RESUMO
Neuroinflammation is causally associated with Alzheimer's disease (AD) pathology. Reactive glia cells secrete various neurotoxic factors that impair neuronal homeostasis eventually leading to neuronal loss. Although the glial activation mechanism in AD has been relatively well studied, how it perturbs intraneuronal signaling, which ultimately leads to neuronal cell death, remains poorly understood. Here, we report that compound stimulation with the neurotoxic factors TNF and glutamate aberrantly activates neuronal TAK1 (also known as MAP3K7), which promotes the pathogenesis of AD in mouse models. Glutamate-induced Ca2+ influx shifts TNF signaling to hyper-activate TAK1 enzymatic activity through Ca2+/calmodulin-dependent protein kinase II, which leads to necroptotic cellular damage. Genetic ablation and pharmacological inhibition of TAK1 ameliorated AD-associated neuronal loss and cognitive impairment in the AD model mice. Our findings provide a molecular mechanism linking cytokines, Ca2+ signaling and neuronal necroptosis in AD.
Assuntos
Doença de Alzheimer , Animais , Camundongos , Doença de Alzheimer/genética , Cálcio , Citocinas/metabolismo , Doenças Neuroinflamatórias , Transdução de Sinais/fisiologiaRESUMO
Mitogen-activated protein kinase kinase kinase 7 (MAP3K7), known as TAK1, is an intracellular signaling intermediate of inflammatory responses. However, a series of mouse Tak1 gene deletion analyses have revealed that ablation of TAK1 does not prevent but rather elicits inflammation, which is accompanied by elevation of reactive oxygen species (ROS). This has been considered a consequence of impaired TAK1-dependent maintenance of tissue integrity. Contrary to this view, here we propose that TAK1 inhibition-induced ROS are an active cellular process that targets intracellular bacteria. Intracellular bacterial effector proteins such as Yersinia's outer membrane protein YopJ are known to inhibit TAK1 to circumvent the inflammatory host responses. We found that such TAK1 inhibition induces mitochondrial-derived ROS, which effectively destroys intracellular bacteria. Two cell death-signaling molecules, caspase 8 and RIPK3, cooperatively participate in TAK1 inhibition-induced ROS and blockade of intracellular bacterial growth. Our results reveal a previously unrecognized host defense mechanism, which is initiated by host recognition of pathogen-induced impairment in a host protein, TAK1, but not directly of pathogens.
Assuntos
Bactérias/crescimento & desenvolvimento , Espaço Intracelular/microbiologia , MAP Quinase Quinase Quinases/metabolismo , Mitocôndrias/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Animais , Caspase 3/metabolismo , Contagem de Colônia Microbiana , Sulfeto de Hidrogênio/farmacologia , MAP Quinase Quinase Quinases/antagonistas & inibidores , Camundongos , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Salmonella/efeitos dos fármacos , Salmonella/crescimento & desenvolvimento , Yersinia/efeitos dos fármacosRESUMO
Aberrant wound healing presents as inappropriate or insufficient tissue formation. Using a model of musculoskeletal injury, we demonstrate that loss of transforming growth factor-ß activated kinase 1 (TAK1) signaling reduces inappropriate tissue formation (heterotopic ossification) through reduced cellular differentiation. Upon identifying increased proliferation with loss of TAK1 signaling, we considered a regenerative approach to address insufficient tissue production through coordinated inactivation of TAK1 to promote cellular proliferation, followed by reactivation to elicit differentiation and extracellular matrix production. Although the current regenerative medicine paradigm is centered on the effects of drug treatment ("drug on"), the impact of drug withdrawal ("drug off") implicit in these regimens is unknown. Because current TAK1 inhibitors are unable to phenocopy genetic Tak1 loss, we introduce the dual-inducible COmbinational Sequential Inversion ENgineering (COSIEN) mouse model. The COSIEN mouse model, which allows us to study the response to targeted drug treatment ("drug on") and subsequent withdrawal ("drug off") through genetic modification, was used here to inactivate and reactivate Tak1 with the purpose of augmenting tissue regeneration in a calvarial defect model. Our study reveals the importance of both the "drug on" (Cre-mediated inactivation) and "drug off" (Flp-mediated reactivation) states during regenerative therapy using a mouse model with broad utility to study targeted therapies for disease. Stem Cells 2019;37:766-778.
Assuntos
Regeneração Óssea/efeitos dos fármacos , Fraturas Ósseas/genética , MAP Quinase Quinase Quinases/genética , Células-Tronco Mesenquimais/enzimologia , Osteoblastos/enzimologia , Cicatrização/genética , Animais , Regeneração Óssea/genética , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , DNA Nucleotidiltransferases/genética , DNA Nucleotidiltransferases/metabolismo , Feminino , Efeito Fundador , Fraturas Ósseas/tratamento farmacológico , Fraturas Ósseas/enzimologia , Fraturas Ósseas/patologia , Regulação da Expressão Gênica , Integrases/genética , Integrases/metabolismo , MAP Quinase Quinase Quinases/antagonistas & inibidores , MAP Quinase Quinase Quinases/deficiência , Masculino , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Osteoblastos/citologia , Osteoblastos/efeitos dos fármacos , Cultura Primária de Células , Inibidores de Proteínas Quinases/farmacologia , Transdução de Sinais , Crânio/efeitos dos fármacos , Crânio/lesões , Crânio/metabolismo , Cicatrização/efeitos dos fármacosRESUMO
BMP signaling plays a critical role in craniofacial development. Augmentation of BMPR1A signaling through neural crest-specific expression of constitutively active Bmpr1a (caBmpr1a) results in craniofacial deformities in mice. To investigate whether deletion of Tak1 may rescue the craniofacial deformities caused by enhanced Smad-dependent signaling through caBMPR1A, we generated embryos to activate transcription of caBmpr1a transgene and ablate Tak1 in neural crest derivatives at the same time. We found that deformities of the double mutant mice showed more severe than those with each single mutation, including median facial cleft and cleft palate. We found higher levels of cell death in the medial nasal and the lateral nasal processes at E10.5 in association with higher levels of p53 in the double mutant embryos. We also found higher levels of pSmad1/5/9 in the lateral nasal processes at E10.5 in the double mutant embryos. Western analyses revealed that double mutant embryos showed similar degrees of upregulation of pSmad1/5/9 with caBmpr1a or Tak1-cKO embryos while the double mutant embryos showed higher levels of phospho-p38 than caBmpr1a or Tak1-cKO embryos at E17.5, but not at E10.5. It suggested that deletion of Tak1 aggravates the craniofacial deformities of the caBmpr1a mutants by increasing p53 and phospho-p38 at different stage of embryogenesis.
Assuntos
Receptores de Proteínas Morfogenéticas Ósseas Tipo I/genética , Morte Celular/genética , Anormalidades Craniofaciais/genética , Estudos de Associação Genética , Genótipo , MAP Quinase Quinase Quinases/genética , Mutação , Animais , Apoptose/genética , Biomarcadores , Receptores de Proteínas Morfogenéticas Ósseas Tipo I/metabolismo , Anormalidades Craniofaciais/diagnóstico , Idade Gestacional , Imuno-Histoquímica , MAP Quinase Quinase Quinases/metabolismo , Camundongos , Camundongos Transgênicos , Fenótipo , Transdução de Sinais , Proteínas Smad/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
Sustained endoplasmic reticulum (ER) stress disrupts normal cellular homeostasis and leads to the development of many types of human diseases, including metabolic disorders. TAK1 (also known as MAP3K7) is a member of the mitogen-activated protein kinase kinase kinase (MAP3K) family and is activated by a diverse set of inflammatory stimuli. Here, we demonstrate that TAK1 regulates ER stress and metabolic signaling through modulation of lipid biogenesis. We found that deletion of Tak1 increased ER volume and facilitated ER-stress tolerance in cultured cells, which was mediated by upregulation of sterol-regulatory-element-binding protein (SREBP)-dependent lipogenesis. In the in vivo setting, central nervous system (CNS)-specific Tak1 deletion upregulated SREBP-target lipogenic genes and blocked ER stress in the hypothalamus. Furthermore, CNS-specific Tak1 deletion prevented ER-stress-induced hypothalamic leptin resistance and hyperphagic obesity under a high-fat diet (HFD). Thus, TAK1 is a crucial regulator of ER stress in vivo, which could be a target for alleviation of ER stress and its associated disease conditions.
Assuntos
Estresse do Retículo Endoplasmático , Hipotálamo/metabolismo , Leptina/metabolismo , MAP Quinase Quinase Quinases/metabolismo , Animais , Gorduras na Dieta/efeitos adversos , Gorduras na Dieta/farmacologia , Hiperfagia/induzido quimicamente , Hiperfagia/genética , Hiperfagia/metabolismo , Hiperfagia/patologia , Hipotálamo/patologia , Leptina/genética , MAP Quinase Quinase Quinases/genética , Camundongos , Camundongos Knockout , Obesidade/induzido quimicamente , Obesidade/genética , Obesidade/metabolismo , Obesidade/patologia , Proteínas de Ligação a Elemento Regulador de Esterol/genética , Proteínas de Ligação a Elemento Regulador de Esterol/metabolismoRESUMO
Transforming growth factor-beta3 (TGF-ß3) plays a critical role in palatal epithelial cells by inducing palatal epithelial fusion, failure of which results in cleft palate, one of the most common birth defects in humans. Recent studies have shown that Smad-dependent and Smad-independent pathways work redundantly to transduce TGF-ß3 signaling in palatal epithelial cells. However, detailed mechanisms by which this signaling is mediated still remain to be elucidated. Here we show that TGF-ß activated kinase-1 (Tak1) and Smad4 interact genetically in palatal epithelial fusion. While simultaneous abrogation of both Tak1 and Smad4 in palatal epithelial cells resulted in characteristic defects in the anterior and posterior secondary palate, these phenotypes were less severe than those seen in the corresponding Tgfb3 mutants. Moreover, our results demonstrate that Trim33, a novel chromatin reader and regulator of TGF-ß signaling, cooperates with Smad4 during palatogenesis. Unlike the epithelium-specific Smad4 mutants, epithelium-specific Tak1:Smad4- and Trim33:Smad4-double mutants display reduced expression of Mmp13 in palatal medial edge epithelial cells, suggesting that both of these redundant mechanisms are required for appropriate TGF-ß signal transduction. Moreover, we show that inactivation of Tak1 in Trim33:Smad4 double conditional knockouts leads to the palatal phenotypes which are identical to those seen in epithelium-specific Tgfb3 mutants. To conclude, our data reveal added complexity in TGF-ß signaling during palatogenesis and demonstrate that functionally redundant pathways involving Smad4, Tak1 and Trim33 regulate palatal epithelial fusion.
Assuntos
MAP Quinase Quinase Quinases/metabolismo , Palato/embriologia , Palato/metabolismo , Transdução de Sinais , Proteína Smad4/metabolismo , Fatores de Transcrição/metabolismo , Fator de Crescimento Transformador beta3/metabolismo , Animais , Apoptose/genética , Fusão Celular , Proliferação de Células , Cruzamentos Genéticos , Embrião de Mamíferos/metabolismo , Ativação Enzimática , Células Epiteliais/metabolismo , Epitélio/metabolismo , Feminino , Deleção de Genes , Regulação da Expressão Gênica no Desenvolvimento , Masculino , Metaloproteinase 13 da Matriz/metabolismo , Camundongos Knockout , Modelos Biológicos , Mutação/genética , Especificidade de Órgãos , Palato/anormalidades , Palato/enzimologiaRESUMO
Autophagic responses are coupled to the activation of the inhibitor of NF-κB kinase (IKK). Here, we report that the essential autophagy mediator Beclin 1 and TGFß-activated kinase 1 (TAK1)-binding proteins 2 and 3 (TAB2 and TAB3), two upstream activators of the TAK1-IKK signalling axis, constitutively interact with each other via their coiled-coil domains (CCDs). Upon autophagy induction, TAB2 and TAB3 dissociate from Beclin 1 and bind TAK1. Moreover, overexpression of TAB2 and TAB3 suppresses, while their depletion triggers, autophagy. The expression of the C-terminal domain of TAB2 or TAB3 or that of the CCD of Beclin 1 competitively disrupts the interaction between endogenous Beclin 1, TAB2 and TAB3, hence stimulating autophagy through a pathway that requires endogenous Beclin 1, TAK1 and IKK to be optimally efficient. These results point to the existence of an autophagy-stimulatory 'switch' whereby TAB2 and TAB3 abandon inhibitory interactions with Beclin 1 to engage in a stimulatory liaison with TAK1.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Reguladoras de Apoptose/metabolismo , Autofagia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas de Membrana/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Reguladoras de Apoptose/genética , Proteína Beclina-1 , Células HeLa , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas de Membrana/genética , Estrutura Terciária de Proteína , Técnicas do Sistema de Duplo-HíbridoRESUMO
Dysregulation in cellular redox systems results in accumulation of reactive oxygen species (ROS), which are causally associated with a number of disease conditions. Transforming growth factor ß-activated kinase 1 (TAK1) is a signaling intermediate of innate immune signaling pathways and is critically involved in the redox regulation in vivo. Ablation of TAK1 causes accumulation of ROS, resulting in epithelial cell death and inflammation. Here we determine the mechanism by which TAK1 kinase is activated in epithelial tissues. TAB1 and TAB2 are structurally unrelated TAK1 binding protein partners. TAB2 is known to mediate polyubiquitin chain-dependent TAK1 activation in innate immune signaling pathways, whereas the role of TAB1 is not defined. We found that epithelial-specific TAB1 and TAB2 double- but not TAB1 or TAB2 single-knockout mice phenocopied epithelial-specific TAK1 knockout mice. We demonstrate that phosphorylation-dependent basal activity of TAK1 is dependent on TAB1. Ablation of both TAB1 and TAB2 diminished the activity of TAK1 in vivo and causes accumulation of ROS in the epithelial tissues. These results demonstrate that epithelial TAK1 activity is regulated through two unique, TAB1-dependent basal and TAB2-mediated stimuli-dependent mechanisms.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Células Epiteliais/enzimologia , MAP Quinase Quinase Quinases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/deficiência , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Ativação Enzimática , Epiderme/enzimologia , Mucosa Intestinal/enzimologia , Queratinócitos/enzimologia , Camundongos , Camundongos Knockout , Oxirredução , Estresse Oxidativo , Fenótipo , Fosforilação , Processamento de Proteína Pós-Traducional , Transdução de SinaisRESUMO
BACKGROUND: The role of Smad-independent TGF-ß signaling in craniofacial development is poorly elucidated. RESULTS: In craniofacial mesenchymal cells, Tak1 regulates both R-Smad C-terminal and linker region phosphorylation in TGF-ß signaling. CONCLUSION: Tak1 plays an irreplaceable role in craniofacial ecto-mesenchyme during embryogenesis. SIGNIFICANCE: Understanding the mechanisms of TGF-ß signaling contributes to knowledge of pathogenetic mechanisms underlying common craniofacial birth defects. Although the importance of TGF-ß superfamily signaling in craniofacial growth and patterning is well established, the precise details of its signaling mechanisms are still poorly understood. This is in part because of the concentration of studies on the role of the Smad-dependent (so-called "canonical") signaling pathways relative to the Smad-independent ones in many biological processes. Here, we have addressed the role of TGF-ß-activated kinase 1 (Tak1, Map3k7), one of the key mediators of Smad-independent (noncanonical) TGF-ß superfamily signaling in craniofacial development, by deleting Tak1 specifically in the neural crest lineage. Tak1-deficient mutants display a round skull, hypoplastic maxilla and mandible, and cleft palate resulting from a failure of palatal shelves to appropriately elevate and fuse. Our studies show that in neural crest-derived craniofacial ecto-mesenchymal cells, Tak1 is not only required for TGF-ß- and bone morphogenetic protein-induced p38 Mapk activation but also plays a role in agonist-induced C-terminal and linker region phosphorylation of the receptor-mediated R-Smads. Specifically, we demonstrate that the agonist-induced linker region phosphorylation of Smad2 at Thr-220, which has been shown to be critical for full transcriptional activity of Smad2, is dependent on Tak1 activity and that in palatal mesenchymal cells TGFßRI and Tak1 kinases mediate both overlapping and distinct TGF-ß2-induced transcriptional responses. To summarize, our results suggest that in neural crest-derived ecto-mesenchymal cells, Tak1 provides a critical point of intersection in a complex dialogue between the canonical and noncanonical arms of TGF-ß superfamily signaling required for normal craniofacial development.
Assuntos
MAP Quinase Quinase Quinases/fisiologia , Crista Neural/citologia , Processamento de Proteína Pós-Traducional , Proteínas Smad/metabolismo , Motivos de Aminoácidos , Animais , Células Cultivadas , Fissura Palatina/enzimologia , Fissura Palatina/genética , Ectoderma/citologia , Feminino , Regulação da Expressão Gênica no Desenvolvimento , Cabeça/embriologia , MAP Quinase Quinase Quinases/deficiência , MAP Quinase Quinase Quinases/genética , Masculino , Mandíbula/anormalidades , Camundongos , Camundongos Transgênicos , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Fosforilação , Proteínas Serina-Treonina Quinases/metabolismo , Receptor do Fator de Crescimento Transformador beta Tipo I , Receptores de Fatores de Crescimento Transformadores beta/metabolismo , Transdução de Sinais , Proteínas Smad Reguladas por Receptor/metabolismo , Proteínas da Superfamília de TGF-beta/fisiologia , Proteína Wnt1/genética , Proteína Wnt1/metabolismoRESUMO
TGF-ß activated kinase 1 (TAK1) is a mediator of various cytokine signaling pathways. Germline deficiency of Tak1 causes multiple abnormalities, including dilated blood vessels at midgestation. However, the mechanisms by which TAK1 regulates vessel formation have not been elucidated. TAK1 binding proteins 1 and 2 (TAB1 and TAB2) are activators of TAK1, but their roles in embryonic TAK1 signaling have not been determined. In the present study, we characterized mouse embryos harboring endothelial-specific deletions of Tak1, Tab1, or Tab2 and found that endothelial TAK1 and TAB2, but not TAB1, were critically involved in vascular formation. TAK1 deficiency in endothelial cells caused increased cell death and vessel regression at embryonic day 10.5 (E10.5). Deletion of TNF signaling largely rescued endothelial cell death in TAK1-deficient embryos at E10.5. However, embryos deficient in both TAK1 and TNF signaling still exhibited dilated capillary networks at E12.5. TAB2 deficiency caused reduced TAK1 activity, resulting in abnormal capillary blood vessels, similar to the compound deficiency of TAK1 and TNF signaling. Ablation of either TAK1 or TAB2 impaired cell migration and tube formation. Our results show that endothelial TAK1 signaling is important for 2 biologic processes in angiogenesis: inhibiting TNF-dependent endothelial cell death and promoting TNF-independent angiogenic cell migration.
Assuntos
Movimento Celular/fisiologia , Células Endoteliais/metabolismo , MAP Quinase Quinase Quinases/metabolismo , Neovascularização Fisiológica/fisiologia , Transdução de Sinais/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Sobrevivência Celular , Embrião de Mamíferos , Citometria de Fluxo , Humanos , Immunoblotting , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Microscopia Confocal , RNA Interferente Pequeno , Veias UmbilicaisRESUMO
The intestinal epithelium is constantly exposed to inducers of reactive oxygen species (ROS), such as commensal microorganisms. Levels of ROS are normally maintained at nontoxic levels, but dysregulation of ROS is involved in intestinal inflammatory diseases. In this article, we report that TGF-ß-activated kinase 1 (TAK1) is a key regulator of ROS in the intestinal epithelium. tak1 gene deletion in the mouse intestinal epithelium caused tissue damage involving enterocyte apoptosis, disruption of tight junctions, and inflammation. Disruption of TNF signaling, which is a major intestinal damage inducer, rescued the inflammatory conditions but not apoptosis or disruption of tight junctions in the TAK1-deficient intestinal epithelium, suggesting that TNF is not a primary inducer of the damage noted in TAK1-deficient intestinal epithelium. We found that TAK1 deficiency resulted in reduced expression of several antioxidant-responsive genes and reduced the protein level of a key antioxidant transcription factor NF-E2-related factor 2, which resulted in accumulation of ROS. Exogenous antioxidant treatment reduced apoptosis and disruption of tight junctions in the TAK1-deficient intestinal epithelium. Thus, TAK1 signaling regulates ROS through transcription factor NF-E2-related factor 2, which is important for intestinal epithelial integrity.
Assuntos
Imunidade nas Mucosas/fisiologia , Mucosa Intestinal/enzimologia , MAP Quinase Quinase Quinases/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Animais , Western Blotting , Epitélio/enzimologia , Epitélio/imunologia , Expressão Gênica , Regulação da Expressão Gênica/imunologia , Imuno-Histoquímica , Mucosa Intestinal/imunologia , MAP Quinase Quinase Quinases/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Fator 2 Relacionado a NF-E2/imunologia , Estresse Oxidativo/imunologia , Espécies Reativas de Oxigênio/imunologia , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Transforming growth factor beta-activated kinase 1 (TAK1) kinase is an indispensable signaling intermediate in tumor necrosis factor (TNF), interleukin 1, and Toll-like receptor signaling pathways. TAK1-binding protein 2 (TAB2) and its closely related protein, TAB3, are binding partners of TAK1 and have previously been identified as adaptors of TAK1 that recruit TAK1 to a TNF receptor signaling complex. TAB2 and TAB3 redundantly mediate activation of TAK1. In this study, we investigated the role of TAB2 by analyzing fibroblasts having targeted deletion of the tab2 gene. In TAB2-deficient fibroblasts, TAK1 was associated with TAB3 and was activated following TNF stimulation. However, TAB2-deficient fibroblasts displayed a significantly prolonged activation of TAK1 compared with wild type control cells. This suggests that TAB2 mediates deactivation of TAK1. We found that a TAK1-negative regulator, protein phosphatase 6 (PP6), was recruited to the TAK1 complex in wild type but not in TAB2-deficient fibroblasts. Furthermore, we demonstrated that both PP6 and TAB2 interacted with the polyubiquitin chains and this interaction mediated the assembly with TAK1. Our results indicate that TAB2 not only activates TAK1 but also plays an essential role in the deactivation of TAK1 by recruiting PP6 through a polyubiquitin chain-dependent mechanism.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , MAP Quinase Quinase Quinases/metabolismo , Sistema de Sinalização das MAP Quinases/fisiologia , Fator de Necrose Tumoral alfa/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Linhagem Celular Transformada , Derme/citologia , Fibroblastos/citologia , Fibroblastos/enzimologia , Deleção de Genes , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Camundongos , NF-kappa B/metabolismo , Fosfoproteínas Fosfatases/metabolismo , Fosforilação/fisiologia , Fator de Necrose Tumoral alfa/farmacologia , Ubiquitina/metabolismo , Ubiquitinação/fisiologiaRESUMO
Recent studies have revealed that TAK1 kinase is an essential intermediate in several innate immune signaling pathways. In this study, we investigated the role of TAK1 signaling in maintaining intestinal homeostasis by generating enterocyte-specific constitutive and inducible gene-deleted TAK1 mice. We found that enterocyte-specific constitutive TAK1-deleted mice spontaneously developed intestinal inflammation as observed by histological analysis and enhanced expression of IL-1beta, MIP-2, and IL-6 around the time of birth, which was accompanied by significant enterocyte apoptosis. When TAK1 was deleted in the intestinal epithelium of 4-wk-old mice using an inducible knockout system, enterocytes underwent apoptosis and intestinal inflammation developed within 2-3 days following the initiation of gene deletion. We found that enterocyte apoptosis and intestinal inflammation were strongly attenuated when enterocyte-specific constitutive TAK1-deleted mice were crossed to TNF receptor 1(-/-) mice. However, these mice later (>14 days) developed ileitis and colitis. Thus, TAK1 signaling in enterocytes is essential for preventing TNF-dependent epithelium apoptosis and the TNF-independent development of ileitis and colitis. We propose that aberration in TAK1 signaling might disrupt intestinal homeostasis and favor the development of inflammatory disease.
Assuntos
Apoptose , Colite/imunologia , Enterócitos/imunologia , Ileíte/imunologia , Mucosa Intestinal/imunologia , MAP Quinase Quinase Quinases/metabolismo , Animais , Colite/metabolismo , Enterócitos/citologia , Enterócitos/metabolismo , Ileíte/metabolismo , Mucosa Intestinal/metabolismo , MAP Quinase Quinase Quinases/imunologia , Camundongos , Camundongos Knockout , Camundongos Mutantes , Transdução de Sinais , Fator de Necrose Tumoral alfa/imunologia , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Estrogens are known to contribute to endothelial function and sympathetic activity, both of which are strongly associated with the pathogenesis of ischemic heart disease. In addition, estrogens improve impaired lipid profile, a risk factor of endothelial dysfunction. In this study, we investigated the effects of OS-0544, a structurally new selective estrogen receptor modulator (SERM), on endothelial function, sympathetic activity, and plasma cholesterol level in ovariectomized (OVX) rats. Female Sprague-Dawley rats were ovariectomized and orally treated with OS-0544 (or OS-0689, the (R)-enantiomer of OS-0544), or 17beta-estradiol (E2) for 4 weeks, starting the next days after ovariectomy or for 1 week, starting 6 weeks after ovariectomy. Ovariectomy significantly increased vasopressin-induced mean blood pressure (AVP-MBP) (57+/-3.3 mm Hg vs. 46+/-3.5 mm Hg, P<0.05) and decreased acetylcholine (Ach)-induced maximum vasorelaxation response (69+/-5.6% vs. 81+/-4.0%, P<0.05). OS-0544 significantly inhibited AVP-MBP elevation (46+/-3.5 mm Hg vs. 57+/-3.3 mm Hg, P<0.05) and decreased Ach-induced maximum vasorelaxation response (90+/-3.3% vs. 69+/-5.6%, P<0.05) in OVX rats. In addition, OS-0689 as well as E2 significantly reduced (up to 67%) the increase in sympathetic activity in OVX rats. Moreover, like E2, OS-0544 significantly decreased plasma cholesterol level in OVX rats. These results demonstrate that OS-0544 has vascular protective effect on vascular function after ovariectomy. It is therefore believed that OS-0544 has vascular protective effect in postmenopausal woman.
Assuntos
Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/fisiologia , Indanos/farmacologia , Ovariectomia , Piperidinas/farmacologia , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Compostos de Espiro/farmacologia , Sistema Nervoso Simpático/efeitos dos fármacos , Acetilcolina/farmacologia , Animais , Pressão Sanguínea/efeitos dos fármacos , Colesterol/sangue , Feminino , Ratos , Ratos Sprague-Dawley , Sistema Nervoso Simpático/fisiologia , Vasodilatação/efeitos dos fármacos , Vasopressinas/farmacologiaRESUMO
RIPK3, a key mediator of necroptosis, has been implicated in the host defense against viral infection primary in immune cells. However, gene expression analysis revealed that RIPK3 is abundantly expressed not only in immune organs but also in the gastrointestinal tract, particularly in the small intestine. We found that orally inoculated Listeria monocytogenes, a bacterial foodborne pathogen, efficiently spread and caused systemic infection in Ripk3-deficient mice while almost no dissemination was observed in wild-type mice. Listeria infection activated the RIPK3-MLKL pathway in cultured cells, which resulted in suppression of intracellular replication of Listeria Surprisingly, Listeria infection-induced phosphorylation of MLKL did not result in host cell killing. We found that MLKL directly binds to Listeria and inhibits their replication in the cytosol. Our findings have revealed a novel functional role of the RIPK3-MLKL pathway in nonimmune cell-derived host defense against Listeria invasion, which is mediated through cell death-independent mechanisms.
Assuntos
Listeria/crescimento & desenvolvimento , Listeriose/prevenção & controle , Necroptose , Proteínas Quinases/fisiologia , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Proteína Serina-Treonina Quinases de Interação com Receptores/fisiologia , Animais , Feminino , Humanos , Listeria/imunologia , Listeria/metabolismo , Listeriose/metabolismo , Listeriose/microbiologia , Listeriose/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteína Serina-Treonina Quinases de Interação com Receptores/genéticaRESUMO
TAK1 binding protein 1 (TAB1) binds and induces autophosphorylation of TGF-beta activating kinase (TAK1). TAK1, a mitogen-activated kinase kinase kinase, is involved in several distinct signaling pathways including non-Smad pathways for TGF-beta superfamily members and inflammatory responses caused by cytokines. Conventional disruption of the murine Tab1 gene results in late gestational lethality showing intraventricular septum defects and underdeveloped lung alveoli. To gain a better understanding of the roles of TAB1 in different tissues, at different stages of development, and in pathological conditions, we generated Tab1 floxed mice in which the loxP sites flank Exons 9 and 10 to remove the C-terminal region of TAB1 protein necessary for activation of TAK1. We demonstrate that Cre-mediated recombination using Sox2-Cre, a Cre line expressed in the epiblast during early embryogenesis, results in deletion of the gene and protein. These homozygous Cre-recombined null embryos display an identical phenotype to conventional null embryos. This animal model will be useful in revealing distinct roles of TAB1 in different tissues at different stages.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Marcação de Genes , Mutação , Animais , Éxons , Integrases/metabolismo , CamundongosRESUMO
The foraging patterns of insects reflect a combination of biotic and abiotic constraints. Pieris rapae (L.) (Lepidoptera: Pieridae) larvae exhibit plasticity in their foraging behavior, and their movements in response to flowers, young foliage, light, and gravity were studied. As predicted for palatable cryptic larvae, young instars of P. rapae exhibited predator avoidance behaviors. First- and second-instar larvae fed underneath the leaves where their eggs were oviposited, while late second- and third-instar larvae migrated away from their feeding damage. Using taxis experiments and choice tests, the direction of larval movement was significantly influenced by a hierarchy of three cues. Third-instar larvae exhibited negative gravitaxis, which could be supplanted by positive trophotaxis to young leaves and flowers. The larvae exhibited a significantly greater attraction to the inflorescence than to young foliage. For both the inflorescence and young foliage, visual cues were sufficient to direct larval movement. Understanding the cues that guide larval foraging may lead to more efficient trap crops for pest management.
Assuntos
Borboletas/fisiologia , Sinais (Psicologia) , Resposta Táctica , Animais , Comportamento Alimentar , Larva/fisiologia , Visão Ocular , Percepção VisualRESUMO
OBJECTIVE: Cutaneous anesthesia in early postoperative period is common after neck dissection even if the cervical nerve (CN) rootlets are preserved. The aim of this study was to evaluate if the preservation of the terminal branches of CNs using sub-sternocleidomastoid (SCM) approach combined with medially placed skin incision can prevent early postoperative anesthesia. MATERIAL AND METHODS: A retrospective chart review was performed on 129 neck dissections in 87 head and neck cancer patients. RESULTS: The early postoperative sensory preservation rates for the ear tab, submandibular, lateral neck, and sub-clavicular areas of CN rootlet-preserved necks (n = 86) were 75.6%, 20.9%, 74.4%, and 86.0%, respectively, compared with 37.2%, 2.3%, 2.3%, and 4.7%, respectively, in CN rootlet-resected necks (n = 43). In CN rootlet-preserved necks, the sub-SCM approach (n = 54) showed 81.5%, 27.8%, 92.6%, and 94.4% preservation rates, respectively, compared with 65.6%, 9.4%, 43.8%, and 71.9%, respectively, using the conventional subplatysmal approach (n = 32). The rates were significantly better in the submandibular, lateral neck, and sub-clavicular areas after sub-SCM approach. CONCLUSIONS: Preservation of CN rootlets is a required element for sensory preservation in neck dissection. The sub-SCM approach can effectively prevent early postoperative cutaneous anesthesia following CN-preserving neck dissection.
Assuntos
Esvaziamento Cervical/métodos , Sensação , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Neoplasias de Cabeça e Pescoço/cirurgia , Humanos , Masculino , Pessoa de Meia-Idade , Esvaziamento Cervical/efeitos adversos , Tratamentos com Preservação do Órgão/métodos , Estudos Retrospectivos , Distúrbios Somatossensoriais/etiologia , Distúrbios Somatossensoriais/prevenção & controleRESUMO
The inhibitory effects of hypertonic conditions on immune responses have been described in clinical studies; however, the molecular mechanism underlying this phenomenon has yet to be defined. Here we investigate osmotic stress-mediated modification of the NF-kappaB pathway, a central signaling pathway in inflammation. We unexpectedly found that osmotic stress could activate IkappaBalpha kinase but did not activate NF-kappaB. Osmotic stress-induced phosphorylated IkappaBalpha was not ubiquitinated, and osmotic stress inhibited interleukin 1-induced ubiquitination of IkappaBalpha and ultimately blocked expression of cytokine/chemokines. Thus, blockage of IkappaBalpha ubiquitination is likely to be a major mechanism for inhibition of inflammation by hypertonic conditions.
Assuntos
Proteínas I-kappa B/metabolismo , Inflamação/metabolismo , NF-kappa B/antagonistas & inibidores , Ubiquitinação/fisiologia , Animais , Células Cultivadas , Humanos , Quinase I-kappa B/metabolismo , Proteínas I-kappa B/antagonistas & inibidores , Interleucina-1/antagonistas & inibidores , Interleucina-1/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Inibidor de NF-kappaB alfa , NF-kappa B/metabolismo , Pressão Osmótica , Transdução de SinaisRESUMO
AMP-activated protein kinase (AMPK) promotes glucose transport, maintains ATP stores, and prevents injury and apoptosis during ischemia. AMPK has several direct molecular targets in the heart but also may interact with other stress-signaling pathways. This study examined the role of AMPK in the activation of the p38 mitogen-activated protein kinase (MAPK). In isolated heart muscles, the AMPK activator 5-aminoimidazole-4-carboxy-amide-1-beta-D-ribofuranoside (AICAR) increased p38 MAPK activation. In AMPK-deficient mouse hearts, expressing a kinase-dead (KD) alpha2 catalytic subunit, p38 MAPK activation was markedly reduced during low-flow ischemia (2.3- versus 7-fold in wild-type hearts, P<0.01) and was similarly reduced during severe no-flow ischemia in KD hearts (P<0.01 versus ischemic wild type). Knockout of the p38 MAPK upstream kinase, MAPK kinase 3 (MKK3), did not affect ischemic activation of either AMPK or p38 MAPK in transgenic mkk3(-/-) mouse hearts. Ischemia increased p38 MAPK recruitment to transforming growth factor-beta-activated protein kinase 1-binding protein 1 (TAB1), a scaffold protein that promotes p38 MAPK autophosphorylation. Moreover, TAB1 was associated with the alpha2 catalytic subunit of AMPK. p38 MAPK recruitment to TAB1/AMPK complexes required AMPK activation and was reduced in ischemic AMPK-deficient transgenic mouse hearts. The potential role of p38 MAPK in mediating the downstream action of AMPK to promote glucose transport was also assessed. The p38 MAPK inhibitor SB203580 partially inhibited both AICAR- and hypoxia-stimulated glucose uptake and GLUT4 translocation. Activation of p38 MAPK by anisomycin also increased glucose transport in heart muscles. Thus, AMPK has an important role in promoting p38 MAPK activation in the ischemic heart by inducing p38 MAPK autophosphorylation through interaction with the scaffold protein TAB1.