NOS抑制剂对缺血再灌注脑组织rCBF及白细胞浸润的 …

目的 应用非选择性ＮＯＳ抑制剂Ｌ－ＮＡＭＥ和选择性ｉＮＯＳ抑制剂ＡＧ治疗鼠大脑中动脉缺血再灌注损伤，通过对脑梗死体积，ｒＣＢＦ和白细胞浸润程度的观察，研究探讨不同类型ＮＯＳ抑制剂治疗脑梗死的机制。方法 采用线栓法制作鼠大脑中动脉缺血再灌注模型，不同缺血及再灌注时间测定脑梗死体积，ｒＣＢＦ，缺血脑组织ＭＰＯ酶活性，结果 应用Ｌ－ＮＡＭＥ（１５ｍｇ／ｋｇ，ｉｐ）不但障碍再灌注后ｒＣＢＦ的恢复，也增加缺     

实验性高血糖大鼠缺血脑组织一氧化氮和中性白细胞浸润的变化

目的　探讨高血糖在缺血再灌注脑损伤中的作用。　　方法　利用链尿酶素诱发大鼠发生高血糖 ,以线栓法复制大鼠大脑中动脉区缺血再灌注模型 ,研究该条件下脑组织损伤、中性白细胞浸润和一氧化氮 (NO)含量变化与血糖的关系。　　结果　高血糖组脑缺血再灌注 1 2h、2 4h和 48h时脑梗死体积、缺血脑组织中性白细胞浸润程度 (MPO活性 )及NO含量均显著高于正常血糖组。　　结论　高血糖加重大鼠脑缺血再灌注损伤 ,其葡萄糖毒性作用机制可能是通过缺血脑组织内中性白细胞和NO的致伤作用实现的。     

PAF受体拮抗剂对缺血/再灌注脑损害的作用研究

目的血小板活化因子(platelet activating factor，PAF)与缺血／再灌注(ischemic reperfusion，IR)脑损害有密切关系，此研究拟用PAF受体拮抗剂WEB2086对PAF、炎性细胞浸润及一氧化氮(NO)在脑IR损害中的作用和机制进行探讨。方法采用线栓法制成大鼠大脑中动脉IR模型，对IR脑组织NO含量、局部脑血流量(rCBF)、中性白细胞髓过氧化物酶(MPO)及脑梗死体积进行测定。结果：PAF受体拮抗剂WEB2086对IR脑组织NO的产生有明显影响．且可明显改善IR脑组织的rCBF和显著降低MPO活性，最终减轻局部IR脑损害。结论PAF受体拮抗剂WEB2086对IR脑组织的保护作用与NO有关。     

大鼠脑缺血-再灌流后脑内MPO活性变化及组织损伤病理进观察

目的 :通过测定大鼠局灶性脑缺血 -再灌流后不同时点脑组织中 MPO活性变化 ,探讨炎症反应与脑缺血 -再灌流损伤的关系。方法 :用线栓法制备大鼠大脑中动脉缺血 -再灌流模型 ,检测缺血 3小时后再灌流不同时点脑组织中 MPO活性、脑梗死体积及光镜病理学变化。结果 :缺血组脑组织有 MPO活性升高、中性粒细胞浸润 ,以再灌流后 4 8、 72小时最为明显 ,脑梗死体积、神经元变性程度随再灌流时间延长而加重。结论 :局灶性缺血脑组织中MPO活性与缺血 -再灌流损伤间具有一定的关系 ,炎症反应是加重脑组织损伤的重要因素。     

局灶性脑缺血和再灌注损伤中一氧化氮合成酶和髓过氧化物酶相互关系研究

目的　研究局灶性脑缺血再灌注期 ,一氧化氮合酶 (NOS)与髓过氧化物酶 (MPO)之间的关系。方法　采用大鼠大脑中动脉线栓 /再灌注模型 ,根据不同再灌注时间对大鼠分组 ,同时设定假手术组 ,取缺血区相邻脑组织分别检测NOS和MPO活性 ,对二者进行相关性分析。结果　假手术组可测得一定的NOS活性 ,再灌注早期 ( 1h)在缺血相邻区即可检测到一定的NOS活性 ,以后随着再灌注时间延长 ,测得NOS活性逐渐升高 ,再灌注 2 4h活性达最高。而假手术组及再灌注早期 ( 1h)代表白细胞浸润程度的MPO活性几乎测不到 ,再灌注早期 6h开始可测到一定的MPO活性 ,以后随着再灌注时间延长 ,测得MPO活性逐渐升高 ,再灌注 2 4hMPO活性达最高。NOS和MPO呈高度正相关。结论　在局灶性脑缺血 /再灌注期NO与白细胞浸润具有相关性     

氨基胍对大鼠局灶性脑缺血再灌注神经细胞损伤的影响

目的　探讨氨基胍 (AG)对脑缺血 再灌注 (IR)神经细胞损伤的影响。方法　用线栓法制作大鼠局灶性脑缺血 再灌注模型 ,动物缺血 2小时后给予腹腔注射AG 10 0mg·kg-1,取不同再灌注时间测定大鼠脑匀浆NOS活性、髓过氧化物酶 (MPO)活性和脑梗死体积。结果　再灌注后 12～ 72小时 ,AG显著降低了iNOS活性 ,且于再灌注后 2 4小时达最大抑制率。再灌注后 2 4～ 72小时 ,AG减少髓过氧化物酶 (MPO)含量。再灌注后 2 4～ 72小时 ,AG减少梗死体积。结论　AG对脑缺血 再灌注神经细胞损伤具有一定的保护作用。     

局灶性鼠脑缺血时一氧化氮及一氧化氮合酶抑制剂的作用机制

目的研究一氧化氮在鼠脑局灶性脑缺血再灌注损伤中的作用。方法用线栓法建立大鼠大脑中动脉区缺血再灌注模型，分别用选择性和非选择性诱导型一氧化氮合酶抑制剂对鼠脑局灶性缺血再灌注过程中脑组织一氧化氮的变化规律及可能作用进行探讨。结果非选择性一氧化氮合酶抑制剂（Ｌ－ＮＡＭＥ）可加重局灶性脑缺血性损害，而选择性诱导型一氧化氮合酶抑制剂（ａｍｉｎｏｇｕａｎｉｄｉｎｅ，ＡＧ）具有明确的脑保护作用。结论不同类型的一氧化氮合酶所产生的一氧化氮在脑局灶性缺血性损害中具有不同的作用。     

尤瑞克林对大鼠局灶性脑缺血再灌注损伤后炎性反应的影响

目的研究尤瑞克林对大鼠局灶性脑缺血再灌注损伤后炎性反应的影响。方法将90只SD大鼠随机分为3组:假手术组,对照组,治疗组。采用线栓法建立大鼠大脑中动脉闭塞再灌注模型,缺血2 h后,拔出线栓,恢复灌注24 h,观察大鼠神经功能缺损症状、脑梗死体积、脑组织中白细胞浸润、MPO活性、IL-1和ICAM-1的表达。结果 (1)假手术组大鼠在神经功能缺损评分、脑梗死体积均低于对照组,有显著的统计学差异(P<0.01);脑组织中白细胞浸润程度、髓过氧化物酶(MPO)活性、ICAM-1和IL-1的表达均较对照组低,统计学差异明显(P<0.01);(2)治疗组与对照组相比,大鼠的神经功能缺损评分低、脑梗死体积小,有显著统计学差异(P<0.01);白细胞浸润程度、MPO活性、ICAM-1和IL-1的表达均较对照组减少(P<0.01)。结论尤瑞克林可通过抑制大鼠脑缺血再灌注损伤后的炎性反应来实现其神经保护作用。     

大剂量甲基强的松龙对大鼠局灶性缺血再灌注脑损伤的影响

目的 探讨大剂量甲基强的松龙（ＭＰ）对局灶性缺血再灌注大鼠脑保护作用的机理。方法采用大鼠局灶性脑缺血再灌注模型,观察缺血前后应用大剂量ＭＰ对大鼠大脑中动脉闭塞侧梗死体积的影响以及脑含水量的变化,同时观察脑组织病理学改变。结果 缺血前后ＭＰ治疗组大脑中动脉供血区脑梗死体积较盐水对照组明显减小（均Ｐ〈０．０１）;缺血性前后ＭＰ治疗组与盐水对照组脑含水量比较无明显差别（均Ｐ〉０．０５）。病理学发现盐水对照组织血管周围可见巨噬细胞浸润,而ＭＰ治疗组无此病理改变。结论 大剂量ＭＰ可改善缺血性脑损伤,其机理与减小缺血区梗死体积、抑制脑组织巨噬细胞浸润有关。     

大鼠脑缺血再灌注后三磷酸腺苷含量和细胞凋亡的变化及药物的影响

目的探讨大鼠脑缺血再灌注后脑ATP含量和脑梗死体积、细胞凋亡的变化及桂哌齐特的影响。方法雄性SD大鼠79只,随机分为假手术组、模型组和桂哌齐特处理组,应用线栓法建立大脑中动脉缺血再灌注模型,采用HPLC法、TUNEL法及TTC染色分别测定脑组织ATP的含量、凋亡细胞数目及脑梗死体积。结果大鼠缺血20、60min和再灌注15、60min脑ATP含量较基础水平降低,再灌注15、60min处理组脑ATP含量较模型组升高;再灌注22h、70h处理组凋亡细胞数目低于模型组,脑梗死体积缩小。结论大鼠局灶性缺血再灌注早期脑ATP含量均降低,梗死灶周边凋亡细胞数目明显增多;桂哌齐特显著阻止脑梗死范围进一步扩大与其改善再灌注后脑组织能量代谢状态抑制细胞凋亡可能有关。     

Involvement of nitric oxide pathway in the acute anticonvulsant effect of melatonin in mice

Melatonin, the major hormone produced by the pineal gland, is shown to have anticonvulsant effects. Nitric oxide (NO) is a known mediator in seizure susceptibility modulation. In the present study, the involvement of NO pathway in the anticonvulsant effect of melatonin in pentylenetetrazole (PTZ)-induced clonic seizures was investigated in mice. Acute intraperitoneal administration of melatonin (40 and 80 mg/kg) significantly increased the clonic seizure threshold induced by intravenous administration of PTZ. This effect was observed as soon as 1 min after injection and lasted for 30 min with a peak effect at 3 min after melatonin administration. Combination of per se non-effective doses of melatonin (10 and 20 mg/kg) and nitric oxide synthase (NOS) substrate L-arginine (30, 60 mg/kg) showed a significant anticonvulsant activity. This effect was reversed by NOS inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME, 30 mg/kg), implying an NO-dependent mechanism for melatonin effect. Pretreatment with L-NAME (30 mg/kg) and N(G)-nitro-L-arginine (L-NNA, 10 mg/kg) inhibited the anticonvulsant property of melatonin (40 and 80 mg/kg) and melatonin 40 mg/kg, respectively. Specific inducible NOS (iNOS) inhibitor aminoguanidine (100 and 300 mg/kg) did not affect the anticonvulsant effect of melatonin, excluding the role of iNOS in this phenomenon, while pretreatment of with 7-NI (50 mg/kg), a preferential neuronal NOS inhibitor, reversed this effect. The present data show an anticonvulsant effect for melatonin in i.v. PTZ seizure paradigm, which may be mediated via NO/L-arginine pathway by constitutively expressed NOS.     

Different effects between 7-nitroindazole and L-NAME on cerebral hemodynamics and hippocampal lesions during kainic acid-induced seizures in newborn rabbits

We examined the effects of 7-nitroindazole (7-NI) and N-omega-nitro-L-arginine methyl ester (L-NAME) on the endogenous nitric oxide (NO) production in vivo, cerebral hemodynamics, and hippocampal lesions to investigate the different roles between endothelial NOS (eNOS) and neuronal NOS (nNOS) during kainic acid (KA)-induced seizures in newborn rabbits. After a pre-treatment with 7-NI (50 mg/kg, i.p.), L-NAME (20 mg/kg, i.v.) or saline (1 ml, i.v.), KA (12 mg/kg, i.v.) was administered. NO production in the brain, regional cerebral blood flow (rCBF), cerebral oxygenation (concentrations of oxyhemoglobin (HbO2), deoxyhemoglobin (HbR), and total hemoglobin (tHb) in the brain tissue), and electroencephalography (EEG) were continuously monitored throughout the experiment lasting at least 60 min after the KA administration. There was a significant increase in NO generation in the brain during KA-induced seizures, which was inhibited by a pre-treatment with 7-NI or L-NAME. KA-induced seizures also increased rCBF significantly, which was inhibited not by 7-NI but by L-NAME. L-NAME pre-treatment caused a significant decrease in HbO2 and a significant increase in HbR during KA-induced seizures, compared with 7-NI and saline pre-treatment. EEG abnormalities and Neuronal damages in the hippocampus were significantly lower in 7-NI- and L-NAME-treated animals respectively, than in saline-treated animals. The present data demonstrated that the selective nNOS inhibitor, 7-NI, attenuated neither rCBF nor cerebral oxygenation during the seizures, while the non-selective NOS (nNOS and eNOS) inhibitor, L-NAME, attenuated both. These findings suggest that NO, probably originating from eNOS, may play an important role in the cerebral circulation. Both 7-NI and L-NAME inhibited the NO production and EEG abnormalities during the seizures that led to less damage to the hippocampus.     

The involvement of endogenous opioids and nitricoxidergic pathway in the anticonvulsant effects of foot-shock stress in mice

The involvement of endogenous opioids and nitric oxide (NO) in the anticonvulsant effects of stress against pentylenetetrazole (PTZ)- or electroconvulsive shock-induced seizures was assessed in mice. The prolonged and intermittent foot-shock stress, which induced opioid-mediated analgesia, had significant protective effects against both seizure types which was reversible by naloxone (0.3, 1 or 2 mg/kg), while brief and continuous foot-shock did not alter the seizure susceptibility. Pre-treatment with non-specific nitric oxide synthase (NOS) inhibitor, N(G)-nitro-L-arginine methyl ester (L-NAME, 1, 2, 5, 10 or 30 mg/kg), but not with specific inducible NOS (iNOS) inhibitor, aminoguanidine (50 or 100 mg/kg), blocked the stress-induced anticonvulsant effects. The lower doses of naloxone (0.3 mg/kg) and L-NAME (2 mg/kg) showed additive effects in blocking the stress-induced anticonvulsant properties. L-arginine at a per se non-effective dose of 20 mg/kg potentiated the stress-induced anticonvulsant properties, an effect which was inhibited by L-NAME but not by aminoguanidine. Furthermore, a low dose of morphine (0.5 mg/kg) showed potentiation with stress in increasing PTZ seizure threshold. This potentiation was reversed by either naloxone or L-NAME at low doses but not by aminoguanidine. Taken together, these results show that NO synthesis, through constitutive but not iNOS, is involved in opioid-dependent stress-induced anticonvulsant effects against electrical and PTZ-induced convulsions.     

脑缺血后cNOSmRNA表达：NO在脑缺血早期的保护作用

在SHR MCAO局灶脑缺血模型基础上,分别测定脑缺血后不同时限NOS活性变化及cNOS mRNA表达水平,并应用外源性NOS抑制剂L-NNA,观察NO代谢变化对脑缺血脑水肿的影响。结果发现脑缺血早期cNOS mRNA表达增加,NOS活性增高,L-NNA则促进脑水肿发生,提示NO在脑缺血早期可能具有保护作用。     

Is intracellular brain pH a dependent factor in NOS inhibition during focal cerebral ischemia?

The interaction between nitric oxide (NO.) and focal cerebral ischemia is multifaceted. Experiments have shown that inhibition of nitric oxide synthase (NOS) either ameliorates or exacerbates focal cerebral ischemia. Recent in vitro experiments have shown that NOS activity is pH-dependent. Previous work from this laboratory has demonstrated that N(G)-nitro-L-arginine-methyl-ester (L-NAME) mitigated cerebral ischemia independent from regional cerebral blood flow (rCBF) changes during moderate focal cerebral ischemia. This study examined the effects of L-NAME inhibition on brain pH(i), rCBF, and NADH redox state during 3 h of severe focal cerebral ischemia. Fifteen fasted rabbits under 1.5% halothane were equally divided into three groups: ischemic controls and two drug groups receiving either 1.0 or 10 mg/kg L-NAME intravenously 30 min prior to ischemia. In the ischemic controls, brain pH(i) declined from 6.95+/-0.04 to 6.60+/-0.05, rCBF declined from 48+/-7 to 10+/-3 ml/100 g/min, and NADH fluorescence increased by 149+/-15% 3 h after onset of ischemia (p<0.01 for all three parameters). L-NAME at either dose did not significantly alter these values. Infarct volume was not significantly different between both the L-NAME treated groups and the ischemic control group. This data suggests that during severe focal cerebral ischemia, NO. mechanisms of injury have a less important punitive role. One possible explanation is that the severity of acidosis secondary to anaerobic metabolism during severe focal cerebral ischemia attenuates NOS activity.     

不同脑缺血和再灌流过程中大鼠脑组织NO含量的动态变化

采用线栓法制成大鼠大脑中动脉梗塞 ( MCAO)模型 ,依 Hb O2 - NO法测定持续性脑缺血和缺血 /再灌流脑组织内 NO含量的变化 ,以探讨不同脑缺血和再灌流过程中 NO的变化规律及其意义。结果 :缺血 3小时受损脑组织 NO水平即增高 ,再灌流后 NO逐步升高 ,而持续性缺血状态下 NO则表现降低后再升高的变化。虽然两组 NO在 7天时均有明显降低 ,但仍高于缺血前水平。认为持续性脑缺血和缺血 /再灌流情况下 NO的变化规律有所不同 ,与缺血脑组织的缺氧及产生 NO所需底物供应缺乏有关 ,且可能与脑组织的损害密切相关     

Nitric oxide mediates cerebral ischemic tolerance in a neonatal rat model of hypoxic preconditioning.

Neuroprotection against cerebral ischemia can be realized if the brain is preconditioned by previous exposure to a brief period of sublethal ischemia. The present study was undertaken to test the hypothesis that nitric oxide (NO) produced from the neuronal isoform of NO synthase (NOS) serves as a necessary signal for establishing an ischemia-tolerant state in brain. A newborn rat model of hypoxic preconditioning was used, wherein exposure to sublethal hypoxia (8% oxygen) for 3 hours renders postnatal day (PND) 6 animals completely resistant to a cerebral hypoxic-ischemic insult imposed 24 hours later. Postnatal day 6 animals were treated 0.5 hour before preconditioning hypoxia with the nonselective NOS inhibitor L-nitroarginine (2 mg/kg intraperitoneally). This treatment, which resulted in a 67 to 81% inhibition of calcium-dependent constitutive NOS activity 0.5 to 3.5 hours after its administration, completely blocked preconditioning-induced protection. However, administration of the neuronal NOS inhibitor 7-nitroindazole (40 mg/kg intraperitoneally) before preconditioning hypoxia, which decreased constitutive brain NOS activity by 58 to 81%, was without effect on preconditioning-induced cerebroprotection, as was pretreatment with the inducible NOS inhibitor aminoguanidine (400 mg/kg intraperitoneally). The protective effects of preconditioning were also not blocked by treating animals with competitive [3-(2-carboxypiperazin-4-yl)propyl-1-phosphonate; 5 mg/kg intraperitoneally] or noncompetitive (MK-801; 1 mg/kg intraperitoneally) N-methyl-D-aspartate receptor antagonists prior to preconditioning hypoxia. These findings indicate that NO production and activity are critical to the induction of ischemic tolerance in this model. However, the results argue against the involvement of the neuronal NOS isoform, activated secondary to a hypoxia-induced stimulation of N-methyl-D-aspartate receptors, and against the involvement of the inducible NOS isoform, but rather suggest that NO produced by the endothelial NOS isoform is required to mediate this profound protective effect.     

Effects of aminoguanidine and cyclooxygenase inhibitors on nitric oxide and prostaglandin production, and nitric oxide synthase and cyclooxygenase expression induced by lipopolysaccharide in the estrogenized rat uterus

BACKGROUND/OBJECTIVE: The aim of our study was first to investigate if there exists an interaction between nitric oxide (NO) and prostaglandin (PG) generation in the estrogenized rat uterus challenged by lipopolysaccharide (LPS), and, secondly, which isoforms of nitric oxide synthase (NOS) and cyclooxygenase (COX) participate in this process. METHODS: To study the effect of LPS and to characterize the isoenzymes involved in the process, specific inhibitors of iNOS (aminoguanidine) and COX-II (meloxicam, nimesulide) and non-specific of COX (indomethacin) were injected intraperitoneally to determine their effect on NO and PG production, and on NOS and COX expression induced by LPS in estrogenized rat uterus. NO production was measured by arginine-citrulline conversion assay and PGE(2)/PGF(2alpha,)by radioconversion. Enzyme expression was evaluated by Western blot analysis. RESULTS: The present work shows that iNOS inhibitor, aminoguanidine, reduced NO and PGE(2)/PGF(2alpha) production induced by LPS injection. Aminoguanidine exerts its effect over the PG metabolism by inhibiting COX-II activity and expression. On the other hand, both indomethacin, a non-selective PG inhibitor, and meloxicam, a COX-II inhibitor, stimulated NO production and reduced PGE(2)/PGF(2alpha) generation. Indomethacin also reduced COX-II and iNOS expression. CONCLUSION: These results indicate that in the estrogenized rat uterus challenged with LPS, PG and NO interact affecting each other's metabolic pathways. The above findings indicate that the interaction between NOS and COX might be important in the regulation of physiopathologic events during pregnancy.