concordant hamster-to-rat aortic xenotransplantation model was used to assess host cellular (lymphocyte activation, mixed lymphocyte reaction [MLR]) and humoral responsiveness (xenoantibody production) as well as histologic xenograft rejection. Recipients were treated for 14 days with FK778, tacrolimus, sirolimus or combination regimens at varying doses. RESULTS: Antibody binding during hyperacute rejection was unaffected by the immunosuppressive treatment, but complement deposition was reduced in the following order: tacrolimus > FK778 approximately sirolimus. FK778 most effectively reduced complement factor 5 in vitro. In untreated rats with hamster aortic xenografts, a large infiltrative response was observed within the grafts with extensive myocyte necrosis. Tacrolimus > FK778 approximately sirolimus dose-dependently diminished xenograft infiltration and in the same order reduced vessel-wall myocyte necrosis. Tacrolimus approximately FK778 > sirolimus reduced in vivo lymphocyte CD25 expression and tacrolimus > FK778 approximately sirolimus diminished MLR. Xenoreactive IgM and IgG antibody production levels were vigorously upregulated a few days after transplantation, but were significantly reduced by tacrolimus > FK778 approximately sirolimus. Combination regimens revealed no significant benefit when compared with the corresponding monotherapy groups. CONCLUSIONS: FK778 mildly interfered with hyperacute rejection and markedly suppressed acute humoral and cellular aortic xenograft rejection. However, T-cell-dependent host responses were most potently suppressed by tacrolimus, and the overall efficacy of FK778 was similar to that of sirolimus.


Assuntos
Alquinos/uso terapêutico , Aorta Abdominal/transplante , Rejeição de Enxerto/prevenção & controle , Isoxazóis/uso terapêutico , Nitrilos/uso terapêutico , Transplante Heterólogo/métodos , Alquinos/farmacocinética , Animais , Anticorpos Anti-Idiotípicos/imunologia , Cricetinae , Modelos Animais de Doenças , Endotélio Vascular/patologia , Citometria de Fluxo , Rejeição de Enxerto/imunologia , Rejeição de Enxerto/patologia , Humanos , Imunoglobulina G/imunologia , Imunoglobulina M/imunologia , Subunidade alfa de Receptor de Interleucina-2/imunologia , Isoxazóis/farmacocinética , Masculino , Nitrilos/farmacocinética , Ratos , Ratos Endogâmicos BN , Ratos Endogâmicos Lew , Linfócitos T/imunologia , Resultado do Tratamento
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