RESUMO
Emerging approaches to treat immune disorders target positive regulatory kinases downstream of antigen receptors with small molecule inhibitors. Here we provide evidence for an alternative approach in which inhibition of the negative regulatory inositol kinase Itpkb in mature T lymphocytes results in enhanced intracellular calcium levels following antigen receptor activation leading to T cell death. Using Itpkb conditional knockout mice and LMW Itpkb inhibitors these studies reveal that Itpkb through its product IP4 inhibits the Orai1/Stim1 calcium channel on lymphocytes. Pharmacological inhibition or genetic deletion of Itpkb results in elevated intracellular Ca2+ and induction of FasL and Bim resulting in T cell apoptosis. Deletion of Itpkb or treatment with Itpkb inhibitors blocks T-cell dependent antibody responses in vivo and prevents T cell driven arthritis in rats. These data identify Itpkb as an essential mediator of T cell activation and suggest Itpkb inhibition as a novel approach to treat autoimmune disease.
Assuntos
Doenças Autoimunes/enzimologia , Doenças Autoimunes/terapia , Linfócitos T CD4-Positivos/metabolismo , Sinalização do Cálcio , Fosfotransferases (Aceptor do Grupo Álcool)/antagonistas & inibidores , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Doenças Autoimunes/patologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Canais de Cálcio/metabolismo , Sinalização do Cálcio/efeitos dos fármacos , Sinalização do Cálcio/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Células HEK293 , Humanos , Fosfatos de Inositol/metabolismo , Células Jurkat , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteína ORAI1 , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Ratos Endogâmicos LewRESUMO
We found the voltage-gated K+ channel Kv12.2 to be a potent regulator of excitability in hippocampal pyramidal neurons. Genetic deletion and pharmacologic block of Kv12.2 substantially reduced the firing threshold of these neurons. Kv12.2-/- (also known as Kcnh3-/-) mice showed signs of persistent neuronal hyperexcitability including frequent interictal spiking, spontaneous seizures and increased sensitivity to the chemoconvulsant pentylenetetrazol.
Assuntos
Epilepsia/fisiopatologia , Canais de Potássio Éter-A-Go-Go/metabolismo , Hipocampo/fisiopatologia , Neurônios/fisiologia , Potenciais de Ação/efeitos dos fármacos , Potenciais de Ação/fisiologia , Animais , Linhagem Celular , Células Cultivadas , Convulsivantes/toxicidade , Epilepsia/induzido quimicamente , Epilepsia/genética , Canais de Potássio Éter-A-Go-Go/antagonistas & inibidores , Canais de Potássio Éter-A-Go-Go/genética , Feminino , Hipocampo/efeitos dos fármacos , Humanos , Técnicas In Vitro , Masculino , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neurônios/efeitos dos fármacos , Pentilenotetrazol/toxicidade , Células Piramidais/efeitos dos fármacos , Células Piramidais/fisiopatologia , Convulsões/induzido quimicamente , Convulsões/genética , Convulsões/fisiopatologia , Gravação em Vídeo , XenopusRESUMO
The intermediate filament protein keratin 8 (K8) is critical for the development of most mouse embryos beyond midgestation. We find that 68% of K8-/- embryos, in a sensitive genetic background, are rescued from placental bleeding and subsequent death by cellular complementation with wild-type tetraploid extraembryonic cells. This indicates that the primary defect responsible for K8-/- lethality is trophoblast giant cell layer failure. Furthermore, the genetic absence of maternal but not paternal TNF doubles the number of viable K8-/- embryos. Finally, we show that K8-/- concepti are more sensitive to a TNF-dependent epithelial apoptosis induced by the administration of concanavalin A (ConA) to pregnant mothers. The ConA-induced failure of the trophoblast giant cell barrier results in hematoma formation between the trophoblast giant cell layer and the embryonic yolk sac in a phenocopy of dying K8-deficient concepti in a sensitive genetic background. We conclude the lethality of K8-/- embryos is due to a TNF-sensitive failure of trophoblast giant cell barrier function. The keratin-dependent protection of trophoblast giant cells from a maternal TNF-dependent apoptotic challenge may be a key function of simple epithelial keratins.
