RESUMO
Akkermansia muciniphila has received great attention because of its beneficial roles in gut health by regulating gut immunity, promoting intestinal epithelial development, and improving barrier integrity. However, A. muciniphila-derived functional molecules regulating gut health are not well understood. Microbiome-secreted proteins act as key arbitrators of host-microbiome crosstalk through interactions with host cells in the gut and are important for understanding host-microbiome relationships. Herein, we report the biological function of Amuc_1409, a previously uncharacterised A. muciniphila-secreted protein. Amuc_1409 increased intestinal stem cell (ISC) proliferation and regeneration in ex vivo intestinal organoids and in vivo models of radiation- or chemotherapeutic drug-induced intestinal injury and natural aging with male mice. Mechanistically, Amuc_1409 promoted E-cadherin/ß-catenin complex dissociation via interaction with E-cadherin, resulting in the activation of Wnt/ß-catenin signaling. Our results demonstrate that Amuc_1409 plays a crucial role in intestinal homeostasis by regulating ISC activity in an E-cadherin-dependent manner and is a promising biomolecule for improving and maintaining gut health.
Assuntos
Verrucomicrobia , beta Catenina , Masculino , Camundongos , Animais , beta Catenina/metabolismo , Verrucomicrobia/metabolismo , Intestinos , Caderinas/metabolismo , AkkermansiaRESUMO
AIMS: The gut microbiota is increasingly recognised as a pivotal regulator of immune system homeostasis and brain health. Recent research has implicated the gut microbiota in age-related cognitive impairment and dementia. Agathobaculum butyriciproducens SR79 T (SR79), which was identified in the human gut, has been reported to be beneficial in addressing cognitive deficits and pathophysiologies in a mouse model of Alzheimer's disease. However, it remains unknown whether SR79 affects age-dependent cognitive impairment. MAIN METHOD: To explore the effects of SR79 on cognitive function during ageing, we administered SR79 to aged mice. Ageing-associated behavioural alterations were examined using the open field test (OFT), tail suspension test (TST), novel object recognition test (NORT), Y-maze alternation test (Y-maze), and Morris water maze test (MWM). We investigated the mechanisms of action in the gut and brain using molecular and histological analyses. KEY FINDINGS: Administration of SR79 improved age-related cognitive impairment without altering general locomotor activity or depressive behaviour in aged mice. Furthermore, SR79 increased mature dendritic spines in the pyramidal cells of layer III and phosphorylation of CaMKIIα in the cortex of aged mice. Age-related activation of astrocytes in the cortex of layers III-V of the aged brain was reduced following SR79 administration. Additionally, SR79 markedly increased IL-10 production and Foxp3 and Muc2 mRNA expression in the colons of aged mice. SIGNIFICANCE: These findings suggest that treatment with SR79 may be a beneficial microbial-based approach for enhancing cognitive function during ageing.
Assuntos
Clostridiales , Transtornos Cognitivos , Disfunção Cognitiva , Camundongos , Humanos , Animais , Idoso , Transtornos Cognitivos/metabolismo , Encéfalo/metabolismo , Envelhecimento/metabolismoRESUMO
Fulminant hepatitis is characterized by rapid and massive immune-mediated liver injury. Dosage-sensitive sex reversal-adrenal hypoplasia congenita critical region on the X chromosome, gene 1 (DAX1; NR0B1) represses the transcription of various genes. Here, we determine whether DAX1 serves as a regulator of inflammatory liver injury induced by concanavalin A (ConA). C57BL/6J (WT), myeloid cell-specific Dax1 knockout (MKO), and hepatocyte-specific Dax1 knockout (LKO) mice received single intravenous administration of ConA. Histopathological changes in liver and plasma alanine aminotransferase and aspartate aminotransferase levels in Dax1 MKO mice were comparable with those in WT mice following ConA administration. Unlike Dax1 MKO mice, Dax1 LKO mice were greatly susceptible to ConA-induced liver injury, which was accompanied by enhanced infiltration of immune cells, particularly CD4+ and CD8+ T cells, in the liver. Factors related to T-cell recruitment, including chemokines and adhesion molecules, significantly increased following enhanced and prolonged phosphorylation of NF-κB p65 in the liver of ConA-administered Dax1 LKO mice. This is the first study to demonstrate that hepatocyte-specific DAX1 deficiency exacerbates inflammatory liver injury via NF-κB p65 activation, thereby causing T-cell infiltration by modulating inflammatory chemokines and adhesion molecules. Our results suggest DAX1 as a therapeutic target for fulminant hepatitis treatment.
Assuntos
Linfócitos T CD8-Positivos , Necrose Hepática Massiva , Camundongos , Animais , NF-kappa B , Camundongos Endogâmicos C57BL , Hepatócitos , Transdução de Sinais , Concanavalina A/toxicidade , Linfócitos T CD4-PositivosRESUMO
Acetaminophen (APAP) is a widely used analgesic and antipyretic drug, but its overdose can cause acute liver failure. The dosage-sensitive sex reversal adrenal hypoplasia congenita critical region on the X chromosome, gene 1 (DAX-1, NR0B1), is an orphan nuclear receptor that acts as a transcriptional co-repressor of various genes. In this study, we identified the role of DAX-1 in APAP-induced liver injury using hepatocyte-specific Dax-1 knockout (Dax-1 LKO) mice. Mouse primary hepatocytes were used as a comparative in vitro study. APAP overdose led to decreased plasma alanine aminotransferase and aspartate aminotransferase levels in Dax-1 LKO mice compared to C57BL/6J (WT) controls, accompanied by reduced liver necrosis. The expression of the genes encoding the enzymes catalyzing glutathione (GSH) synthesis and metabolism and antioxidant enzymes was increased in the livers of APAP-treated Dax-1 LKO mice. The rapid recovery of GSH levels in the mitochondrial fraction of APAP-treated Dax-1 LKO mice led to reduced reactive oxygen species levels, resulting in the inhibition of the prolonged JNK activation. The hepatocyte-specific DAX-1 deficiency increased the protein expression of nuclear factor erythroid 2-related factor 2 (Nrf2) compared with WT controls after APAP administration. These results indicate that DAX-1 deficiency in hepatocytes protects against APAP-induced liver injury by Nrf2-regulated antioxidant defense.
Assuntos
Antipiréticos , Doença Hepática Induzida por Substâncias e Drogas , Receptor Nuclear Órfão DAX-1 , Fator 2 Relacionado a NF-E2 , Acetaminofen/toxicidade , Alanina Transaminase/metabolismo , Animais , Antioxidantes/metabolismo , Aspartato Aminotransferases/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/genética , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/prevenção & controle , Proteínas Correpressoras/metabolismo , Receptor Nuclear Órfão DAX-1/genética , Glutationa/metabolismo , Hepatócitos/metabolismo , Fígado/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Receptores Nucleares Órfãos/metabolismo , Espécies Reativas de Oxigênio/metabolismoRESUMO
Autism spectrum disorder (ASD) is a neurodevelopment disorder characterized by deficits in social interaction and restrictive, repetitive, and stereotypical patterns of behavior. However, there is no pharmacological drug that is currently used to target these core ASD symptoms. Sodium phenylbutyrate (NaPB) is a well-known long-term treatment of urea cycle disorders in children. In this study, we assessed the therapeutic effects of NaPB, which is a chemical chaperone as well as histone deacetylase inhibitor on a BTBR T + Itpr3tf/J (BTBR) mice model of ASD. We found that acute and chronic treatment of NaPB remarkably improved, not only core ASD symptoms, including repetitive behaviors and sociability deficit, but also cognitive impairment in the BTBR mice. NaPB substantially induced histone acetylation in the brain of the BTBR mice. Intriguingly, the therapeutic effects of NaPB on autistic-like behaviors, such as repetitive behaviors, impaired sociability, and cognitive deficit also showed in the valproic acid (VPA)-induced mouse model of autism. In addition, pentylenetetrazole (PTZ)-induced seizure was significantly attenuated by NaPB treatment in C57BL/6J and BTBR mice. These findings suggest that NaPB may provide a novel therapeutic approach for the treatment of patients with ASD.
Assuntos
Transtorno do Espectro Autista/tratamento farmacológico , Disfunção Cognitiva/tratamento farmacológico , Asseio Animal/efeitos dos fármacos , Fenilbutiratos/uso terapêutico , Comportamento Social , Comportamento Estereotipado/efeitos dos fármacos , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Transtorno do Espectro Autista/induzido quimicamente , Transtorno do Espectro Autista/psicologia , Encéfalo/efeitos dos fármacos , Encéfalo/fisiologia , Disfunção Cognitiva/psicologia , Modelos Animais de Doenças , Feminino , Asseio Animal/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos , Camundongos Transgênicos , Fenilbutiratos/farmacologia , Comportamento Estereotipado/fisiologia , Ácido Valproico/toxicidadeRESUMO
The dopamine precursor 3,4dihydroxyphenyl lalanine (LDOPA) is the most widely used symptomatic treatment for Parkinson's disease (PD); however, its prolonged use is associated with LDOPAinduced dyskinesia in more than half of patients after 10 years of treatment. The present study investigated whether cotreatment with ßLapachone, a natural compound, and LDOPA has protective effects in a 6hydroxydopamine (6OHDA)induced mouse model of PD. Unilateral 6OHDAlesioned mice were treated with vehicle or ßLapachone (10 mg/kg/day) and LDOPA for 11 days. Abnormal involuntary movements (AIMs) were scored on days 5 and 10. ßLapachone (10 mg/kg) cotreatment with LDOPA decreased the AIMs score on both days 5 and 10. ßLapachone was demonstrated to have a beneficial effect on the axial and limb AIMs scores on day 10. There was no significant suppression in dopamine D1 receptorrelated and ERK1/2 signaling in the DAdenervated striatum by ßLapachonecotreatment with LDOPA. Notably, ßLapachonecotreatment with LDOPA increased phosphorylation at the Ser9 site of glycogen synthase kinase 3ß (GSK3ß), indicating suppression of GSK3ß activity in both the unlesioned and 6OHDAlesioned striata. In addition, astrocyte activation was markedly suppressed by ßLapachonecotreatment with LDOPA in the striatum and substantia nigra of the unilateral 6OHDA model. These findings suggest that ßLapachone cotreatment with LDOPA therapy may have therapeutic potential for the suppression or management of the development of LDOPAinduced dyskinesia in patients with PD.
Assuntos
Discinesias , Levodopa/efeitos adversos , Naftoquinonas/farmacologia , Oxidopamina/efeitos adversos , Doença de Parkinson Secundária , Animais , Discinesias/tratamento farmacológico , Discinesias/metabolismo , Discinesias/patologia , Levodopa/farmacologia , Masculino , Camundongos , Oxidopamina/farmacologia , Doença de Parkinson Secundária/induzido quimicamente , Doença de Parkinson Secundária/tratamento farmacológico , Doença de Parkinson Secundária/metabolismo , Doença de Parkinson Secundária/patologiaRESUMO
The growth arrest and DNA damage-inducible beta (Gadd45ß) protein have been associated with various cellular functions, but its role in progressive renal disease is currently unknown. Here, we examined the effect of Gadd45ß deletion on cell proliferation and apoptosis, inflammation, and renal fibrosis in an early chronic kidney disease (CKD) mouse model following unilateral ureteral obstruction (UUO). Wild-type (WT) and Gadd45ß-knockout (KO) mice underwent either a sham operation or UUO and the kidneys were sampled eight days later. A histological assay revealed that ablation of Gadd45ß ameliorated UUO-induced renal injury. Cell proliferation was higher in Gadd45ß KO mouse kidneys, but apoptosis was similar in both genotypes after UUO. Expression of pro-inflammatory cytokines after UUO was down-regulated in the kidneys from Gadd45ß KO mice, whereas UUO-mediated immune cell infiltration remained unchanged. The expression of pro-inflammatory cytokines in response to LPS stimulation decreased in bone marrow-derived macrophages from Gadd45ß KO mice compared with that in WT mice. Importantly, UUO-induced renal fibrosis was ameliorated in Gadd45ß KO mice unlike in WT mice. Gadd45ß was involved in TGF-ß signalling pathway regulation in kidney fibroblasts. Our findings demonstrate that Gadd45ß plays a crucial role in renal injury and may be a therapeutic target for the treatment of CKD.
Assuntos
Antígenos de Diferenciação/genética , Deleção de Genes , Nefropatias/etiologia , Obstrução Ureteral/complicações , Animais , Antígenos de Diferenciação/metabolismo , Apoptose/genética , Biomarcadores , Biópsia , Linhagem Celular , Proliferação de Células , Citocinas/metabolismo , Modelos Animais de Doenças , Suscetibilidade a Doenças , Fibrose , Imuno-Histoquímica , Inflamação , Mediadores da Inflamação , Nefropatias/metabolismo , Nefropatias/patologia , Masculino , Camundongos , Camundongos Knockout , Insuficiência Renal Crônica/etiologia , Insuficiência Renal Crônica/metabolismo , Insuficiência Renal Crônica/patologia , Fator de Crescimento Transformador beta/metabolismoRESUMO
Parkinson's disease (PD) is a neurodegenerative disease characterized by the progressive loss of dopaminergic neurons in the substantia nigra, leading to motor symptoms. Despite the remarkable improvements in the management of PD in recent decades, many patients remain significantly disabled. Metformin is a primary medication for the management of type 2 diabetes. We previously showed that co-treatment with metformin and 3,4-dihydroxyphenyl-l-alanine (l-DOPA) prevented the development of l-DOPA-induced dyskinesia in a 6-hydroxydopamine (6-OHDA)-lesioned animal model of PD. However, effects of metformin on PD- and aging-induced genes in reactive astrocytes remain unknown. In this study, we assessed the effect of metformin on motor function, neuroprotection, and reactive astrocytes in the 6-OHDA-induced PD animal model. In addition, the effects of metformin on the genes expressed by specific types of astrocytes were analyzed in PD model and aged mice. Here, we showed that metformin treatment effectively improves the motor symptoms in the 6-OHDA-induced PD mouse model, whereas metformin had no effect on tyrosine hydroxylase-positive neurons. The activation of AMPK and BDNF signaling pathways was induced by metformin treatment on the 6-OHDA-lesioned side of the striatum. Metformin treatment caused astrocytes to alter reactive genes in a PD animal model. Moreover, aging-induced genes in reactive astrocytes were effectively regulated or suppressed by metformin treatment. Taken together, these results suggest that metformin should be evaluated for the treatment of Parkinson's disease and related neurologic disorders characterized by astrocyte activation.
Assuntos
Envelhecimento/fisiologia , Astrócitos/efeitos dos fármacos , Astrócitos/fisiologia , Corpo Estriado/fisiologia , Metformina/administração & dosagem , Doença de Parkinson/fisiopatologia , Animais , Corpo Estriado/efeitos dos fármacos , Modelos Animais de Doenças , Feminino , Masculino , Camundongos Endogâmicos C57BL , Atividade Motora/efeitos dos fármacosRESUMO
The NAD(P)H:quinone oxidoreductase 1 (NQO1) gene encodes a cytosolic flavoenzyme that catalyzes the two-electron reduction of quinones to hydroquinones. A polymorphic form of NQO1 is associated with mood disorders such as schizophrenia. However, the role of NQO1 in dopaminergic system has not yet been elucidated. To determine the role of NQO1 in the dopaminergic system, we investigated pharmaco-behavioral effects of d-amphetamine using NQO1-deficienct mice. According to our comparative study involving NQO1+/+ and NQO1-/- mice, NQO1 deficiency increased d-amphetamine-induced psychomotor activity and psychological dependency compared to wild-type mice. Basal and d-amphetamine-induced dopamine levels were also enhanced by NQO1 deficiency. In NQO1-/- mice, neural activation induced by d-amphetamine was higher in dorsolateral striatum, but not in dorsomedial and ventral striata. Although protein level of CaMKIIα, which is a key player in amphetamine-induced dopamine efflux, was decreased in striata of NQO1-/- mice, phosphorylation of CaMKIIα was markedly enhanced in NQO1-/- mice compared to wild-type mice. Interestingly, experiments with pharmacological antagonist showed that D2 antagonist-induced suppression of locomotion required activation of NQO1. Moreover, the rewarding effect in response to D1 agonist was increased by NQO1 deficiency. These results suggest that striatal NQO1 is of considerable interest to understand the mechanism of dopaminergic regulation of psychiatric disorders.
Assuntos
Corpo Estriado/metabolismo , Dextroanfetamina/farmacologia , Dopamina/metabolismo , NAD(P)H Desidrogenase (Quinona)/deficiência , Receptores de Dopamina D1/metabolismo , Receptores de Dopamina D2/metabolismo , Animais , Corpo Estriado/efeitos dos fármacos , Antagonistas dos Receptores de Dopamina D2/farmacologia , Locomoção/efeitos dos fármacos , Locomoção/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de Dopamina D1/agonistas , Receptores de Dopamina D1/antagonistas & inibidores , Receptores de Dopamina D2/agonistasRESUMO
Humulus japonicus (HJ) is a widely used herbal medicine in Asia with antioxidative, antimicrobial, and antiinflammatory effects. We investigated the potential therapeutic effects of HJ in rheumatoid arthritis (RA) using a mouse model of collageninduced arthritis (CIA) and a lipopolysaccharidestimulated murine macrophage cell line (RAW 264.7). The CIA mice were administered 300 mg/kg HJ orally starting 3 days prior to second immunization. The clinical and histopathological findings were assessed in the paw of CIA mice. The levels of autoantibodies and inflammatory markers were determined in the plasma and cell culture supernatant, respectively. The expression at mRNA and protein levels was analyzed by reverse transcription quantitativePCR and western blot analysis, respectively. HJ significantly decreased the gross arthritic scores and paw swelling in CIA mice. Furthermore, synovial inflammation, cartilage destruction, and bone erosion were markedly reduced by HJ. It also decreased the expression of inflammatory enzymes in both the paw of mice and RAW 264.7 cells. Moreover, the expression of genes related to all macrophages and proinflammatory M1 macrophage were significantly decreased, whereas the expression of antiinflammatory M2 macrophage marker was markedly increased in the paw of HJtreated CIA mice. In addition, HJ suppressed the levels of plasma antitype II collagen antibody following the decreased expression of T helper type 1 (Th1) and Th2 cellassociated surface markers and cytokines in the paw. HJ also significantly inhibited the expression of IL6 both in vitro and in vivo, followed by reduced STAT3 phosphorylation and expression in the paw of CIA mice. Finally, the expression of osteoclastrelated genes was decreased in the paw of HJtreated CIA mice. These findings suggest that HJ can play a role in suppressing the development of CIA by overall regulation of articular inflammation. This study should provide new insights into the use of HJ as a therapeutically effective natural product against RA.
Assuntos
Anti-Inflamatórios/uso terapêutico , Artrite Experimental/tratamento farmacológico , Artrite Reumatoide/tratamento farmacológico , Humulus , Extratos Vegetais/uso terapêutico , Animais , Anti-Inflamatórios/química , Artrite Experimental/imunologia , Artrite Reumatoide/imunologia , Autoanticorpos/imunologia , Modelos Animais de Doenças , Humulus/química , Inflamação/tratamento farmacológico , Inflamação/imunologia , Mediadores da Inflamação/imunologia , Masculino , Camundongos , Extratos Vegetais/química , Células RAW 264.7RESUMO
Clusterin is a glycoprotein that is expressed in most human tissues and found in body fluids. In our previous studies we demonstrated that clusterin has a protective effect against hepatic lipid accumulation and renal fibrosis; however, the role of clusterin in hepatic fibrosis is unknown. Here, we examined whether clusterin had protective effects against hepatic fibrosis using in vitro and in vivo models. Clusterin was upregulated in the livers of human cirrhotic patients and in thioacetamide (TAA)-induced and bile duct ligation mouse models of liver fibrosis. Loss and overexpression of clusterin promoted and attenuated hepatic fibrosis after TAA injection, respectively. In addition, we found that clusterin attenuates hepatic fibrosis by inhibiting the activation of hepatic stellate cells and Smad3 signaling pathways. Thus, clusterin plays an important role in hepatic fibrosis.
Assuntos
Clusterina/genética , Células Estreladas do Fígado/citologia , Cirrose Hepática/metabolismo , Proteína Smad3/metabolismo , Tioacetamida/efeitos adversos , Animais , Linhagem Celular , Clusterina/metabolismo , Modelos Animais de Doenças , Regulação para Baixo , Técnicas de Inativação de Genes , Células Estreladas do Fígado/metabolismo , Humanos , Cirrose Hepática/induzido quimicamente , Cirrose Hepática/genética , Masculino , Camundongos , Transdução de Sinais , Regulação para CimaRESUMO
Clinical prescription of cisplatin, one of the most widely used chemotherapeutic agents, is limited by its side effects, particularly tubular injury-associated nephrotoxicity. Since details of the underlying mechanisms are not fully understood, we investigated the role of pyruvate dehydrogenase kinase (PDK) in cisplatin-induced acute kidney injury. Among the PDK isoforms, PDK4 mRNA and protein levels were markedly increased in the kidneys of mice treated with cisplatin, and c-Jun N-terminal kinase activation was involved in cisplatin-induced renal PDK4 expression. Treatment with the PDK inhibitor sodium dichloroacetate (DCA) or genetic knockout of PDK4 attenuated the signs of cisplatin-induced acute kidney injury, including apoptotic morphology of the kidney tubules along with numbers of TUNEL-positive cells, cleaved caspase-3, and renal tubular injury markers. Cisplatin-induced suppression of the mitochondrial membrane potential, oxygen consumption rate, expression of electron transport chain components, cytochrome c oxidase activity, and disruption of mitochondrial morphology were noticeably improved in the kidneys of DCA-treated or PDK4 knockout mice. Additionally, levels of the oxidative stress marker 4-hydroxynonenal and mitochondrial reactive oxygen species were attenuated, whereas superoxide dismutase 2 and catalase expression and glutathione synthetase and glutathione levels were recovered in DCA-treated or PDK4 knockout mice. Interestingly, lipid accumulation was considerably attenuated in DCA-treated or PDK4 knockout mice via recovered expression of peroxisome proliferator-activated receptor-α and coactivator PGC-1α, which was accompanied by recovery of mitochondrial biogenesis. Thus, PDK4 mediates cisplatin-induced acute kidney injury, suggesting that PDK4 might be a therapeutic target for attenuating cisplatin-induced acute kidney injury.
Assuntos
Injúria Renal Aguda/prevenção & controle , Cisplatino , Túbulos Renais/enzimologia , Proteínas Serina-Treonina Quinases/deficiência , Injúria Renal Aguda/enzimologia , Injúria Renal Aguda/genética , Injúria Renal Aguda/patologia , Animais , Apoptose , Caspase 3/metabolismo , Células Cultivadas , Modelos Animais de Doenças , Metabolismo Energético , Inibidores Enzimáticos/farmacologia , Regulação Enzimológica da Expressão Gênica , Predisposição Genética para Doença , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Túbulos Renais/efeitos dos fármacos , Túbulos Renais/ultraestrutura , Masculino , Potencial da Membrana Mitocondrial , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias/enzimologia , Mitocôndrias/patologia , Biogênese de Organelas , Estresse Oxidativo , Fenótipo , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/genética , Piruvato Desidrogenase Quinase de Transferência de Acetil , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fatores de TempoRESUMO
The hallmark of renal tubulointerstitial fibrosis is the accumulation of myofibroblasts and extracellular matrix proteins. Fyn, a member of the Src family of kinases, has diverse biological functions including regulation of mitogenic signaling and proliferation and integrin-mediated interaction. Src family proteins promote pulmonary fibrosis by augmenting transforming growth factor-ß signaling, but their role in renal fibrosis is less understood. We observed upregulation of Fyn in a renal fibrosis model induced by unilateral ureteral obstruction. Upon ureteral obstruction, Fyn-deficient mice exhibited attenuated renal fibrosis relative to wild-type mice. Furthermore, obstruction-induced renal expression of type I collagen, fibronectin, α-smooth muscle actin, and plasminogen activator inhibitor-1 was suppressed. Pharmacologic inhibition of Fyn blocked induction of extracellular matrix proteins in kidney cell lines. Importantly, the attenuation of renal fibrosis by Fyn deficiency was not accompanied by changes in the Smad pathway. Rather, the antifibrotic effect of Fyn deficiency was associated with downregulation of signal transducer and activator of transcription 3 (STAT3). Small, interfering RNA targeting STAT3 in Fyn-deficient cells further suppressed α-smooth muscle actin expression, whereas a STAT3 activator partially restored plasminogen activator inhibitor-1 expression, indicating that STAT3 signaling is critically involved in this process. Thus, Fyn plays an important role in renal fibrosis. Hence, Fyn kinase inhibitors may be therapeutically useful against renal fibrosis.
Assuntos
Nefroesclerose/metabolismo , Proteínas Proto-Oncogênicas c-fyn/metabolismo , Fator de Transcrição STAT3/metabolismo , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Caderinas/metabolismo , Receptores ErbB/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Humanos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Nefroesclerose/etiologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-fyn/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-fyn/genética , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Obstrução Ureteral/complicações , Quinases da Família src/metabolismoRESUMO
Hepatic steatosis is associated with increased insulin resistance and tricarboxylic acid (TCA) cycle flux, but decreased ketogenesis and pyruvate dehydrogenase complex (PDC) flux. This study examined whether hepatic PDC activation by inhibition of pyruvate dehydrogenase kinase 2 (PDK2) ameliorates these metabolic abnormalities. Wild-type mice fed a high-fat diet exhibited hepatic steatosis, insulin resistance, and increased levels of pyruvate, TCA cycle intermediates, and malonyl-CoA but reduced ketogenesis and PDC activity due to PDK2 induction. Hepatic PDC activation by PDK2 inhibition attenuated hepatic steatosis, improved hepatic insulin sensitivity, reduced hepatic glucose production, increased capacity for ß-oxidation and ketogenesis, and decreased the capacity for lipogenesis. These results were attributed to altered enzymatic capacities and a reduction in TCA anaplerosis that limited the availability of oxaloacetate for the TCA cycle, which promoted ketogenesis. The current study reports that increasing hepatic PDC activity by inhibition of PDK2 ameliorates hepatic steatosis and insulin sensitivity by regulating TCA cycle anaplerosis and ketogenesis. The findings suggest PDK2 is a potential therapeutic target for nonalcoholic fatty liver disease.
Assuntos
Fígado Gorduroso/enzimologia , Fígado Gorduroso/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Ciclo do Ácido Cítrico/genética , Ciclo do Ácido Cítrico/fisiologia , Dieta Hiperlipídica/efeitos adversos , Fígado Gorduroso/etiologia , Glucose/metabolismo , Resistência à Insulina , Lipogênese/fisiologia , Fígado/metabolismo , Fígado/patologia , Masculino , Malonil Coenzima A/metabolismo , Camundongos , Camundongos Knockout , Ácido Oxaloacético/metabolismo , Proteínas Serina-Treonina Quinases/deficiência , Proteínas Serina-Treonina Quinases/genética , Piruvato Desidrogenase Quinase de Transferência de Acetil , Complexo Piruvato Desidrogenase/metabolismo , Ácido Pirúvico/metabolismoRESUMO
Vascular calcification, a pathologic response to defective calcium and phosphate homeostasis, is strongly associated with cardiovascular mortality and morbidity. In this study, we have observed that pyruvate dehydrogenase kinase 4 (PDK4) is upregulated and pyruvate dehydrogenase complex phosphorylation is increased in calcifying vascular smooth muscle cells (VSMCs) and in calcified vessels of patients with atherosclerosis, suggesting that PDK4 plays an important role in vascular calcification. Both genetic and pharmacological inhibition of PDK4 ameliorated the calcification in phosphate-treated VSMCs and aortic rings and in vitamin D3-treated mice. PDK4 augmented the osteogenic differentiation of VSMCs by phosphorylating SMAD1/5/8 via direct interaction, which enhances BMP2 signaling. Furthermore, increased expression of PDK4 in phosphate-treated VSMCs induced mitochondrial dysfunction followed by apoptosis. Taken together, our results show that upregulation of PDK4 promotes vascular calcification by increasing osteogenic markers with no adverse effect on bone formation, demonstrating that PDK4 is a therapeutic target for vascular calcification.
Assuntos
Proteínas Quinases/genética , Proteínas Quinases/metabolismo , Proteína Smad1/metabolismo , Proteína Smad5/metabolismo , Proteína Smad8/metabolismo , Calcificação Vascular/metabolismo , Animais , Apoptose/genética , Biomarcadores , Remodelação Óssea/genética , Diferenciação Celular , Células Cultivadas , Modelos Animais de Doenças , Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Masculino , Camundongos , Camundongos Knockout , Mitocôndrias/genética , Mitocôndrias/metabolismo , Músculo Liso Vascular/citologia , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/citologia , Miócitos de Músculo Liso/metabolismo , Osteogênese/genética , Fosforilação , Ligação Proteica , Inibidores de Proteínas Quinases/farmacologia , Proteínas Quinases/químicaRESUMO
OBJECTIVE: Vascular calcification which refers to ectopic mineralization in vascular cells is associated with several conditions, such as chronic kidney disease, atherosclerosis, and diabetes mellitus. Estrogen-related receptor (ERR)γ is a member of the orphan nuclear receptor superfamily, which plays diverse roles in regulating homeostatic and metabolic processes. However, the role of ERRγ in vascular calcification has not been investigated to date. The aim of the present study was to examine the role of ERRγ in vascular calcification. APPROACH AND RESULTS: Vascular calcification was induced by treating rat aortic vascular smooth muscle cells with calcification medium. ERRγ expression in vascular smooth muscle cells was induced during calcification medium-induced vascular calcification. Adenovirus-mediated overexpression of ERRγ in vascular smooth muscle cells resulted in the upregulation of the expression of osteogenic genes, including runt-related transcription factor 2, osteopontin, and Msx2, and the downregulation of α-smooth muscle actin. Adenovirus-mediated overexpression of ERRγ induced bone morphogenetic protein 2 (BMP2) expression, leading to increased phosphorylation of the intracellular BMP2 effector proteins SMAD1/5/8. Collectively, these data suggested that ERRγ promotes dedifferentiation of vascular smooth muscle cells to an osteogenic phenotype during the vascular calcification process. Inhibition of endogenous ERRγ expression or activity using a specific siRNA or the selective inverse agonist GSK5182 attenuated vascular calcification and osteogenic gene expression in vitro and in vivo. CONCLUSIONS: The present results indicate that ERRγ plays a key role in vascular calcification by upregulating the BMP2 signaling pathway, suggesting that inhibition of ERRγ is a potential therapeutic strategy for the prevention of vascular calcification.
Assuntos
Doenças da Aorta/metabolismo , Proteína Morfogenética Óssea 2/metabolismo , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Osteogênese , Receptores de Estrogênio/metabolismo , Calcificação Vascular/metabolismo , Animais , Aorta/metabolismo , Aorta/patologia , Doenças da Aorta/genética , Doenças da Aorta/patologia , Doenças da Aorta/prevenção & controle , Sítios de Ligação , Proteína Morfogenética Óssea 2/genética , Desdiferenciação Celular , Linhagem Celular , Modelos Animais de Doenças , Masculino , Camundongos Endogâmicos C57BL , Músculo Liso Vascular/efeitos dos fármacos , Músculo Liso Vascular/patologia , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/patologia , Regiões Promotoras Genéticas , Interferência de RNA , Ratos , Ratos Sprague-Dawley , Receptores de Estrogênio/antagonistas & inibidores , Receptores de Estrogênio/genética , Transdução de Sinais , Tamoxifeno/análogos & derivados , Tamoxifeno/farmacologia , Fatores de Tempo , Transfecção , Regulação para Cima , Calcificação Vascular/genética , Calcificação Vascular/patologia , Calcificação Vascular/prevenção & controleRESUMO
Ischemic retinopathies causing overexpression of pro-angiogenic factors, including vascular endothelial growth factor (VEGF), are the most common cause of blindness. Thus, understanding the pathophysiology of targetable pathways that regulate retinal VEGF is of great interest. A conserved binding site for estrogen-related receptor γ (ERRγ) has been identified in the promoter of the Vegfa gene. ERRγ is a constitutively active orphan nuclear receptor and its expression is increased by hypoxic stimuli in metabolically active tissues. This study evaluated the role of ERRγ in the ischemic retina and the anti-VEGF potential of GSK5182, a selective inverse agonist of ERRγ. In an oxygen-induced retinopathy (OIR) mouse model, immunohistochemistry showed significantly increased ERRγ expression in the ganglion cell layer at postnatal day (P) 17. In a ganglion cell line (RGC-5), mRNA and protein levels of ERRγ were increased by desferrioxamine treatment and hypoxic conditions (1% O2). Transient transfection of RGC-5 cells revealed that ERRγ regulated Vegfa expression and this was inhibited by GSK5182. Intravitreal injection of GSK5182 into the OIR model at P14 inhibited retinal Vegfa mRNA expression at P17. GSK5182 suppresses hypoxia-induced VEGF expression via ERRγ; therefore, ERRγ could be a treatment target for ischemic retinopathies.
Assuntos
Hipóxia/metabolismo , Receptores de Estrogênio/biossíntese , Retina/patologia , Fator A de Crescimento do Endotélio Vascular/biossíntese , Animais , Linhagem Celular , Camundongos , Camundongos Endogâmicos C57BL , Ratos , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Dimethyl fumarate (DMF) has several pharmacological benefits including immunomodulation and prevention of fibrosis, which are dependent on the NF-E2-related factor 2 (Nrf2) antioxidant pathways. Therefore, we hypothesized that DMF could attenuate vascular calcification via Nrf2 activation. Vascular calcification induced by hyperphosphataemia was significantly inhibited by DMF in vascular smooth muscle cells (VSMCs) in a dose-dependent manner. DMF-mediated Nrf2 upregulation was accompanied by the reduced expressions of genes related with osteoblast-like phenotype based on promoter activity, mRNA and protein expression, and von Kossa staining. Likewise, Nrf2 overexpression significantly decreased the formation of calcium deposit similar to the level of osteogenic staining in VSMCs, and DMF with Nrf2 knockdown failed to attenuate hyperphosphatemia induced vascular calcification. Furthermore, DMF significantly attenuated the calcification of ex vivo ring culture from both rat common carotid artery and mouse thoracic aorta as well as in vivo mouse model of Vitamin D3-induced calcification consistent with the increased Nrf2 protein levels in early stage of calcification by DMF. In conclusion, our data support that DMF stimulates Nrf2 activity to attenuate hyperphosphatamia in vitro or Vitamin D3-induced in vivo vascular calcification, which would be a beneficial effect on vascular diseases induced by oxidative stress such as vascular calcification.
Assuntos
Fumaratos/farmacologia , Músculo Liso Vascular/efeitos dos fármacos , Fator 2 Relacionado a NF-E2/metabolismo , Calcificação Vascular/tratamento farmacológico , Animais , Aorta Torácica/efeitos dos fármacos , Aorta Torácica/patologia , Cálcio/metabolismo , Artéria Carótida Primitiva/efeitos dos fármacos , Artéria Carótida Primitiva/patologia , Fumarato de Dimetilo , Relação Dose-Resposta a Droga , Fumaratos/administração & dosagem , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Músculo Liso Vascular/citologia , Miócitos de Músculo Liso/efeitos dos fármacos , Miócitos de Músculo Liso/metabolismo , Estresse Oxidativo/efeitos dos fármacos , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Regulação para Cima/efeitos dos fármacos , Calcificação Vascular/patologiaRESUMO
Excessive proliferation of vascular smooth muscle cells (VSMCs) and incomplete re-endothelialization is a major clinical problem limiting the long-term efficacy of percutaneous coronary angioplasty. We tested if dimethylfumarate (DMF), an anti-psoriasis drug, could inhibit abnormal vascular remodeling via NF-E2-related factor 2 (Nrf2)-NAD(P)H quinone oxidoreductase 1 (NQO1) activity. DMF significantly attenuated neointimal hyperplasia induced by balloon injury in rat carotid arteries via suppression of the G1 to S phase transition resulting from induction of p21 protein in VSMCs. Initially, DMF increased p21 protein stability through an enhancement in Nrf2 activity without an increase in p21 mRNA. Later on, DMF stimulated p21 mRNA expression through a process dependent on p53 activity. However, heme oxygenase-1 (HO-1) or NQO1 activity, well-known target genes induced by Nrf2, were dispensable for the DMF induction of p21 protein and the effect on the VSMC proliferation. Likewise, DMF protected endothelial cells from TNF-α-induced apoptosis and the dysfunction characterized by decreased eNOS expression. With knock-down of Nrf2 or NQO1, DMF failed to prevent TNF-α-induced cell apoptosis and decreased eNOS expression. Also, CD31 expression, an endothelial specific marker, was restored in vivo by DMF. In conclusion, DMF prevented abnormal proliferation in VSMCs by G1 cell cycle arrest via p21 upregulation driven by Nrf2 and p53 activity, and had a beneficial effect on TNF-α-induced apoptosis and dysfunction in endothelial cells through Nrf2-NQO1 activity suggesting that DMF might be a therapeutic drug for patients with vascular disease.