ABSTRACT
Abstract Introduction: Diabetes mellitus (DM) is a chronic disease characterized by hyperglycemia that leads to diabetic nephropathy (DN). We showed that P2X7, a purinergic receptor, was highly expressed in DM; however, when oxidative stress was controlled, renal NO recovered, and the activation of this receptor remained significantly reduced. The aim of this study was to assess the influence of NO on the P2X7 and apoptosis in mouse immortalized mesangial cells (MiMC) cultured in high glucose (HG) medium. Methods: MiMCs were cultured with DMEM and exposed to normal glucose (NG), mannitol (MA), or HG. Cell viability was assessed by an automated counter. Supernatants were collected for NO quantification, and proteins were extracted for analysis of NO synthases (iNOS and eNOS), caspase-3, and P2X7. Results: Cell viability remained above 90% in all groups. There was a significant increase in the proliferation of cells in HG compared to MA and NG. NO, iNOS, caspase-3, and P2X7 were significantly increased in HG compared to NG and MA, with no changes in eNOS. We observed that there was a strong and significant correlation between P2X7 and NO. Discussion: The main finding was that the production of NO by iNOS was positively correlated with the increase of P2X7 in MCs under HG conditions, showing that there is a common stimulus between them and that NO interacts with the P2X7 pathway, contributing to apoptosis in experimental DM. These findings could be relevant to studies of therapeutic targets for the prevention and/or treatment of hyperglycemia-induced kidney damage to delay DN progression.
Resumo Introdução: Diabetes mellitus (DM) é uma doença crônica caracterizada por hiperglicemia levando à nefropatia diabética (ND). Mostramos que P2X7, um receptor purinérgico, foi altamente expresso na DM; entretanto, quando o estresse oxidativo foi controlado, o NO renal recuperou-se, e a ativação deste receptor permaneceu significativamente reduzida. Este estudo objetivou avaliar a influência do NO no P2X7 e a apoptose em células mesangiais imortalizadas de camundongos (CMiC) cultivadas em meio de glicose elevada (GE). Métodos: CMiCs foram cultivadas em meio DMEM e expostas à glicose normal (GN), manitol (MA), ou GE. A viabilidade celular foi avaliada por contador automático. Sobrenadantes foram coletados para quantificação de NO, e foram extraídas proteínas para análise de NO sintases (iNOS e eNOS), caspase-3, e P2X7. Resultados: A viabilidade celular permaneceu acima de 90% em todos os grupos. Houve aumento significativo na proliferação de células na GE comparado com MA e GN. NO, iNOS, caspase-3 e P2X7 foram significativamente aumentados na GE comparados com GN e MA, sem alterações na eNOS. Observamos que houve correlação forte e significativa entre P2X7 e NO. Discussão: O principal achado foi que a produção de NO pela iNOS foi positivamente correlacionada com aumento de P2X7 em CMs sob condições de GE, mostrando que existe um estímulo comum entre eles e que o NO interage com a via do P2X7, contribuindo para apoptose na DM experimental. Estes achados podem ser relevantes para estudos de alvos terapêuticos para a prevenção e/ou tratamento de danos renais induzidos por hiperglicemia para retardar a progressão da ND.
ABSTRACT
OBJECTIVE: To investigate the protective effect of the decoction of tartary buckwheat flavonoids on focal cerebral is-chemia-reperfusion injury of rats. MEHTODS: Fifty male SD rats will be randomly divided into normal group, cerebral ischemia-reperfusion group (model group), high dose of tartary buckwheat flavonoids group, middle dose of tartary buckwheat flavonoids group and low dose of tartary buckwheat flavonoids group, each groups of 10 rats. A rat model of the right middle cerebral artery occlusion/reper-fusion(MCAO) was established by the filament method. After being operated, treatment-group rats will be administered 100,75 and 50 mg · kg-1 of the decoction of tartary buckwheat flavonoids three times a day for 7 consecutive days, after administrated for 7 d, rats in each group will undergo neurobehavioral tests. Expressions of cerebral inducible nitric oxide synthase (iNOS) and endothelial nitric oxide synthase (eNOS) were measured by SP immunohistochemistry. The optical density value (OD) was measured by imaging analysis, and the percentage of cells with iNOS and eNOS positive expression was analyzed under light microscope. RESULTS: Compared with ischemia-reperfusion group, neurological function score increased in the decoction of tartary buckwheat flavonoids groups. Treatment groups had lower expression level of iNOS but higher expression level of eNOS than those in the model groups (P < 0.01, P < 0.05). The number of neurons of Hippocampal CA1 was increased (P < 0.05). CONCLUSION: Tartary buckwheat flavonoids can improve neurological function and decrease the expression of iNOS and increase the expression of eNOS in cerebral ischemia-reperfusion rats, which may contribute to the protection of neural function.
ABSTRACT
Las enfermedades sistémicas crónicas afectan el músculo esquelético, siendo la inflamación y el estrés oxidativo algunos de los mecanismos involucrados. El efecto de la hipertensión arterial esencial sobre el músculo esquelético no es bien conocido. Se estudiaron los músculos soleo y extensor digitorum longus (EDL) de ratas espontáneamente hipertensas (SHR), comparadas con las controles normotensas Wistar Kyoto (WKY). Se determinaron los niveles de nitritos y nitratos en µmoles/mg-proteína; las sintasas del óxido nítrico: endotelial (eNOS); neuronal (nNOS); e inducible (iNOS), nitrotirosina y el factor de necrosis tumoral-alfa (TNF-α) en ng/mg-proteína. En las SHR, en el soleo y el EDL respectivamente, se incrementó la nitrotirosina (24,4 ± 5,0 vs. 3,3 ± 0,3, p<0,001; 20,2 ± 4,3 vs. 4,5 ± 0,4, p<0,0037), iNOS (26,6 ± 3,7 vs. 8,3 ± 0,9; 21,3 ± 3,7 vs. 11,0 ± 0,8 ambos p<0,0001), y TNF-α (2,2 ± 0,5 vs. 0,6 ± 0,1, p<0,05; 1,9 ± 0,2 vs. 0,6 ± 0,1, p<0,02); hubo disminución de eNOS en el soleo (20,6 ± 1,4 vs. 30,3 ± 1,2, p<0,00001); de nNOS (soleo 16,8 ± 1,4 vs. 20,7 ± 1,8, p< 0,05; EDL 13,6 ± 1,3 vs. 21,9 ± 1,8, p<0,005) y de nitrito en el EDL (5,8 ± 0,3 vs. 7,1 ± 0,5, p<0,026). En las SHR se observó correlación positiva entre TNF-α vs. nitrotirosina: soleo (r=0,798; p<0,031) y tendencia en EDL (r=0,739; p<0,057); iNOS vs. nitrotirosina (soleo: r=0,908 p<0,0001; EDL: r=0,707; p=0,01), tendencia entre TNF-α vs. iNOS en EDL (r=0,736; p=0,059); y correlación negativa entre eNOS vs. nitrotirosina en soleo (r=-0,816; p=0,0012). En conclusión, las SHR presentan un proceso inflamatorio muscular, evidenciado por el incremento de TNF-α, nitrotirosina, e iNOS. La disminución de las sintasas constitutivas, con incremento de la iNOS es evidencia de la disfunción endotelial.
Systemic diseases affect skeletal muscle, and inflammation and oxidative stress are some of the involved mechanisms. There is scarce information about the effects of essential hypertension on skeletal muscle. The soleus and extensor digitorum longus (EDL) muscles of spontaneously hypertensive rats (SHR) were studied compared to control Wistar Kyoto (WKY) rats. The levels of nitrite and nitrate in µmol/mg-protein; endothelial (eNOS), neuronal (nNOS), and inducible (iNOS) nitric oxide synthases, nitrotyrosine and tumour necrosis factor alpha (TNF-α) in ng/mg-protein were determined. Compared with controls, the SHR showed increased levels of nitrotyrosine (soleus 24.4 ± 5.0 vs. 3.3±0.3, p<0.001; EDL 20.2 ± 4.3 vs. 4.5 ± 0.4, p<0.0037), iNOS (soleus 26.6 ± 3.7 vs. 8.3 ± 0.9; EDL 21.3 ± 3.7 vs. 11.0 ± 0.8, both p<0.0001) and TNF-α (soleus 2.2 ± 0.5 vs. 0.6 ± 0.1, p<0.05; EDL 1.9 ± 0.2 vs. 0.6 ± 0.1, p<0.02). A decrease of eNOS was found in soleus muscle (20.6 ± 1.4 vs. 30.3 ± 1.2, p<0.00001); of nNOS (soleus 16.8 ± 1.4 vs. 20.7 ± 1.8, p< 0.05; EDL 13.6 ± 1.3 vs. 21.9 ± 1.8, p<0.005) and nitrite in EDL (5.8 ± 0.3 vs. 7.1 ± 0.5, p<0.026).There was a positive correlation between TNF-α vs. nitrotyrosine in soleus (r=0.798; p<0.031) and a tendency in EDL (r=0.739; p=0.059); iNOS vs. nitrotyrosine (soleus: r=0.908; p<0.0001; EDL: r=0.707; p<0.01), a tendency between TNF-α and iNOS (EDL: r=0.736; p<0.059); and a negative correlation between eNOS vs. nitrotyrosine in soleus muscle (r=-0.816; p<0.0012). In conclusion, in skeletal muscles of SHR an inflammatory process was found evidenced by the increase in TNF-α, nitrotyrosine and iNOS. The decreased levels of constitutive synthases, together with the higher level of iNOS, are indicative of endothelial dysfunction.
Subject(s)
Animals , Male , Rats , Hypertension/metabolism , Muscle, Skeletal/metabolism , Oxidative Stress , Endothelium, Vascular/physiopathology , Muscle, Skeletal/chemistry , Myositis/metabolism , Nitric Oxide Synthase Type I/analysis , Nitric Oxide Synthase Type II/analysis , Nitric Oxide Synthase Type III/analysis , Rats, Inbred SHR , Rats, Inbred WKY , Tumor Necrosis Factor-alpha/analysis , Tyrosine/analysis , Tyrosine/analogs & derivativesABSTRACT
@#Objective To study the effect of electrical stimulation of vagus nerve on inflammatory response in septic shock rats. Methods SD rats were randomly divided into 5 groups: Group Ⅰ was the sham group, group Ⅱ with the cecal ligation and puncture (CLP) and the vagus nerve were isolated but not transected, group Ⅲ with bilateral cervical vagotomy following CLP, group Ⅳ with bilateral cervical vagotomy after CLP and the left vagus nerve trunks were stimulated with bipolar electrodes, group Ⅴ with bilateral cervical vagotomy after CLP and the right vagus nerve trunks were stimulated. The common carotid artery pressure was monitored, and the plasma tumor necrosis factor α (TNF-α), nitric oxide synthases (NOS) and nitric oxide (NO) were measured 2 h after stimulation. Results The mean arterial blood pressure (MAP) gradually decreased and the concentration of plasma TNF-α, NOS and NO significantly increased after CLP. Electrical stimulation of the left and right vagus nerve significantly increased the MAP and decreased the plasma TNF-α, NOS and NO levels. Conclusion Direct electrical stimulation of the left and right vagus nerve can significantly improve the blood pressure and reduced plasma TNF-α, NOS and NO levels during septic shock, which may play a role in anti-shock in rats.
ABSTRACT
The present study was aimed at investigating whether FK506 alters the regulation of nitric oxide(NO) system. Male Sprague-Dawley rats were treated with FK506(1 mg/kg/day, i.m.) for 3 weeks. Control group was without treatment of FK506. Plasma levels and urinary excretion of NO metabolites(nitrite/nitrate, NOx) were measured. The protein expression of NO synthases(NOS) and tissue contents of NOx were determined in the kidney and thoracic aorta. The aorta was also examined of its changes in isometric tension in responses to acetylcholine and sodium nitroprusside. The arterial pressure did not significantly differ between FK506-treated and control groups. Plasma NOx levels remained unaltered, while urinary NOx excretion was significantly decreased in FK 506-treated group. Tissue contents of NOx were significantly decreased, although the expression of ecNOS and iNOS proteins was significantly altered neither in the kidney nor in the aorta. Acetylcholine-induced relaxation of the isolated aortic ring was significantly attenuated, whereas sodium nitroprusside-induced relaxation was not significantly affected. These results suggest that FK506 decreases the tissue contents of NO, without significantly affecting the expression of NOS.
Subject(s)
Animals , Humans , Male , Rats , Acetylcholine , Aorta , Aorta, Thoracic , Arterial Pressure , Kidney , Nitric Oxide , Nitroprusside , Plasma , Rats, Sprague-Dawley , Relaxation , Sodium , TacrolimusABSTRACT
The present study was aimed at investigating whether the development of hypertension is related with an altered expression of nitric oxide synthases (NOS) in the kidney. By Western blot analysis, the expression of bNOS and ecNOS isoforms was determined in the kidney of deoxycorticosterone acetate (DOCA)-salt and two-kidney, one clip (2K1C) rats. In DOCA-salt hypertension, the expression of both bNOS and ecNOS was decreased, along with tissue contents of nitrites. In 2K1C hypertension, the nitrite content of the clipped kidney was decreased along with ecNOS levels, whereas neither the nitrite content nor the expression of NOS isoforms was significantly altered in the contralateral non-clipped kidney. These results suggest that the development of hypertension is associated with an altered renal expression of NOS and nitric oxide generation in DOCA-salt and 2K1C rats.
Subject(s)
Animals , Rats , Blotting, Western , Desoxycorticosterone , Hypertension , Kidney , Nitric Oxide , Nitrites , Protein IsoformsABSTRACT
OBJECTIVE:To investigate the effects of selenium(Se)and/or vitamin E(VE)on the NO and NOS in heart,liver,kidney and serum of experimental hyperlipidemic rats.METHODS:SD rats were divided into5groups,administreated by Se and/or VE.After4weeks,the NO contents and NOS activities in heart,liver,kidney and serum were assayed by NO kit and NOS kit respectively.RESULTS:NO contents and NOS activities could be reduced in heart,liver,but increased in serum and kidney by high-fat feed(HFF).Meanwhile,VE and/or Se could increase the NO contents in all the experimental samples and NOS activities in heart,liver and kidney(P