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1.
Biomedicines ; 8(9)2020 Sep 21.
Artigo em Inglês | MEDLINE | ID: mdl-32967121

RESUMO

Apurinic/apyrimidinic endonuclease 1/redox factor-1 (APE1/Ref-1) is involved in DNA base repair and reducing activity. However, the role of APE1/Ref-1 in atherosclerosis is unclear. Herein, we investigated the role of APE1/Ref-1 in atherosclerotic apolipoprotein E (ApoE-/-) mice fed with a Western-type diet. We found that serologic APE1/Ref-1 was strongly correlated with vascular inflammation in these mice. Neutrophil/lymphocyte ratio (NLR), endothelial cell/macrophage activation, and atherosclerotic plaque formation, reflected by atherosclerotic inflammation, were increased in the ApoE-/- mice fed with a Western-type diet. APE1/Ref-1 expression was upregulated in aortic tissues of these mice, and was co-localized with cells positive for cluster of differentiation 31 (CD31) and galectin-3, suggesting endothelial cell/macrophage expression of APE1/Ref-1. Interestingly, APE1/Ref-1 plasma levels of ApoE-/- mice fed with a Western-type diet were significantly increased compared with those of the mice fed with normal diet (15.76 ± 3.19 ng/mL vs. 3.51 ± 0.50 ng/mL, p < 0.05), and were suppressed by atorvastatin administration. Correlation analysis showed high correlation between plasma APE1/Ref-1 levels and NLR, a marker of systemic inflammation. The cut-off value for APE1/Ref-1 for predicting atherosclerotic inflammation at 4.903 ng/mL showed sensitivity of 100% and specificity of 91%. We conclude that APE1/Ref-1 expression is upregulated in aortic endothelial cells/macrophages of atherosclerotic mice, and that plasma APE1/Ref-1 levels could predict atherosclerotic inflammation.

2.
Free Radic Biol Med ; 139: 16-23, 2019 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-31100475

RESUMO

Apurinic/apyrimidinic endonuclease/redox factor-1 (Ref-1), a multifunctional protein secreted from stimulated cells, has been identified as a new serological biomarker. Despite recent reports on the role of Ref-1 in inflammation, the biological function of secreted Ref-1 remains unknown, especially in vivo. This study aimed to evaluate the possible roles of secreted Ref-1 in lipopolysaccharide-induced systemic inflammation in vivo. We generated a secretory Ref-1 adenoviral vector system, AdPPT-LS-Ref-1, by conjugation of preprotrypsin leading sequence (PPT-LS) with full-length Ref-1 sequences. Expression of tumor necrosis factor-α (TNF-α)-induced vascular cell adhesion molecule-1 (VCAM-1) in endothelial cells and lipopolysaccharide (LPS)-induced cyclooxygenase-2 in Raw264.7 cells was inhibited by secretory Ref-1, and this inhibitory effect was abrogated following neutralization of Ref-1 with anti-Ref-1 antibody. Plasma Ref-1 levels following administration of AdPPT-LS-Ref-1 (2 × 109 ifu, i.p.) for 24 h were substantially higher than those recorded following administration of Adßgal (84.6 ±â€¯7.2 ng/ml vs. 4.4 ±â€¯1.5 ng/ml). Treatment with LPS (10 mg/kg, i.v. for 6 h) markedly increased VCAM-1 expression, cathepsin or myeloperoxidase activity, which were significantly suppressed by treatment with AdPPT-LS-Ref-1. Furthermore, LPS-induced cytokines, such as TNF-α, interleukin (IL)-1ß, IL-6, and monocyte chemoattractant protein 1, were significantly inhibited in AdPPT-LS-Ref-1-treated mice. However, LPS-induced myeloperoxidase activities were not suppressed by treatment with the redox mutant of secretory Ref-1, AdPPT-LS-Ref-1(C65A/C93A), or wild-type AdRef-1. Collectively, these results suggest that secreted Ref-1 has anti-inflammatory properties and that its redox cysteine residue is associated with the anti-inflammatory activity in vivo. Furthermore, our findings indicate that secretory Ref-1 may be useful as a therapeutic biomolecule against systemic inflammation.


Assuntos
Anti-Inflamatórios não Esteroides/farmacologia , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/genética , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/farmacologia , Lipopolissacarídeos/antagonistas & inibidores , Sepse/terapia , Adenoviridae/genética , Adenoviridae/metabolismo , Animais , Anti-Inflamatórios não Esteroides/metabolismo , Catepsinas/genética , Catepsinas/metabolismo , Quimiocina CCL2/genética , Quimiocina CCL2/metabolismo , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/antagonistas & inibidores , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/metabolismo , Regulação da Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/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 , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Interleucina-6/genética , Interleucina-6/metabolismo , Lipopolissacarídeos/administração & dosagem , Masculino , Camundongos , Camundongos Endogâmicos ICR , Peroxidase/genética , Peroxidase/metabolismo , Precursores de Proteínas/genética , Precursores de Proteínas/metabolismo , Células RAW 264.7 , Sepse/induzido quimicamente , Sepse/genética , Sepse/patologia , Tripsina/genética , Tripsina/metabolismo , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Molécula 1 de Adesão de Célula Vascular/genética , Molécula 1 de Adesão de Célula Vascular/metabolismo
3.
Sci Rep ; 8(1): 8701, 2018 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-29880821

RESUMO

Triple-negative breast cancer (TNBC) represents a relatively small proportion of all BCs but a relatively large proportion of BC-related death. Thus, more effective therapeutic strategies are needed for the management of TNBC. We demonstrated that the stimulation of apoptosis by the binding of secreted acetylated-apurinic apyrimidinic endonuclease 1/redox factor-1 (Ac-APE1/Ref-1) to the receptor for advanced glycation end products (RAGE) was essential for TNBC cell death in response to hyperacetylation. The aim of the present study was to assess the potential therapeutic efficacy of secretory Ac-APE1/Ref-1 in orthotopic TNBC xenografts in vivo. We found that hyperacetylation in xenografts caused secretion of Ac-APE1/Ref-1 into the blood, where the factor bound directly to RAGE in hyperacetylated tumor tissues. Hyperacetylation in the TNBC xenografts induced strong inhibition of tumor growth and development, leading to apoptotic cell death, accompanied by increased RAGE expression and generation of reactive oxygen species. Tissues exhibited markedly higher counts of apoptotic bodies, a reduced proliferation index, and reduced neovascularization compared with control tumors. Ac-APE1/Ref-1-stimulated apoptosis was markedly reduced in RAGE-knockdown tumors compared with RAGE-overexpressing tumors, even in the presence of hyperacetylation. The function of secreted Ac-APE1/Ref-1 was confirmed in other hyperacetylated TNBCs xenografts using BT-549 and MDA-MB-468 cells, demonstrating its relevance as an anti-cancer molecule.


Assuntos
Apoptose , Proliferação de Células , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/metabolismo , Neoplasias de Mama Triplo Negativas/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Acetilação , Animais , Linhagem Celular Tumoral , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/genética , Feminino , Xenoenxertos , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Transplante de Neoplasias , Receptor para Produtos Finais de Glicação Avançada/genética , Receptor para Produtos Finais de Glicação Avançada/metabolismo , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/patologia , Proteínas Supressoras de Tumor/genética
4.
Int J Mol Sci ; 19(3)2018 Mar 12.
Artigo em Inglês | MEDLINE | ID: mdl-29534512

RESUMO

Anthocyanins, the most prevalent flavonoids in red/purple fruits and vegetables, are known to improve immune responses and reduce chronic disease risks. In this study, the anti-inflammatory activities of an anthocyanin-rich extract from red Chinese cabbage (ArCC) were shown based on its inhibitory effects in cultured endothelial cells and hyperlipidemic apolipoprotein E-deficient mice. ArCC treatment suppressed monocyte adhesion to tumor necrosis factor-α-stimulated endothelial cells. This was validated by ArCC's ability to downregulate the expression and transcription of endothelial adhesion molecules, determined by immunoblot and luciferase promoter assays, respectively. The regulation of adhesion molecules was accompanied by transcriptional inhibition of nuclear factor-κB, which restricted cytoplasmic localization as shown by immunocytochemistry. Administration of ArCC (150 or 300 mg/kg/day) inhibited aortic inflammation in hyperlipidemic apolipoprotein E-deficient mice, as shown by in vivo imaging. Immunohistochemistry and plasma analysis showed that the aortas from these mice exhibited markedly lower leukocyte infiltration, reduced plaque formation, and lower concentrations of blood inflammatory cytokines than those observed in the control mice. The results suggest that the consumption of anthocyanin-rich red Chinese cabbage is closely correlated with lowering the risk of vascular inflammatory diseases.


Assuntos
Antocianinas/análise , Apolipoproteínas E/genética , Aterosclerose/tratamento farmacológico , Brassica/química , Endotélio Vascular/efeitos dos fármacos , Extratos Vegetais/farmacologia , Animais , Aorta/efeitos dos fármacos , Aorta/metabolismo , Aorta/patologia , Aterosclerose/metabolismo , Aterosclerose/patologia , Linhagem Celular Tumoral , Citocinas/sangue , Endotélio Vascular/metabolismo , Células HEK293 , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Masculino , Camundongos , Extratos Vegetais/química , Extratos Vegetais/uso terapêutico , Molécula 1 de Adesão de Célula Vascular/genética , Molécula 1 de Adesão de Célula Vascular/metabolismo
5.
Int J Mol Sci ; 18(10)2017 Sep 25.
Artigo em Inglês | MEDLINE | ID: mdl-28946662

RESUMO

Vascular calcification plays a role in the pathogenesis of atherosclerosis, diabetes, and chronic kidney disease; however, the role of apurinic/apyrimidinic endonuclease 1/redox factor-1 (APE1/Ref-1) in inorganic phosphate (Pi)-induced vascular smooth muscle cell (VSMC) calcification remains unknown. In this study, we investigated the possible role of APE1/Ref-1 in Pi-induced VSMC calcification. We observed that Pi decreased endogenous APE1/Ref-1 expression and promoter activity in VSMCs, and that adenoviral overexpression of APE1/Ref-1 inhibited Pi-induced calcification in VSMCs and in an ex vivo organ culture of a rat aorta. However, a redox mutant of APE1/Ref-1(C65A/C93A) did not reduce Pi-induced calcification in VSMCs, suggesting APE1/Ref-1-mediated redox function against vascular calcification. Additionally, APE1/Ref-1 overexpression inhibited Pi-induced intracellular and mitochondrial reactive oxygen species production, and APE1/Ref-1 overexpression resulted in decreased Pi-induced lactate dehydrogenase activity, pro-apoptotic Bax levels, and increased anti-apoptotic Bcl-2 protein levels. Furthermore, APE1/Ref-1 inhibited Pi-induced osteoblastic differentiation associated with alkaline phosphatase activity and inhibited Pi-exposure-induced loss of the smooth muscle phenotype. Our findings provided valuable insights into the redox function of APE1/Ref-1 in preventing Pi-induced VSMC calcification by inhibiting oxidative stress and osteoblastic differentiation, resulting in prevention of altered osteoblastic phenotypes in VSMCs.


Assuntos
Calcificação Fisiológica/genética , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/genética , Músculo Liso Vascular/citologia , Miócitos de Músculo Liso/metabolismo , Osteoblastos/metabolismo , Fenótipo , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Calcificação Fisiológica/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/genética , Células Cultivadas , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Inativação Gênica , Masculino , Mitocôndrias/metabolismo , Músculo Liso Vascular/patologia , Mutação , Miócitos de Músculo Liso/efeitos dos fármacos , Osteoblastos/efeitos dos fármacos , Oxirredução , Fosfatos/metabolismo , Fosfatos/farmacologia , Interferência de RNA , Ratos , Espécies Reativas de Oxigênio/metabolismo , Calcificação Vascular/genética , Calcificação Vascular/metabolismo , Calcificação Vascular/patologia
6.
Korean J Physiol Pharmacol ; 21(4): 377-384, 2017 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-28706451

RESUMO

Activation of protein kinase C (PKC) is closely linked with endothelial dysfunction. However, the effect of PKCßII on endothelial dysfunction has not been characterized in cultured endothelial cells. Here, using adenoviral PKCßII gene transfer and pharmacological inhibitors, the role of PKCßII on endothelial dysfucntion was investigated in cultured endothelial cells. Phorbol 12-myristate 13-acetate (PMA) increased reactive oxygen species (ROS), p66shc phosphorylation, intracellular adhesion molecule-1, and monocyte adhesion, which were inhibited by PKCßi (10 nM), a selective inhibitor of PKCßII. PMA increased the phosphorylation of CREB and manganese superoxide dismutase (MnSOD), which were also inhibited by PKCßi. Gene silencing of CREB inhibited PMA-induced MnSOD expression, suggesting that CREB plays a key role in MnSOD expression. Gene silencing of PKCßII inhibited PMA-induced mitochondrial ROS, MnSOD, and ICAM-1 expression. In contrast, overexpression of PKCßII using adenoviral PKCßII increased mitochondrial ROS, MnSOD, ICAM-1, and p66shc phosphorylation in cultured endothelial cells. Finally, PKCßII-induced ICAM-1 expression was inhibited by Mito-TEMPO, a mitochondrial ROS scavenger, suggesting the involvement of mitochondrial ROS in PKC-induced vascular inflammation. Taken together, the results suggest that PKCßII plays an important role in PMA-induced endothelial dysfunction, and that the inhibition of PKCßII-dependent p66shc signaling acts as a therapeutic target for vascular inflammatory diseases.

7.
J Med Food ; 20(5): 511-518, 2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28504909

RESUMO

Brassica rapa L. ssp. pekinensis, commonly known as Chinese cabbage, is a cruciferous vegetable traditionally consumed in east Asia. Although its habitual consumption could account for the low incidence of chronic vascular inflammation, the therapeutic and protective potential of phytochemicals derived from Chinese cabbage has been poorly studied. In this study, we identified the phenolic compounds, kaempferol and quercetin, from the ethanol extract of Chinese cabbage (EtCC). We show for the first time that EtCC contains effective phytochemicals that suppress tumor necrosis factor (TNF)-α-induced inflammatory response in human umbilical vein endothelial cells. The EtCC inhibited TNF-α-induced monocyte adhesion to endothelial cells in a dose-dependent manner. The antiadhesive activity of EtCC directly correlated with downregulation of expression and transcription of vascular cell adhesion molecule-1 (VCAM-1). It was caused by an Nrf-2-dependent mechanism, leading to activation of antioxidant responsive element-driven promoter. Taken together, these results suggest that EtCC inhibits the expression of TNF-α-induced adhesion molecules through the indirect transcriptional modulation of VCAM-1 in endothelial cells. In conclusion, regular consumption of vegetables containing dietary phytochemicals might be a potential therapeutic strategy to protect against various stresses, to prevent several pathological conditions, and to treat chronic vascular inflammation, such as atherosclerosis.


Assuntos
Brassica rapa/química , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/imunologia , Extratos Vegetais/farmacologia , Fator de Necrose Tumoral alfa/imunologia , Humanos , Molécula 1 de Adesão Intercelular/genética , Molécula 1 de Adesão Intercelular/imunologia , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Extratos Vegetais/isolamento & purificação , Fator de Necrose Tumoral alfa/genética , Molécula 1 de Adesão de Célula Vascular/genética , Molécula 1 de Adesão de Célula Vascular/imunologia
8.
Sci Rep ; 6: 23015, 2016 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-26964514

RESUMO

Apurinic apyrimidinic endonuclease 1/Redox factor-1 (APE1/Ref-1) is a multifunctional protein with redox activity and is proved to be secreted from stimulated cells. The aim of this study was to evaluate the functions of extracellular APE1/Ref-1 with respect to leading anti-inflammatory signaling in TNF-α-stimulated endothelial cells in response to acetylation. Treatment of TNF-α-stimulated endothelial cells with an inhibitor of deacetylase that causes intracellular acetylation, considerably suppressed vascular cell adhesion molecule-1 (VCAM-1). During TSA-mediated acetylation in culture, a time-dependent increase in secreted APE1/Ref-1 was confirmed. The acetyl moiety of acetylated-APE1/Ref-1 was rapidly removed based on the removal kinetics. Additionally, recombinant human (rh) APE1/Ref-1 with reducing activity induced a conformational change in rh TNF-α receptor 1 (TNFR1) by thiol-disulfide exchange. Following treatment with the neutralizing anti-APE1/Ref-1 antibody, inflammatory signals via the binding of TNF-α to TNFR1 were remarkably recovered, leading to up-regulation of reactive oxygen species generation and VCAM-1, in accordance with the activation of p66(shc) and p38 MAPK. These results strongly indicate that anti-inflammatory effects in TNF-α-stimulated endothelial cells by acetylation are tightly linked to secreted APE1/Ref-1, which inhibits TNF-α binding to TNFR1 by reductive conformational change, with suggestion as an endogenous inhibitor of vascular inflammation.


Assuntos
DNA Liase (Sítios Apurínicos ou Apirimidínicos)/genética , Inflamação/genética , Receptores Tipo I de Fatores de Necrose Tumoral/genética , Fator de Necrose Tumoral alfa/metabolismo , Acetilação , Anti-Inflamatórios/administração & dosagem , Citoplasma/efeitos dos fármacos , Citoplasma/genética , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/química , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/metabolismo , Células Endoteliais/efeitos dos fármacos , Inibidores de Histona Desacetilases/administração & dosagem , Humanos , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Inflamação/patologia , Oxirredução , Ligação Proteica , Conformação Proteica/efeitos dos fármacos , Receptores Tipo I de Fatores de Necrose Tumoral/química , Receptores Tipo I de Fatores de Necrose Tumoral/metabolismo , Proteínas Repressoras/genética , Fator de Necrose Tumoral alfa/administração & dosagem , Molécula 1 de Adesão de Célula Vascular/genética , Proteínas Quinases p38 Ativadas por Mitógeno/genética
9.
Integr Med Res ; 5(2): 131-139, 2016 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-28462108

RESUMO

BACKGROUND: Ulmus davidiana var. japonica Rehder (UD) has long been used in traditional folk medicine in Asia. This study is designed to investigate the antiadhesive activity of the ethanol extract of UD (UDE) and its underlying mechanisms in cultured endothelial cells. METHODS: The dried root bark of UD was extracted with 80% (v/v) ethanol. The antiadhesive activity of the UDE was investigated in cultured human umbilical vein endothelial cells and human embryonic kidney epithelial 293T (HEK 293T) cells stably transfected with pGL3-vascular cell adhesion molecule (VCAM)-1-luc. Monocyte adhesion in endothelial cells was induced by tumor necrosis factor-alpha (TNF-α), and the protective effects of UDE on monocyte-endothelial cell adhesion, VCAM-1 expression, reactive oxygen species production, and nuclear factor-κB activity were determined. RESULTS: Exposure to UDE at a concentration of 3-30 µg/mL for 24 hours produced no detectable cytotoxicity in human umbilical vein endothelial cells, but it significantly inhibited TNF-α-induced monocyte adhesion and VCAM-1 expression. TNF-α treatment of HEK 293T/VCAM-1-luc cells resulted in increased luciferase activity of the VCAM-1 promoter, which was inhibited by treatment with UDE. Additionally, TNF-α-induced reactive oxygen species generation, nuclear translocation of nuclear factor-κB, and IκBα degradation in human umbilical vein endothelial cells were effectively reduced by treatment with 30 µg/mL of UDE. CONCLUSION: Our results indicated that UDE treatment inhibited TNF-α-induced monocyte adhesion in endothelial cells, suggesting that UD may reduce vascular endothelial inflammation.

10.
Korean J Physiol Pharmacol ; 19(5): 467-72, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26330760

RESUMO

Histone deacetylase (HDAC) has been recognized as a potentially useful therapeutic target for cardiovascular disorders. However, the effect of the HDAC inhibitor, trichostatin A (TSA), on vasoreactivity and hypertension remains unknown. We performed aortic coarctation at the inter-renal level in rats in order to create a hypertensive rat model. Hypertension induced by abdominal aortic coarctation was significantly suppressed by chronic treatment with TSA (0.5 mg/kg/day for 7 days). Nicotinamide adenine dinucleotide phosphate-driven reactive oxygen species production was also reduced in the aortas of TSA-treated aortic coarctation rats. The vasoconstriction induced by angiotensin II (Ang II, 100 nM) was inhibited by TSA in both endothelium-intact and endothelium-denuded rat aortas, suggesting that TSA has mainly acted in vascular smooth muscle cells (VSMCs). In cultured rat aortic VSMCs, Ang II increased p66shc phosphorylation, which was inhibited by the Ang II receptor type I (AT1R) inhibitor, valsartan (10 µM), but not by the AT2R inhibitor, PD123319. TSA (1~10 µM) inhibited Ang II-induced p66shc phosphorylation in VSMCs and in HEK293T cells expressing AT1R. Taken together, these results suggest that TSA treatment inhibited vasoconstriction and hypertension via inhibition of Ang II-induced phosphorylation of p66shc through AT1R.

11.
Cancer Res Treat ; 47(4): 823-33, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-25672588

RESUMO

PURPOSE: Apurinic/apyrimidinic endonuclease 1/redox factor-1 (APE1/Ref-1) is a multifunctional protein that shows elevated expression in a number of cancers. We attempted to determine whether serum APE1/Ref-1 is elevated in patients with bladder cancer. MATERIALS AND METHODS: Serum APE1/Ref-1 levels were determined using enzyme-linked immunosorbent assay in serum from patients with bladder cancer who had not received chemotherapy or radiotherapy (n=51) and non-tumor controls (n=55). The area under the receiver operating characteristic area under the curve was applied to determine the correlation between clinical factors and the serum levels of APE1/Ref-1. RESULTS: Serum levels of APE1/Ref-1 in bladder cancer patients were significantly elevated compared to those of the control group (3.548 ± 0.333 ng/100 µL [n=51] for bladder cancer vs. 1.547 ± 0.319 ng/100 µL [n=55] for the control group), with a sensitivity and specificity of 93% and 59%, respectively. Serum APE1/Ref-1 levels are associated with tumor stage, grade, muscle invasion, and recurrence. CONCLUSION: Serum APE1/Ref-1 might be useful as a potential serologic biomarker for bladder cancer.


Assuntos
Biomarcadores Tumorais/sangue , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/sangue , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/metabolismo , Neoplasias da Bexiga Urinária/sangue , Neoplasias da Bexiga Urinária/diagnóstico , Idoso , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/genética , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Sensibilidade e Especificidade , Testes Sorológicos
12.
Mitochondrion ; 17: 42-9, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24861944

RESUMO

Protein kinase C (PKC) induces mitochondrial dysfunction, which is an important pathological factor in cardiovascular diseases. The role of apurinic/apyrimidinic endonuclease-1/redox factor-1 (APE1/Ref-1) on PKC-induced mitochondrial dysfunction has not been variously investigated. In this study, phorbol 12-myristate 13-acetate (PMA), an activator of protein kinase C, induced mitochondrial hyperpolarization and reactive oxygen species generation and also increased mitochondrial translocation of APE1/Ref-1. APE1/Ref-1 overexpression suppressed PMA-induced mitochondrial dysfunction. In contrast, gene silencing of APE1/Ref-1 increased the sensitivity of mitochondrial dysfunction. Moreover, mitochondrial targeting sequence (MTS)-fused APE1/Ref-1 more effectively suppressed PMA-induced mitochondrial dysfunctions. These results suggest that mitochondrial APE1/Ref-1 is contributed to the protective role to protein kinase C-induced mitochondrial dysfunction in endothelial cells.


Assuntos
DNA Liase (Sítios Apurínicos ou Apirimidínicos)/metabolismo , Células Endoteliais/fisiologia , Mitocôndrias/fisiologia , Proteína Quinase C/metabolismo , Animais , Potencial da Membrana Mitocondrial , Camundongos , Espécies Reativas de Oxigênio/metabolismo
13.
Mol Cells ; 36(5): 439-45, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-24213673

RESUMO

Apurinic/apyrimidinic endonuclease1/redox factor-1 (APE1/Ref-1) is a multifunctional protein involved in base excision DNA repair and transcriptional regulation of gene expression. APE1/Ref-1 is mainly localized in the nucleus, but cytoplasmic localization has also been reported. However, the functional role of cytoplasmic APE1/Ref-1 and its redox cysteine residue are still unknown. We investigated the role of cytoplasmic APE1/Ref-1 on tumor necrosis factor-α (TNF-α)-induced vascular cell adhesion molecule-1 (VCAM-1) expressions in endothelial cells. Endogenous APE1/Ref-1 was mainly observed in the nucleus, however, cytoplasmic APE1/Ref-1 was increased by TNF-α. Cytoplasmic APE1/Ref-1 expression was not blunted by cycloheximide, a protein synthesis inhibitor, suggesting cytoplasmic translocation of APE1/Ref-1. Transfection of an N-terminus deletion mutant APE1/Ref-1(29-318) inhibited TNF-α-induced VCAM-1 expression, indicating an anti-inflammatory role for APE1/Ref-1 in the cytoplasm. In contrast, redox mutant of APE1/Ref-1 (C65A/C93A) transfection led to increased TNF-α-induced VCAM-1 expression. Our findings suggest cytoplasmic APE1/Ref-1 localization and redox cysteine residues of APE1/Ref-1 are associated with its anti-inflammatory activity in endothelial cells.


Assuntos
Anti-Inflamatórios/metabolismo , Cisteína/metabolismo , Citoplasma/metabolismo , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Molécula 1 de Adesão de Célula Vascular/metabolismo , Núcleo Celular/genética , Núcleo Celular/metabolismo , Cicloeximida/farmacologia , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/genética , Regulação da Expressão Gênica , Células Endoteliais da Veia Umbilical Humana , Humanos , Mutação , Oxirredução , Espécies Reativas de Oxigênio/metabolismo , Transfecção
14.
Korean Circ J ; 43(5): 340-2, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23755081

RESUMO

We describe a 64-year-old male patient with panhypopituitarism who experienced polymorphic ventricular tachycardia (VT) associated with long QT intervals. The panhypopituitarism developed as a sequelae of radiation therapy administered 20 years prior to his current presentation and was recently aggravated by urinary tract infection with sepsis. In this case, polymorphic VT was resistant to conventional therapy (including magnesium infusion), and QT prolongation and T wave inversion were normalized after the administration of steroid and thyroid hormones. Thyroid hormone is generally known to be associated with torsades de pointes (TdP), but steroid or other hormones may also provoke TdP. Hormonal disorders should be considered as a cause of polymorphic VT with long QT intervals. Some arrhythmias can be life-threatening, and they can be prevented with supplementation of the insufficient hormone.

15.
Biochem Biophys Res Commun ; 435(3): 403-7, 2013 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-23665318

RESUMO

Apurinic/apyrimidinic endonuclease 1/Redox factor-1 (APE1/Ref-1) can be acetylated via post-translational modification. We investigated the effect of an inhibitor of histone deacetylases on the extracellular release of APE1/Ref-1 in HEK293 cells. Trichostatin A (TSA), an inhibitor of histone deacetylases, induced APE1/Ref-1 secretion without changing cell viability. In a fluorescence quantitative assay, the secreted APE1/Ref-1 was estimated to be about 10 ng/mL in response to TSA (1 µM). However, TSA did not induce the secretion of lysine-mutated APE1/Ref-1 (K6R/K7R). TSA also caused nuclear to cytoplasmic translocation of APE1/Ref-1. Taken together, these findings suggest that APE1/Ref-1 is a protein whose secretion is governed by lysine acetylation.


Assuntos
DNA Liase (Sítios Apurínicos ou Apirimidínicos)/metabolismo , Inibidores de Histona Desacetilases/farmacologia , Ácidos Hidroxâmicos/farmacologia , Acetilação/efeitos dos fármacos , Transporte Ativo do Núcleo Celular/efeitos dos fármacos , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/química , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/genética , Espaço Extracelular/enzimologia , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Células HEK293 , Humanos , Lisina/química , Mutagênese Sítio-Dirigida , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo
16.
Biochem Biophys Res Commun ; 435(4): 621-6, 2013 Jun 14.
Artigo em Inglês | MEDLINE | ID: mdl-23685156

RESUMO

Apurinic/apyrimidinic endonuclease1/Redox factor-1 (APE1/Ref-1) is a multifunctional protein involved in base excision DNA repair and in transcriptional regulation of gene expression. We investigated whether APE1/Ref-1 increased in plasma of endotoxemic rats. Lipopolysaccharide (LPS) was used to induce endotoxemia in rats. Administration of LPS (10 mg/kg, i.p.) significantly induced plasma nitrite production and tumor necrosis factor-α (TNF-α). A 37 kDa immunoreactive band was detected in cell-free plasma of LPS-treated rats using anti-APE1/Ref-1, which reached a maximum at 12 h after the LPS injection. The 37 kDa immunoreactive band was identified as rat APE1/Ref-1 by liquid chromatography/tandem mass spectrometry. Interestingly, treatment with recombinant human APE1/Ref-1 protein (2-5 µg/ml for 18 h) inhibited TNF-α-induced vascular cell adhesion molecule-1 expression in human umbilical vein endothelial cells. Taken together, the level of plasma APE1/Ref-1 increased in LPS-induced endotoxemic rats, suggesting that plasma APE1/Ref-1 might serve as a serological biomarker for endotoxemia.


Assuntos
DNA Liase (Sítios Apurínicos ou Apirimidínicos)/sangue , Endotoxemia/sangue , Animais , Biomarcadores/sangue , Endotoxemia/induzido quimicamente , Endotoxemia/diagnóstico , Lipopolissacarídeos , Masculino , Ratos , Ratos Sprague-Dawley
17.
Nutr Res Pract ; 7(1): 9-14, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-23422838

RESUMO

Bamboo leaves (Phyllostachys pubescens Mazel ex J. Houz (Poacea)) have a long history of food and medical applications in Asia, including Japan and Korea. They have been used as a traditional medicine for centuries. We investigated the mechanism of anti-inflammatory activity of a bamboo leaf extract (BLE) on tumor necrosis factor-alpha (TNF-α)-induced monocyte adhesion in human umbilical vein endothelial cells (HUVECs). Exposure of HUVECs to BLE did not inhibit cell viability or cause morphological changes at concentrations ranging from 1 µg/ml to 1 mg/ml. Treatment with 0.1 mg/ml BLE caused 63% inhibition of monocyte adhesion in TNF-α-activated HUVECs, which was associated with 38.4% suppression of vascular cell adhesion molecule-1 expression. Furthermore, TNF-α-induced reactive oxygen species generation was decreased to 47.9% in BLE treated TNF-α-activated HUVECs. BLE (0.05 mg/ml) also caused about 50% inhibition of interleukin-6 secretion from lipopolysaccharide-stimulated monocyte. The results indicate that BLE may be clinically useful as an anti-inflammatory or anti-oxidant for human cardiovascular disease including atherosclerosis.

18.
FEBS Lett ; 586(9): 1349-55, 2012 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-22616995

RESUMO

Peripheral benzodiazepine receptor (PBR) is a multifunctional protein mainly found on the outer mitochondrial membrane. PBR expression is increased by tumor necrosis factor-α (TNF-α) in endothelial cells. Adenoviral overexpression of PBR inhibits monocyte adhesion, VCAM-1, and ICAM-1 expression in TNF-α-activated endothelial cells. Rotenone, cyclosporine A, and bongkrekic acid suppress TNF-α-induced VCAM-1 expression. Overexpression of PBR inhibits voltage-dependent anion channel-1 (VDAC-1) expression and the silencing of PBR increases VDAC-1 expression in endothelial cells. Moreover, TNF-α-induced VCAM-1 expression is suppressed by VDAC-1 gene silencing. PBR overexpression significantly decreases TNF-α-induced mitochondrial reactive oxygen species and MnSOD expression. These results suggest that PBR can inhibit endothelial activation and this action is related to the inhibition of mitochondrial ROS and/or VDAC-1 expression in endothelial cells.


Assuntos
Regulação para Baixo , Células Endoteliais/metabolismo , Receptores de GABA/metabolismo , Canal de Ânion 1 Dependente de Voltagem/metabolismo , Adenoviridae/genética , Adesão Celular/efeitos dos fármacos , Citocinas/metabolismo , Regulação para Baixo/efeitos dos fármacos , Células Endoteliais/citologia , Células Endoteliais/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/citologia , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Molécula 1 de Adesão Intercelular/metabolismo , Mitocôndrias/metabolismo , Monócitos/citologia , Monócitos/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Receptores de GABA/genética , Superóxido Dismutase/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Regulação para Cima/efeitos dos fármacos , Molécula 1 de Adesão de Célula Vascular/metabolismo , Canal de Ânion 1 Dependente de Voltagem/genética
19.
Cardiovasc Res ; 91(3): 502-9, 2011 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-21467074

RESUMO

AIMS: Phosphorylation of the adaptor protein p66shc is essential for p66shc-mediated oxidative stress. We investigated the role of the reducing protein/DNA repair enzyme apurinic/apyrimidinic endonuclease1 (APE1) in modulating protein kinase CßII (PKCßII)-mediated p66shc phosphorylation in cultured endothelial cells and PKC-mediated vasoconstriction of arteries. METHODS AND RESULTS: Oxidized low-density lipoprotein (oxLDL)induced p66shc phosphorylation at serine 36 residue and PKCßII phosphorylation in mouse endothelial cells. Adenoviral overexpression of APE1 resulted in reduction of oxLDL-induced p66shc and PKCßII phosphorylation. Phorbol 12-myristate 13-acetate (PMA), which stimulates PKCs, induced p66shc phosphorylation and this was inhibited by a selective PKCßII inhibitor. Adenoviral overexpression of PKCßII also increased p66shc phosphorylation. Overexpression of APE1 suppressed PMA-induced p66shc phosphorylation. Moreover, PMA-induced p66shc phosphorylation was augmented in cells in which APE1 was knocked down. PMA increased cytoplasmic APE1 expression, compared with the basal condition, suggesting the role of cytoplasmic APE1 against p66shc phosphorylation. Finally, vasoconstriction induced by phorbol-12,13, dibutylrate, another PKC agonist, was partially inhibited by transduction of Tat-APE1 into arteries. CONCLUSION: APE1 suppresses oxLDL-induced p66shc activation in endothelial cells by inhibiting PKCßII-mediated serine phosphorylation of p66shc, and mitigates vasoconstriction induced by activation of PKC.


Assuntos
DNA Liase (Sítios Apurínicos ou Apirimidínicos)/metabolismo , Células Endoteliais/enzimologia , Proteína Quinase C/metabolismo , Proteínas Adaptadoras da Sinalização Shc/metabolismo , Vasoconstrição , Animais , Células Cultivadas , DNA Liase (Sítios Apurínicos ou Apirimidínicos)/genética , Células Endoteliais/efeitos dos fármacos , Ativação Enzimática , Ativadores de Enzimas/farmacologia , Humanos , Lipoproteínas LDL/metabolismo , Camundongos , Fosforilação , Proteína Quinase C/antagonistas & inibidores , Proteína Quinase C/genética , Proteína Quinase C beta , Inibidores de Proteínas Quinases/farmacologia , Transporte Proteico , Ratos , Ratos Sprague-Dawley , Proteínas Recombinantes/metabolismo , Transdução de Sinais , Proteína 1 de Transformação que Contém Domínio 2 de Homologia de Src , Fatores de Tempo , Transdução Genética , Transfecção , Vasoconstrição/efeitos dos fármacos
20.
Korean J Physiol Pharmacol ; 15(6): 339-44, 2011 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-22359471

RESUMO

Ulmus davidiana var. japonica Rehder (Urticales: Ulmaceae) (UD) is a tree widespread in northeast Asia. It is traditionally used for anticancer and anti-inflammatory therapy. The present study investigated the effect of an ethanol extract of UD on vascular tension and its underlying mechanism in rats. The dried root bark of UD was ground and extracted with 80% ethanol. The prepared UD extract was used in further analysis. The effect of UD on the cell viability, vasoreactivity and hemodynamics were investigated using propidium iodide staining in cultured cells, isometric tension recording and blood pressure analysis, respectively. Low dose of UD (10~100µg/ml) did not affect endothelial cell viability, but high dose of UD reduced cell viability. UD induced vasorelaxation in the range of 0.1~10µg/ml with an ED(50) value of 2µg/ml. UD-induced vasorelaxation was completely abolished by removal of the endothelium or by pre-treatment with L-NAME, an inhibitor of nitric oxide synthase. UD inhibited calcium influx induced by phenylephrine and high K(+) and also completely abolished the effect of L-NAME. Intravenous injection of UD extracts (10~100 mg/kg) decreased arterial and ventricular pressure in a dose-dependent manner. Moreover, UD extracts reduced the ventricular contractility (+dP/dt) in anesthetized rats. However, UD-induced hypotensive actions were minimized in L-NAME-treated rats. Taken together, out results showed that UD induced vasorelaxation and has antihypertensive properties, which may be due the activation of nitric oxide synthase in endothelium.

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