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1.
J Cell Physiol ; 234(5): 6667-6678, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30317589

RESUMO

Organ-organ crosstalk is involved in homeostasis. Gastrointestinal symptoms are common in patients with renal failure. The aim of this study was to elucidate the relationship between gastrointestinal motility and gastrointestinal symptoms in chronic kidney disease. We performed studies in C57BL/6 mice with chronic kidney disease after 5/6 nephrectomy. Gastrointestinal motility was evaluated by assessing the ex vivo responses of ileum and distal colon strips to electrical field stimulation. Feces were collected from mice, and the composition of the gut microbiota was analyzed using 16S ribosomal RNA sequencing. Mice with chronic kidney disease after 5/6 nephrectomy showed a decreased amount of stool, and this constipation was correlated with a suppressed contraction response in ileum motility and decreased relaxation response in distal colon motility. Spermine, one of the uremic toxins, inhibited the contraction response in ileum motility, but four types of uremic toxins showed no effect on the relaxation response in distal colon motility. The 5/6 nephrectomy procedure disturbed the balance of the gut microbiota in the mice. The motility dysregulation and constipation were resolved by antibiotic treatments. The expression levels of interleukin 6, tumor necrosis factor-α, and iNOS in 5/6 nephrectomy mice were increased in the distal colon but not in the ileum. In addition, macrophage infiltration in 5/6 nephrectomy mice was increased in the distal colon but not in the ileum. We found that 5/6 nephrectomy altered gastrointestinal motility and caused constipation by changing the gut microbiota and causing colonic inflammation. These findings indicate that renal failure was remarkably associated with gastrointestinal dysregulation.


Assuntos
Microbioma Gastrointestinal/fisiologia , Motilidade Gastrointestinal/fisiologia , Nefrectomia , Insuficiência Renal Crônica/microbiologia , Animais , Colo/microbiologia , Colo/patologia , Colo/cirurgia , Trato Gastrointestinal/microbiologia , Inflamação/patologia , Masculino , Camundongos Endogâmicos C57BL , Nefrectomia/métodos , Insuficiência Renal Crônica/cirurgia
2.
Biol Pharm Bull ; 41(2): 182-189, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29386478

RESUMO

Interleukin (IL)-19 is a member of the IL-10 family of interleukins and is an immuno-modulatory cytokine produced by the main macrophages. The gastrointestinal tissues of IL-19 knockout mice show exacerbated experimental colitis mediated by the innate immune system and T cells. There is an increasing focus on the interaction and relationship of IL-19 with the function of T cells. Contact hypersensitivity (CHS) is T cell-mediated cutaneous inflammation. Therefore, we asked whether IL-19 causes CHS. We investigated the immunological role of IL-19 in CHS induced by 1-fluoro-2,4-dinitrofluorobenzene as a hapten. IL-19 was highly expressed in skin exposed to the hapten, and ear swelling was increased in IL-19 knockout mice. The exacerbation of the CHS response in IL-19 knockout mice correlated with increased levels of IL-17 and IL-6, but no alterations were noted in the production of interferon (IFN)γ and IL-4 in the T cells of the lymph nodes. In addition to the effect on T cell response, IL-19 knockout mice increased production of inflammatory cytokines. These results show that IL-19 suppressed hapten-dependent skin inflammation in the elicitation phase of CHS.


Assuntos
Dermatite de Contato/metabolismo , Interleucinas/agonistas , Linfonodos/metabolismo , Pele/metabolismo , Animais , Células Cultivadas , Dermatite de Contato/sangue , Dermatite de Contato/imunologia , Dermatite de Contato/patologia , Dinitrofluorbenzeno/análogos & derivados , Dinitrofluorbenzeno/toxicidade , Orelha , Regulação da Expressão Gênica/efeitos dos fármacos , Haptenos/toxicidade , Imunidade Inata/efeitos dos fármacos , Imuno-Histoquímica , Interleucina-10 , Interleucina-17/agonistas , Interleucina-17/genética , Interleucina-17/metabolismo , Interleucina-6/agonistas , Interleucina-6/genética , Interleucina-6/metabolismo , Interleucinas/sangue , Interleucinas/genética , Interleucinas/metabolismo , Linfonodos/efeitos dos fármacos , Linfonodos/imunologia , Linfonodos/patologia , Camundongos Endogâmicos BALB C , Camundongos Knockout , RNA Mensageiro/metabolismo , Pele/efeitos dos fármacos , Pele/imunologia , Pele/patologia , Baço/efeitos dos fármacos , Baço/imunologia , Baço/metabolismo , Baço/patologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T/metabolismo , Linfócitos T/patologia , Células Th1/efeitos dos fármacos , Células Th1/imunologia , Células Th1/metabolismo , Células Th1/patologia
3.
Int Immunopharmacol ; 55: 205-215, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-29272817

RESUMO

Macrophages are important cells that need to be controlled at the site of inflammation. Several factors are involved in chronic inflammation and its timely resolution. Free fatty acids drive the inflammatory response in macrophages and contribute to the vicious cycle of the inflammatory response. However, the identity of the uptake pathways of fatty acids is not fully clear in macrophages and how the inflammatory responses are regulated by the uptake of fatty acids remain poorly understood. We investigated the relationship between fatty acid transport protein (FATP) and the inflammatory response signaling pathway in macrophages as the first report. The FATP family has composed six isoforms, FATP1-6. We found that FATP1 is the most highly expressed isoform in macrophages. Forced expression of FATP1 enhanced production of inflammatory cytokines, such as TNFα and IL-6 concomitant with the increased uptake of fatty acids, increased level of ceramide, and increased phosphorylation of c-Jun N-terminal kinase (JNK). The enhancement by FATP1 was abolished by treatment with a JNK inhibitor, NF-κB inhibitor, or ceramide synthesis inhibitor. siRNA-mediated knockdown of FATP1 strongly inhibited the production of TNFα and IL-6. Similarly, an inhibitor of FATP1 inhibited the production of TNFα and IL-6. Finally, an inhibitor of FATP1 attenuated the production of inflammatory cytokines in bronchoalveolar lavage fluid in an LPS-induced acute lung injury in vivo mouse model. In summary, we propose that FATP1 is an important regulator of inflammatory response signaling in macrophages. Our findings suggest that ceramide-JNK signaling is important to terminate or sustain inflammation.


Assuntos
Lesão Pulmonar Aguda/tratamento farmacológico , Proteínas de Transporte de Ácido Graxo/metabolismo , Inflamação/imunologia , Macrófagos/fisiologia , Lesão Pulmonar Aguda/induzido quimicamente , Animais , Ceramidas/metabolismo , Proteínas de Transporte de Ácido Graxo/antagonistas & inibidores , Proteínas de Transporte de Ácido Graxo/genética , Ácidos Graxos/metabolismo , Interleucina-6/metabolismo , MAP Quinase Quinase 4/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Células RAW 264.7 , RNA Interferente Pequeno/genética , Fator de Necrose Tumoral alfa/metabolismo
4.
Neurochem Int ; 119: 171-177, 2018 10.
Artigo em Inglês | MEDLINE | ID: mdl-29108865

RESUMO

Insulin resistance in brain has been reported in Alzheimer's diseases (AD). Insulin signaling is important for homeostasis in brain function and reported to be disturbed in neurons leading to tau phosphorylation and neurofibrillary tangles. Many investigations of insulin in neurons have been reported; however, it has not been reported whether astrocytes also produce insulin. In the present study, we assessed the expression of insulin in astrocytes cultured from rat embryonic brain and the effects of amyloid ß1-42 (Aß) and lipopolysaccharide (LPS) on the expression. We found that astrocytes expressed preproinsulin mRNAs and insulin protein, and that Aß or LPS decreased these expressions. Antioxidants, glutathione and N-acetylcysteine, restored the decreases in insulin mRNA expression by Aß and by LPS. Insulin protein was detected in astrocyte conditioned medium. These results suggest that astrocytes express and secrete insulin. Oxidative stress might be involved in the decreased insulin expression by Aß or LPS. The insulin decrease by Aß in astrocytes could be a novel disturbing mechanism for brain insulin signaling in AD.


Assuntos
Peptídeos beta-Amiloides/metabolismo , Antioxidantes/farmacologia , Astrócitos/metabolismo , Insulina/metabolismo , Doença de Alzheimer/metabolismo , Animais , Células Cultivadas , Neurônios/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Fosforilação/efeitos dos fármacos , Precursores de Proteínas/metabolismo , Ratos Wistar , Transdução de Sinais/efeitos dos fármacos , Proteínas tau/metabolismo
6.
J Biol Chem ; 292(11): 4727-4742, 2017 03 17.
Artigo em Inglês | MEDLINE | ID: mdl-28167533

RESUMO

Glycolytic glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a multifunctional protein that also mediates cell death under oxidative stress. We reported previously that the active-site cysteine (Cys-152) of GAPDH plays an essential role in oxidative stress-induced aggregation of GAPDH associated with cell death, and a C152A-GAPDH mutant rescues nitric oxide (NO)-induced cell death by interfering with the aggregation of wild type (WT)-GAPDH. However, the detailed mechanism underlying GAPDH aggregate-induced cell death remains elusive. Here we report that NO-induced GAPDH aggregation specifically causes mitochondrial dysfunction. First, we observed a correlation between NO-induced GAPDH aggregation and mitochondrial dysfunction, when GAPDH aggregation occurred at mitochondria in SH-SY5Y cells. In isolated mitochondria, aggregates of WT-GAPDH directly induced mitochondrial swelling and depolarization, whereas mixtures containing aggregates of C152A-GAPDH reduced mitochondrial dysfunction. Additionally, treatment with cyclosporin A improved WT-GAPDH aggregate-induced swelling and depolarization. In doxycycline-inducible SH-SY5Y cells, overexpression of WT-GAPDH augmented NO-induced mitochondrial dysfunction and increased mitochondrial GAPDH aggregation, whereas induced overexpression of C152A-GAPDH significantly suppressed mitochondrial impairment. Further, NO-induced cytochrome c release into the cytosol and nuclear translocation of apoptosis-inducing factor from mitochondria were both augmented in cells overexpressing WT-GAPDH but ameliorated in C152A-GAPDH-overexpressing cells. Interestingly, GAPDH aggregates induced necrotic cell death via a permeability transition pore (PTP) opening. The expression of either WT- or C152A-GAPDH did not affect other cell death pathways associated with protein aggregation, such as proteasome inhibition, gene expression induced by endoplasmic reticulum stress, or autophagy. Collectively, these results suggest that NO-induced GAPDH aggregation specifically induces mitochondrial dysfunction via PTP opening, leading to cell death.


Assuntos
Morte Celular , Gliceraldeído-3-Fosfato Desidrogenases/metabolismo , Mitocôndrias/patologia , Óxido Nítrico/metabolismo , Estresse Oxidativo , Agregados Proteicos , Fator de Indução de Apoptose/metabolismo , Linhagem Celular Tumoral , Citocromos c/metabolismo , Humanos , Mitocôndrias/metabolismo , Proteínas de Transporte da Membrana Mitocondrial/metabolismo
7.
Biochem Biophys Res Commun ; 484(2): 385-389, 2017 03 04.
Artigo em Inglês | MEDLINE | ID: mdl-28130107

RESUMO

Synthesis of poly(ADP-ribose) (PAR) is catalyzed by PAR polymerase-1 (PARP-1) in neurons. PARP1 plays a role in various types of brain damage in neurodegenerative disorders. In neurons, overactivation of PARP-1 during oxidative stress induces robust PAR formation, which depletes nicotinamide adenine dinucleotide levels and leads to cell death. However, the role of the newly-formed PAR in neurodegenerative disorders remains elusive. We hypothesized that the effects of PAR could occur in the extracellular space after it is leaked from damaged neurons. Here we report that extracellular PAR (EC-PAR) functions as a neuroprotective molecule by inducing the synthesis of glial cell line-derived neurotrophic factor (GDNF) in astrocytes during neuronal cell death, both in vitro and in vivo. In primary rat astrocytes, exogenous treatment with EC-PAR produced GDNF but not other neurotrophic factors. The effect was concentration-dependent and did not affect cell viability in rat C6 astrocytoma cells. Topical injection of EC-PAR into rat striatum upregulated GDNF levels in activated astrocytes and improved pathogenic rotation behavior in a unilateral 6-hydroxydopamine model of Parkinson disease in rats. These findings indicate that EC-PAR acts as a neurotrophic enhancer by upregulating GDNF levels. This effect protects the remaining neurons following oxidative stress-induced brain damage, such as that seen with Parkinson disease.


Assuntos
Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Poli Adenosina Difosfato Ribose/metabolismo , Animais , Astrócitos/metabolismo , Linhagem Celular Tumoral , Células Cultivadas , Modelos Animais de Doenças , Espaço Extracelular/metabolismo , Técnicas In Vitro , Fatores de Crescimento Neural/metabolismo , Doença de Parkinson/terapia , Ratos , Ratos Wistar , Regulação para Cima
8.
J Vet Med Sci ; 79(2): 403-411, 2017 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-27928109

RESUMO

The Na+/Ca2+ exchanger (NCX) is a bidirectional transporter that is controlled by membrane potential and transmembrane gradients of Na+ and Ca2+. To reveal the functional role of NCX on gastrointestinal motility, we have previously used NCX1 and NCX2 heterozygote knockout mice (HET). We found that NCX1 and NCX2 play important roles in the motility of the gastric fundus, ileum and distal colon. Therefore, we believed that NCX1 and NCX2 play an important role in transport of intestinal contents. Here, we investigated the role of NCX in a mouse model of drug-induced diarrhea. The fecal consistencies in NCX1 HET and NCX2 HET were assessed using a diarrhea induced by magnesium sulfate, 5-hydroxytryptamine (5-HT) and prostaglandin E2 (PGE2). NCX2 HET, but not NCX1 HET, exacerbated magnesium sulfate-induced diarrhea by increasing watery fecals. Likewise, 5-HT-induced diarrheas were exacerbated in NCX2 HET, but not NCX1 HET. However, NCX1 HET and NCX2 HET demonstrated PGE2 induced diarrhea similar to those of wild-type mice (WT). As well as the result of the distal colon shown previously, in the proximal and transverse colons of WT, the myenteric plexus layers and the longitudinal and circular muscle layers were strongly immunoreactive to NCX1 and NCX2. In this study, we demonstrate that NCX2 has important roles in development of diarrhea.


Assuntos
Colo/metabolismo , Diarreia/metabolismo , Motilidade Gastrointestinal , Trocador de Sódio e Cálcio/metabolismo , Animais , Diarreia/induzido quimicamente , Fezes , Técnicas In Vitro , Sulfato de Magnésio , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout
9.
Naunyn Schmiedebergs Arch Pharmacol ; 390(3): 261-268, 2017 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-27942772

RESUMO

Inflammatory bowel disease results from chronic dysregulation of the mucosal immune system and aberrant activation of both the innate and adaptive immune responses. IL-19 is a member of the IL-10 family, and IL-10 plays an important role in inflammatory bowel disease. We have previously shown that IL-19 knockout mice are more susceptible to innate-mediated colitis. Next, we ask whether IL-19 contributes to T cells-mediated colitis. Here, we investigated the role of IL-19 in a mouse model of Th2 cell-mediated colitis. Inflammatory responses in IL-19-deficient mice were assessed using a Th2-mediated colitis induced by oxazolone. The colitis was evaluated by analyzing the body weight loss and histology of the colon. Lymph node cells were cultured in vitro to determine cytokine production. IL-19 knockout mice exacerbated oxazolone-induced colitis by stimulating the transport of inflammatory cells into the colon, and by increasing IgE production and the number of circulating eosinophil. The exacerbation of oxazolone-induced colonic inflammation following IL-19 knockout mice was accompanied by an increased production of IL-4 and IL-9, but no changes in the expression of IL-5 and IL-13 in lymph node cells. IL-19 plays an anti-inflammatory role in the Th2-mediated colitis model, suggesting that IL-19 may represent a potential therapeutic target for reducing colonic inflammation.


Assuntos
Colite/prevenção & controle , Colo/metabolismo , Mediadores da Inflamação/metabolismo , Interleucina-10/metabolismo , Células Th2/metabolismo , Animais , Células Cultivadas , Colite/induzido quimicamente , Colite/genética , Colite/metabolismo , Colo/imunologia , Colo/patologia , Modelos Animais de Doenças , Predisposição Genética para Doença , Mediadores da Inflamação/imunologia , Interleucina-10/deficiência , Interleucina-10/genética , Linfonodos/imunologia , Linfonodos/metabolismo , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Oxazolona , Fenótipo , Fatores de Proteção , Células Th2/imunologia , Fatores de Tempo
10.
J Pharmacol Sci ; 132(3): 181-186, 2016 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-27816547

RESUMO

In gastric smooth muscles, the released Ca2+ activates the contractile proteins and Ca2+ taken up from the cytosol cause relaxation. The Na+/Ca2+ exchanger (NCX) is an antiporter membrane protein that controls Ca2+ influx and efflux across the membrane. However, the possible relation of NCX in gastric fundus motility is largely unknown. Here, we investigated electric field stimulation (EFS)-induced relaxations in the circular muscles of the gastric fundus in smooth muscle-specific NCX1 transgenic mice (Tg). EFS caused a bi-phasic response, transient and sustained relaxation. The sustained relaxation prolonged for an extended period after the end of the stimulus. EFS-induced transient relaxation and sustained relaxation were greater in Tg than in wild-type mice (WT). Disruption of nitric oxide component by N-nitro-l-arginine, EFS-induced transient and sustained relaxations caused still marked in Tg compared to WT. Inhibition of PACAP by antagonist, EFS-induced sustained relaxation in Tg was not seen, similar to WT. Nevertheless, transient relaxation remained more pronounced in Tg than in WT. Next, we examined responses to NO and PACAP in smooth muscles. The magnitudes of NOR-1, which generates NO, and PACAP-induced relaxations were greater in Tg than in WT. In this study, we demonstrate that NCX1 regulates gastric fundus motility.


Assuntos
Fundo Gástrico/fisiologia , Trocador de Sódio e Cálcio/biossíntese , Animais , Estimulação Elétrica , Fundo Gástrico/metabolismo , Técnicas In Vitro , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Relaxamento Muscular/fisiologia , Músculo Liso/metabolismo , Músculo Liso/fisiologia
11.
Naunyn Schmiedebergs Arch Pharmacol ; 389(10): 1081-90, 2016 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-27411318

RESUMO

The Na(+)/Ca(2+) exchanger (NCX) is a plasma membrane transporter that is involved in regulating intracellular Ca(2+) concentrations in various tissues. The physiological roles by which NCX influences gastrointestinal motility are incompletely understood, although its role in the heart, brain, and kidney has been widely investigated. In this study, we focused on the functions of the NCX isoforms, NCX1 and NCX2, in the motility of the ileum in the gastrointestinal tract. We investigated the response to electric field stimulation (EFS) in the longitudinal smooth muscle of the ileum obtained from wild-type mice (WT), NCX1-heterozygote knockout mice (NCX1 HET), NCX2 HET and smooth muscle-specific NCX1.3 transgenic mice (NCX1.3 Tg). EFS induced a phasic contraction that persisted during EFS and a tonic contraction that occurred after the end of EFS. We found that the amplitudes of the phasic and tonic contractions were significantly smaller in NCX2 HET, but not in NCX1 HET, compared to WT. Moreover, the magnitudes of acetylcholine (ACh)- and substance P (SP)-induced contractions of NCX2 HET, but not of NCX1 HET, were smaller compared to WT. In contrast, the amplitudes of the phasic and tonic contractions were greater in NCX1.3 Tg compared to WT. Similar to EFS, the magnitude of ACh-induced contraction was greater in NCX1.3 Tg than in WT. Taken together, our findings indicated that NCX1 and NCX2 play important roles in ileal motility and suggest that NCX1 and NCX2 regulate the motility in the ileum by controlling the sensitivity of smooth muscles to ACh and SP.


Assuntos
Motilidade Gastrointestinal , Íleo/metabolismo , Contração Muscular , Músculo Liso/metabolismo , Trocador de Sódio e Cálcio/metabolismo , Acetilcolina/farmacologia , Animais , Estimulação Elétrica , Motilidade Gastrointestinal/efeitos dos fármacos , Genótipo , Íleo/efeitos dos fármacos , Íleo/inervação , Técnicas In Vitro , Camundongos Endogâmicos C57BL , Camundongos Knockout , Contração Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Músculo Liso/inervação , Fenótipo , Trocador de Sódio e Cálcio/genética , Substância P/farmacologia
12.
Nitric Oxide ; 53: 13-21, 2016 Feb 29.
Artigo em Inglês | MEDLINE | ID: mdl-26725192

RESUMO

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH), a homotetrameric enzyme involved in a key step of glycolysis, also has a role in mediating cell death under nitrosative stress. Our previous reports suggest that nitric oxide-induced intramolecular disulfide-bonding GAPDH aggregation, which occurs through oxidation of the active site cysteine (Cys-152), participates in a mechanism to account for nitric oxide-induced death signaling in some neurodegenerative/neuropsychiatric disorders. Here, we demonstrate a rescue strategy for nitric oxide-induced cell death accompanied by GAPDH aggregation in a mutant with a substitution of Cys-152 to alanine (C152A-GAPDH). Pre-incubation of purified wild-type GAPDH with C152A-GAPDH under exposure to nitric oxide inhibited wild-type GAPDH aggregation in a concentration-dependent manner in vitro. Several lines of structural analysis revealed that C152A-GAPDH extensively interfered with nitric oxide-induced GAPDH-amyloidogenesis. Overexpression of doxycycline-inducible C152A-GAPDH in SH-SY5Y neuroblastoma significantly rescued nitric oxide-induced death, concomitant with the decreased formation of GAPDH aggregates. Further, both co-immunoprecipitation assays and simulation models revealed a heterotetramer composed of one dimer each of wild-type GAPDH and C152A-GAPDH. These results suggest that the C152A-GAPDH mutant acts as a dominant-negative molecule against GAPDH aggregation via the formation of this GAPDH heterotetramer. This study may contribute to a new therapeutic approach utilizing C152A-GAPDH against brain damage in nitrosative stress-related disorders.


Assuntos
Cisteína/metabolismo , Gliceraldeído-3-Fosfato Desidrogenases/metabolismo , Óxido Nítrico/farmacologia , Domínio Catalítico/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Cisteína/genética , Relação Dose-Resposta a Droga , Gliceraldeído-3-Fosfato Desidrogenases/genética , Humanos , Agregados Proteicos/efeitos dos fármacos , Relação Estrutura-Atividade , Células Tumorais Cultivadas
13.
Naunyn Schmiedebergs Arch Pharmacol ; 389(1): 63-72, 2016 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-26475617

RESUMO

The muscular layer in the GI tract consists of an inner circular muscular layer and an outer longitudinal muscular layer. Acetylcholine (ACh) is the representative neurotransmitter that causes contractions in the gastrointestinal tracts of most animal species. There are many reports of muscarinic receptor-mediated contraction of longitudinal muscles, but few studies discuss circular muscles. The present study detailed the contractile response in the circular smooth muscles of the mouse ileum. We used small muscle strips (0.2 mm × 1 mm) and large muscle strips (4 × 4 mm) isolated from the circular and longitudinal muscle layers of the mouse ileum to compare contraction responses in circular and longitudinal smooth muscles. The time to peak contractile responses to carbamylcholine (CCh) were later in the small muscle strips (0.2 × 1 mm) of circular muscle (5.7 min) than longitudinal muscles (0.4 min). The time to peak contractile responses to CCh in the large muscle strips (4 × 4 mm) were also later in the circular muscle (3.1 min) than the longitudinal muscle (1.4 min). Furthermore, a muscarinic M2 receptor antagonist and gap junction inhibitor significantly delayed the time to peak contraction of the large muscle strips (4 × 4 mm) from the circular muscular layer. Our findings indicate that muscarinic M2 receptors in the circular muscular layer of mouse ileum exert a previously undocumented function in gut motility via the regulation of gap junctions.


Assuntos
Carbacol/farmacologia , Agonistas Colinérgicos/farmacologia , Íleo/efeitos dos fármacos , Contração Muscular/efeitos dos fármacos , Músculo Liso/efeitos dos fármacos , Receptor Muscarínico M2/fisiologia , Animais , Junções Comunicantes/fisiologia , Íleo/fisiologia , Técnicas In Vitro , Masculino , Camundongos Endogâmicos C57BL , Contração Muscular/fisiologia , Músculo Liso/fisiologia
14.
Biosci Biotechnol Biochem ; 80(3): 547-53, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26540606

RESUMO

Ferulic acid (FA) has been reported to exhibit protective effects against amyloid-ß (Aß)-induced neurodegeneration in vitro and in vivo. Recently, we developed two water-soluble FA derivatives: 1-feruloyl glycerol and 1-feruloyl diglycerol. In this study, we examined the neuroprotective effects of these water-soluble FA derivatives on Aß-induced neurodegeneration both in vitro and in vivo. FA and water-soluble FA derivatives inhibited Aß aggregation and destabilized pre-aggregated Aß to a similar extent. Furthermore, water-soluble FA derivatives, as well as FA, inhibited Aß-induced neuronal cell death in cultured neuronal cells. In in vivo experiments, oral administration of water-soluble FA derivatives to mice improved Aß-induced dysmnesia assessed by contextual fear conditioning test and protected hippocampal neurons against Aß-induced neurotoxicity. This study provides useful evidence suggesting that water-soluble FA derivatives are expected to be effective neuroprotective agents.


Assuntos
Peptídeos beta-Amiloides/fisiologia , Morte Celular/fisiologia , Ácidos Cumáricos/química , Neurônios/citologia , Peptídeos beta-Amiloides/metabolismo , Animais , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Solubilidade , Água
15.
Int Immunopharmacol ; 29(2): 468-475, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26476684

RESUMO

Inflammatory bowel disease results from chronic dysregulation of the mucosal immune system and aberrant activation of both the innate and adaptive immune responses. Interleukin (IL)-19, a member of the IL-10 family, functions as an anti-inflammatory cytokine. Here, we investigated the contribution of IL-19 to intestinal inflammation in a model of T cell-mediated colitis in mice. Inflammatory responses in IL-19-deficient mice were assessed using the 2,4,6-trinitrobenzene sulfonic acid (TNBS) model of acute colitis. IL-19 deficiency aggravated TNBS-induced colitis and compromised intestinal recovery in mice. Additionally, the exacerbation of TNBS-induced colonic inflammation following genetic ablation of IL-19 was accompanied by increased production of interferon-gamma, IL-12 (p40), IL-17, IL-22, and IL-33, and decreased production of IL-4. Moreover, the exacerbation of colitis following IL-19 knockout was also accompanied by increased production of CXCL1, G-CSF and CCL5. Using this model of induced colitis, our results revealed the immunopathological relevance of IL-19 as an anti-inflammatory cytokine in intestinal inflammation in mice.


Assuntos
Colite/induzido quimicamente , Colite/genética , Inflamação/induzido quimicamente , Inflamação/genética , Interleucina-10/genética , Animais , Colite/patologia , Doença de Crohn/induzido quimicamente , Doença de Crohn/patologia , Citocinas/antagonistas & inibidores , Citocinas/biossíntese , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Recuperação de Função Fisiológica , Linfócitos T/imunologia , Ácido Trinitrobenzenossulfônico
16.
Biochem Biophys Res Commun ; 467(2): 373-6, 2015 Nov 13.
Artigo em Inglês | MEDLINE | ID: mdl-26431872

RESUMO

The glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) has multiple functions, including mediating oxidative stress-induced neuronal cell death. This process is associated with disulfide-bonded GAPDH aggregation. Some reports suggest a link between GAPDH and the pathogenesis of several oxidative stress-related diseases. However, the pathological significance of GAPDH aggregation in disease pathogenesis remains unclear due to the lack of an effective GAPDH aggregation inhibitor. In this study, we identified a GAPDH aggregation inhibitor (GAI) peptide and evaluated its biological profile. The decapeptide GAI specifically inhibited GAPDH aggregation in a concentration-dependent manner. Additionally, the GAI peptide did not affect GAPDH glycolytic activity or cell viability. The GAI peptide also exerted a protective effect against oxidative stress-induced cell death in SH-SY5Y cells. This peptide could potentially serve as a tool to investigate GAPDH aggregation-related neurodegenerative and neuropsychiatric disorders and as a possible therapy for diseases associated with oxidative stress-induced cell death.


Assuntos
Inibidores Enzimáticos/farmacologia , Gliceraldeído-3-Fosfato Desidrogenases/antagonistas & inibidores , Neurônios/efeitos dos fármacos , Oligopeptídeos/farmacologia , Agregados Proteicos/efeitos dos fármacos , Sequência de Aminoácidos , Animais , Apoptose/efeitos dos fármacos , Domínio Catalítico , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Inibidores Enzimáticos/síntese química , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Gliceraldeído-3-Fosfato Desidrogenases/química , Gliceraldeído-3-Fosfato Desidrogenases/genética , Gliceraldeído-3-Fosfato Desidrogenases/metabolismo , Glicólise/efeitos dos fármacos , Humanos , Modelos Moleculares , Dados de Sequência Molecular , Neurônios/metabolismo , Neurônios/patologia , Oligopeptídeos/síntese química , Oxirredução , Estresse Oxidativo , Células PC12 , Ligação Proteica , Ratos , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo
17.
J Biol Chem ; 290(43): 26072-87, 2015 Oct 23.
Artigo em Inglês | MEDLINE | ID: mdl-26359500

RESUMO

Alzheimer disease (AD) is a progressive neurodegenerative disorder characterized by loss of neurons and formation of pathological extracellular deposits induced by amyloid-ß peptide (Aß). Numerous studies have established Aß amyloidogenesis as a hallmark of AD pathogenesis, particularly with respect to mitochondrial dysfunction. We have previously shown that glycolytic glyceraldehyde-3-phosphate dehydrogenase (GAPDH) forms amyloid-like aggregates upon exposure to oxidative stress and that these aggregates contribute to neuronal cell death. Here, we report that GAPDH aggregates accelerate Aß amyloidogenesis and subsequent neuronal cell death both in vitro and in vivo. Co-incubation of Aß40 with small amounts of GAPDH aggregates significantly enhanced Aß40 amyloidogenesis, as assessed by in vitro thioflavin-T assays. Similarly, structural analyses using Congo red staining, circular dichroism, and atomic force microscopy revealed that GAPDH aggregates induced Aß40 amyloidogenesis. In PC12 cells, GAPDH aggregates augmented Aß40-induced cell death, concomitant with disruption of mitochondrial membrane potential. Furthermore, mice injected intracerebroventricularly with Aß40 co-incubated with GAPDH aggregates exhibited Aß40-induced pyramidal cell death and gliosis in the hippocampal CA3 region. These observations were accompanied by nuclear translocation of apoptosis-inducing factor and cytosolic release of cytochrome c from mitochondria. Finally, in the 3×Tg-AD mouse model of AD, GAPDH/Aß co-aggregation and mitochondrial dysfunction were consistently detected in an age-dependent manner, and Aß aggregate formation was attenuated by GAPDH siRNA treatment. Thus, this study suggests that GAPDH aggregates accelerate Aß amyloidogenesis, subsequently leading to mitochondrial dysfunction and neuronal cell death in the pathogenesis of AD.


Assuntos
Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Gliceraldeído-3-Fosfato Desidrogenases/metabolismo , Peptídeos beta-Amiloides/biossíntese , Animais , Humanos , Camundongos , Camundongos Transgênicos , Microscopia de Força Atômica , Mitocôndrias/fisiologia , Células PC12 , Ratos
18.
J Biol Chem ; 290(23): 14493-503, 2015 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-25882840

RESUMO

In addition to its role in DNA repair, nuclear poly(ADP-ribose) polymerase-1 (PARP-1) mediates brain damage when it is over-activated by oxidative/nitrosative stress. Nonetheless, it remains unclear how PARP-1 is activated in neuropathological contexts. Here we report that PARP-1 interacts with a pool of glyceradehyde-3-phosphate dehydrogenase (GAPDH) that translocates into the nucleus under oxidative/nitrosative stress both in vitro and in vivo. A well conserved amino acid at the N terminus of GAPDH determines its protein binding with PARP-1. Wild-type (WT) but not mutant GAPDH, that lacks the ability to bind PARP-1, can promote PARP-1 activation. Importantly, disrupting this interaction significantly diminishes PARP-1 overactivation and protects against both brain damage and neurological deficits induced by middle cerebral artery occlusion/reperfusion in a rat stroke model. Together, these findings suggest that nuclear GAPDH is a key regulator of PARP-1 activity, and its signaling underlies the pathology of oxidative/nitrosative stress-induced brain damage including stroke.


Assuntos
Encéfalo/patologia , Gliceraldeído-3-Fosfato Desidrogenases/metabolismo , Infarto da Artéria Cerebral Média/metabolismo , Infarto da Artéria Cerebral Média/patologia , Estresse Oxidativo , Poli(ADP-Ribose) Polimerases/metabolismo , Sequência de Aminoácidos , Animais , Encéfalo/irrigação sanguínea , Encéfalo/enzimologia , Encéfalo/metabolismo , Linhagem Celular , Núcleo Celular/enzimologia , Núcleo Celular/metabolismo , Núcleo Celular/patologia , Ativação Enzimática , Gliceraldeído-3-Fosfato Desidrogenases/análise , Humanos , Infarto da Artéria Cerebral Média/enzimologia , Masculino , Modelos Moleculares , Dados de Sequência Molecular , Nitrocompostos/análise , Nitrocompostos/metabolismo , Poli(ADP-Ribose) Polimerase-1 , Poli(ADP-Ribose) Polimerases/análise , Ratos , Ratos Wistar
19.
J Biol Chem ; 290(1): 56-64, 2015 Jan 02.
Artigo em Inglês | MEDLINE | ID: mdl-25391652

RESUMO

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) plays roles in both energy maintenance, and stress signaling by forming a protein complex with seven in absentia homolog 1 (Siah1). Mechanisms to coordinate its glycolytic and stress cascades are likely to be very important for survival and homeostatic control of any living organism. Here we report that apoptosis signal-regulating kinase 1 (ASK1), a representative stress kinase, interacts with both GAPDH and Siah1 and is likely able to phosphorylate Siah1 at specific amino acid residues (Thr-70/Thr-74 and Thr-235/Thr-239). Phosphorylation of Siah1 by ASK1 triggers GAPDH-Siah1 stress signaling and activates a key downstream target, p300 acetyltransferase in the nucleus. This novel mechanism, together with the established S-nitrosylation/oxidation of GAPDH at Cys-150, provides evidence of how the stress signaling involving GAPDH is finely regulated. In addition, the present results imply crosstalk between the ASK1 and GAPDH-Siah1 stress cascades.


Assuntos
Gliceraldeído-3-Fosfato Desidrogenase (Fosforiladora)/metabolismo , MAP Quinase Quinase Quinase 5/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Recombinantes de Fusão/metabolismo , Transdução de Sinais/genética , Ubiquitina-Proteína Ligases/metabolismo , Sequência de Aminoácidos , Sítios de Ligação , Regulação da Expressão Gênica , Gliceraldeído-3-Fosfato Desidrogenase (Fosforiladora)/genética , Células HEK293 , Humanos , Peróxido de Hidrogênio/farmacologia , MAP Quinase Quinase Quinase 5/genética , Dados de Sequência Molecular , Proteínas Nucleares/genética , Estresse Oxidativo , Fosforilação/efeitos dos fármacos , Ligação Proteica/efeitos dos fármacos , Proteínas Recombinantes de Fusão/genética , Ubiquitina-Proteína Ligases/genética
20.
Pharmacology ; 94(5-6): 230-8, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25427675

RESUMO

Na(+)/Ca(2+) exchanger 1 (NCX1) is a plasma membrane transporter involved in regulating intracellular Ca(2+) concentrations. NCX1 is critical for Ca(2+) regulation in cardiac muscle, vascular smooth muscle and nerve fibers. However, little is known about the physiological role of NCX1 in gastrointestinal motility. To determine the role of NCX1 in gastrointestinal tissues, we examined electric field stimulation (EFS)-induced responses in the longitudinal smooth muscle of the distal colon in smooth muscle-specific NCX1 transgenic mice (Tg). Tg show that NCX1 protein was overexpressed in the distal colon at a level twofold greater than that of endogenous NCX1. We found that the amplitudes of EFS-induced relaxation that persisted during EFS were greater in Tg than in wild-type mice (WT). Under the nonadrenergic, noncholinergic condition, the EFS-induced relaxation in Tg was also greater than that in WT. Inhibition of NO synthase, CO synthase, soluble guanylate cyclase (sGC), and protein kinase G (PKG) all attenuated the enhanced relaxation in Tg, demonstrating the importance of NCX1 in NO/sGC/PKG signaling. The action of NOR-1, an NO donor, induced enhanced relaxation in Tg compared with that in WT. Unlike NOR-1, pituitary adenylate cyclase-activating peptide and vasoactive intestinal peptide induced a similar relaxation in Tg compared with that in WT. In this study, we demonstrate that NCX1 plays an important role in smooth muscle motility in the mouse distal colon.


Assuntos
Colo/fisiologia , Trocador de Sódio e Cálcio/fisiologia , Animais , Motilidade Gastrointestinal/fisiologia , Hidroxilaminas/farmacologia , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Relaxamento Muscular , Doadores de Óxido Nítrico/farmacologia , Trocador de Sódio e Cálcio/genética , Trocador de Sódio e Cálcio/metabolismo
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