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1.
Ann Dermatol ; 35(4): 275-284, 2023 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-37550228

RESUMO

BACKGROUND: Janus kinase (Jak) 3 has recently been shown as a beneficial target for the treatment of chronic inflammatory diseases, such as psoriasis and alopecia areata. The role of Jak3 in tissue repair and remodeling is emerging. OBJECTIVE: This study aimed to investigate the role of Jak3 signaling in the remodeling of the sebaceous gland (SG) during skin wound repair, and the development of in vitro SGs. METHODS: Mouse skin tissue (ICR mouse) was obtained from the recovered skin eight days after a 4 mm biopsy punch wound. To observe the role of Jak3, the selective inhibitors WHI-p131 and PF06651600 was administered. Formation of in vitro SG was examined using primary sebocyte cultures obtained postnatally from 3-day-old mice. RESULTS: The data showed that SGs showed highly positive signals with anti-isolectin B4, which also used for detection of angiogenetic vessels and the basal epidermis. Isolectin B4 could be a good indicator of SGs. The Jak3 inhibitors significantly reduced the area and volume of SG remodeling with reduced expression of p-Jak3. In addition, the area of cultured intact SG in vitro was significantly decreased in a concentration-dependent manner by Jak3 inhibition. CONCLUSION: These data showed that Jak3 signaling is a potent regulator to develop SGs. Jak3 inhibition did not decrease the number of sebocytes in SGs but decreased the area and volume of SG remodeling. Therefore, Jak3 inhibition may be a potential target for the treatment of SG hyperplasia and associated skin diseases.

2.
Part Fibre Toxicol ; 18(1): 42, 2021 11 25.
Artigo em Inglês | MEDLINE | ID: mdl-34819099

RESUMO

BACKGROUND: Nanoparticles have been utilized in brain research and therapeutics, including imaging, diagnosis, and drug delivery, owing to their versatile properties compared to bulk materials. However, exposure to nanoparticles leads to their accumulation in the brain, but drug development to counteract this nanotoxicity remains challenging. To date, concerns have risen about the potential toxicity to the brain associated with nanoparticles exposure via penetration of the brain blood barrier to address this issue. METHODS: Here the effect of silica-coated-magnetic nanoparticles containing the rhodamine B isothiocyanate dye [MNPs@SiO2(RITC)] were assessed on microglia through toxicological investigation, including biological analysis and integration of transcriptomics, proteomics, and metabolomics. MNPs@SiO2(RITC)-induced biological changes, such as morphology, generation of reactive oxygen species, intracellular accumulation of MNPs@SiO2(RITC) using transmission electron microscopy, and glucose uptake efficiency, were analyzed in BV2 murine microglial cells. Each omics data was collected via RNA-sequencing-based transcriptome analysis, liquid chromatography-tandem mass spectrometry-based proteome analysis, and gas chromatography- tandem mass spectrometry-based metabolome analysis. The three omics datasets were integrated and generated as a single network using a machine learning algorithm. Nineteen compounds were screened and predicted their effects on nanotoxicity within the triple-omics network. RESULTS: Intracellular reactive oxygen species production, an inflammatory response, and morphological activation of cells were greater, but glucose uptake was lower in MNPs@SiO2(RITC)-treated BV2 microglia and primary rat microglia in a dose-dependent manner. Expression of 121 genes (from 41,214 identified genes), and levels of 45 proteins (from 5918 identified proteins) and 17 metabolites (from 47 identified metabolites) related to the above phenomena changed in MNPs@SiO2(RITC)-treated microglia. A combination of glutathione and citrate attenuated nanotoxicity induced by MNPs@SiO2(RITC) and ten other nanoparticles in vitro and in the murine brain, protecting mostly the hippocampus and thalamus. CONCLUSIONS: Combination of glutathione and citrate can be one of the candidates for nanotoxicity alleviating drug against MNPs@SiO2(RITC) induced detrimental effect, including elevation of intracellular reactive oxygen species level, activation of microglia, and reduction in glucose uptake efficiency. In addition, our findings indicate that an integrated triple omics approach provides useful and sensitive toxicological assessment for nanoparticles and screening of drug for nanotoxicity.


Assuntos
Nanopartículas , Dióxido de Silício , Animais , Citratos , Ácido Cítrico , Glutationa , Fenômenos Magnéticos , Camundongos , Microglia , Nanopartículas/toxicidade , Ratos , Dióxido de Silício/toxicidade
3.
Neurochem Res ; 44(1): 147-153, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-29357018

RESUMO

Regulation of glutamate metabolism via glutamate dehydrogenase (GDH) might be the promising therapeutic approach for treating neurodegenerative disorders. In the central nervous system, glutamate functions both as a major excitatory neurotransmitter and as a key intermediate metabolite for neurons. GDH converts glutamate to α-ketoglutarate, which serves as a TCA cycle intermediate. Dysregulated GDH activity in the central nervous system is highly correlated with neurological disorders. Indeed, studies conducted with mutant mice and allosteric drugs have shown that deficient or overexpressed GDH activity in the brain can regulate whole body energy metabolism and affect early onset of Parkinson's disease, Alzheimer's disease, temporal lobe epilepsy, and spinocerebellar atrophy. Moreover, in strokes with excitotoxicity as the main pathophysiology, mice that overexpressed GDH exhibited smaller ischemic lesion than mice with normal GDH expression. In additions, GDH activators improve lesions in vivo by increasing α-ketoglutarate levels. In neurons exposed to an insult in vitro, enhanced GDH activity increases ATP levels. Thus, in an energy crisis, neuronal mitochondrial activity is improved and excitotoxic risk is reduced. Consequently, modulating GDH activity in energy-depleted conditions could be a sound strategy for maintaining the mitochondrial factory in neurons, and thus, protect against metabolic failure.


Assuntos
Sistemas de Liberação de Medicamentos/tendências , Glutamato Desidrogenase/metabolismo , Ácido Glutâmico/metabolismo , Doenças Neurodegenerativas/metabolismo , Fármacos Neuroprotetores/administração & dosagem , Trifosfato de Adenosina/metabolismo , Animais , Metabolismo Energético/efeitos dos fármacos , Metabolismo Energético/fisiologia , Indução Enzimática/efeitos dos fármacos , Indução Enzimática/fisiologia , Humanos , Doenças Neurodegenerativas/tratamento farmacológico , Neuroproteção/efeitos dos fármacos , Neuroproteção/fisiologia
4.
Neuroreport ; 28(14): 929-935, 2017 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-28817455

RESUMO

During spinal cord development, endogenous progenitors expressing nestin can migrate into the target and differentiate into neurons and other glial cells. Microglial cells can also be derived from nestin progenitor cells, even in the adult brain. Knockdown of Jak kinase 3 (Jak3) signaling can increase neurogenesis with longer neurite outgrowth in cortical progenitor cells. This study investigated the effect of Jak3 signaling on differentiation from nestin progenitor cells using E13.5 spinal progenitor cell cultures. In growth factors-enriched culture, developing neurons could not survive after several days and also a significant proportion of nestin-expressing cells transformed into ameboid Iba1 microglial cells, which increased exponentially after 5 days. This microgliogenesis was predominantly regulated by Jak3 signaling, which was confirmed by transcription factors responsible for microgliogenesis, and microglial migration and phagocytosis, such as Pu.1, Irf8, and Runx1. Jak3 inhibition also significantly increased the Tuj1 growing neurites with little microglial activation. These results indicated that neuronal and microglial cell differentiation was regulated primarily by Jak3 signaling and the developing neurons and neurite outgrowth might also be regulated by Jak3-dependent microglial activity.


Assuntos
Diferenciação Celular/fisiologia , Janus Quinase 3/metabolismo , Microglia/metabolismo , Nestina/metabolismo , Células-Tronco Neurais/metabolismo , Animais , Proteínas de Ligação ao Cálcio/metabolismo , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Peptídeos e Proteínas de Sinalização Intercelular/administração & dosagem , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Janus Quinase 3/antagonistas & inibidores , Janus Quinase 3/genética , Camundongos Endogâmicos ICR , Proteínas dos Microfilamentos/metabolismo , Microglia/efeitos dos fármacos , Células-Tronco Neurais/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Medula Espinal/efeitos dos fármacos , Medula Espinal/embriologia , Medula Espinal/metabolismo , Traumatismos da Medula Espinal/metabolismo
5.
Neuroscience ; 340: 487-500, 2017 01 06.
Artigo em Inglês | MEDLINE | ID: mdl-27845178

RESUMO

Deregulation of glutamate homeostasis is associated with degenerative neurological disorders. Glutamate dehydrogenase (GDH) is important for glutamate metabolism and plays a central role in expanding the pool of tricarboxylic acid (TCA) cycle intermediate alpha-ketoglutarate (α-KG), which improves overall bioenergetics. Under high energy demand, maintenance of ATP production results in functionally active mitochondria. Here, we tested whether the modulation of GDH activity can rescue ischemia/reperfusion-induced neuronal death in an in vivo mouse model of middle artery occlusion and an in vitro oxygen/glucose depletion model. Iodoacetate, an inhibitor of glycolysis, was also used in a model of energy failure, remarkably depleting ATP and α-KG. To stimulate GDH activity, the GDH activator 2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid and potential activator beta-lapachone were used. The GDH activators restored α-KG and ATP levels in the injury models and provided potent neuroprotection. We also found that beta-lapachone increased glutamate utilization, accompanied by a reduction in extracellular glutamate. Thus, our hypothesis that mitochondrial GDH activators increase α-KG production as an alternative energy source for use in the TCA cycle under energy-depleted conditions was confirmed. Our results suggest that increasing GDH-mediated glutamate oxidation represents a new therapeutic intervention for neurodegenerative disorders, including stoke.


Assuntos
Isquemia Encefálica/tratamento farmacológico , Encéfalo/efeitos dos fármacos , Glutamato Desidrogenase/metabolismo , Naftoquinonas/farmacologia , Fármacos Neuroprotetores/farmacologia , Traumatismo por Reperfusão/tratamento farmacológico , Animais , Astrócitos/efeitos dos fármacos , Astrócitos/enzimologia , Astrócitos/patologia , Encéfalo/enzimologia , Encéfalo/patologia , Isquemia Encefálica/enzimologia , Isquemia Encefálica/patologia , Morte Celular/efeitos dos fármacos , Morte Celular/fisiologia , Células Cultivadas , Técnicas de Cocultura , Modelos Animais de Doenças , Infarto da Artéria Cerebral Média , Masculino , Camundongos Endogâmicos ICR , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/enzimologia , Neurônios/efeitos dos fármacos , Neurônios/enzimologia , Neurônios/patologia , Distribuição Aleatória , Traumatismo por Reperfusão/enzimologia , Traumatismo por Reperfusão/patologia
6.
Phys Ther ; 96(6): 808-17, 2016 06.
Artigo em Inglês | MEDLINE | ID: mdl-26586863

RESUMO

BACKGROUND: Low-intensity ultrasound (LIUS) can reduce pain and improve function in arthritic joints. Neutrophils are first-line actors in host defense that recruit macrophages. Dead neutrophils are removed during resolution of inflammation. Delayed neutrophil clearance can lead to extended inflammation or even chronic autoimmune disease. Although neutrophil extracellular traps (NETs) in arthritic tissue are involved in the pathogenesis of arthritis, their functional role has not been clarified. OBJECTIVES: This study aimed to investigate the effect of LIUS on synovial inflammation and its resolution via neutrophil clearance. METHODS: Synovitis was induced by intra-articular injection of complete Freund's adjuvant (CFA) into the left knee joint of 58 adult male Sprague-Dawley rats. Low-intensity ultrasound (1 MHz, 200 mW/cm(2)) was applied for 10 minutes daily. Neutrophil clearance was assessed with the expression of myeloperoxidase (MPO). In addition, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining and NET formation in the synovium were observed. In neutrophil and macrophage cultures from peripheral blood, the effect of NET clearance by LIUS was investigated. RESULTS: In CFA-induced synovitis, MPO-positive neutrophils peaked after 2 to 3 days, filling the inflammatory core. Monocytes and macrophages in the periphery later infiltrated the core and were reduced thereafter. Low-intensity ultrasound reduced synovial hyperplasia and induced earlier MPO clearance. Neutrophils in the core of the inflamed synovium exhibited NET formation, which LIUS increased. Low-intensity also induced NETs in peripheral polymorphonuclear cells in an intensity-dependent manner and potentiated phorbol myristate acetate (PMA)-induced NETosis. The PMA-induced NETs were cleared by macrophages; clearance was enhanced by LIUS. LIMITATIONS: The effect of LIUS on CFA-induced inflammation was observed only during the acute phase. Although the effect of LIUS on NETosis in the in vitro neutrophil culture system was clear, the in vivo NETosis cannot be quantified. CONCLUSIONS: Neutrophil extracellular traps act in inflammatory synovitis, and LIUS enhanced the NETs and resulted in neutrophil clearance by enhancing the phagocytosis of macrophages, which might be a factor underlying the therapeutic effect of LIUS in arthritic synovium.


Assuntos
Armadilhas Extracelulares , Neutrófilos , Membrana Sinovial/patologia , Sinovite/terapia , Terapia por Ultrassom/métodos , Animais , Apoptose/efeitos da radiação , Células Cultivadas , Armadilhas Extracelulares/efeitos da radiação , Hiperplasia/terapia , Macrófagos/fisiologia , Masculino , Neutrófilos/enzimologia , Peroxidase/metabolismo , Fagocitose/efeitos da radiação , Ratos Sprague-Dawley , Sinovite/induzido quimicamente , Ondas Ultrassônicas
7.
Int Immunopharmacol ; 26(1): 203-11, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25843256

RESUMO

Fructose-1,6-bisphosphate (FBP) is a glycolytic intermediate with salutary effects in various brain injury models, but its neuroprotective mechanism is incompletely understood. In this study, we examined the effects of FBP on the expression of adhesion molecules in cerebrovascular endothelial cells and explored the possible mechanisms therein involved. FBP significantly down-regulated lipopolysaccharide (LPS)-induced expression of adhesion molecules and leukocyte adhesion to brain endothelial cells and inhibited NF-κB activity, which is implicated in the expression of adhesion molecules. FBP abrogated ICAM-1 expression and NF-κB activation induced by macrophage-activating lipopeptide 2-kDa (MALP-2) or overexpression of MyD88 or TRAF6. FBP suppressed TRAF6-induced phosphorylation of TAK1, IKKß and IκBα, but fail to affect NF-κB activity induced by ectopic expression of IKKß. In addition, LPS-induced IRAK-1 phosphorylation was inhibited by FBP, suggesting the presence of multiple molecular targets of FBP in MyD88-dependent signaling pathway. FBP significantly attenuated ICAM-1 expression and NF-κB activity induced by poly[I:C] or overexpression of TRIF or TBK1. FBP significantly repressed the expression of interferon-ß (IFN-ß) and the activation of IFN regulatory factor 3 (IRF3) induced by LPS, poly[I:C] or overexpression of TRIF or TBK1, but fail to affect IRF3 activity induced by ectopic expression of constitutively active IRF3. Overall, our results demonstrate that FBP modulates both MyD88- and TRIF-dependent signaling pathways of TLR4 and subsequent inflammatory responses in brain endothelial cells, providing insight into its neuroprotective mechanism in brain injury associated with inflammation.


Assuntos
Encéfalo/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Frutosedifosfatos/farmacologia , Molécula 1 de Adesão Intercelular/biossíntese , Fármacos Neuroprotetores/farmacologia , Receptor 4 Toll-Like/metabolismo , Animais , Encéfalo/irrigação sanguínea , Encéfalo/imunologia , Adesão Celular/efeitos dos fármacos , Adesão Celular/imunologia , Linhagem Celular , Células Endoteliais/imunologia , Células Endoteliais/metabolismo , Ensaio de Imunoadsorção Enzimática , Frutosedifosfatos/administração & dosagem , Humanos , Lipopolissacarídeos/farmacologia , Camundongos , Fármacos Neuroprotetores/administração & dosagem , Transdução de Sinais/efeitos dos fármacos , Células U937
8.
Eur J Pharmacol ; 754: 11-8, 2015 May 05.
Artigo em Inglês | MEDLINE | ID: mdl-25704611

RESUMO

Inflammation has been implicated in the pathogenesis of various cerebral diseases. Thus, control of brain inflammation is regarded as one of the important therapeutic strategies for the treatment of neurodegenerative diseases such as Alzheimer׳s disease and stroke. Isobavachalcone, a flavonoid from Psoralea corylifolia, is known to possess a wide spectrum of biological activities and is expected to be useful in preventing or treating neurodegenerative diseases. However, very little is known regarding its effects on cerebral inflammation. In this study, we examined the effect of isobavachalcone on leukocyte adhesion and intercellular adhesion molecule-1 (ICAM-1) expression in brain endothelial cells activated with lipopolysaccharide (LPS) and explored the possible mechanisms involved. Isobavachalcone significantly down-regulated LPS-induced ICAM-1 expression and leukocyte-endothelial cell adhesion and suppressed NF-κB activity which is implicated in the expression of ICAM-1. It attenuated ICAM-1 expression as well as NF-κB transcriptional activity induced by macrophage-activating lipopeptide 2-kDa (MALP-2) or polyriboinosinic polyribocytidylic acid (poly[I:C]). Isobavachalcone also down-regulated LPS or poly[I:C]-induced expression of IFN-ß, which can indirectly activate NF-κB. These data imply that isobavachalcone can modulate both MyD88-dependent and TRIF-dependent signaling of toll-like receptor 4 (TLR4). Taken together, our data suggest that isobavachalcone inhibits LPS-induced ICAM-1 expression and leukocyte adhesion to brain endothelial cell by blocking TLR4 signaling and thus, has the potential to ameliorate neuronal injury in brain diseases associated with inflammation.


Assuntos
Encéfalo/citologia , Chalconas/farmacologia , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Molécula 1 de Adesão Intercelular/metabolismo , Transdução de Sinais/efeitos dos fármacos , Receptor 4 Toll-Like/antagonistas & inibidores , Animais , Encéfalo/efeitos dos fármacos , Adesão Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Regulação para Baixo/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Interleucina-6/metabolismo , Lipopeptídeos/antagonistas & inibidores , Lipopeptídeos/farmacologia , Lipopolissacarídeos/farmacologia , Camundongos , NF-kappa B/metabolismo , Poli I-C/antagonistas & inibidores , Poli I-C/farmacologia
9.
J Ultrasound Med ; 33(6): 949-57, 2014 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-24866602

RESUMO

OBJECTIVES: To determine whether low-intensity ultrasound (US) can reduce red blood cell (RBC) edema and, if so, whether the US activity is associated with aquaporin 1 (AQP-1), a water channel in the cell membrane. METHODS: Red blood cell edema was induced by gramicidin D treatment at 40 ng/mL for 20 minutes and evaluated by a hematocrit assay. Low-intensity continuous wave US at 1 MHz was applied to RBCs for the last 10 minutes of gramicidin D treatment. To determine whether US activity was associated with AQP-1, RBCs were treated with 40 µM mercuric chloride (HgCl(2)), an AQP-1 inhibitor, for 20 minutes at the time of gramicidin D treatment. Posttreatment morphologic changes in RBCs were observed by actin staining with phalloidin. RESULTS: Red blood cell edema increased significantly with gramicidin D at 20 (1.8%), 40 (6.7%), 60 (16.7%), and 80 (11.3%) ng/mL, reaching a peak at 60 ng/mL, compared to the control group (20 ng/mL, P = .019; 40, 60, and 80 ng/mL, P < .001). No significant RBC hemolysis was observed in any group. Edema induced by gramicidin D at 40 ng/mL was significantly reduced by US at 30 (3.4%; P = .003), 70 (4.4%; P = .001), and 100 (2.9%; P = .001) mW/cm(2). Subsequent experiments showed that edema reduction by US ranged from 7% to 10%. Cotreatment with HgCl(2) partially reversed the US effect and showed a significantly different level of edema compared to gramicidin D-alone and US-cotreated groups (P = .001). These results were confirmed by microscopic observation of RBC morphologic changes. CONCLUSIONS: Low-intensity US could reduce gramicidin D-induced RBC edema, and its effect appeared to at least partly involve regulation of AQP-1 activity. These results suggest that low-intensity US can be used as an alternative treatment to control edema and related disorders.


Assuntos
Aquaporina 1/metabolismo , Água Corporal/metabolismo , Tamanho Celular/efeitos da radiação , Eritrócitos/citologia , Eritrócitos/efeitos da radiação , Gramicidina/farmacologia , Terapia por Ultrassom/métodos , Animais , Aquaporina 1/efeitos da radiação , Tamanho Celular/efeitos dos fármacos , Células Cultivadas , Eritrócitos/efeitos dos fármacos , Eritrócitos/fisiologia , Ondas de Choque de Alta Energia , Ativação do Canal Iônico/efeitos dos fármacos , Ativação do Canal Iônico/efeitos da radiação , Masculino , Osmorregulação/efeitos dos fármacos , Osmorregulação/efeitos da radiação , Doses de Radiação , Ratos , Ratos Sprague-Dawley
10.
Exp Neurol ; 249: 95-103, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-24005111

RESUMO

The immature brain is prone to seizure; however, the mechanism underlying this vulnerability has not been clarified. Febrile seizure is common in young children, and the use of non-steroidal anti-inflammatory drugs for febrile seizure is not recommended. In previous studies, we established that prostaglandin (PG) F2α, a product of cyclooxygenase (COX), acts as an endogenous anticonvulsant in the adult mouse. Therefore, we assumed that COX-2 activity was involved with seizure susceptibility in early life. In the present study, immature mice (postnatal day 9) were far more prone to kainic acid (KA)-induced seizures than mature mice (after postnatal day 35). Seizure activity began later in immature mice, but was more severe and was unaffected by a potent COX inhibitor, indomethacin; in contrast, indomethacin aggravated seizure activity in mature mice. Immature mouse brains exhibited little basal COX-2 expression and little KA-induced COX-2 induction, while KA-induced COX-2 expression and PGF2α release were prominent in mature brains. During brain development, COX expression was increased and glycosylated in an age-dependent manner, which was necessary for COX enzyme activity. Intracisternal PGF2α administration also reduced KA-induced seizure activity and mortality. Taken together, low COX activity and the resulting deficiency of PGF2α may be an essential cause of increased seizure susceptibility in the immature brain.


Assuntos
Encéfalo/crescimento & desenvolvimento , Encéfalo/metabolismo , Ciclo-Oxigenase 2/biossíntese , Dinoprosta/biossíntese , Convulsões/metabolismo , Fatores Etários , Animais , Animais Recém-Nascidos , Encéfalo/efeitos dos fármacos , Células Cultivadas , Dinoprosta/administração & dosagem , Suscetibilidade a Doenças , Glicosilação , Camundongos , Regulação para Cima/fisiologia
11.
JAKSTAT ; 2(2): e23282, 2013 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-24058805

RESUMO

Myogenic differentiation plays an important role in muscle regeneration and is regulated by two transcription factor families, MRFs and MEF2, which induce differentiation of myoblasts through expression of the muscle-specific gene, myogenin. In addition, many intracellular signaling pathways are also involved in myogenic differentiation, including p38 MAPK, ERK/MAPK and PI3K/AKT. The JAK-STAT pathway is activated by various cytokines and positively or negatively regulates the differentiation of myoblasts. JAK1 plays a notable role in proliferation; whereas, JAK2 and JAK3 function mainly in differentiation. The STATs, molecules downstream of JAK, regulate myogenesis. With JAK1, STAT1 promotes proliferation, while STAT3 has a dual effect on proliferation and differentiation. The JAK-STAT negative regulator, SOCS, is also associated with myogenesis; although, its role is controversial. In this review, we will discuss the role of the JAK-STAT pathway on myogenic differentiation.

12.
Arch Pharm Res ; 36(9): 1149-59, 2013 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-23604722

RESUMO

Fructose-1,6-bisphosphate (FBP), a glycolytic intermediate, has neuroprotective effects in various brain injury models. However, its effects on blood-brain barrier (BBB) are largely unknown. In this study, we investigated the effects of FBP on lipopolysaccharide (LPS)-induced BBB dysfunction in in vitro BBB model comprising co-culture of mouse brain endothelial cell line, bEnd.3 and mouse primary astrocyte and explored its action mechanism therein involved. LPS induced the impairment of endothelial permeability and transendothelial electrical resistance (TEER). The functional changes were confirmed by alterations in immunostaining for junctional proteins occludin, ZO-1 and VE-cadherin, such as the loss of cortical staining pattern and appearance of intercellular gaps in endothelial cells. Co-administration of FBP alleviated the deleterious effects of LPS on BBB permeability and TEER in a dose dependent manner. And also FBP inhibited the LPS-induced changes in the distribution of endothelial junctional proteins, resulting in the better preservation of monolayer integrity. FBP suppressed the production of reactive oxygen species (ROS) but did not affect cyclooxygenase-2 expression and prostaglandin E2 production in endothelial cells stimulated with LPS. Taken together, these data suggest that FBP could ameliorate LPS-induced BBB dysfunction through the maintenance of junctional integrity, which might be mediated by downregulation of ROS production.


Assuntos
Astrócitos/efeitos dos fármacos , Barreira Hematoencefálica/efeitos dos fármacos , Córtex Cerebral/efeitos dos fármacos , Endotélio Vascular/efeitos dos fármacos , Frutosedifosfatos/farmacologia , Fármacos Neuroprotetores/farmacologia , Animais , Animais Recém-Nascidos , Astrócitos/citologia , Astrócitos/metabolismo , Barreira Hematoencefálica/citologia , Barreira Hematoencefálica/metabolismo , Linhagem Celular , Permeabilidade da Membrana Celular/efeitos dos fármacos , Células Cultivadas , Córtex Cerebral/irrigação sanguínea , Córtex Cerebral/citologia , Córtex Cerebral/metabolismo , Técnicas de Cocultura , Endotélio Vascular/citologia , Endotélio Vascular/metabolismo , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Lipopolissacarídeos/antagonistas & inibidores , Lipopolissacarídeos/toxicidade , Camundongos , Camundongos Endogâmicos ICR , Ratos , Espécies Reativas de Oxigênio/antagonistas & inibidores , Espécies Reativas de Oxigênio/metabolismo , Proteínas de Junções Íntimas/metabolismo , Junções Íntimas/efeitos dos fármacos , Junções Íntimas/metabolismo
13.
Carcinogenesis ; 34(7): 1543-50, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23508637

RESUMO

Genomic analyses have revealed the enormous heterogeneity in essentially all cancer types. However, the identification of precise subtypes, which are biologically informative and clinically useful, remains a challenge. The application of integrative analysis of multilayered genomic profiles to define the chromosomal regions of genomic copy number alterations with concomitant transcriptional deregulation is posited to provide a promising strategy to identify driver targets. In this study, we performed an integrative analysis of the DNA copy numbers and gene expression profiles of hepatocellular carcinoma (HCC). By comparing DNA copy numbers between HCC subtypes based on gene expression pattern, we revealed the DNA copy number alteration with concordant gene expression changes at 6p21-p24 particularly in the HCC subtype of aggressive phenotype without expressing stemness genes. Among the genes at 6p21-p24, we identified IER3 as a potential driver. The clinical utility of IER3 copy numbers was demonstrated by validating its clinical correlation with independent cohorts. In addition, short hairpin RNA-mediated knock-down experiment revealed the functional relevance of IER3 in liver cancer progression. In conclusion, our results suggest that genomic copy number alterations with transcriptional deregulation at 6p21-p24 identify an aggressive HCC phenotype and a novel functional biomarker.


Assuntos
Carcinoma Hepatocelular/genética , Cromossomos Humanos Par 6/genética , Variações do Número de Cópias de DNA , Regulação Neoplásica da Expressão Gênica , Transcrição Gênica , Proteínas Reguladoras de Apoptose/genética , Proteínas Reguladoras de Apoptose/metabolismo , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Carcinoma Hepatocelular/patologia , Proliferação de Células , Hibridização Genômica Comparativa , Progressão da Doença , Células Hep G2 , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Fenótipo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transcriptoma
14.
Br J Pharmacol ; 169(3): 604-18, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23317035

RESUMO

BACKGROUND AND PURPOSE: Prostaglandin E2 (PGE2) has been implicated in the regulation of adhesion molecules, leukocyte adhesion and infiltration into inflamed site. However, the underlying mechanism therein involved remains ill-defined. In this study, we explored its cellular mechanism of action in the regulation of the intercellular adhesion molecule-1 (ICAM-1) expression in the brain endothelial cells. EXPERIMENTAL APPROACH: bEnd.3 cells, the murine cerebrovascular endothelial cell line and primary mouse brain endothelial cells were treated with PGE2 with or without agonists/antagonists of PGE2 receptors and associated signalling molecules. ICAM-1 expression, Akt phosphorylation and activity of NF-κB were determined by reverse transcription polymerase chain reaction (RT-PCR), immunoblot analysis, luciferase assay and immunocytochemistry. KEY RESULTS: PGE2 significantly up-regulated the expression of ICAM-1, which was blocked by EP4 antagonist (ONO-AE2-227) and knock-down of EP4. PGE2 effects were mimicked by forskolin, dibutyryl cAMP (dbcAMP) and an exchange protein directly activated by cAMP (Epac) activator (8-Cpt-cAMP) but not a protein kinase A activator (N6-Bnz-cAMP). PGE2-induced ICAM-1 expression was reduced by knock-down of Epac1. A PI3K specific inhibitor (LY294002), Akt inhibitor VIII (Akti) and NF-κB inhibitors (Bay-11-7082 and MG-132) attenuated the induction of ICAM-1 by PGE2. PGE2, dbcAMP and 8-Cpt-cAMP induced the phosphorylation of Akt, IκB kinase and IκBα and the translocation of p65 to the nucleus and increased NF-κB dependent reporter gene activity, which was diminished by Akti. CONCLUSION AND IMPLICATIONS: Our findings suggest that PGE2 induces ICAM-1 expression via EP4 receptor and Epac/Akt/NF-κB signalling pathway in bEnd.3 brain endothelial cells, supporting its pathophysiological role in brain inflammation.


Assuntos
Encéfalo/irrigação sanguínea , Dinoprostona/metabolismo , Endotélio Vascular/metabolismo , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Molécula 1 de Adesão Intercelular/metabolismo , Receptores de Prostaglandina E Subtipo EP4/agonistas , Sistemas do Segundo Mensageiro , Animais , Encéfalo/citologia , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Adesão Celular/efeitos dos fármacos , Linhagem Celular , Células Cultivadas , AMP Cíclico/análogos & derivados , AMP Cíclico/farmacologia , AMP Cíclico/fisiologia , Endotélio Vascular/citologia , Endotélio Vascular/efeitos dos fármacos , Endotélio Vascular/imunologia , Inibidores Enzimáticos/farmacologia , Inativação Gênica , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Monócitos/citologia , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Monócitos/metabolismo , Receptores de Prostaglandina E Subtipo EP4/antagonistas & inibidores , Receptores de Prostaglandina E Subtipo EP4/genética , Receptores de Prostaglandina E Subtipo EP4/metabolismo , Sistemas do Segundo Mensageiro/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacos , Vasodilatadores/farmacologia
15.
Cell Signal ; 24(3): 742-9, 2012 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-22120524

RESUMO

Skeletal muscle differentiation is regulated by transcription factors, including members of the myogenic regulatory factor (MRF) family and many signaling pathways. The JAK1 and JAK2 pathways are known to each have different effects on myoblast proliferation and differentiation; however, the role of JAK3 in myoblast differentiation remains unclear. In this study, we investigated the effect of JAK3 inhibition on myogenic differentiation in the C2C12 mouse myoblast cell line. During myogenic differentiation, treatment with the JAK3 inhibitor WHIp154 significantly increased the number of MHC-positive multinucleated myotubes and the expressions of myosin heavy chain (MHC), myogenin (MGN), MyoD, and myogenic enhancer factor 2 (MEF2). Knockdown of the JAK3 gene using siJAK3 also significantly increased MHC, MGN and MyoD mRNA expressions as well as insulin-like growth factor-II (IGF-II) gene expression. During differentiation, JAK3 was initially activated and later decreased. Differentiation decreased STAT1, which was further decreased by WHIp154. In contrast, STAT3 gradually was elevated during differentiation, and was increased by JAK3 inhibition. Moreover, we found that up-regulation of AKT activity and down-regulation of ERK activity cooperated to accelerate myogenic differentiation. Taken together, these data indicate that JAK3 inhibition potently facilitates myoblast differentiation through antagonistic STAT1/STAT3 activities. Additionally, JAK3 inhibition induced precocious differentiation and played important roles for terminal differentiation, including fusion, which is involved with regulation of AKT and ERK pathways.


Assuntos
Diferenciação Celular , Janus Quinase 3/metabolismo , Mioblastos/citologia , Mioblastos/enzimologia , Animais , Linhagem Celular , Fator de Crescimento Insulin-Like II/genética , Fator de Crescimento Insulin-Like II/metabolismo , Janus Quinase 3/antagonistas & inibidores , Sistema de Sinalização das MAP Quinases , Camundongos , Proteína MyoD/genética , Proteína MyoD/metabolismo , Fatores de Regulação Miogênica/genética , Fatores de Regulação Miogênica/metabolismo , Miogenina/genética , Miogenina/metabolismo , Cadeias Pesadas de Miosina/genética , Cadeias Pesadas de Miosina/metabolismo , Quinazolinas/química , Quinazolinas/farmacologia , Interferência de RNA , RNA Interferente Pequeno/metabolismo , Fator de Transcrição STAT1/metabolismo , Fator de Transcrição STAT3/metabolismo
16.
Stem Cells ; 28(10): 1816-28, 2010 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-20979137

RESUMO

Neuronal precursor cells (NPCs) are temporally regulated and have the ability to proliferate and differentiate into mature neurons, oligodendrocytes, and astrocytes in the presence of growth factors (GFs). In the present study, the role of the Jak pathway in brain development was investigated in NPCs derived from neurosphere cultures using Jak2 and Jak3 small interfering RNAs and specific inhibitors. Jak2 inhibition profoundly decreased NPC proliferation, preventing further differentiation into neurons and glial cells. However, Jak3 inhibition induced neuronal differentiation accompanied by neurite growth. This phenomenon was due to the Jak3 inhibition-mediated induction of neurogenin (Ngn)2 and NeuroD in NPCs. Jak3 inhibition induced NPCs to differentiate into scattered neurons and increased the expression of Tuj1, microtubule associated protein 2 (MAP2), Olig2, and neuroglial protein (NG)2, but decreased glial fibrillary acidic protein (GFAP) expression, with predominant neurogenesis/polydendrogenesis compared with astrogliogenesis. Therefore, Jak2 may be important for NPC proliferation and maintenance, whereas knocking-down of Jak3 signaling is essential for NPC differentiation into neurons and oligodendrocytes but does not lead to astrocyte differentiation. These results suggest that NPC proliferation and differentiation are differentially regulated by the Jak pathway.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Janus Quinase 3/metabolismo , Neurônios/citologia , Neurônios/metabolismo , Células-Tronco/citologia , Células-Tronco/metabolismo , Animais , Western Blotting , Encéfalo/embriologia , Encéfalo/metabolismo , Diferenciação Celular/genética , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Imuno-Histoquímica , Janus Quinase 2/antagonistas & inibidores , Janus Quinase 2/genética , Janus Quinase 2/metabolismo , Janus Quinase 3/antagonistas & inibidores , Janus Quinase 3/genética , Camundongos , Neurogênese/efeitos dos fármacos , Neurogênese/genética , Neurônios/efeitos dos fármacos , Quinazolinas/farmacologia , RNA Interferente Pequeno/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células-Tronco/efeitos dos fármacos , Tirfostinas/farmacologia
17.
Cardiovasc Res ; 87(1): 119-26, 2010 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-20154065

RESUMO

AIMS: Because apoptotic death plays a critical role in cardiomyocyte loss during ischaemic heart injury, a detailed understanding of the mechanisms involved is likely to have a substantial impact on the optimization and development of treatment strategies. The goal of this study was to assess gene profiling during ischaemia/hypoxia and to evaluate the functions of ischaemia/hypoxia-responsive genes in in vivo and in vitro ischaemia/hypoxia-induced cardiomyocyte apoptosis models. METHODS AND RESULTS: DNA microarray analysis and real-time polymerase chain reaction were performed on hearts obtained from an in vivo rat transient ischaemia model and on neonatal rat cardiomyocytes from an in vitro hypoxia model. Three genes, namely Ddit4, Gadd45beta and Atf3, were found to be up-regulated in vivo and in vitro. Using loss-of-function and gain-of-function techniques, the functions of these ischaemia/hypoxia-responsive genes were evaluated. Ischaemia/hypoxia-induced cardiomyocyte apoptosis was remarkably attenuated by the small interfering RNA-mediated down-regulation of Gadd45beta in vivo and in vitro, whereas ectopic Gadd45beta expression significantly aggravated hypoxia-induced apoptosis in vitro. CONCLUSION: These results suggest that Gadd45beta is a key player in ischaemia/hypoxia-induced apoptotic cardiomyocyte death, and that strategies based on its inhibition might be of benefit in the treatment of acute ischaemic heart disease.


Assuntos
Antígenos de Diferenciação/genética , Apoptose , Isquemia Miocárdica/genética , Miócitos Cardíacos/metabolismo , Fator 3 Ativador da Transcrição/genética , Animais , Animais Recém-Nascidos , Antígenos de Diferenciação/metabolismo , Apoptose/genética , Hipóxia Celular , Linhagem Celular , Modelos Animais de Doenças , Perfilação da Expressão Gênica/métodos , Técnicas de Silenciamento de Genes , Hemodinâmica , Masculino , Isquemia Miocárdica/metabolismo , Isquemia Miocárdica/patologia , Isquemia Miocárdica/fisiopatologia , Miócitos Cardíacos/patologia , Análise de Sequência com Séries de Oligonucleotídeos , Interferência de RNA , Ratos , Ratos Sprague-Dawley , Proteínas Repressoras/genética , Fatores de Tempo , Fatores de Transcrição , Transfecção , Regulação para Cima
18.
Neurosci Lett ; 468(3): 254-8, 2010 Jan 14.
Artigo em Inglês | MEDLINE | ID: mdl-19900507

RESUMO

Blood-brain barrier (BBB) dysfunction contributes to the pathophysiology of cerebrovascular diseases such as stroke. In the present study, we investigated the role of PKC isoforms in aglycemic hypoxia-induced hyperpermeability using an in vitro model of the BBB consisting of mouse bEnd.3 cells. PKCbetaII and PKCdelta isoforms were activated during aglycemic hypoxia. CGP53353, a specific PKCbetaII inhibitor, significantly attenuated aglycemic hypoxia-induced BBB hyperpermeability and disruption of occludin and zonula occludens-1 (ZO-1), indicating a deleterious role of PKCbetaII in the regulation of BBB permeability during aglycemic hypoxia. Conversely, rottlerin, a specific PKCdelta inhibitor, exacerbated BBB hyperpermeability and tight junction (TJ) disruption during aglycemic hypoxia, indicating a protective role of PKCdelta against aglycemic hypoxia-induced BBB hyperpermeability. Furthermore, disruption of TJ proteins during aglycemic hypoxia was attenuated by PKCbetaII DN and PKCdelta WT overexpression, and aggravated by PKCbetaII WT and PKCdelta DN overexpression. These results suggest that PKCbetaII and PKCdelta counter-regulate BBB permeability during aglycemic hypoxia.


Assuntos
Barreira Hematoencefálica/metabolismo , Glucose/metabolismo , Proteína Quinase C-delta/fisiologia , Proteína Quinase C/fisiologia , Animais , Permeabilidade Capilar , Hipóxia Celular , Linhagem Celular , Isoenzimas/antagonistas & inibidores , Isoenzimas/fisiologia , Camundongos , Proteína Quinase C/antagonistas & inibidores , Proteína Quinase C beta , Proteína Quinase C-delta/antagonistas & inibidores , Ratos , Junções Íntimas/fisiologia
19.
J Pharmacol Sci ; 110(3): 405-9, 2009 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-19609071

RESUMO

In this study, we examined the effect of Cd on the expression of vascular cell adhesion molecule-1 (VCAM-1) and its mechanisms in bEnd.3 cells. The treatment with Cd increased protein and mRNA expressions of VCAM-1 and increased the phosphorylations of p38, JNK, and ERK. The Cd-induced VCAM-1 expression was significantly suppressed by either a specific p38 mitogen-activated protein kinase (MAPK) inhibitor (SB202190) or a JNK inhibitor (SP600125), but not by an ERK inhibitor (U0126). These results suggest that Cd induces the expression of VCAM-1, at least in part, via p38 and JNK pathways in bEnd.3 cells.


Assuntos
Cádmio/farmacologia , Endotélio Vascular/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Molécula 1 de Adesão de Célula Vascular/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Encéfalo/citologia , Humanos , Camundongos
20.
J Neurosci ; 29(18): 5974-84, 2009 May 06.
Artigo em Inglês | MEDLINE | ID: mdl-19420263

RESUMO

Neural-cadherin (N-cadherin), a member of the classical cadherin family of transmembrane glycoproteins, mediates cellular recognition and cell-cell adhesion through calcium-dependent homophilic interactions and plays important roles in the development and maintenance of the nervous system. Metalloproteinase is known to cleave N-cadherin, which is further cleaved by gamma-secretase. The intracellular domain of N-cadherin interacts with beta-catenin, and beta-catenin stability is critical for cell-cell adhesion and cell survival. In the present study, we showed that N-cadherin is cleaved specifically by calpain, resulting in the generation of a novel 110 kDa fragment. The cleavage occurred in ischemic brain lesions and in vitro neural cells in the presence of NMDA and ionomycin, and was restored by calpain inhibitors but not matrix metalloproteinase or gamma-secretase inhibitors. Calpain directly cleaved N-cadherin in in vitro calpain assays, and calpain inhibitors prevented its cleavage in a dose-dependent manner. Using N-cadherin deletion mutants, we found that calpain cleavage sites exist in at least four regions of the cytoplasmic domain. Treatment with NMDA induced neuronal death, and it suppressed the expression of surface N-cadherin and the N-cadherin/beta-catenin interaction, effects that were prevented by calpain inhibitor. Furthermore, calpain-mediated N-cadherin cleavage significantly affected cell-cell adhesion, AKT signaling, the N-cadherin/beta-catenin interaction and the Wnt target gene expressions through the accumulation of nuclear beta-catenin.


Assuntos
Lesões Encefálicas/metabolismo , Caderinas/metabolismo , Calpaína/metabolismo , Animais , Animais Recém-Nascidos , Biotinilação/métodos , Lesões Encefálicas/patologia , Caderinas/genética , Cálcio/metabolismo , Calpaína/farmacologia , Células Cultivadas , Modelos Animais de Doenças , Maleato de Dizocilpina/farmacologia , Embrião de Mamíferos , Endocitose/efeitos dos fármacos , Endocitose/fisiologia , Inibidores Enzimáticos/farmacologia , Agonistas de Aminoácidos Excitatórios/farmacologia , Antagonistas de Aminoácidos Excitatórios/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas de Fluorescência Verde/genética , Técnicas In Vitro , Ionomicina/farmacologia , Ionóforos/farmacologia , Camundongos , Modelos Biológicos , Mutação/genética , N-Metilaspartato/farmacologia , Neurônios/efeitos dos fármacos , Estrutura Terciária de Proteína/efeitos dos fármacos , Estrutura Terciária de Proteína/fisiologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/fisiologia , Frações Subcelulares/efeitos dos fármacos , Transfecção/métodos , Proteínas Wnt/metabolismo , beta Catenina/metabolismo
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