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1.
BMC Nephrol ; 15: 203, 2014 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-25522739

RESUMO

BACKGROUND: AKI frequently develops in sepsis patients, significantly decreasing the overall prognosis. There are currently no diagnostic markers available which reliably predict the prognosis of sepsis-associated AKI. Recently, ATP content of CD4+ T cells (ATP_CD4) has been shown to correlate with survival in sepsis. The aim of the study was to determine ATP_CD4 in sepsis-associated AKI. METHODS: Thirty-three patients with sepsis were prospectively analyzed for ATP_CD4 at three different time points. Results were related to survival, renal recovery, and further clinical/laboratory findings. RESULTS: ATP_CD4 tended to lower in concentration at 48 h after onset of sepsis in those patients with complete renal recovery. There were no differences between patients with no AKI and those with AKI of different severity (AKIN 1-3). Urinary NGAL did not correlate with renal prognosis. CONCLUSION: ATP_CD4 may serve as risk predictor in sepsis-associated AKI. Lower concentrations may indicate a higher chance of complete renal recovery in sepsis.


Assuntos
Injúria Renal Aguda/diagnóstico , Trifosfato de Adenosina/análise , Linfócitos T CD4-Positivos/química , Sepse/complicações , Injúria Renal Aguda/complicações , Proteínas de Fase Aguda/urina , Idoso , Idoso de 80 Anos ou mais , Biomarcadores/análise , Progressão da Doença , Feminino , Humanos , Lipocalina-2 , Lipocalinas/urina , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos Prospectivos , Proteínas Proto-Oncogênicas/urina , Sepse/diagnóstico
2.
PLoS One ; 9(8): e105703, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25144233

RESUMO

BACKGROUND: T-cell exhaustion seems to play a critical role in CD8+ T-cell dysfunction during chronic viral infections. However, up to now little is known about the mechanisms underlying CD4+ T-cell dysfunction during chronic hepatitis B virus (CHB) infection and the role of inhibitory molecules such as programmed death 1 (PD-1) for CD4+ T-cell failure. METHODS: The expression of multiple inhibitory molecules such as PD-1, CTLA-4, TIM-3, CD244, KLRG1 and markers defining the grade of T-cell differentiation as CCR7, CD45RA, CD57 and CD127 were analyzed on virus-specific CD4+ T-cells from peripheral blood using a newly established DRB1*01-restricted MHC class II Tetramer. Effects of in vitro PD-L1/2 blockade were defined by investigating changes in CD4+ T-cell proliferation and cytokine production. RESULTS: CD4+ T-cell responses during chronic HBV infection was characterized by reduced Tetramer+CD4+ T-cell frequencies, effector memory phenotype, sustained PD-1 but low levels of CTLA-4, TIM-3, KLRG1 and CD244 expression. PD-1 blockade revealed individualized patterns of in vitro responsiveness with partly increased IFN-γ, IL-2 and TNF-α secretion as well as enhanced CD4+ T-cell expansion almost in treated patients with viral control. CONCLUSION: HBV-specific CD4+ T-cells are reliably detectable during different courses of HBV infection by MHC class II Tetramer technology. CD4+ T-cell dysfunction during chronic HBV is basically linked to strong PD-1 upregulation but absent coregulation of multiple inhibitory receptors. PD-L1/2 neutralization partly leads to enhanced CD4+ T-cell functionality with heterogeneous patterns of CD4+ T-cell rejunivation.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Receptores Coestimuladores e Inibidores de Linfócitos T/imunologia , Regulação da Expressão Gênica/imunologia , Hepatite B Crônica/imunologia , Receptor de Morte Celular Programada 1/imunologia , Adolescente , Adulto , Idoso , Linfócitos T CD4-Positivos/patologia , Feminino , Hepatite B Crônica/patologia , Humanos , Masculino , Pessoa de Meia-Idade
3.
BMC Anesthesiol ; 13(1): 36, 2013 Oct 29.
Artigo em Inglês | MEDLINE | ID: mdl-24168374

RESUMO

BACKGROUND: AKI significantly worsens prognosis of hospitalized patients. This is particularly the case in patients with sepsis. The risk for aquiring sepsis is significantly increased in malignant diseases. Aim of the present retrospective study was to analyze outcomes of tumor patients with sepsis and AKI. METHODS: One-thousand and seventeen patients, treated at the ICU of the Department of Nephrology and Rheumatology of the University Hospital Göttingen from 2009 to 2011 were retrospectively analyzed for mortality, sepsis, AKI, need for renal replacement therapy (dialysis) and malignancies. RESULTS: AKI occurred significantly more frequent in septic than in non-septic patients and in tumor as oposed to non-tumor patients. Mortaliy rates were higher in the respective latter groups. Mortality increased even further if patients suffered from a malignant disease with sepsis and AKI. Mortality rates peaked if dialysis treatment became mandatory. In non-solid tumors 100% of the patients died if they suffered drom sepsis and AKI. This was not the case in solid malignancies (mortality rate 56%). CONCLUSIONS: We conclude that prognosis of tumor patients with AKI and sepsis is very poor. Mortality increases to almost 70% if diaylsis therapy is initiated. Non-solid tumors are associated with a 100% mortality if sepsis and AKI conincide.

4.
J Med Case Rep ; 5: 598, 2011 Dec 29.
Artigo em Inglês | MEDLINE | ID: mdl-22206706

RESUMO

INTRODUCTION: Thrombotic thrombocytopenic purpura-hemolytic uremic syndrome is a life-threatening condition with various etiopathogeneses. Without therapy approximately 90% of all patients die from the disease. CASE PRESENTATION: We report the case of a 17-year-old Caucasian woman with widespread hematomas and headache. Due to hemolytic anemia, thrombocytopenia, and schistocytosis, thrombotic thrombocytopenic purpura-hemolytic uremic syndrome was suspected and plasma exchange therapy was initiated immediately. Since her thrombocyte level did not increase during the first week of therapy, plasma treatment had to be intensified to a twice-daily schedule. Further diagnostics showed markedly reduced activities of both ADAMTS-13 (a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13 - also known as von Willebrand factor-cleaving protease) and factor H. Test results for antibodies against both proteins were positive. While plasma exchange therapy was continued, rituximab was given once weekly for four consecutive weeks. After the last dose, thrombocytes and activities of ADAMTS-13 and factor H increased into the normal range. Our patient improved and was discharged from the hospital. CONCLUSIONS: Since no clinical symptoms/laboratory findings indicated a malignant or specific autoimmune-mediated disorder, the diagnosis made was thrombotic thrombocytopenic purpura-hemolytic uremic syndrome due to idiopathic combined, autoantibody-mediated ADAMTS-13/factor H deficiency.

5.
Gastroenterology ; 141(4): 1422-31, 1431.e1-6, 2011 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-21763239

RESUMO

BACKGROUND & AIMS: Inhibitory receptors such as programmed death 1 (PD-1) and cytotoxic T lymphocyte-associated antigen (CTLA)-4 mediate CD8+ T-cell exhaustion during chronic viral infection, but little is known about roles in dysfunction of CD4+ T cells. METHODS: We investigated the functions of inhibitory molecules on hepatitis C virus (HCV)-, influenza-, and Epstein-Barr virus (EBV)-specific CD4+ T cells in patients with chronic infections compared with patients with resolved HCV infection and healthy donors. Expression of PD-1, CTLA-4, CD305, and CD200R were analyzed on HCV-specific CD4+ T cells, isolated from peripheral blood using major histocompatibility complex class II tetramers. We investigated the effects of in vitro inhibition of various inhibitory pathways on proliferation and cytokine production by CD4+ T cells, and we compared these effects with those from inhibition of interleukin (IL)-10 and transforming growth factor (TGF)-ß1. RESULTS: PD-1 and CTLA-4 were up-regulated on virus-specific CD4+ T cells from patients with chronic HCV infections. PD-1 expression was lower on influenza- than on HCV-specific CD4+ T cells from subjects with chronic HCV infection, whereas CTLA-4 was expressed at similar levels, independent of their specificity. CD305 and CD200R were up-regulated in HCV resolvers. Blockade of PD-L1/2, IL-10, and TGF-ß1 increased expansion of CD4+ T cells in patients with chronic HCV, whereas inhibition of IL-10 and TGF-ß1 was most effective in restoring HCV-specific production of interferon gamma, IL-2, and tumor necrosis factor α. CONCLUSIONS: We characterized expression of inhibitory molecules on HCV-, influenza-, and EBV-specific CD4+ T cells and the effects of in vitro blockade on CD4+ T-cell expansion and cytokine production. Inhibition of PD-1, IL-10, and TGF-ß1 is most efficient in restoration of HCV-specific CD4+ T cells.


Assuntos
Antígenos CD/metabolismo , Linfócitos T CD4-Positivos/imunologia , Proliferação de Células , Hepacivirus/imunologia , Hepatite C Crônica/imunologia , Ativação Linfocitária , Anticorpos Neutralizantes , Antígenos CD/imunologia , Antígenos de Superfície/metabolismo , Linfócitos T CD4-Positivos/virologia , Antígeno CTLA-4/metabolismo , Estudos de Casos e Controles , Células Cultivadas , Feminino , Alemanha , Hepacivirus/genética , Hepatite C Crônica/diagnóstico , Herpesvirus Humano 4/imunologia , Humanos , Interferon gama/metabolismo , Interleucina-10/imunologia , Interleucina-10/metabolismo , Interleucina-2/metabolismo , Masculino , Pessoa de Meia-Idade , Receptores de Orexina , Orthomyxoviridae/imunologia , Receptor de Morte Celular Programada 1/metabolismo , RNA Viral/sangue , Receptores de Superfície Celular/metabolismo , Fator de Crescimento Transformador beta1/imunologia , Fator de Crescimento Transformador beta1/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Carga Viral
6.
PLoS One ; 6(2): e16797, 2011 Feb 08.
Artigo em Inglês | MEDLINE | ID: mdl-21347433

RESUMO

Cellular immune responses during acute Hepatitis C virus (HCV) and HIV infection are a known correlate of infection outcome. Viral adaptation to these responses via mutation(s) within CD8+ T-cell epitopes allows these viruses to subvert host immune control. This study examined HCV evolution in 21 HCV genotype 1-infected subjects to characterise the level of viral adaptation during acute and early HCV infection. Of the total mutations observed 25% were within described CD8+ T-cell epitopes or at viral adaptation sites. Most mutations were maintained into the chronic phase of HCV infection (75%). The lack of reversion of adaptations and high proportion of silent substitutions suggests that HCV has structural and functional limitations that constrain evolution. These results were compared to the pattern of viral evolution observed in 98 subjects during a similar phase in HIV infection from a previous study. In contrast to HCV, evolution during acute HIV infection is marked by high levels of amino acid change relative to silent substitutions, including a higher proportion of adaptations, likely reflecting strong and continued CD8+ T-cell pressure combined with greater plasticity of the virus. Understanding viral escape dynamics for these two viruses is important for effective T cell vaccine design.


Assuntos
Adaptação Fisiológica/genética , Evolução Molecular , Hepacivirus/genética , Hepacivirus/fisiologia , Hepatite C/virologia , Doença Aguda , Alelos , Epitopos/imunologia , Genótipo , Antígenos HLA/imunologia , Hepacivirus/patogenicidade , Hepatite C/diagnóstico , Hepatite C/imunologia , Hepatite C Crônica/diagnóstico , Hepatite C Crônica/imunologia , Hepatite C Crônica/virologia , Humanos , Mutação , Prognóstico , Linfócitos T/imunologia , Linfócitos T/virologia
7.
Gastroenterology ; 137(4): 1280-8.e1-6, 2009 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-19596013

RESUMO

BACKGROUND & AIMS: Down-regulation of hepatitis C virus (HCV)-specific CD4(+) T-cell responses is a hallmark of chronic viral persistence in acute hepatitis C. FOXP3(+)CD25(+)CD4(+) regulatory T cells can modulate HCV-specific immune responses in vitro, but the role of virus-specific regulatory T cells in the pathogenesis of chronic viral persistence is unknown. METHODS: Two novel HLA-DR15 tetramers were synthesized to study the kinetics and phenotype of FOXP3(+)-expressing HCV-specific CD4(+) T cells from 10 patients with acute hepatitis C and 15 patients with chronic hepatitis C. RESULTS: In acute hepatitis C, generally only a low percentage of HCV-specific CD4(+) T cells expressed FOXP3(+) (mean of 2.5% in patients with self-limited acute hepatitis C vs 2.4% in patients with evolving chronic hepatitis C). Although distinct but short-lived increases in virus-specific FOXP3(+)CD4(+) T cells occurred in 3 patients (30%, 26%, and 7% of tet(+) CD4(+) T cells, respectively), these did not correlate with the evolution of chronic hepatitis C. HCV-specific FOXP3(+)CD4(+) T cells displayed a distinct phenotype, with only 10% expressing CD25 and 40% being CD127low. Interestingly, this phenotype of FOXP3(+)CD4(+) T cells was already expanded in bulk CD4(+) T cells in patients with chronic hepatitis C. CONCLUSIONS: Although short-lived increases in HCV-specific FOXP3(+)CD4(+) T cells occur during the course of acute hepatitis C, we could not demonstrate an association of HCV-specific regulatory T cells and persistent viremia.


Assuntos
Fatores de Transcrição Forkhead/metabolismo , Hepacivirus/imunologia , Hepatite C Crônica/imunologia , Hepatite C/imunologia , Linfócitos T Reguladores/imunologia , Doença Aguda , Adulto , Idoso , Proliferação de Células , Células Cultivadas , Progressão da Doença , Feminino , Antígenos HLA-DR/imunologia , Cadeias HLA-DRB1 , Hepatite C/diagnóstico , Anticorpos Anti-Hepatite C/sangue , Hepatite C Crônica/diagnóstico , Humanos , Imunofenotipagem , Subunidade alfa de Receptor de Interleucina-2/metabolismo , Subunidade alfa de Receptor de Interleucina-7/metabolismo , Cinética , Masculino , Pessoa de Meia-Idade , Linfócitos T Reguladores/virologia , Viremia/imunologia
9.
Eur J Gastroenterol Hepatol ; 20(8): 778-83, 2008 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-18617783

RESUMO

OBJECTIVES: Recurrent hepatitis C virus (HCV) infection after liver transplantation (LT) is a major cause of transplant failure in HCV-positive patients. We retrospectively assessed the efficacy and safety of antiviral therapy and determined the factors influencing sustained virologic response (SVR) in LT recipients. METHODS: Between 1998 and 2007, we treated 36 LT recipients for hepatitis C cirrhosis and subsequent HCV recurrence (27 genotype 1 and 9 genotypes 2/3) with pegylated interferon alpha-2a (180 microg/week), pegylated interferon alpha-2b (1.5 microg/kg per week), or standard interferon alpha-2b (3 MIU 3X/week) plus ribavirin (600-1200 mg/day) for 48 weeks. RESULTS: SVR was achieved in seven of 27 (26%) of genotype 1 patients versus nine of nine (100%) genotype 2/3 patients (P=0.0001). Early virologic response at week 12 was associated with permanent viral clearance. Side effects included cytopenia and acute hearing loss, but rate of therapy withdrawal and dose reduction was low. CONCLUSION: Combination therapy in patients with HCV reinfection after LT yields an excellent SVR rate in genotype 2/3 patients, but remains unsatisfactory in genotype 1 patients. Virologic response at week 12 (early virologic response) can determine whether therapy should be continued or not.


Assuntos
Antivirais/uso terapêutico , Hepacivirus/genética , Hepatite C/tratamento farmacológico , Transplante de Fígado , Adulto , Antivirais/efeitos adversos , Esquema de Medicação , Quimioterapia Combinada , Feminino , Genótipo , Sobrevivência de Enxerto , Hepacivirus/efeitos dos fármacos , Hepatite C/cirurgia , Humanos , Imunossupressores/uso terapêutico , Interferon-alfa/efeitos adversos , Interferon-alfa/uso terapêutico , Masculino , Pessoa de Meia-Idade , Recidiva , Estudos Retrospectivos , Ribavirina/efeitos adversos , Ribavirina/uso terapêutico , Análise de Sobrevida , Resultado do Tratamento
10.
Cytokine ; 41(3): 209-16, 2008 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-18191408

RESUMO

In this study, we analyzed if IL-22 displays, similar to other IL-10 like cytokines such as IL-28A, antiviral properties in hepatic cells. Using RT-PCR and immunoblotting, we demonstrated that hepatic cell lines and primary hepatocytes express the functional IL-22 receptor complex consisting of IL-22R1 and IL-10R2. Hepatic IL-22 mRNA expression as measured by quantitative PCR was up-regulated in autoimmune and viral hepatitis compared to cholestatic liver diseases, while IL-22 serum levels did not differ significantly between patients with viral hepatitis and normal controls. IL-22 did not significantly change the expression levels of IFN-alpha/-beta and of the antiviral proteins MxA and 2',5'-OAS. Consequently, it had in comparison to IFN-alpha no relevant antiviral activity in in vitro models of HCV replication and infection. Taken together, hepatic IL-22 expression is up-regulated in viral hepatitis but IL-22 does not directly regulate antiviral proteins and has, in contrast to IFN-alpha, no effect on HCV replication.


Assuntos
Doenças Autoimunes/imunologia , Hepatite C/imunologia , Hepatócitos/imunologia , Interleucinas/metabolismo , Linhagem Celular , Humanos , Immunoblotting , Subunidade beta de Receptor de Interleucina-10/genética , Subunidade beta de Receptor de Interleucina-10/metabolismo , Interleucinas/sangue , Interleucinas/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Interleucina/genética , Receptores de Interleucina/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Regulação para Cima , Interleucina 22
11.
PLoS One ; 2(7): e649, 2007 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-17653276

RESUMO

BACKGROUND: CD4+ T cell help is critical in maintaining antiviral immune responses and such help has been shown to be sustained in acute resolving hepatitis C. In contrast, in evolving chronic hepatitis C CD4+ T cell helper responses appear to be absent or short-lived, using functional assays. METHODOLOGY/PRINCIPAL FINDINGS: Here we used a novel HLA-DR1 tetramer containing a highly targeted CD4+ T cell epitope from the hepatitis C virus non-structural protein 4 to track number and phenotype of hepatitis C virus specific CD4+ T cells in a cohort of seven HLA-DR1 positive patients with acute hepatitis C in comparison to patients with chronic or resolved hepatitis C. We observed peptide-specific T cells in all seven patients with acute hepatitis C regardless of outcome at frequencies up to 0.65% of CD4+ T cells. Among patients who transiently controlled virus replication we observed loss of function, and/or physical deletion of tetramer+ CD4+ T cells before viral recrudescence. In some patients with chronic hepatitis C very low numbers of tetramer+ cells were detectable in peripheral blood, compared to robust responses detected in spontaneous resolvers. Importantly we did not observe escape mutations in this key CD4+ T cell epitope in patients with evolving chronic hepatitis C. CONCLUSIONS/SIGNIFICANCE: During acute hepatitis C a CD4+ T cell response against this epitope is readily induced in most, if not all, HLA-DR1+ patients. This antiviral T cell population becomes functionally impaired or is deleted early in the course of disease in those where viremia persists.


Assuntos
Linfócitos T CD4-Positivos/virologia , Hepacivirus/isolamento & purificação , Hepatite C/imunologia , Linfócitos T Auxiliares-Indutores/virologia , Doença Aguda , Sequência de Aminoácidos , Sequência de Bases , Primers do DNA , Epitopos de Linfócito T/imunologia , Feminino , Genótipo , Antígeno HLA-DR1/química , Antígeno HLA-DR1/imunologia , Hepacivirus/genética , Humanos , Imunidade Celular , Fígado/imunologia , Fígado/virologia , Masculino , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/imunologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteínas Virais/análise , Proteínas Virais/química , Proteínas Virais/imunologia
12.
Vaccine ; 25(26): 4960-6, 2007 Jun 21.
Artigo em Inglês | MEDLINE | ID: mdl-17466417

RESUMO

BACKGROUND/AIMS: In hepatitis B virus infection, viral elimination is dependent on an efficient antiviral T cell response which is not detectable in chronic hepatitis B. Therefore, new therapeutic concepts focus on T cell activation, such as epitope-based T cell-targeted vaccines. However, with the development of peptide-based vaccines in mind, viral mutations frequently described in hepatitis B within known immunodominant helper epitopes may have an influence on peptide selection. METHODS: Mutant peptides within immunodominant epitopes (aa 1-20, aa 91-105, and aa 143-157) at position 12, 14, 93, 97, 147, 151, 153, and 155 were tested with peripheral blood mononuclear and specific clone cells for their ability to induce proliferation, produce cytokines, induce T cell receptor down-regulation or antagonize wild-type activity of the hepatitis B core antigen-specific CD4+ T cell clones. RESULTS: Five variants could not induce T cell proliferation or cytokine production when the variants were presented alone. Coincubation with wild-type epitopes leads to T cell activation showing that the variants do not act as T cell receptor antagonists for hepatitis B virus-specific CD4+ T cells. In contrast, five other variants and wild-type peptides stimulated CD4+ T cell proliferation and production of Th1 cytokines. CONCLUSIONS: Our data demonstrate that frequently occurring mutations within immunodominant epitopes have rather a nonstimulatory than a strengthening effect and thus should not included in a vaccine.


Assuntos
Epitopos/imunologia , Vacinas contra Hepatite B/genética , Vacinas contra Hepatite B/imunologia , Proteínas do Nucleocapsídeo/genética , Proteínas do Nucleocapsídeo/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Proliferação de Células , Células Clonais , Citocinas/metabolismo , Regulação para Baixo , Citometria de Fluxo , Hepatite B/patologia , Antígenos de Superfície da Hepatite B/genética , Humanos , Monócitos/imunologia , Monócitos/metabolismo , Mutação/genética , Mutação/fisiologia , Proteínas do Nucleocapsídeo/farmacologia , Peptídeos/genética , Peptídeos/farmacologia , Receptores de Antígenos de Linfócitos T/biossíntese , Receptores de Antígenos de Linfócitos T/genética
13.
Am J Physiol Gastrointest Liver Physiol ; 292(4): G1019-28, 2007 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-17204547

RESUMO

The IL-10-like cytokine IL-22 is produced by activated T cells. In this study, we analyzed the role of this cytokine system in hepatic cells. Expression studies were performed by RT-PCR and quantitative PCR. Signal transduction was analyzed by Western blot experiments and ELISA. Cell proliferation was measured by MTS and [(3)H]thymidine incorporation assays. Hepatocyte regeneration was studied in in vitro restitution assays. Binding of IL-22 to its receptor complex expressed on human hepatic cells and primary human hepatocytes resulted in the activation of MAPKs, Akt, and STAT proteins. IL-22 stimulated cell proliferation and migration, which were both significantly inhibited by the phosphatidylinositol 3-kinase inhibitor wortmannin. IL-22 increased the mRNA expression of suppressor of cytokine signaling (SOCS)-3 and the proinflammatory cytokines IL-6, IL-8, and TNF-alpha. SOCS-1/3 overexpression abrogated IL-22-induced STAT activation and decreased IL-22-mediated liver cell regeneration. Hepatic IL-22 mRNA expression was detectable in different forms of human hepatitis, and hepatic IL-22 mRNA levels were increased in murine T cell-mediated hepatitis in vivo following cytomegalovirus infection, whereas no significant differences were seen in an in vivo model of ischemia-reperfusion injury. In conclusion, IL-22 promotes liver cell regeneration by increasing hepatic cell proliferation and hepatocyte migration through the activation of Akt and STAT signaling, which is abrogated by SOCS-1/3 overexpression.


Assuntos
Hepatócitos/metabolismo , Interleucinas/metabolismo , Regeneração Hepática , Fígado/metabolismo , Transdução de Sinais , Proteínas Supressoras da Sinalização de Citocina/metabolismo , Animais , Técnicas de Cultura de Células , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Hepatectomia , Hepatite/metabolismo , Hepatócitos/efeitos dos fármacos , Humanos , Mediadores da Inflamação/metabolismo , Interleucinas/farmacologia , Fígado/citologia , Fígado/cirurgia , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Mensageiro/metabolismo , Receptores de Interleucina/genética , Receptores de Interleucina/metabolismo , Fatores de Transcrição STAT/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteína 1 Supressora da Sinalização de Citocina , Proteína 3 Supressora da Sinalização de Citocinas , Proteínas Supressoras da Sinalização de Citocina/genética , Linfócitos T/metabolismo , Fatores de Tempo , Transfecção , Regulação para Cima , Interleucina 22
14.
Kidney Int ; 68(1): 96-109, 2005 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-15954899

RESUMO

BACKGROUND: The peptide hormone relaxin has been demonstrated to exert antifibrotic effects in renal and extrarenal tissues. The aims of this study were to identify potential anti-fibrotic effects of relaxin on human renal fibroblasts in vitro and to analyze their mechanisms. METHODS: All experiments were performed in established renal fibroblast cell lines and in primary cortical fibroblasts. Effects of relaxin were analyzed on cell proliferation, apoptosis, activation of renal fibroblasts, synthesis and secretion of collagen type I and fibronectin, as well as on the secretion of matrix metalloproteinases (MMPs). Effects on transforming growth factor-beta1 (TGF-beta1) receptor binding were analyzed by flow cytometry and on TGF-beta1 signal transduction by immunoblot analyses for Smad4 and 7, translocation from cytosol to nucleus for Smad2 and 3 as well as for phosphorylated and unphosphorylated forms of p38, c-Jun NH2 terminal kinase (JNK) and extracellular-regulated protein kinase (ERK). Finally, specific siRNAs for Smad2 and 3 were applied to assess the signal transduction pathway. RESULTS: After stimulation with relaxin, tyrosine phosphorylation of a 220 kD protein was demonstrated, indicating interaction with the receptor. Relaxin had only modest inhibitory effects on cell proliferation, and no effects on apoptosis. Conversely, relaxin exerted robust effects on TGF-beta1-induced fibroblast to myofibroblast transformation as well as on matrix synthesis and secretion even at the smallest dose tested. The secretion of MMP-2 and MMP-9 was induced noticeably by all investigated relaxin concentrations. TGF-beta1 receptor binding was not influenced by relaxin; however, it prevented Smad2 phosphorylation, translocation to nucleus, and complex formation between Smad2 and 3 indicating a possible interaction with TGF-beta1 signaling. These findings were corroborated by studies using siRNAs to Smad2 and 3 where siRNA to Smad2 but not to Smad3 inhibited the TGF-beta1 induction of fibronectin synthesis. There was no influence of relaxin on intracellular Smad3, Smad4, and Smad7 translocation or phosphorylation of mitogen-activated protein (MAP) kinases. CONCLUSION: Relaxin is a potent inhibitor of TGF-beta1-induced extracellular matrix (ECM) synthesis and secretion as well as fibroblast activation. Furthermore, it induces ECM degradation by induction of MMP-2 and MMP-9. These effects are mediated, at least in part, by inhibition of TGF-beta1 signaling.


Assuntos
Proteínas de Ligação a DNA/antagonistas & inibidores , Fibroblastos/efeitos dos fármacos , Rim/patologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Relaxina/farmacologia , Transativadores/antagonistas & inibidores , Apoptose/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Linhagem Celular , Proteínas de Ligação a DNA/metabolismo , Matriz Extracelular/metabolismo , Matriz Extracelular/patologia , Fibroblastos/metabolismo , Fibroblastos/patologia , Fibrose , Humanos , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Necrose , Fosforilação/efeitos dos fármacos , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Proteína Smad2 , Proteína Smad3 , Transativadores/metabolismo , Fator de Crescimento Transformador beta/farmacologia , Fator de Crescimento Transformador beta1 , Tirosina/metabolismo
15.
Hepatology ; 41(3): 643-51, 2005 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-15726647

RESUMO

Chronic evolution of acute hepatitis C (aHC) occurs in more than 80% of patients but can frequently be prevented by early treatment with interferon (IFN)-alpha. Plasmacytoid dendritic cells (pDCs) are the major endogenous IFN-alpha producers, but their role in aHC is unknown. In this study, frequency, phenotype, and pDC function were analyzed in 13 patients with aHC and 32 patients with chronic hepatitis C (cHC) compared with 20 healthy controls, 33 sustained responders to antiviral treatment, 14 patients with acute hepatitis B (aHB), and 21 patients with nonviral inflammatory disease. In aHC, pDCs in the peripheral blood were significantly reduced compared with healthy controls (median, 0.1% vs. 0.36%, P < .0005) and were inversely correlated to alanine aminotransferase levels (r = -0.823; P < .005). Circulating pDCs in aHC were immature, as determined via reduced expression of HLA-DR and CCR7, and produced little amounts of IFN-alpha (median, 3.5 pg/50,000 peripheral blood mononuclear cells [PBMCs] vs. 498.4 pg/50,000 PBMCs in healthy controls; P < .0005). Less pronounced changes were present in cHC (median, 0.17%, 28.0 pg/50,000 PBMCs IFN-alpha, respectively). However, a significantly reduced frequency and IFN-alpha production was also found in self-limited aHB (median 0.1%, 8.6 pg/50,000 PBMCs) and in patients with nonviral inflammatory disease (median 0.19%, 7.5 pg/50,000 PBMCs). In conclusion, in aHC frequency and IFN-alpha-producing capacity of peripheral blood pDCs are dramatically reduced and inversely correlated with the degree of liver inflammation. In cHC there is incomplete recovery of pDC function, which, however, could be solely due to the chronic inflammatory state.


Assuntos
Células Dendríticas/fisiologia , Hepatite C Crônica/imunologia , Hepatite C/imunologia , Interferon-alfa/biossíntese , Doença Aguda , Adulto , Idoso , Feminino , Antígenos HLA-DR/análise , Humanos , Masculino , Pessoa de Meia-Idade , Receptores CCR7 , Receptores de Quimiocinas/análise
16.
Hepatology ; 41(3): 595-602, 2005 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-15726657

RESUMO

Induction of cytochrome P450 3A (CYP3A) has been suggested as a mechanism of action of ursodeoxycholic acid (UDCA) in cholestasis. CYP3A is of key importance in human drug metabolism, being involved in presystemic extraction of more than 50% of all drugs currently available and of various endogenous compounds. Therefore, we compared the induction potential of UDCA with that of the prototypical inducer rifampicin in a human model study with the CYP3A substrates budesonide and cortisol. Twelve patients with early-stage primary biliary cirrhosis and eight healthy volunteers were treated with UDCA (15 mg/kg daily) for 3 weeks and subsequently with rifampicin (600 mg/d) for 1 week. Extensive pharmacokinetic profiling of oral budesonide (3 mg) was performed by determination of budesonide and phase I metabolites (6beta-hydroxybudesonide, 16alpha-hydroxyprednisolone) in plasma and urine at baseline and at the end of each treatment. In parallel, urinary 6beta-hydroxycortisol, a validated marker of CYP3A induction, was determined. UDCA did not affect biotransformation of budesonide and urinary excretion of 6beta-hydroxycortisol either in patients or in healthy volunteers. Ratios of areas under plasma concentration-time curves (AUC(0-12 h) during UDCA/AUC(0-12 h) before UDCA) of both metabolites were not higher than those of budesonide itself. In contrast, administration of rifampicin markedly induced CYP3A metabolism, resulting in abolished budesonide plasma levels and high urinary excretion of 6beta-hydroxycortisol. Metabolite formation was enhanced by rifampicin, but not by UDCA (e.g., AUC(16alpha-hydroxyprednisolone)/AUC(budesonide) in patients: baseline, 8.6 +/- 3.9; UDCA, 10.7 +/- 7.1; rifampicin, 527.0 +/- 248.7). In conclusion, UDCA is not a relevant inducer of CYP3A enzymes in humans.


Assuntos
Hidrocarboneto de Aril Hidroxilases/fisiologia , Hidrocortisona/análogos & derivados , Cirrose Hepática Biliar/metabolismo , Oxirredutases N-Desmetilantes/fisiologia , Ácido Ursodesoxicólico/farmacologia , Adulto , Idoso , Área Sob a Curva , Budesonida/farmacocinética , Citocromo P-450 CYP3A , Feminino , Humanos , Hidrocortisona/urina , Masculino , Pessoa de Meia-Idade
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