Effect of aspirin on prostanoids and nitric oxide production in streptozotocin-diabetic rats with ischemic retinopathy

The importance in experimental diabetic retinopathy of prostacyclin and nitric oxide (NO), as well as the possible effect of acetylsalicylic acid (ASA), are well known. To investigate the effect of two doses of aspirin in the prevention of retinal ischemia in streptozotocin-diabetic rats, we compared nondiabetic rats and diabetic rats after 1, 2 and 3 months of diabetes, and diabetic rats treated with 2 mg or 10 mg ASA/kg per day p.o. from the first day of diabetes.The parameters determined after 1, 2 and 3 months of development were platelet aggregation, thromboxane B(2) (TxB(2)) production, 6-keto-prostaglandin F(1)(alpha) (stable metabolite of prostacyclin), NO, plasma nitrites/nitrates, and percentage retinal surface occupied by horseradish peroxidase (HRP)-permeable vessels. In diabetic rats platelet aggregation and thromboxane concentration were increased, and prostacyclin, NO and area occupied by HRP-permeable vessels were decreased.Acetylsalicylic acid reduced platelet aggregation, and lowered thromboxane production by 82%-99%. Prostacyclin production was inhibited by 92%-95% with 10 mg ASA/kg per day, and by 8%-20% with 2 mg ASA/kg per day. In diabetic rats NO production increased after 2 and 3 months of treatment to levels seen in nondiabetic rats. The reduction in HRP-permeable retinal surface decreased from a maximum of 87% in DR to 51% after treatment with 2 mg ASA/kg per day, and to 62% after 10 mg ASA/kg per day. We conclude that ASA (2 mg/kg per day and 10 mg/kg per day) increased NO production in streptozotocin-diabetic rats and reduced the degree of retinal ischemia.     

阿斯匹林与尼莫地平合用对血小板聚集的影响

目的观察阿斯匹林与尼莫地平合用对兔体外和大鼠半离体血小板聚集的影响。方法用比浊法测量血小板的聚集率。结果在体外和半离体实验中，两药合用均有浓度（剂量）依赖性地抑制由二磷酸腺苷（ADP）和胶原诱导的血小板聚集，合用后Q值均大于1。结论阿斯匹林与尼莫地平合用有协同抑制血小板聚集作用。     

阿司匹林与绞股蓝皂苷合用对血小板聚集的影响

目的：观察阿司匹林与绞股蓝皂苷合用对大鼠体外和体内血小板聚集的影响。方法：用比浊法测定血小板的聚集率和用血栓法测定血栓湿重。结果：在体内、外的实验中,两药合用均有剂量依赖性地抑制由ADP和胶原诱导的血小板聚集,合用后Q值均大于1。结论：阿司匹林与绞股蓝皂苷合用协同抑制血小板聚集作用。     

Effects of endothelial nitric oxide synthase gene polymorphisms on platelet function,nitric oxide release,and interactions with estradiol

Impaired platelet-derived nitric oxide (NO) contributes to acute coronary syndromes by enhancing platelet recruitment and thrombus formation. Polymorphic variants of the endothelial NO synthase (eNOS) gene have been associated with cardiovascular diseases. To examine whether eNOS variants affect platelet-derived NO and platelet function, and to assess the effects of estradiol on platelet function, we studied platelets from 47 healthy caucasians who were genotyped for eNOS polymorphisms in the promoter region (T-786 C), in intron 4, and in exon 7 (Glu298Asp). Platelet aggregation, platelet-derived NO and superoxide production were measured in control samples and samples pretreated with 17-alpha-estradiol (10 nmol/l). The occurrence of variants in the promoter region (P = 0.002) or in exon 7 (P = 0.007), but not in intron 4 (P > 0.05), were associated with lower levels of platelet-derived NO. An increased (P = 0.047) release of superoxide was observed with platelets from subjects with the variant in the promoter region, but not with other eNOS genetic variants. The eNOS gene polymorphisms did not affect ADP-induced platelet aggregation (P > 0.05). However, estradiol significantly increased platelet aggregation (P = 0.004), and platelet-derived superoxide (P = 0.047) in individuals homozygous for the variant in exon 7, but not in subject with other genotypes. These data demonstrate that the eNOS variants in the promoter region and in exon 7 decrease platelet-derived NO and that estradiol significantly increases platelet aggregation in homozygous for the variant in exon 7 but not in subjects with other genotypes, suggesting that eNOS variants may influence the thrombotic response.     

Co-induction of nitric oxide synthase and cyclo-oxygenase: interactions between nitric oxide and prostanoids.

1. Lipopolysaccharide (LPS) co-induces nitric oxide synthase (iNOS) and cyclo-oxygenase (COX-2) in J774.2 macrophages. Here we have used LPS-activated J774.2 macrophages to investigate the effects of exogenous or endogenous nitric oxide (NO) on COX-2 in both intact and broken cell preparations. NOS activity was assessed by measuring the accumulation of nitrite using the Griess reaction. COX-2 activity was assessed by measuring the formation of 6-keto-prostaglandin F1 alpha (6-keto-PGF1 alpha) by radioimmunoassay. Western blot analysis was used to determine the expression of COX-2 protein. We have also investigated whether endogenous NO regulates the activity and/or expression of COX in vivo by measuring NOS and COX activity in the lung and kidney, as well as release of prostanoids from the perfused lung of normal and LPS-treated rats. 2. Incubation of cultured murine macrophages (J774.2 cells) with LPS (1 microgram ml-1) for 24 h caused a time-dependent accumulation of nitrite and 6-keto-PGF1 alpha in the cell culture medium which was first significant after 6 h. The formation of both 6-keto-PGF1 alpha and nitrite elicited by LPS was inhibited by cycloheximide (1 microM) or dexamethasone (1 microM). Western blot analysis showed that J774.2 macrophages contained COX-2 protein after LPS administration, whereas untreated cells contained no COX-2. 3. The accumulation of 6-keto-PGF1 alpha in the medium of LPS-activated J774.2 macrophages was concentration-dependently inhibited by chronic (24 h) exposure to sodium nitroprusside (SNP; 1-1000 microM).(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of low-dose aspirin treatment on human platelet aggregation

1. The effect of aspirin 50 mg/day during 28 days on human platelet aggregation (PAG) induced by ADP and collagen has been studied in 12 healthy volunteers. 2. The results show that aspirin treatment reduces both ADP and collagen-induced PAG (P less than 0.01). 3. Maximal inhibition of PAG appears in the second week of treatment for ADP-induced PAG (41.1%) and in the fourth week for collagen-induced PAG (50.2%). No significant differences exist between the controls taken along the treatment during the 28 days of the study. 4. The data suggest that the low-dose of aspirin tested provides adequate PAG inhibition to be used in the prophylaxis of thromboembolism patients, thus avoiding intolerances to higher dosages.     

Inhibitory effects of copper-aspirin complex on platelet aggregation

目的：研究阿司匹林铜（ＣｕＡｓｐ）对血小板聚集性的影响及其机制．方法：用Ｂｏｒｎ氏法测定ＣｕＡｓｐ对兔血小板聚集性的影响．用荧光光度法和放射免疫法观察ＣｕＡｓｐ对兔血小板５羟色胺的释放和ＴＸＢ２的产生及血浆中ＴＸＢ２和６ｋｅｔｏＰＧＦ１α水平的影响．结果：ＣｕＡｓｐ体外呈浓度依赖性抑制花生四烯酸（ＡＡ）诱导的血小板聚集和５羟色胺的释放（ＩＣ５０分别为１７和１９μｍｏｌ·Ｌ－１，９５％可信限为９－３３和１０－３０μｍｏｌ·Ｌ－１），且抑制ＴＸＢ２的产生（Ｐ＜００５）．ＣｕＡｓｐ１０ｍｇ·ｋｇ－１灌胃选择性抑制ＡＡ诱导聚集．降低血浆ＴＸＢ２，同时升高６ｋｅｔｏＰＧＦ１α的水平（Ｐ＜００５）．结论：ＣｕＡｓｐ体内外均有比Ａｓｐ更强的抗血小板聚集作用．     

The effect of endothelium-derived nitric oxide on ex vivo whole blood platelet aggregation in man

Summary The effects of changes in endogenous endothelium-derived nitric oxide (NO) on forearm blood flow and ex vivo platelet aggregation have been studied in 7 healthy volunteers.Measurements were made of forearm blood flow and ex vivo collagen-stimulated platelet aggregation during unilateral brachial artery infusions of saline, acetylcholine (ACh), N^G monomethyl-L-arginine (L-NMMA), and prostacyclin (PGI₂). The uninfused arm acted as a control.Forearm blood flow was increased by ACh, an agent which stimulates NO release, and decreased by L-NMMA, an agent which stereospecifically inhibits NO synthesis.Collagen-stimulated platelet aggregation measured ex vivo in whole blood draining the infused arm was unaltered by either ACh or L-NMMA. Conversely, PGI₂, an agent which acts independently of NO, caused an increase in forearm blood flow which was accompanied by significant inhibition of platelet aggregation.The results suggest that release of endothelium-derived NO in quantities sufficient to cause substantial changes in blood vessel tone does not lead to changes in platelet aggregation in the blood flowing through the vessels. It is, however, still possible that endothelium-derived NO modulates platelet activity at the level of the endothelium. Present address: Department of Medicine and Therapeutics, University of Aberdeen Polwarth Building, Forrester Hill, Aberdeen, UK     

Effect of oral aspirin dose on platelet aggregation and vascular prostacyclin (PGI2) synthesis in humans and rabbits

Large doses of oral aspirin inhibit platelet aggregation and vascular synthesis of the antiaggregatory vasodilator prostaglandin I2 (PGI2) by irreversibly acetylating the cyclooxygenase enzyme. In order to determine if one can achieve selective inactivation of platelet cyclooxygenase using oral doses of aspirin, we studied human and rabbit platelet aggregation and rabbit aortic synthesis of PGI2 before and 3 hr after various doses of aspirin. In rabbits, lower doses of aspirin produced a major inhibition of platelet aggregation and a minor inhibition of PGI2 synthesis, while higher doses of aspirin inhibited both platelet aggregation and vascular PGI2 synthesis. In humans, we found that a dose equivalent to approximately 1/4 of one 300 mg aspirin tablet consistently produced a major inhibition of cyclooxygenase-dependent platelet aggregation in a pattern similar to the inhibition of rabbit platelet aggregation where the majority of rabbit PGI2 synthetic capacity was not inhibited. In another rabbit study, we found that it takes the vasculature over 24 hr to return to control PGI2 synthetic capacity following a single, high dose of oral aspirin. In conclusion, we speculate that approximately 1/4 of an aspirin tablet, which inhibits a major portion of cyclooxygenase-dependent human platelet aggregation, may not inhibit a major portion of vascular cyclooxygenase-dependent PGI2 synthesis and may be more efficacious as an antithrombotic agent in man than are higher doses of aspirin.     

Effect of clonixin and aspirin on platelet aggregation in human volunteers.

The effect of clonixin and aspirin on platelet function was assessed in healthy volunteers. Both drugs inhibited secondary platelet aggregation and prolonged bleeding time, but the effect of clonixin was significantly less than that of aspirin. Hemorrhagic complications are less likely after clonixin than after aspirin.     

Inhibition of rat platelet aggregation by the diazeniumdiolate nitric oxide donor MAHMA NONOate

1. Inhibition of rat platelet aggregation by the nitric oxide (NO) donor MAHMA NONOate (Z-1-N-methyl-N-[6-(N-methylammoniohexyl)amino]diazen-1-ium-1,2-diolate) was investigated. The aims were to compare its anti-aggregatory effect with vasorelaxation, to determine the effects of the soluble guanylate cyclase inhibitor, ODQ (1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one), and to investigate the possible role of activation of sarco-endoplasmic reticulum calcium-ATPase (SERCA), independent of soluble guanylate cyclase, using thapsigargin. 2 MAHMA NONOate concentration-dependently inhibited sub-maximal aggregation responses to collagen (2-10 micro g ml(-1)) and adenosine diphosphate (ADP; 2 micro M) in platelet rich plasma. It was (i). more effective at inhibiting aggregation induced by collagen than by ADP, and (ii). less potent at inhibiting platelet aggregation than relaxing rat pulmonary artery. 3. ODQ (10 micro M) caused only a small shift (approximately half a log unit) in the concentration-response curve to MAHMA NONOate irrespective of the aggregating agent. 4. The NO-independent activator of soluble guanylate cyclase, YC-1 (3-(5'-hydroxymethyl-2'-furyl)-1-benzyl indazole; 1-100 micro M), did not inhibit aggregation. The cGMP analogue, 8-pCPT-cGMP (8-(4-chlorophenylthio)guanosine 3'5' cyclic monophosphate; 0.1-1 mM), caused minimal inhibition. 5. On collagen-aggregated platelets responses to MAHMA NONOate (ODQ 10 micro M present) were abolished by thapsigargin (200 nM). On ADP-aggregated platelets thapsigargin caused partial inhibition. 6. Results with S-nitrosoglutathione (GSNO) resembled those with MAHMA NONOate. Glyceryl trinitrate and sodium nitroprusside were poor inhibitors of aggregation. 7. Thus inhibition of rat platelet aggregation by MAHMA NONOate (like GSNO) is largely ODQ-resistant and, by implication, independent of soluble guanylate cyclase. A likely mechanism of inhibition is activation of SERCA.     

Effects of pentoxifylline and its metabolites on platelet aggregation in whole blood from healthy humans

It is known that pentoxifylline inhibits platelet aggregation in vitro, but the effects from pentoxifylline and its main metabolites: 3,7-dimetyl-1(5 hydroxyhexyl)xanthine (R-M1 and S-M1), 3,7-dimetyl -1(4-carboxybutyl)xanthine (M4), 3,7-dimetyl -1(3-carboxypropyl)xanthine (M5), on platelet aggregation in whole blood in vitro and in vivo have not been studied. We found that pentoxifylline, rac-M1, R-M1, S-M1 and M4 significantly inhibit ADP induced platelet aggregation in whole blood in vitro in a concentration-dependent manner, R-M1 being the most potent followed by rac-M1, S-M1, pentoxifylline, and M4. In this series of experiments the effects on aggregation induced ATP-release were less pronounced and were only significant after treatment with pentoxifylline, rac-M1 and R-M1, but the potency order appears to be the same. Since the metabolites are not available for use in humans, and also since each substance would be extensively metabolised in vivo, we made an attempt to estimate the relative contribution of each substance to the total effect of pentoxifylline in vivo. Previously published concentrations of pentoxifylline and these metabolites in humans, after administration of pentoxifylline, were used in combination with the potency ratios from this study. The findings from these calculations were that the main effect in vivo comes from S-M1 followed by pentoxifylline, the other metabolites contribute less than 10% each. In conclusion: in the following potency order R-M1, rac-M1, pentoxifylline, S-M1 and M4 all have significant effects on platelet aggregation in whole blood in vitro. However, it appears that the main effects in vivo are caused by S-M1 and pentoxifylline.     

血栓弹力图评价阿司匹林与氯吡格雷对脑梗死患者血小板聚集功能的影响

目的：研究通过血栓弹力图（TEG）比较阿司匹林与氯吡格雷对血小板聚集功能的影响,探讨TEG 在脑梗死患者抗血小板药物个体化治疗中的意义。方法采用 TEG 检测100例急性脑梗死患者抗血小板药物治疗后花生四烯酸（AA）和腺苷二磷酸（ADP）受体途径诱导的血小板抑制率,患者抗血小板药物治疗包括阿司匹林组（n ＝50）、氯吡格雷组（n ＝50）。比较两组血小板抑制率、患者神经功能评分、卒中复发率。结果TEG 显示阿司匹林组的血小板抑制率为（85．23±21．98）％,显著高于氯吡格雷组的（47．31±22.37）％（t ＝7．340,P ＝0．005）;TEG 显示阿司匹林与氯吡格雷血小板抑制率为良好的患者,神经功能恢复更佳（P ＜0．05）,其中阿司匹林抑制率显示良好的患者1年内卒中复发率更低（χ2＝4．460,P ＝0．035;χ2＝7.232,P ＝0．007）。结论TEG 对脑梗死患者抗血小板个体化治疗具有指导作用,可用于监测药物疗效以及评估患者预后。     

香菇多糖对巨噬细胞一氧化氮和一氧化氮合酶活性的影响

目的研究香菇多糖(LTN)诱导巨噬细胞的一氧化氮(NO)生成和一氧化氮合酶(iNOS)的活性,探讨LTN的免疫调节作用机理.方法采用Griess反应和荧光法测定不同剂量的LTN作用小鼠腹腔巨噬细胞后NO的生成量和iNOS活性.观察mRNA转录抑制剂、蛋白质合成抑制剂和iNOS抑制剂对巨噬细胞NO的生成和iNOS活性的影响.结果LTN能使小鼠腹腔巨噬细胞NO生成增加,iNOS活性增高,并呈作用剂量依赖关系.3种抑制剂均能抑制LTN诱导的小鼠腹腔巨噬细胞N0的生成和iNOS活性.结论LTN能刺激小鼠腹腔巨噬细胞提高iNOS活性和NO的生成.提示LTN的免疫调节作用机制可能与LTN刺激巨噬细胞NO生成有关.     

香菇多糖对巨噬细胞一氧化氮和一氧化氮合酶活性的影响

目的研究香菇多糖(LTN)诱导巨噬细胞的一氧化氮(NO)生成和一氧化氮合酶(iNOS)的活性,探讨LTN的免疫调节作用机理.方法采用Griess反应和荧光法测定不同剂量的LTN作用小鼠腹腔巨噬细胞后NO的生成量和iNOS活性.观察mRNA转录抑制剂、蛋白质合成抑制剂和iNOS抑制剂对巨噬细胞NO的生成和iNOS活性的影响.结果LTN能使小鼠腹腔巨噬细胞NO生成增加,iNOS活性增高,并呈作用剂量依赖关系.3种抑制剂均能抑制LTN诱导的小鼠腹腔巨噬细胞N0的生成和iNOS活性.结论LTN能刺激小鼠腹腔巨噬细胞提高iNOS活性和NO的生成.提示LTN的免疫调节作用机制可能与LTN刺激巨噬细胞NO生成有关.     

Effects of gamma-butyrobetaine and mildronate on nitric oxide production in lipopolysaccharide-treated rats

Production of nitric oxide was measured in lipopolysaccharide-treated rats (10 mg/kg, 4 hr) using the electron paramagnetic resonance method. As compared to the control animals, the nitric oxide level in liver of lipopolysaccharide-treated rats increased from 27.6+/-4.7 to 1485+/-129 ng/g tissue, in heart from 4.8+/-0.7 to 271+/-26 ng/g tissue, in blood from 33.6+/-12.4 to 638+/-136 ng/g tissue, in kidney from 3.3+/-0.5 to 356+/-31 ng/g tissue, in brain cortex from 46.0+/-3.4 to 227+/-27 ng/g tissue, in cerebellum from 27.7+/-2.6 to 218+/-30 ng/g tissue, and in testes from 13.8+/-1.1 to 86+/-8 ng/g tissue. Administration of the antiischaemic drug, mildronate (120 mg/kg) caused a significant twofold decrease of the nitric oxide level in brain cortex and cerebellum 1 hr after drug administration. Its natural analogue gamma-butyrobetaine (30 mg/kg) triggered a twofold decrease of the nitric oxide concentration in all studied tissues 30 min. after the administration. Nitric oxide reached the initial level 2 hr later. Neither mildronate nor gamma-butyrobetaine could inhibit the inducible nitric oxide synthase in vitro. Analogues of gamma-butyrobetaine appear to be prospective drugs for the treatment of circulatory complications of sepsis.     

普伐他汀对人内皮祖细胞一氧化氮合成的影响

目的观察普伐他汀对人内皮祖细胞（EPCs）一氧化氮（NO）合成的影响。方法密度梯度离心法获取外周血单个核细胞,培养7d后,收集贴壁细胞并分别加入普伐他汀,10μmol/L及100μmol/L干预48h,免疫组化、荧光显微镜和流式细胞仪鉴定EPC,用RT-PCR方法测定对细胞内皮型一氧化氮合酶（eNOS）mRNA表达的影响,并用硝酸还原酶法测定培养液中一氧化氮（NO）的水平。结果普伐他汀组的人内皮祖细胞eNOS mRNA的表达、NO的合成明显增加。结论普伐他汀可增加人内皮祖细胞eNOS mRNA的表达和NO的合成     

Dose-dependent aspirin hydrolysis and platelet aggregation in patients with atherosclerosis.

In a double blind, randomized trial, the effects of aspirin (1, 5, and 15 mg/kg) were compared with the changes in platelet aggregation at 6 and 24 hours after dosage. It is found that there is a negative correlation between aspirin hydrolysis velocity in blood and capability of aspirin to decrease platelet aggregation with ADP and collagen in patients with atherosclerosis. Relationship between these parameters depends on aspirin dosage. The correlation was more marked for low doses of aspirin. It is suggested that the effect of aspirin in low dosage on platelet aggregation might be ineffective in many patients without control of aspirin hydrolysis velocity in blood.