RESUMEN
Thionamides, inhibitors of the thyroid peroxidase-mediated iodination, are clinically used in the treatment of hyperthyroidism. However, the use of antithyroid drugs is associated with immunomodulatory effects, and recent studies with thionamide-related heterocyclic thioderivates demonstrated direct anti-inflammatory and immunosuppressive properties. Using primary human T-lymphocytes, we show that the heterocyclic thionamides carbimazole and propylthiouracil inhibit synthesis of the proinflammatory cytokines tumor necrosis factor (TNF)alpha and interferon (IFN)gamma. In addition, DNA binding of nuclear factor (NF)-kappaB, a proinflammatory transcription factor that regulates both TNFalpha and IFNgamma synthesis, and NF-kappaB-dependent reporter gene expression were reduced. Abrogation of NF-kappaB activity was accompanied by reduced phosphorylation and proteolytic degradation of inhibitor of kappaB (IkappaB)alpha, the inhibitory subunit of the NF-kappaB complex. Carbimazole inhibited NF-kappaB via the small GTPase Rac-1, whereas propylthiouracil inhibited the phosphorylation of IkappaBalpha by its kinase inhibitor of kappaB kinase alpha. Methimazole had no effect on NF-kappaB induction, demonstrating that drug potency correlated with the chemical reactivity of the thionamide-associated sulfur group. Taken together, our data demonstrate that thioureylenes with a common, heterocyclic structure inhibit inflammation and immune function via the NF-kappaB pathway. Our results may explain the observed remission of proinflammatory diseases upon antithyroid therapy in hyperthyroid patients. The use of related thioureylenes may provide a new therapeutic basis for the development and application of anti-inflammatory compounds.
Asunto(s)
Antitiroideos/farmacología , Quinasa I-kappa B/antagonistas & inhibidores , FN-kappa B/antagonistas & inhibidores , Proteína de Unión al GTP rac1/antagonistas & inhibidores , Antitiroideos/química , Humanos , Quinasa I-kappa B/metabolismo , Células Jurkat , Leucocitos Mononucleares/efectos de los fármacos , Leucocitos Mononucleares/metabolismo , FN-kappa B/metabolismo , Compuestos de Sulfhidrilo/química , Compuestos de Sulfhidrilo/farmacología , Proteína de Unión al GTP rac1/metabolismoRESUMEN
Treatment of hyperthyroidism by thionamides is associated with immunomodulatory effects, but the mechanism of thionamide-induced immunosuppression is unclear. Here we show that thionamides directly inhibit interleukin-2 cytokine expression, proliferation, and the activation (CD69 expression) of primary human T lymphocytes. Inhibition of immune function was associated with a repression of DNA binding of the cooperatively acting immunoregulatory transcription factors activator protein 1 (AP-1) and nuclear factor of activated T-cells (NFAT). Likewise, thionamides block the GTPase p21Ras, the mitogen-activated protein kinases, and impair the calcineurin/calmodulin-dependent NFAT dephosphorylation and nuclear translocation. The potency of inhibition correlated with the chemical reactivity of the thionamide-associated sulfur group. Taken together, our data demonstrate that thio-derivates with a common heterocyclic thioureylene-structure mediate a direct suppression of immune functions in T-cells via inhibition of the AP-1/NFAT pathway. Our observations may also explain the clinical and pathological resolution of some secondary, calcineurin, and mitogen-activated protein kinase-associated diseases upon thionamide treatment in hyperthyroid patients. This offers a new therapeutic basis for the development and application of heterocyclic thio-derivates.
Asunto(s)
Inmunosupresores/farmacología , Factores de Transcripción NFATC/antagonistas & inhibidores , Factores de Transcripción NFATC/fisiología , Transducción de Señal/fisiología , Linfocitos T/fisiología , Tiobarbitúricos/farmacología , Factor de Transcripción AP-1/antagonistas & inhibidores , Factor de Transcripción AP-1/fisiología , Proliferación Celular/efectos de los fármacos , Humanos , Inmunosupresores/química , Células Jurkat , Transducción de Señal/efectos de los fármacos , Relación Estructura-Actividad , Linfocitos T/efectos de los fármacos , Tiobarbitúricos/químicaRESUMEN
The involvement of p38 in fundamental physiological processes and the fact that deregulation often leads to disease indicates the potential impact of p38 dependent mechanisms. Here we demonstrate a new pathway that includes the induction of the mitogen activated protein kinase p38 by protein kinase C and results in a specific phosphorylation of c-Jun in T-lymphocytes. P38 directly phosphorylates c-Jun within its transactivation domain at serine 63 and serine 73 and thus posttranscriptionally affects the presence of DNA-bound phosphorylated c-Jun, a prerequisite for activator protein 1 dependent gene transcription. Moreover, DNA-binding activity of c-Fos, FosB, and JunB were also dependent on the p38 protein kinase activity, whereas JunD, Fra-1 and Fra-2 were not affected. Although we show that stress induced mitogen activated protein kinases share c-Jun as a substrate for phosphorylation, p38 mediated effects could not be rescued by the c-Jun N-terminal kinases. This demonstrates that the protein kinase p38 plays a unique and non-redundant role in posttranslational c-Jun regulation. The induction of a p38 dependent c-Jun phosphorylation was comparable in both CD4(+) and CD8(+) T-cells, proposing a ubiquitous pathway that is not linked to T-cell subtype and effector function. In contrast, ATF-2 was predominantly phosphorylated in CD8(+) T-cells. Different cell lines show p38-dependent c-Jun phosphorylation upon phorbol ester induction but there is evidence that the simian virus 40 large T-antigen may interfere with this pathway.
Asunto(s)
Proteínas Proto-Oncogénicas c-jun/metabolismo , Factor de Transcripción AP-1/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Animales , Anticuerpos/farmacología , Antígenos CD28/inmunología , Complejo CD3/inmunología , Línea Celular , Línea Celular Tumoral , ADN/metabolismo , Dactinomicina/farmacología , Inhibidores Enzimáticos/farmacología , Humanos , Isoenzimas/antagonistas & inhibidores , Isoenzimas/metabolismo , Proteínas Quinasas JNK Activadas por Mitógenos/antagonistas & inhibidores , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Ratones , Células 3T3 NIH , Fosforilación/efectos de los fármacos , Unión Proteica/efectos de los fármacos , Proteína Quinasa C/metabolismo , Proteínas Proto-Oncogénicas c-fos/metabolismo , Serina/metabolismo , Subgrupos de Linfocitos T/metabolismo , Linfocitos T/efectos de los fármacos , Linfocitos T/metabolismo , Acetato de Tetradecanoilforbol/análogos & derivados , Acetato de Tetradecanoilforbol/farmacología , Proteínas Quinasas p38 Activadas por Mitógenos/antagonistas & inhibidoresRESUMEN
BACKGROUND: Thrombotic microangiopathies (TMA) in adults such as thrombotic thrombocytopenic purpura (TTP) and hemolytic uremic syndrome (HUS) are life-threatening disorders if untreated. Clinical presentation is highly variable and prognostic factors for clinical course and outcome are not well established. METHODS: We performed a retrospective observational study of 62 patients with TMA, 22 males and 40 females aged 16 to 76 years, treated with plasma exchange at one center to identify clinical risk factors for the development of renal insufficiency. RESULTS: On admission, 39 of 62 patients (63%) had acute renal failure (ARF) with 32 patients (52%) requiring dialysis treatment. High systolic arterial pressure (SAP, pâ=â0.009) or mean arterial pressure (MAP, pâ=â0.027) on admission was associated with acute renal failure. Patients with SAP>140 mmHg on admission had a sevenfold increased risk of severe kidney disease (OR 7.464, CI 2.097-26.565). MAP>100 mmHg indicated a fourfold increased risk for acute renal failure (OR 4.261, CI 1.400-12.972). High SAP, diastolic arterial pressure (DAP), and MAP on admission were also independent risk factors for persistent renal insufficiency with the strongest correlation for high MAP. Moreover, a high C-reactive protein (CRP) level on admission correlated with renal failure in the course of the disease (pâ=â0.003). At discharge, renal function in 11 of 39 patients (28%) had fully recovered, 14 patients (23%) remained on dialysis, and 14 patients (23%) had non-dialysis-dependent chronic kidney disease. Seven patients (11%) died. We identified an older age as risk factor for death. CONCLUSIONS: High blood pressure as well as high CRP serum levels on admission are associated with renal insufficiency in TMA. High blood pressure on admission is also a strong predictor of sustained renal insufficiency. Thus, adult TMA patients with high blood pressure may require special attention to prevent persistent renal failure.
Asunto(s)
Lesión Renal Aguda/diagnóstico , Lesión Renal Aguda/etiología , Indicadores de Salud , Microangiopatías Trombóticas/diagnóstico , Lesión Renal Aguda/mortalidad , Adolescente , Adulto , Anciano , Biomarcadores/análisis , Femenino , Humanos , Masculino , Persona de Mediana Edad , Admisión del Paciente/estadística & datos numéricos , Pronóstico , Estudios Retrospectivos , Factores de Riesgo , Microangiopatías Trombóticas/complicaciones , Microangiopatías Trombóticas/mortalidad , Adulto JovenRESUMEN
The conserved Hippo signaling pathway regulates organ size in Drosophila melanogaster and mammals and has an essential role in tumor suppression and the control of cell proliferation. Recent studies identified activators of Hippo signaling, but antagonists of the pathway have remained largely elusive. In this paper, we show that NPHP4, a known cilia-associated protein that is mutated in the severe degenerative renal disease nephronophthisis, acts as a potent negative regulator of mammalian Hippo signaling. NPHP4 directly interacted with the kinase Lats1 and inhibited Lats1-mediated phosphorylation of the Yes-associated protein (YAP) and TAZ (transcriptional coactivator with PDZ-binding domain), leading to derepression of these protooncogenic transcriptional regulators. Moreover, NPHP4 induced release from 14-3-3 binding and nuclear translocation of YAP and TAZ, promoting TEA domain (TEAD)/TAZ/YAP-dependent transcriptional activity. Consistent with these data, knockdown of NPHP4 negatively affected cellular proliferation and TEAD/TAZ activity, essentially phenocopying loss of TAZ function. These data identify NPHP4 as a negative regulator of the Hippo pathway and suggest that NPHP4 regulates cell proliferation through its effects on Hippo signaling.
Asunto(s)
Neoplasias de la Mama/metabolismo , Proliferación Celular , Riñón/metabolismo , Proteínas/metabolismo , Transducción de Señal , Proteínas 14-3-3/metabolismo , Aciltransferasas , Animales , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Proteínas de Ciclo Celular , Línea Celular Tumoral , Femenino , Genes Reporteros , Células HEK293 , Humanos , Inmunoprecipitación , Riñón/patología , Proteínas Nucleares/metabolismo , Fosforilación , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas/genética , Interferencia de ARN , Proteínas Recombinantes de Fusión/metabolismo , Factores de Transcripción/metabolismo , TransfecciónRESUMEN
Barbiturates are known to suppress protective immunity, and their therapeutic use is associated with nosocomial infections. Although barbiturates inhibit T cell proliferation, differentiation, and cytokine synthesis, only thiobarbiturates markedly reduce the activation of immune regulatory transcription factors such as nuclear factor-kappaB and nuclear factor of activated T cells. In this study, we investigated barbiturate-mediated effects on the regulation of the transcription factor activator protein 1 (AP-1) in primary T lymphocytes. We show that both thiobarbiturates and their oxy-analogs inhibit AP-1-dependent gene expression and AP-1 complex formation at clinically relevant doses. Furthermore, mitogen-activated protein (MAP) kinase activity, which transcriptionally and posttranslationally regulates AP-1 complex formation, is suppressed by most barbiturates. CD3/CD28- or phorbol 12-myristate 13-acetate (PMA)/ionomycin-induced p38 and extracellular signal-regulated kinase 1/2 phosphorylation or c-jun NH2-terminal kinase (JNK) 1/2 kinase activity was significantly diminished by pentobarbital, thiamylal, secobarbital, or methohexital treatment. These barbiturates also inhibited the initiators of the MAP kinase cascade, the small G proteins ras and rac-1, and prevented binding to their partners raf-1 and PAK, respectively. Thiopental, unlike the other barbiturates, only reduced ras and JNK activity upon direct CD3/CD28 receptor engagement. Contrarily, upon PMA/ionomycin stimulation, thiopental blocked AP-1-dependent gene expression independently of the small G protein ras and MAP kinases, thus suggesting an additional, unknown mechanism of AP-1 regulation. In conclusion, our results contribute to the explanation of a clinically manifested immune suppression in barbiturate-treated patients and support the idea of a MAP kinase-independent regulation of AP-1 by PKC and calcium in human T cells.