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1.
Life Sci ; 235: 116840, 2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31494171

RESUMO

AIMS: Ovarian ischemia as a consequence of torsion constitutes a gynecologic emergency affecting females during reproductive age. Its management by detorsion results in ovarian ischemia-reperfusion (IR) injury. Thus, a conservative treatment with detorsion is highly recommended. Therefore, we attempted to investigate the effect and underlying mechanisms of angiotensin 1-7 (Ang-(1-7)) treatment against ovarian IR injury. MAIN METHODS: Female rats were included into: Sham group; Ang-(1-7) (300 µg/kg, i.p.) group; ovarian IR groups with and without Ang-(1-7) treatment. We determined ovarian Ang-(1-7), malondialdehyde (MDA) and nitric oxide (NO) in addition to serum total anti-oxidant capacity (TAC) levels. Ovarian gene expressions of angiotensin converting enzyme 2 (ACE2), Mas receptor, tumor necrosis factor alpha (TNF-α) and B-cell leukemia/lymphoma-2 (BCL-2) were estimated. Furthermore, histopathological changes and ovarian expressions of nuclear factor kappa B (NF-κB), inducible and endothelial nitric oxide synthases (iNOS and eNOS) were done. KEY FINDINGS: Treatment of ovarian IR rats with Ang-(1-7) led to marked improvement of ovarian damage through histological examination which was accompanied with marked increase in ovarian Ang-(1-7) level and expressions of ACE2 and Mas receptor, decrease in MDA and NO levels and expressions of NF-kB, iNOS and TNF-α with increase in serum TAC levels and ovarian expressions of eNOS and BCL-2. SIGNIFICANCE: Our results proved the protective effect of Ang-(1-7) against ovarian IR injury in rats and this may be attributed to ACE2/Ang (1-7)/Mas axis which showed anti-oxidant, anti-inflammatory and anti-apoptotic effects. Therefore, Ang-(1-7) can be used in the future for treatment of ovarian IR injury.


Assuntos
Angiotensina I/farmacologia , Ovário/efeitos dos fármacos , Fragmentos de Peptídeos/farmacologia , Traumatismo por Reperfusão/prevenção & controle , Animais , Antioxidantes/metabolismo , Feminino , Expressão Gênica/efeitos dos fármacos , Malondialdeído/metabolismo , NF-kappa B/biossíntese , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo II/biossíntese , Óxido Nítrico Sintase Tipo III/biossíntese , Ovário/lesões , Ovário/metabolismo , Peptidil Dipeptidase A/biossíntese , Proteínas Proto-Oncogênicas/biossíntese , Proteínas Proto-Oncogênicas c-bcl-2/biossíntese , Ratos , Receptores Acoplados a Proteínas-G/biossíntese , Soro/metabolismo , Fator de Necrose Tumoral alfa/biossíntese
2.
Biomed Res ; 40(4): 145-152, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31413235

RESUMO

We investigated the mechanisms by which propiverine hydrochloride influenced bladder activity in rats with pelvic venous congestion (PC) and urinary frequency. To create PC rats, female rats were anesthetized with isoflurane and the bilateral common iliac veins and bilateral uterine veins were ligated. At 4 weeks after ligation, we assessed voiding behaviour, locomotor activity, and urinary 8-hydroxydeoxyguanosine (8-OHdG) and nitric oxide metabolites (NOx). We also performed cystometry and measured mRNAs for nitric oxide synthase (NOS) and several receptors in the bladder wall. PC rats showed a decrease in locomotor activity and an increased frequency of urination. There was a decrease in endothelial NOS (eNOS), M3, and TRPV1 mRNA expression in the bladder wall, as well as an increase in inducible NOS (iNOS) mRNA. Administration of propiverine to PC rats increased locomotor activity to the level in sham rats, improved bladder function, decreased urinary 8-OHdG excretion, and increased urinary NOx excretion. In addition, propiverine increased neuronal NOS (nNOS) mRNA expression, and decreased expression of iNOS, M3 and TRPV1 mRNA in the bladder wall. Therefore, propiverine not only improved bladder dysfunction through its previously reported actions (anti-muscarinic effect, Ca antagonist effect, and inhibition of noradrenaline re-uptake), but also by reducing inflammation.


Assuntos
Benzilatos/farmacologia , Hiperemia/tratamento farmacológico , Doenças da Bexiga Urinária/tratamento farmacológico , Bexiga Urinária/fisiopatologia , Animais , Modelos Animais de Doenças , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Hiperemia/metabolismo , Hiperemia/patologia , Hiperemia/fisiopatologia , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Inflamação/patologia , Inflamação/fisiopatologia , Óxido Nítrico Sintase Tipo I/biossíntese , Óxido Nítrico Sintase Tipo III/biossíntese , Ratos , Ratos Sprague-Dawley , Canais de Cátion TRPV/biossíntese , Bexiga Urinária/metabolismo , Bexiga Urinária/patologia , Doenças da Bexiga Urinária/metabolismo , Doenças da Bexiga Urinária/patologia , Doenças da Bexiga Urinária/fisiopatologia
3.
Environ Pollut ; 252(Pt A): 317-329, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31158660

RESUMO

Fine dust (FD) is a form of air pollution and is responsible for a wide range of diseases. Specially, FD is associated with several cardiovascular diseases (CVDs); long-term exposure to FD was shown to decrease endothelial function, but the underlying mechanism remains unclear. We investigated whether exposure to FD causes premature senescence-associated endothelial dysfunction in endothelial cells (ECs) isolated from porcine coronary arteries. The cells were treated with different concentrations of FD and senescence associated-beta galactosidase (SA-ß-gal) activity, cell cycle progression, expression of endothelial nitric oxide synthase (eNOS), oxidative stress level, and vascular function were evaluated. We found that FD increased SA-ß-gal activity, caused cell cycle arrest, and increased oxidative stress, suggesting the premature induction of senescence; on the other hand, eNOS expression was downregulated and platelet aggregation was enhanced. FD exposure impaired vasorelaxation in response to bradykinin and activated the local angiotensin system (LAS), which was inhibited by treatment with the antioxidant N-acetyl cysteine (NAC) and angiotensin II receptor type 1 (AT1) antagonist losartan (LOS). NAC and LOS also suppressed FD-induced SA-ß-gal activity, increased EC proliferation and eNOS expression, and improved endothelial function. These results demonstrate that FD induces premature senescence of ECs and is associated with increased oxidative stress and activation of LAS. This study can serve as a pharmacological target for prevention and/or treatment of air pollution-associated CVD.


Assuntos
Poluição do Ar/efeitos adversos , Angiotensinas/metabolismo , Senescência Celular/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/fisiologia , Material Particulado/farmacologia , Receptor Tipo 1 de Angiotensina/metabolismo , Acetilcisteína/farmacologia , Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Animais , Antioxidantes/metabolismo , Plaquetas/citologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Vasos Coronários/citologia , Endotélio Vascular/citologia , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Losartan/farmacologia , Óxido Nítrico Sintase Tipo III/biossíntese , Oxirredução , Estresse Oxidativo/efeitos dos fármacos , Suínos , beta-Galactosidase/antagonistas & inibidores , beta-Galactosidase/metabolismo
4.
J Biochem Mol Toxicol ; 33(7): e22332, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-30974023

RESUMO

Alpha-naphthylthiourea (ANTU), a rodenticide induces lung toxicity. Chrysin a flavonoid possesses antioxidant, anti-inflammatory, and antihypertensive potential. The aim of this study was to evaluate the efficacy of chrysin against ANTU-induced pulmonary edema (PE) and pulmonary arterial hypertension (PAH) in laboratory rats. Sprague-Dawley rats were used to induce PE (ANTU, 10 mg/kg, ip) and PAH (ANTU, 5 mg/kg, ip, 4 weeks). Animals were treated with chrysin (10, 20, and 40 mg/kg) and various biochemical, molecular, and histological parameters were evaluated. Acute administration of ANTU induces PE revealed by significant (P < 0.05) increase in relative lung weight, pleural effusion volume, lung edema, bronchoalveolar lavage fluid cell counts, total protein, 5-hydroxytryptamine (5-HT), lactate dehydrogenase (LDH), and γ-glutamyl transferase (GGT), whereas pretreatment with chrysin (20 and 40 mg/kg, ip) significantly (P < 0.05) attenuated these ANTU-induced biochemical and histological alterations. Repeated administration of ANTU caused induction of PAH evaluated by significant (P < 0.05) alterations in electrocardiographic, hemodynamic changes, and left ventricular function, whereas chrysin (20 and 40 mg/kg, p.o.) treatment significantly (P < 0.05) attenuated these alterations. ANTU-induced hematological and serum biochemical (aspartate transaminase, alanine transaminase, LDH, and creatinine kinase MB) alterations were significantly (P < 0.05) inhibited by chrysin. It also significantly (P < 0.05) decreased elevated levels of oxido-nitrosative stress in the right ventricle (RV) and lung. Chrysin significantly (P < 0.05) attenuated downregulated endothelial nitric oxide synthase and upregulated vascular endothelial growth factor messenger RNA and protein expressions both in the RV and pulmonary artery. Chrysin inhibited ANTU-induced PE and PAH via modulation of inflammatory responses (5-HT, LDH, and GGT), oxido-nitrosative stress, and VEGF and eNOs levels.


Assuntos
Flavonoides/farmacologia , Hipertensão Pulmonar , Óxido Nítrico Sintase Tipo III/biossíntese , Edema Pulmonar , Tioureia/análogos & derivados , Fator A de Crescimento do Endotélio Vascular/biossíntese , Animais , Hipertensão Pulmonar/induzido quimicamente , Hipertensão Pulmonar/metabolismo , Hipertensão Pulmonar/patologia , Hipertensão Pulmonar/prevenção & controle , Pulmão/metabolismo , Pulmão/patologia , Masculino , Artéria Pulmonar/metabolismo , Artéria Pulmonar/patologia , Edema Pulmonar/induzido quimicamente , Edema Pulmonar/metabolismo , Edema Pulmonar/patologia , Edema Pulmonar/prevenção & controle , Ratos , Ratos Sprague-Dawley , Tioureia/efeitos adversos , Tioureia/farmacologia
5.
Exp Eye Res ; 181: 150-156, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30716330

RESUMO

Dysfunctional vascular endothelial nitric oxide synthase (eNOS) has been proposed to play a main pathophysiological role in various ocular diseases. The aim of the present study was to test the hypothesis that the chronic lack of eNOS impairs endothelium-dependent vasodilation in retinal arterioles. The relevance of eNOS for mediating vascular responses was studied in retinal vascular preparations from eNOS-deficient mice (eNOS-/-) and wild-type controls in vitro. Changes in luminal diameter in response to vasoactive agents were measured by videomicroscopy. The thromboxane mimetic, U46619, induced similar concentration-dependent constriction of retinal arterioles in eNOS-/- and wild-type mice. Responses to the endothelium-independent vasodilator, nitroprusside, did not differ between both mouse genotypes, either. In contrast, responses to the endothelium-dependent vasodilator, acetylcholine, were blunted in eNOS-/- mice. Non-isoform-selective blockade of either nitric oxide synthase (NOS) or cyclooxygenase (COX) alone did not affect responses to acetylcholine. However, combined blockade of both enzyme families markedly attenuated cholinergic vasodilation. Also, combined blockade of COX and neuronal NOS (nNOS) blunted acetylcholine-induced vasodilation, while combined COX and inducible NOS (iNOS) inhibition had no effect. Simultaneous NOS and COX-1 blockade did not affect cholinergic vasodilation, whereas combined NOS and COX-2 inhibition markedly reduced vasodilation to acetylcholine. These findings are the first to demonstrate that the chronic lack of eNOS is associated with moderate endothelial dysfunction in retinal arterioles. However, eNOS-deficiency is partially compensated by nNOS and COX-2 metabolites, which are reciprocally regulated.


Assuntos
Arteríolas/fisiopatologia , Regulação da Expressão Gênica , Óxido Nítrico Sintase Tipo III/genética , Artéria Retiniana/fisiopatologia , Doenças Retinianas/fisiopatologia , Vasodilatação/fisiologia , Animais , Arteríolas/metabolismo , Modelos Animais de Doenças , Endotélio Vascular/metabolismo , Endotélio Vascular/fisiopatologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Óxido Nítrico Sintase Tipo III/biossíntese , RNA/genética , Artéria Retiniana/metabolismo , Doenças Retinianas/genética , Doenças Retinianas/metabolismo
6.
Am J Physiol Cell Physiol ; 316(4): C481-C491, 2019 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-30673304

RESUMO

The inadequate trophoblast invasion is associated with the development of preeclampsia (PE). Considering that annexin A4 (ANXA4) enhances tumor invasion, we aimed to explore the functional role of ANXA4 in trophoblast cells and to examine the underlying mechanism. ANXA4 expression in PE placentas was analyzed using immunohistochemistry and Western blotting. Cell proliferation, invasion, and apoptosis were determined using a MTT assay, Transwell assay, and flow cytometry, respectively. The expression levels of matrix metalloproteinase (MMP)-2, MMP-9, phosphoinositide 3-kinase (PI3K), Akt, phosphorylated (p)-Akt, and phosphorylated endothelial nitric oxide synthase (p-eNOS) were detected by Western blotting. Placentas were prepared for pathological examination using hematoxylin and eosin staining and apoptosis determination using the TUNEL method. Expression of ANXA4, PI3K, p-Akt and p-eNOS was downregulated in human PE placentas and PE placenta-derived extravillous cytotrophoblasts (EVCTs). Furthermore, ANXA4 overexpression promoted cell proliferation and invasion, inhibited cell apoptosis, and upregulated protein expression of PI3K, p-Akt, and p-eNOS in human trophoblast cells HTR-8/SVneo and JEG-3. By contrast, ANXA4 knockdown exerted the opposite effects. Furthermore, inhibition of the PI3K/Akt pathway by LY294002 abrogated the ANXA4 overexpression-mediated effects on trophoblast behavior. Furthermore, eNOS knockdown abrogated the ANXA4 overexpression-induced promotion of cell invasion and MMP2/9 expression. Additionally, in N-nitro-l-arginine methyl ester (l-NAME)-induced PE rats, ANXA4 overexpression alleviated PE progression, accompanied by an increase in expression of PI3K, p-Akt, and p-eNOS in rat placentas. Our findings demonstrate that ANXA4 expression is downregulated in PE. ANXA4 may promote trophoblast invasion via the PI3K/Akt/eNOS pathway.


Assuntos
Anexina A4/biossíntese , Óxido Nítrico Sintase Tipo III/biossíntese , Fosfatidilinositol 3-Quinases/biossíntese , Pré-Eclâmpsia/metabolismo , Proteínas Proto-Oncogênicas c-akt/biossíntese , Trofoblastos/metabolismo , Animais , Células Cultivadas , Feminino , Humanos , Pré-Eclâmpsia/patologia , Gravidez , Ratos , Ratos Sprague-Dawley , Transdução de Sinais/fisiologia , Trofoblastos/patologia
7.
J Cereb Blood Flow Metab ; 39(2): 332-341, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-28840777

RESUMO

Our goal was to examine whether in utero exposure to alcohol impaired reactivity of cerebral arterioles during development. We fed Sprague-Dawley dams a liquid diet with or without alcohol (3% ethanol) for the duration of pregnancy (21-23 days). Around 4-6 weeks after birth, we examined reactivity of cerebral arterioles to eNOS- (ADP) and nNOS-dependent (NMDA) agonists in the offspring. We found that in utero exposure to alcohol attenuated responses of cerebral arterioles to ADP and NMDA, but not to nitroglycerin in rats exposed to alcohol in utero. L-NMMA reduced responses to agonists in control rats, but not in rats exposed to alcohol in utero. Treatment of dams with apocynin for the duration of pregnancy rescued the impairment in reactivity to ADP and NMDA in the offspring. Protein expression of NOX-2 and NOX-4 was increased in alcohol rats compared to control rats. We also found an increase in superoxide levels in the cortex of rats exposed to alcohol in utero. Our findings suggest that in utero exposure to alcohol impairs eNOS and nNOS reactivity of cerebral arterioles via a chronic increase in oxidative stress.


Assuntos
Arteríolas , Córtex Cerebral , Etanol/efeitos adversos , Exposição Materna/efeitos adversos , Estresse Oxidativo/efeitos dos fármacos , Efeitos Tardios da Exposição Pré-Natal , Acetofenonas/farmacologia , Difosfato de Adenosina/farmacologia , Animais , Arteríolas/metabolismo , Arteríolas/patologia , Arteríolas/fisiopatologia , Córtex Cerebral/irrigação sanguínea , Córtex Cerebral/metabolismo , Córtex Cerebral/patologia , Córtex Cerebral/fisiopatologia , Doença Crônica , Etanol/farmacologia , Feminino , Masculino , NADPH Oxidase 2/biossíntese , NADPH Oxidase 4/biossíntese , Óxido Nítrico Sintase Tipo I/biossíntese , Óxido Nítrico Sintase Tipo III/biossíntese , Gravidez , Efeitos Tardios da Exposição Pré-Natal/metabolismo , Efeitos Tardios da Exposição Pré-Natal/patologia , Efeitos Tardios da Exposição Pré-Natal/fisiopatologia , Ratos , Ratos Sprague-Dawley , Superóxidos/metabolismo , ômega-N-Metilarginina/farmacologia
8.
J Clin Invest ; 129(2): 531-545, 2019 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-30422822

RESUMO

Hutchinson-Gilford progeria syndrome (HGPS) is a premature aging disorder characterized by accelerated cardiovascular disease with extensive fibrosis. It is caused by a mutation in LMNA leading to expression of truncated prelamin A (progerin) in the nucleus. To investigate the contribution of the endothelium to cardiovascular HGPS pathology, we generated an endothelium-specific HGPS mouse model with selective endothelial progerin expression. Transgenic mice develop interstitial myocardial and perivascular fibrosis and left ventricular hypertrophy associated with diastolic dysfunction and premature death. Endothelial cells show impaired shear stress response and reduced levels of endothelial nitric oxide synthase (eNOS) and NO. On the molecular level, progerin impairs nucleocytoskeletal coupling in endothelial cells through changes in mechanoresponsive components at the nuclear envelope, increased F-actin/G-actin ratios, and deregulation of mechanoresponsive myocardin-related transcription factor-A (MRTFA). MRTFA binds to the Nos3 promoter and reduces eNOS expression, thereby mediating a profibrotic paracrine response in fibroblasts. MRTFA inhibition rescues eNOS levels and ameliorates the profibrotic effect of endothelial cells in vitro. Although this murine model lacks the key anatomical feature of vascular smooth muscle cell loss seen in HGPS patients, our data show that progerin-induced impairment of mechanosignaling in endothelial cells contributes to excessive fibrosis and cardiovascular disease in HGPS patients.


Assuntos
Células Endoteliais/metabolismo , Hipertrofia Ventricular Esquerda/metabolismo , Lamina Tipo A/biossíntese , Mecanotransdução Celular , Miocárdio/metabolismo , Elementos de Resposta , Transativadores/metabolismo , Animais , Modelos Animais de Doenças , Células Endoteliais/patologia , Fibrose , Humanos , Hipertrofia Ventricular Esquerda/genética , Hipertrofia Ventricular Esquerda/patologia , Lamina Tipo A/genética , Camundongos , Camundongos Transgênicos , Miocárdio/patologia , Óxido Nítrico/biossíntese , Óxido Nítrico/genética , Óxido Nítrico Sintase Tipo III/biossíntese , Óxido Nítrico Sintase Tipo III/genética , Transativadores/genética
9.
Biomed Pharmacother ; 109: 886-891, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30551542

RESUMO

Piceatannol (3,3',4,5'-trans-trihydroxystilbene) is a natural polyphenols compound that occurs hydroxylated analogue of resveratrol showing widely biological activities. Previous studies have demonstrated its functions on anti-cancer, neuroprotection and cardioprotection. However, few studies have clarified the benefits of piceatannol on cardiomyocytes except its anti-oxidative effect based on the original property of polyphenols. Here we apply H9c2 cardiomyocytes to study the cardioprotective mechanisms of piceatannol in vitro. We firstly verify its anti-peroxidation effect by using H2O2-induced in vitro model. Then, flow cytometry results show piceatannol reduce cellular apoptosis by enhancing Bcl-2 expressions in immunoblot analysis. Meantime, piceatannol decreases H2O2-induced excessive ROS and calcium overloading, and prevents mitochondrial depolarization. Most importantly, piceatannol pretreatment can regulate PI3K-Akt-eNOS signaling pathway to alleviate peroxidative injury. Immunoblot analysis of PI3K, Akt, p-Akt and eNOS shows H2O2 significantly reduces expressions of these proteins. Pretreatment of piceatannol evidently increases their expressions and decreases iNOS expression, implying piceatannol can upregulate PI3K-Akt-eNOS signaling to protect cardiomyocytes from peroxidative injury.


Assuntos
Miócitos Cardíacos/efeitos dos fármacos , Óxido Nítrico Sintase Tipo III/biossíntese , Fosfatidilinositol 3-Quinases/biossíntese , Proteínas Proto-Oncogênicas c-akt/biossíntese , Transdução de Sinais/efeitos dos fármacos , Estilbenos/farmacologia , Animais , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Relação Dose-Resposta a Droga , Miócitos Cardíacos/metabolismo , Proteínas Proto-Oncogênicas c-akt/agonistas , Ratos , Transdução de Sinais/fisiologia
10.
J Biochem Mol Toxicol ; 33(2): e22245, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30431688

RESUMO

Salvianolic acid (SA) is known for improving blood circulation, scavenging hydroxyl radicals, and preventing platelet aggregation. The research explored whether SA can protect against cardiovascular disease induced by high glucose conditions. Our results indicate that SA significantly increases cells viability and nitric oxide levels while decreasing reactive oxygen species generation. SA upregulated the expression levels of Bcl-2 and decreased the levels of Bax, cleaved caspase-3, and cleaved caspase-9. Furthermore, the expression levels of Sirtuin 1 (Sirt1) and p-endothelial nitric oxide synthase (eNOS) were markedly increased in response to SA treatment. Moreover, exposure of human umbilical vein endothelial cells to Ex527 resulted in reducing expression of p-eNOS. However, the beneficial effects of SA were abolished partially when Ex527 was added. These findings suggest that SA can be used as a potential therapeutic to protect against high glucose-induced endothelial injury by modulating Sirt1-eNOS pathway.


Assuntos
Alcenos/farmacologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Glucose/metabolismo , Células Endoteliais da Veia Umbilical Humana/enzimologia , Óxido Nítrico Sintase Tipo III/biossíntese , Polifenóis/farmacologia , Transdução de Sinais/efeitos dos fármacos , Sirtuína 1/biossíntese , Células Endoteliais da Veia Umbilical Humana/patologia , Humanos
11.
Scand J Surg ; 108(1): 67-75, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30319041

RESUMO

BACKGROUND:: The saphenous vein is the most frequently used bypass conduit for vascular reconstructions, which may need to be stored for a prolonged time. The aim of this study was to compare the effect of different preservation solutions on the morphology of saphenous veins during the long-term cold storage. DESIGN:: An in vitro study. MATERIAL AND METHODS:: Saphenous vein samples, collected from 29 patients undergoing varicose vein surgery, were stored at +4°C in (1) 10% formalin, (2) isotonic saline with heparin and antibiotic, (3) phosphate-buffered saline, (4) 2.5% glutaraldehyde + phosphate-buffered saline, and (5) Custodiol (histidine-tryptophan-ketoglutarate). Changes in the vein wall were histologically investigated up to day 35. Possible retention of the capacity of endothelial function was evaluated by immunohistochemical detection of endothelial nitric oxide synthase. RESULTS:: Formalin as the control medium well preserved the vein wall morphology, but endothelial nitric oxide synthase immunostaining was very weak. Phosphate-buffered saline and isotonic saline with heparin and antibiotic poorly preserved vein wall morphology. Phosphate-buffered saline endothelial nitric oxide synthase staining decreased dramatically throughout the study period. Compared to phosphate-buffered saline, stronger isotonic saline with heparin and antibiotic endothelial nitric oxide synthase staining was noted at day 35 (p < 0.001). Custodiol and glutaraldehyde better preserved vein morphology compared to ISHA and PBS at day 5 and later (p < 0.001), but compared to stronger isotonic saline with heparin and antibiotic their endothelial nitric oxide synthase staining was weaker. CONCLUSION:: In terms of preserving the morphology of saphenous veins, phosphate-buffered saline and isotonic saline with heparin and antibiotic were the poorest, while Custodiol and glutaraldehyde were the best. Demonstrating good retention of endothelial nitric oxide synthase staining throughout the study period, isotonic saline with heparin and antibiotic seems to have the best potential to retain vein wall functionality, despite relatively poor morphological preservation.


Assuntos
Aloenxertos , Fármacos Cardiovasculares/administração & dosagem , Soluções Farmacêuticas/administração & dosagem , Veia Safena , Preservação de Tecido/métodos , Aloenxertos/efeitos dos fármacos , Aloenxertos/metabolismo , Aloenxertos/patologia , Temperatura Baixa , Humanos , Imuno-Histoquímica , Técnicas In Vitro , Óxido Nítrico Sintase Tipo III/biossíntese , Distribuição Aleatória , Veia Safena/efeitos dos fármacos , Veia Safena/metabolismo , Veia Safena/patologia , Fatores de Tempo , Varizes/metabolismo , Varizes/patologia , Varizes/cirurgia
12.
Anticancer Res ; 38(11): 6099-6106, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30396924

RESUMO

BACKGROUND/AIM: Budding uninhibited by benzimidazole-related 1 (BUBR1) and endothelial nitric oxide synthase (eNOS) are related to aging and angiogenesis. This study examined the effect of low BUBR1 expression on eNOS expression in vivo, in vitro, and human gastric cancer tissues. MATERIALS AND METHODS: Human umbilical vein endothelial cells (HUVECs) were passaged to investigate the effect of aging on BUBR1 and eNOS expression; expression of eNOS and phospho-eNOS protein was assessed in BUBR1 siRNA-transfected HUVECs. Additionally, guanosine 3',5' cyclic monophosphate (cGMP) and eNOS protein levels were measured in BUBR1-insufficient mice (Bubr1L/-). BUBR1 and eNOS expression levels were also evaluated in human gastric cancer tissues. RESULTS: BUBR1 and eNOS, but not p-eNOS, levels were reduced significantly in aged and BUBR1 siRNA-transfected HUVECs. Additionally, cGMP production and the eNOS protein level were reduced in Bubr1L/- mice. Human gastric cancer tissues with low BUBR1 expression showed no eNOS expression. CONCLUSION: A decrease in BUBR1 reduced eNOS bioavailability through a pathway other than eNOS phosphorylation.


Assuntos
Óxido Nítrico Sintase Tipo III/biossíntese , Proteínas Serina-Treonina Quinases/deficiência , Neoplasias Gástricas/enzimologia , Fatores Etários , Animais , Proteínas de Ciclo Celular/deficiência , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Senescência Celular/fisiologia , GMP Cíclico/biossíntese , Células Endoteliais da Veia Umbilical Humana , Humanos , Imuno-Histoquímica , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Óxido Nítrico Sintase Tipo III/metabolismo , Fosforilação , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , RNA Interferente Pequeno/genética , Transfecção , Fator A de Crescimento do Endotélio Vascular/farmacologia
13.
Int J Biol Macromol ; 120(Pt A): 66-72, 2018 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30076931

RESUMO

This study aims to explore the role and mechanism of lncRNA SNHG5 in spinal cord injury (SCI). The interaction between SNHG5 and Krüppel-like factor 4 (KLF4) was verified by RNA pull-down and RNA immunoprecipitation (RIP) assay. Rat neural function was evaluated by BBB and BMS scores. Results showed that GFAP and Iba-1 (specific proteins for astrocytes and microglia respectively) were upregulated in spinal cord of SCI rats. Simultaneously, spinal cord also expressed substantially higher levels of SNHG5, KLF4 and eNOS (endothelial Nitric Oxide Synthase) than sham group. In traumatically injured astrocytes and microglia, SNHG5 overexpression increased cells viability, which was significantly inhibited by SNHG5 knockdown. KLF4 is a directly target for SNHG5 and is positively regulated by SNHG5. The knockdown of KLF4 effectively decreased astrocytes and microglia viability induced by SHNG5 overexpression and attenuated the pcDNA-SNHG5-mediated repression of the apoptosis. In SCI rats, the injection of Lenti-SNHG5 reduced BBB and BMS scores and also enhanced the protein expression of KLF4, eNOS, GFAP and Iba-1. In summary, our data suggested that SNHG5 promotes SCI via increasing the viability of astrocytes and microglia. The mechanism by which SNHG5 works is its directive interaction to KLF4 and contribution to eNOS upregulation.


Assuntos
Astrócitos/metabolismo , Fatores de Transcrição Kruppel-Like/biossíntese , Microglia/metabolismo , RNA Longo não Codificante/biossíntese , Traumatismos da Medula Espinal/metabolismo , Regulação para Cima , Animais , Astrócitos/patologia , Sobrevivência Celular/genética , Técnicas de Silenciamento de Genes , Fatores de Transcrição Kruppel-Like/genética , Microglia/patologia , Óxido Nítrico Sintase Tipo III/biossíntese , Óxido Nítrico Sintase Tipo III/genética , RNA Longo não Codificante/genética , Ratos , Ratos Sprague-Dawley , Traumatismos da Medula Espinal/genética , Traumatismos da Medula Espinal/patologia
14.
J Am Heart Assoc ; 7(14)2018 07 11.
Artigo em Inglês | MEDLINE | ID: mdl-29997131

RESUMO

BACKGROUND: In vitro studies suggest that nephron nitric oxide synthase 3 (NOS3) modulates tubule Na+ transport. METHODS AND RESULTS: To assess nephron NOS3 relevance in vivo, knockout (KO) mice with doxycycline-inducible nephron-wide deletion of NOS3 were generated. During 1 week of salt loading, KO mice, as compared with controls, had higher arterial pressure and Na+ retention, a tendency towards reduced plasma renin concentration, and unchanged glomerular filtration rate. Chronic high salt-treated KO mice had modestly decreased total NCC and total SPAK/OSR1 versus controls, however percent phosphorylation of NCC (at T53) and of SPAK/OSR1 was increased. In contrast, total and phosphorylated NKCC2 (at T96/101) were suppressed by 50% each in KO versus control mice after chronic salt intake. In response to an acute salt load, KO mice had delayed urinary Na+ excretion versus controls; this delay was completely abolished by furosemide, partially reduced by hydrochlorothiazide, but not affected by amiloride. After 4 hours of an acute salt load, phosphorylated and total NCC were elevated in KO versus control mice. Acute salt loading did not alter total NKCC2 or SPAK/OSR1 in KO versus control mice but increased the percent phosphorylation of NKCC2 (at T96/101 and S126) and SPAK/OSR1 in KO versus control mice. CONCLUSIONS: These findings indicate that nephron NOS3 is involved in blood pressure regulation and urinary Na+ excretion during high salt intake. Nephron NOS3 appears to regulate NKCC2 and NCC primarily during acute salt loading. These effects of NOS3 may involve SPAK/OSR1 as well as other pathways.


Assuntos
Pressão Sanguínea/fisiologia , DNA/genética , Regulação da Expressão Gênica , Hipertensão/genética , Néfrons/metabolismo , Óxido Nítrico Sintase Tipo III/genética , Sódio/metabolismo , Animais , Modelos Animais de Doenças , Taxa de Filtração Glomerular , Hipertensão/metabolismo , Hipertensão/fisiopatologia , Masculino , Camundongos , Camundongos Knockout , Óxido Nítrico Sintase Tipo III/biossíntese , Transdução de Sinais , Cloreto de Sódio na Dieta/efeitos adversos
15.
Proc Natl Acad Sci U S A ; 115(29): E6900-E6909, 2018 07 17.
Artigo em Inglês | MEDLINE | ID: mdl-29967172

RESUMO

Neurons of the medullary reticular nucleus gigantocellularis (NGC) and their targets have recently been a focus of research on mechanisms supporting generalized CNS arousal (GA) required for proper cognitive functions. Using the retro-TRAP method, we characterized transcripts enriched in NGC neurons which have projections to the thalamus. The unique expression and activation of the endothelial nitric oxide (eNOS) signaling pathway in these cells and their intimate connections with blood vessels indicate that these neurons exert direct neurovascular coupling. Production of nitric oxide (NO) within eNOS-positive NGC neurons increases after environmental perturbations, indicating a role for eNOS/NO in modulating environmentally appropriate levels of GA. Inhibition of NO production causes dysregulated behavioral arousal after exposure to environmental perturbation. Further, our findings suggest interpretations for associations between psychiatric disorders and mutations in the eNOS locus.


Assuntos
Nível de Alerta/fisiologia , Encéfalo , Circulação Cerebrovascular/fisiologia , Neurônios/metabolismo , Óxido Nítrico Sintase Tipo III , Transdução de Sinais/fisiologia , Animais , Encéfalo/irrigação sanguínea , Encéfalo/citologia , Encéfalo/metabolismo , Loci Gênicos , Camundongos , Camundongos Transgênicos , Neurônios/citologia , Óxido Nítrico Sintase Tipo III/biossíntese , Óxido Nítrico Sintase Tipo III/genética
16.
Pharmacol Rep ; 70(4): 746-752, 2018 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-29936361

RESUMO

BACKGROUND: Diabetes mellitus is a metabolic disorder characterized by chronic hyperglycemia accompanied by impaired vascular and endothelial function. Activation of ATP-sensitive potassium (KATP) channels can protect endothelial function against hypertension and hyperglycemia. KMUP-1, a xanthine derivative, has been demonstrated to modulate K+-channel activity in smooth muscles. This study investigated protective mechanisms of KMUP-1 in impaired mesenteric artery (MA) reactivity in streptozotocin (STZ)-induced diabetic rats. METHODS: Rats were divided into three groups: control, STZ (65 mg/kg, ip) and STZ + KMUP-1 (5 or 10 mg/kg/day, ip). MA reactivity was measured by dual wire myograph. MA smooth muscle cells (MASMCs) were enzymatically dissociated and the KATP currents recorded by a whole-cell patch-clamp technique. RESULTS: STZ decreased MA KATP currents in a time-course dependent manner and achieved steady inhibition at day 14. In the MASMCs of STZ-treated rats, KMUP-1 partially recovered the KATP currents, suggesting that vascular KATP channels were activated by KMUP-1. K+ (80 mM KCl)-induced MA contractions in STZ-treated rats were higher than those of control rats. KMUP-1 significantly attenuated STZ-stimulated MA contractions in response to high K+, suggesting that KMUP-1 may partly restore the vascular reactivity of MAs. In addition, STZ decreased the expression of endothelial nitric oxide synthase (eNOS) and this effect was reversed by KMUP-1, suggesting that KMUP-1 could improve STZ-induced vascular endothelial dysfunction. CONCLUSION: KMUP-1 prevents STZ impairment of MA reactivity, eNOS levels and KATP channels, and accordingly protects against vascular dysfunction in diabetic rats.


Assuntos
Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/fisiopatologia , Canais KATP/metabolismo , Artérias Mesentéricas/fisiopatologia , Piperidinas/farmacologia , Doenças Vasculares/complicações , Doenças Vasculares/prevenção & controle , Xantinas/farmacologia , Animais , Diabetes Mellitus Experimental/metabolismo , Relação Dose-Resposta a Droga , Masculino , Potenciais da Membrana/fisiologia , Artérias Mesentéricas/metabolismo , Miócitos de Músculo Liso/fisiologia , Óxido Nítrico Sintase Tipo III/biossíntese , Potássio/farmacologia , Ratos , Fatores de Tempo , Doenças Vasculares/metabolismo , Doenças Vasculares/fisiopatologia
17.
Physiol Res ; 67(Suppl 1): S137-S147, 2018 06 27.
Artigo em Inglês | MEDLINE | ID: mdl-29947534

RESUMO

We examined the upregulation of ET-1/ETBR/eNOS signaling in renoprotective effect of vitamin D in kidney fibrosis model in mice using unilateral ureteral obstruction (UUO). One group was treated with intraperitoneal injection of 0.125 mg/kg of Calcitriol (UUO+VD). Vascular remodeling was quantified based on lumen area and lumen/wall area ratio (LWAR) of intrarenal arteries using Sirius Red staining. ET-1, ETBR, eNOS, CD31 and VEGF mRNA expressions were quantified using qRT-PCR. Focusing on endothelin-1 (ET-1) signaling in endothelial cells (EC), siRNA of ET-1 was performed in human umbilical vein endothelial cells (HUVEC) for reducing ET-1 expression. Then HUVECs were treated with and without 100 nM Calcitriol treatment in hypoxic and normoxic conditions to elucidate ET-1/eNOS signaling. Our in vivo study revealed vascular remodeling and renal ischemia attenuation after Calcitriol treatment. Vascular remodeling was attenuated in the UUO+VD group as shown by increasing lumen areas and LWAR in intrarenal arteries. These findings were associated with significant higher CD31 and VEGF mRNA expression compared to the UUO group. Vitamin D treatment also increased ET-1, ETBR and eNOS mRNA expressions. Our in vitro study demonstrated Calcitriol induced ET-1 and eNOS mRNA expressions upregulation in HUVEC under normoxic and hypoxic condition. Meanwhile, siRNA for ET-1 inhibited the upregulation of eNOS mRNA expression after Calcitriol treatment. Vitamin D ameliorates kidney fibrosis through attenuating vascular remodeling and ischemia with upregulating ET-1/ETBR and eNOS expression.


Assuntos
Endotelina-1/biossíntese , Nefropatias/metabolismo , Óxido Nítrico Sintase Tipo III/biossíntese , Receptor de Endotelina B/biossíntese , Remodelação Vascular/efeitos dos fármacos , Vitamina D/farmacologia , Animais , Fibrose , Células Endoteliais da Veia Umbilical Humana , Humanos , Isquemia/tratamento farmacológico , Isquemia/metabolismo , Nefropatias/tratamento farmacológico , Masculino , Camundongos , RNA Mensageiro/biossíntese , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/fisiologia , Remodelação Vascular/fisiologia , Vitamina D/uso terapêutico
18.
Life Sci ; 205: 145-154, 2018 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-29733850

RESUMO

AIMS: The study aimed to investigate the protective effect of chronic intermittent hypobaric hypoxia (CIHH) on endothelium function and relaxation of mesenteric artery in metabolism syndrome (MS) rats. MAIN METHODS: Male adult Sprague-Dawley rats were randomly divided into control (CON), CIHH (treated with 28-days hypobaric hypoxia simulating an altitude of 5000 m, 6 h daily), MS (induced by high fat diet and 10% fructose water feeding), and MS + CIHH groups. Body weight, systolic arterial pressure, blood biochemical and the endothelium dependent relaxation (EDR) of mesenteric arteries were measured. The expression of phosphor-endothelial nitric oxide synthase (p-eNOS), endoplasmic reticulum (ER) stress-related proteins and autophagy-related proteins in mesenteric arteries was assayed. KEY FINDINGS: The MS rats displayed hypertension, obesity, metabolic abnormity and insulin resistance, EDR was attenuated, p-eNOS expression was down-regulated, the expressions of ER stress-related proteins were up-regulated, and autophagy dysfunction occurred. All aforementioned abnormalities in MS rats were ameliorated in MS + CIHH rats. Furthermore, the improvement of CIHH on EDR and p-eNOS was cancelled by the ER stress inducer, and the autophagy inhibitor. SIGNIFICANCE: In conclusion CIHH protects endothelium function and enhances relaxation in mesenteric arteries of MS rats through improving autophagy function, reducing ER stress and up-regulating p-eNOS.


Assuntos
Autofagia , Endotélio Vascular/patologia , Hipóxia/patologia , Síndrome Metabólica/patologia , Síndrome Metabólica/terapia , Pressão do Ar , Animais , Pressão Arterial , Peso Corporal , Dieta Hiperlipídica , Estresse do Retículo Endoplasmático , Masculino , Artérias Mesentéricas/patologia , Óxido Nítrico Sintase Tipo III/biossíntese , Ratos , Ratos Sprague-Dawley
19.
Mol Med Rep ; 18(1): 1181-1187, 2018 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-29845232

RESUMO

MicroRNA (miR)­24 has been reported to associate with various diseases by acting on different signaling pathways. The present study aimed to elucidate the association between miR­24 expression levels and vasospasm following subarachnoid hemorrhage (SAH), and its underlying mechanism. An miR online database was searched, identifying endothelial nitric oxide synthase (NOS3) as a potential target gene of miR­24. A luciferase reporter assay performed to investigate the regulatory association between miR­24 and NOS3 revealed that miR­24 bound to the NOS3 3' untranslated region and inhibited NOS3 expression. Reverse transcription­quantitative polymerase chain reaction and western blot analysis were performed to investigate the miR­24 and NOS3 expression levels in samples from patients with SAH, and demonstrated a negative correlation between the two. In addition, miR­24 expression levels were increased in SAH patients with vasospasm compared with those without, whereas the opposite results were observed for NOS3. Vascular smooth muscle cells (VSMCs) transfected with an miR­24 inhibitor exhibited increased expression levels of NOS3, whereas those transfected with an miR­24 mimic or NOS3 small interfering RNA exhibited reduced expression levels of NOS3, compared with the control. These results indicated a negative regulatory association between miR­24 and NOS3. Downregulation of NOS3 may induce vasospasm following SAH, which may be due to the upregualtion of miR­24 in VSMCs.


Assuntos
Regulação Enzimológica da Expressão Gênica , MicroRNAs/biossíntese , Óxido Nítrico Sintase Tipo III/biossíntese , Hemorragia Subaracnóidea/metabolismo , Regulação para Cima , Vasoespasmo Intracraniano/metabolismo , Adulto , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Hemorragia Subaracnóidea/complicações , Hemorragia Subaracnóidea/patologia , Vasoespasmo Intracraniano/etiologia , Vasoespasmo Intracraniano/patologia
20.
Angiogenesis ; 21(4): 711-724, 2018 11.
Artigo em Inglês | MEDLINE | ID: mdl-29737439

RESUMO

The nitric oxide (NO) secreted by vascular endothelium is required for the maintenance of cardiovascular homeostasis. Diminished release of NO generated by endothelial NO synthase contributes to endothelial dysfunction. Hypoxia and ischemia reduce endothelial eNOS expression via posttranscriptional mechanisms that result in NOS3 transcript destabilization. Here, we examine whether microRNAs contribute to this mechanism. We followed the kinetics of hypoxia-induced changes in NOS3 mRNA and eNOS protein levels in primary human umbilical vein endothelial cells (HUVECs). Utilizing in silico predictive protocols to identify potential miRNAs that regulate eNOS expression, we identified miR-200b as a candidate. We established the functional miR-200b target sequence within the NOS3 3'UTR, and demonstrated that manipulation of the miRNA levels during hypoxia using miR-200b mimics and antagomirs regulates eNOS levels, and established that miR-200b physiologically limits eNOS expression during hypoxia. Furthermore, we demonstrated that the specific ablation of the hypoxic induction of miR-200b in HUVECs restored eNOS-driven hypoxic NO release to the normoxic levels. To determine whether miR-200b might be the only miRNA that had this effect, we utilized Next Generation Sequencing (NGS) to follow hypoxia-induced changes in the miRNA levels in HUVECS and found 83 novel hypoxamiRs, with two candidate miRNAs besides miR-200b that could potentially influence eNOS levels. Taken together, the data establish miR-200b-eNOS regulation as a first hypoxamiR-based mechanism that limits NO bioavailability during hypoxia in endothelial cells, and show that hypoxamiRs could become useful therapeutic targets for cardiovascular diseases and other hypoxic-related diseases including various types of cancer.


Assuntos
Regulação Enzimológica da Expressão Gênica , Células Endoteliais da Veia Umbilical Humana/metabolismo , MicroRNAs/metabolismo , Óxido Nítrico Sintase Tipo III/biossíntese , Óxido Nítrico/metabolismo , Hipóxia Celular , Células HEK293 , Humanos
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