ABSTRACT
Objective To investigate the correlation between inducible nitric oxide synthase (iNOS) gene polymorphism and ischemic stroke in Chinese Han population.Methods Patients with first-ever stroke and the age-and sex-matched healthy controls were enrolled in the study.Taqman probe fluorescence quantitative polymerase chain reaction technique was used to detect the genotype distribution of rs2779248 C/T and rs1137933 C/T polymorphisms.Results A total of 246 patients with ischemic stroke and 246 controls were enrolled.The distribution frequencies of rs2779248 CC,CT and TT genotypes in the patient group were 57.7％,36.6％,and 5.7％,respectively,and in the control group were 68.7％,28.0％,and 3.3％,respectively.The T allele frequency of the patient group was significantly higher than that of the control group (24.0％ vs.17.3％;P =0.015).Multivariate logistic regression analysis showed that after adjusting for the risk factors including age,sex,hypertension,and diabetes,the risk of ischemic stroke in the CT + TT genotype carriers was 1.64 times of the CC genotype carriers (odds ratio 1.64,95％ confidence interval 1.07-2.51;P=0.022).The distribution frequencies of rs1137933 CC,CT and TT in the patient group were 58.1％,37.8％,and 4.1％,respectively and in the control group were 68.3％,29.3％,and 2.4％,respectively.The T allele frequency of the patient group was significantly higher than that of the control group (23.0％ vs.17.1％;P =0.013).Multivariate logistic regression analysis showed that after adjusting for traditional risk factors,the risk of ischemic stroke of patients with the TT + CT genotype carriers was 1.60 times of the CC genotype carriers (odds ratio 1.60,95％ confidence interval 1.05-2.46;P=0.030).Conclusions The rs2779248 C/T and rs1137933 C/T polymorphisms in iNOS gene may be associated with the risk of onset of ischemic stroke in Chinese Han population.
ABSTRACT
Objective To investigate the effect of sinomenine on the levels of nuclear factor kappa B (NF-κB) and inducible nitric oxide synthase (iNOS) during renal ischemia-reperfusion (I/R) in rats.Methods Seventy-two healthy male Wistar rats, aged 6-9 weeks, weighing 180-220 g, were randomly assigned into 4 groups (n =18 each) using a random number table: control group (group C), sinomenine group (group SIN), group I/R, and sinomenine+I/R group (group SIN+I/R).The animals were anesthetized with 10％ chloral hydrate 350 mg/kg.The left renal pedicles were clamped with atraumatic microclips for 45 min followed by reperfusion, and the right kidney was removed immediately after onset of reperfusion to establish the model of renal I/R injury.Sinomenine 60 mg/kg was injected intraperitoneally at 30 min before reperfusion in group SIN +I/R, and at the corresponding time point in group SIN.At 6, 8 and 12 h of reperfusion, the blood samples were drawn by cardiac puncture for measurement of serum creatinine (Cr) and blood urea nitrogen (BUN) concentrations.The left renal specimens were obtained for examination of pathological changes (with light microscopes) and for determination of the rate of NF-κB-positive cells and iNOS expression in renal tissues (by immunohistochemistry).Results Compared with group C, the concentrations of serum Cr and BUN, and rate of NF-κB-positive cells were significantly increased, and the expression of iNOS was up-regulated in I/R and SIN+I/R groups, and no significant change was found in the parameters mentioned above in group SIN.Compared with group I/R, the concentrations of serum Cr and BUN, and rate of NF-κB-positive cells were significantly decreased, and the expression of iNOS was down-regulated in group SIN+I/R.The microscopic examination showed that the pathological changes of kidney were significantly attenuated in group SIN+I/R compared with group I/R.Conclusion The mechanism by which sinomenine attenuates renal I/R injury is related to inhibited activity of NF-κB and down-regulated expression of iNOS in rats.
ABSTRACT
Objective To evaluate the effects of curcumin pretreatment on the activity of inducible nitric oxide synthase (iNOS) during lung injury induced by intestinal ischemia-reperfusion (Ⅰ/R) in rats.Methods Twenty-four pathogen-free female Sprague-Dawley rats,weighing 180-220 g,were randomly divided into 3 groups (n =8 each) using a random number table:sham operation group (group Sham);intestinal Ⅰ/R group (group Ⅱ/R);curcumin pretreatment group (group Cur).A rat model of lung injury induced by intestinal Ⅰ/R which was produced by occlusion of superior mesenteric artery for 75 min followed by reperfusion was established.At 5 days before Ⅰ/R,curcumin 200 mg/kg (in 20 mg/ml of normal saline) was given through a gastric tube in group Cur,while the equal volume of normal saline was given instead in Sham and Ⅱ/R groups.The rats were sacrificed at 4 h of reperfusion,and the pulmonary specimens were obtained for determination of wet/dry lung weight ratio (W/D ratio),NO content (by using nitrate reductase method) and iNOS activity (using colorimetric method) and for examination of pathological changes (with light microscope).The pathological changes of the lung were scored.Results Compared with group Sham,the pathological scores,W/D ratio,NO content and iNOS activity were significantly increased in Ⅱ/ R and Cur groups.Compared with Ⅱ/R group,the pathological scores,W/D ratio,NO content and iNOS activity were significantly decreased in group Cur.The pathological changes of lungs were significantly attenuated in group Cur as compared with H/R group.Conclusion The mechanism by which curcumin pretreatment attenuates lung injury induced by intestinal Ⅰ/R is related to decrease in iNOS activity in rats.
ABSTRACT
ObjectiveTo investigate the effects of simvastatin preconditioning on the expression of inducible and endothelial nitric oxide synthase ( iNOS,eNOS) in thoracic aorta in a rat model of sepsis.Methods Eighty pathogen-free female Wistar rats aged 4 months weighing 200-250 g were randomly divided into 4 groups:group normal control (group Ⅰ,n =8) ; group sham operation (group Ⅱ,n =8) ; group sepsis (group Ⅲ,n =32) and group simvastatin preconditioning (group Ⅳ,n =32).Sepsis was induced by cecal ligation and puncture (CLP) in groups Ⅲ and Ⅳ.In group Ⅳ simvastatin 20 mg/kg was given via a gastric tube once a day for 2 weeksbefore CLP.The thoracic aorta specimens were taken at 3,6,24 and 48 h after CLP (n =8 at each time point)for detection of iNOS and eNOS protein expression by Western blot analysis.ResultsCLP significantly up-regulated iNOS expression and down-regulated eNOS expression in group Ⅲ as compared with groups Ⅰ and Ⅱ.Simvastatin pretreatment significantly attenuated CLP-induced increase in iNOS expression and decrease in eNOS expression in group Ⅳ as compared with group Ⅲ.ConclusionSimvastatin preconditioning can protect vascular endothelial cells from septic injury by down-regulating iNOS expression and up-regulating eNOS expression in vascular endothelial cells.
ABSTRACT
ObjectiveTo investigate the effect of propofol on endothelial nitric oxide synthase (eNOS) and inducible nitric oxide synthase (iNOS) expression in thoracic aorta of hypertensive rats.MethodsHealthy SD rats of both sexes weighing 240-280 g were used in this study.Hypertension was induced by subcutaneous deoxycorticosterone 25 mg/kg twice a week for consecutive 7 weeks.Sixty-four hypertensive rats were randomly divided into four groups (n ＝ 16 each):hypertension group (group H),low,medium and high dose propofol group ( groups P1,P2,P3 ).Groups P1,P2 and P3 received infusion of propofol at a rate of 20,30 and 40 mg* kg- 1 · h- 1 for 3 h respectively,while group H received equal volume normal saline instead of propofol.Mean arterial pressure (MAP) was monitored and recorded before,1 h and 3 h after the start of propofol or normal saline infusion.All animals were sacrificed at 3 h of intravenous administration.Blood samples were collected by taking out the eyeballs for determination of serum NO concentrations by nitrate reductase method.The expression of eNOS mRNA,iNOS mBNA was determined by reverse transcription-polymerase chain reaction.The expression of eNOS and iNOS protein was determined by Western blot.ResultsCompared to group H,MAP was decreased significantly,the serum NO concentrations were increased significantly,the expression of eNOS mRNA and protein in thoracic aorta was up-regulated,and the expression of iNOS mRNA and protein in thoracic aorta was down-regulated in a dose-dependent manner in groups P1,P2 and P3 ( P < 0.05 or 0.01 ).ConclusionPropofol can down-regulate iNOS expression and up-regudate eNOS expression in endothelial cells of thoracic aorta and promote NO release in hypertensive rats,Which is the mechanism of propofol decreasing pressure.
ABSTRACT
ObjectiveTo evaluate the effect of aminoguanidine on expression of inducible nitric oxide synthase (iNOS) in neurons in the small intestinal nerve plexus of starved rats.MethodsNinety male SD rats weighing 230-270 g were randomly divided into 3 groups:group normal control (group C,n =10) ; group starvation (group S,n=40) and group starvation + aminoguanidine (group A,n =40).The animals were allowed free access to water but no food during starvation in S and A groups.In group A the animals were given aminoguanidine 150 mg·kg-1 ·d-1 intraperitoneally during starvation.Ten animals were sacrificed at 3,5,7 and 9 d of starvation respectively and intestine specimens were taken for determination of ratio of intestinal transit using dextran blue-2000 as indicator.Then the specimens of intestinal myenteric nerve plexus of ileum were collected and stained by histochemistry with nicotinamide-adenine dinucleotide phosphate-d for determination of iNOS expression.ResultsStarvation significantly reduced the small intestinal transit and increased iNOS expression in neurons in the myenteric nerve plexus of small intestine in proportion to days of starvation in group S compared with group C.Intraperitoneal aminoguanidine significantly attenuated the starvation-induced changes in intestinal transit and iNOS expression.ConclusionAminoguanidine can attenuate the up-regulation of the expression of iNOS in neurons in the myenteric nerve plexus of small intestine induced by starvation and is helpful in promoting the intestinal transit in starved rats.
ABSTRACT
ObjectiveTo evaluate the efffects dexamethasone (DEX) administration at different times on intestinal ischemia-reperfusion(I/R) injury and inducible nitric oxide synthase(iNOS) activity in mice.Methods Tirty-five healthy male Kunming mice weighing 20-24 g were randomly divided into 5 groups( n =7 each): Sham operation group (group Ⅰ ); intestinal I/R group (group Ⅱ ); DEX administration before ischemia group (group Ⅲ ); DEX administration during ischemia group ( group Ⅳ) and DEX administration at the begining of reperfusion group (group Ⅴ ).Intestinal I/R injury was induced by clamping the superior mesenteric artery for 30 min.Normal saline10 mg/kg,DEX 10 mg/kg was injected iv at 30 min before ischemia in groups Ⅱ and Ⅲ respectively.DEX 10 mg/kg was injected iv at 5 min of ischemia in groupⅣ and immediately at the begining of reperfusion in group Ⅴ.The mice were sacrificed at 3 h of reperfusion,and then the small intestinal tissues were taken for determination of intestinal pathological score( Chiu score),iNOS activity and nitric oxide (NO) content.ResultsChiu score was significantly higher in groups Ⅱ - Ⅴ,and iNOS activity and NO content were sinificantly higher in groups Ⅱ,Ⅳ and Ⅴ than in group Ⅰ ( P ＜ 0.05).Chiu score,iNOS activity and NO content were sinificantly lower in group Ⅲ,and were higher in group Ⅴ than in group Ⅱ ( P ＜ 0.05).There was no significant difference in the indexes mentioned above between groups Ⅱ and Ⅳ ( P ＞ 0.05).ConclusionDEX administration before ischemia can reduce intestinal I/R injury by inhibiting iNOS activity; DEX administration during ischemia has no effcet on intestinal I/R injury and iNOS activity; DEX administration at the begining of reperfusion aggravates intestinal I/R injury by enhancing iNOS activity.
ABSTRACT
Objective To investigate the role of nuclear factor-kappa B (NF-κB)signaling pathway in propofol-induced suppression of up-regulation of inducible nitric oxide synthase (iNOS) gene expression in LPSstimulated RAW264.7 cells.Methods RAW264.7 cells were purchased from cell bank of Chinese Academy of Sciences and cultured in DMEM culture medium containing 10％ fetal bovine serum.The cells were seeded in 6 cm diameter dishes (3 ml/dish) or in 6-well plates (2 ml/well) with a density of 5 × 105/ml and randomly divided into 3 groups ( n =18): normal control group (group C),group LPS (group L)and group LPS + propofol (group LP).The cells were incubated with LPS 1 μg/ml in groups L and LP.Propofol 50μmol/L was added to the culture medium at 2 h before LPS in group LP.Cells were harvested at 30 min after being stimulated with LPS.Phosphorylation of IκB kinase(p-IKK) and NF-κB activity were detected by Western blot.The expression of iNOS mRNA was determined after 6 h exposure of the cells to LPS.Results LPS significantly up-regulated the expression of p-IKK and iNOS mRNA and increased NF-κB activity in group L as compared with group C.Propofol pretreatment significantly attenuated the effects of LPS on p-IKK,iNOS mRNA expression and NF-κB activity.Conclusion NF-κB signaling pathway is involved in the propofol-induced suppression of up-regulation of iNOS mRNA expression in LPS-stimulated RAW264.7 cells.
ABSTRACT
Objective To explore the expression of nitric oxide synthases including neuronal nitric oxide synthases(nNOS), inducible nitric oxide synthases(iNOS), endothelial nitric oxide synthases (eNOS) in neurogenic bladder tissues, and analyze it's producing feature and significance. Methods There were 30 cases with neurogenic bladder(18 males, 12 females). The average age was 6.3±3.1 years. All patients appeared with myelodysplasia, urinary and fecal incontinence in different degree. Twenty-six cases were manifested with hyperreflexia bladders, and all patients were treated with surgical procedures. During operation, collected bladder tissue samples including tissues of apex vesicae and tissues of bladder neck, and all tissues were enveloped with mineral wax. All tissues were detected for nNOS, iNOS, and eNOS respectively in tissues of apex vesicae and tissues of bladder neck,and with normal bladder tissues as control group (bladder tissues of hypospadia, 10 cases), and according to clinical features, to explore the expression of NOS, and to analyze the relationship among them. Results In normal apex vesicae tissues, all cases stained with nNOS, and distributed among bundles of smooth muscles, and surface of smooth muscles and interstitial tissue, histochemica;score (HS) 2.8-4.0 and 1.2-2.7. There were no stained cells in bladder tissues of iNOS, and HS was very low, HS:0-0. 4 and 0-0.1 ;eNOS mainly distributed in interstitial tissues in rarefaction manners, and mainly in vascular endothelial cell (VEC), and smooth muscles had no stainings the most expression among them was nNOS, and mainly distributed in bladder neck tissues. In neurogenic bladder tissues, the main expression of NOS type was iNOS, and nNOS decreased significantly. eNOS mainly expressed in VEC among interstitial tissues, and had no staining in smooth muscle cells and collagenoblast and rarefaction of microvessel in bladder tissues, and microvessel density decreased significantly than normal bladder tissues. Microvessal density(MVD) in bladder tisssus (6. 8± 3.2/100per square) was less than that in normal tissues (16.7±6.3/100 per square). Conclusions In normal bladder tissues, nNOS mainly distributes in bladder neck and urethra, and nitric oxide mainly derives from nNOS. Much more matrix fibers, fewer nitrogenergic nerves, and less nNOS expression are seen in neurogenic bladder interstitial tissue. There are more iNOS expressions in bladder tissues,and NO is mainly derived from iNOS, and it may play an important role in pathological bladder tissues, especially in fibrosis of bladder wall. eNOS may be considered as angiopoietic labeling, and may evaluate the blood supply of bladder.
ABSTRACT
Objective To investigate the genetic association between the inducible nitric oxide synthase (NOS) 2A gene and stroke with a history of coronary artery disease ( CAD). Methods 708 patients with stroke and 235 healthy controls were recruited in this study, and the stroke group was delaminated into 2 subgroups according to the history of CAD. SNP rs28944190, an A to C base change located in intron 22 of the gene, was used as a genetic marker. PCR-based restriction fragment length polymorphism analysis was applied to genotype rs28944190 (Hac Ⅲ site). Results The x2 test showed no association between patients with stroke and healthy controls. Of 708 patients, 94 had a history of CAD and the frequency of allele C of rs28944190 was significantly higher in patients with a history of CAD than those without (23.9% vs 16.6%, x2 =5.629, df= 1, P =0.018, OR = 1.580, 95% CI 1.083—2.306), especially in male patients (x2 = 8. 592, df= 1, P = 0. 003, OR = 1. 983, 95% CI 1. 255—3. 134). The frequency of genotype AA + AC of rs28944190 was significantly higher in patients with a history of CAD than those without such a history (47.9% vs 30. 8%, x2 = 10. 761, df= 1, P = 0. 001, OR = 2. 065, 95% CI 1.34—3.19), especially in male patients (x2 = 15. 762, df= 1, P =0. 000, OR =2. 985, 95% CI 1.74—5. 12). Conclusion The present study suggests that the NOS2A gene is unlikely to contribute to the etiology of stroke.
ABSTRACT
Objective To observe the protection of human keratinocyte cell line, HaCaT cell, from UVA damage by resveratrol and its possible mechanism. Methods HaCaT cells were incubated with or without 0.01 mmol/L or 0.1 mmol/L resveratrol after exposure to 5 J/cm2 UVA irradiation. Unirradiated HaCaT cells-without the treatment with resveratrol served as the control. After another 24-hour culture, MTT assay was used to detect the proliferation of cells, RT-PCR and Western-blot to measure the iNOS mRNA and protein expression respectively, electron microscopic technique to observe the changes in cell ultrastructure. Results After irradiation with UVA of 5 J/cm2, the proliferation of HaCaT cells decreased with the absorbance at 490 nm descending from 0.889±0.083 to 0.542±0.004, while a significant increase was observed in the relative expression level of iNOS mRNA and protein in HaCaT cells (1.532±0.041 vs 0.009±0.003, 1.331 ±0.046 vs 0.003±0.001, both P < 0.05) with the presence of typical apoptotic cells. The treatment with 0.01 and 0.1 mmol/L resveratrol significantly promoted the proliferation of irradiated cells with the absorbance at 490 nm being 0.753±0.435 and 0.892±0.173 respectively, but inhibited the mRNA (0.853±0.038 vs 1.532±0.041, 0.392±0.033 vs 1.532±0.041, both P< 0.05) and protein expression level (0.809±0.018 vs 1.331±0.046, 0.412±0.026 vs 1.331±0.046, both P< 0.05) of iNOS in irradiated cells, and the resveratrol of 0.1 mmol/L was more effective than that ofO.01 mmol/L in all tested parameters (P< 0.05). Furthermore, no apoptofic cells or necrotic cells were observed in irradiated ceils incubated with resveratrol. Conclusion Resveratrol effectively protects HaCaT cells from UVA damage, which may be related to the inhibition of UVA-induced iNOS expression by resveratrol.