Modulation of adjuvant arthritis by endogenous nitric oxide.

1. The role of endogenous nitric oxide (NO) in adjuvant arthritis in Lewis rats has been studied by use of L-arginine, the amino acid from which NO is synthesized, and NG-nitro-L-arginine methyl ester (L-NAME), an inhibitor of NO synthase. Prolonged modulation (35 days) of the L-arginine: NO pathway in rats was achieved by dissolving test compounds in the drinking water (L-arginine: 3, 10 and 30 mg ml-1; L-NAME: 0.1, 1 and 10 mg ml-1). 2. Arthritis was exacerbated by L-arginine and suppressed by L-NAME in a dose-related fashion. Combined treatment with L-NAME (1 mg ml-1) and L-arginine (30 mg ml-1) did not modify the arthritis. 3. Reduced weight gain, which is a feature of adjuvant arthritis, was modified by these compounds so that L-arginine reduced weight gain whereas L-NAME increased weight gain compared with that in control animals. 4. D-Arginine (30 mg ml-1), NG-nitro-D-arginine methyl ester (D-NAME: 1 mg ml-1) and L-lysine (30 mg ml-1), an amino acid not involved in the generation of NO, were without effect on either arthritis or body weight gain. 5. Antigen-stimulated proliferation of T-lymphocytes as well as generation of nitrite (NO2-) and release of acid phosphatase from macrophages were all enhanced in L-arginine-treated arthritic rats and reduced in L-NAME-treated animals. 6. These results suggest that endogenous NO modulates adjuvant arthritis, possibly by interfering with the activation of T-lymphocytes and/or macrophages.     

Dual effect of nitric oxide in articular inflammatory pain in zymosan-induced arthritis in rats

The contribution of nitric oxide (NO) to articular pain in arthritis induced by zymosan (1 mg, intra articular) in rats was assessed by measuring articular incapacitation (AI). Systemic treatment with the non-selective NO synthase (NOS) inhibitor L-NAME (10 - 100 mg kg(-1) i.p.) or with the selective iNOS inhibitors aminoguanidine (AG; 10 - 100 mg kg(-1) i.p.) or 1400W (0.5 - 1 mg kg(-1) s.c.) inhibited the AI induced by injection of zymosan 30 min later. Local (intra articular) treatment with the NOS inhibitors (L-NAME or AG, 0.1 - 1 micromol; 1400W, 0.01 (micromol) 30 min before zymosan also inhibited the AI. Systemic or local treatment with the NOS inhibitors (L-NAME; AG, 100 mg kg(-1) i.p. or 0.1 micromol joint(-1); 1400W, 1 mg kg(-1) s.c. or 0.01 micromol joint(-1)), 2 h after zymosan did not affect the subsequent AI. Local treatment with the NO donors SNP or SIN-1, 2 h after zymosan did inhibit AI. L-NAME and AG, given i.p. inhibited nitrite but not prostaglandin E(2) (PGE(2)) levels in the joints. L-NAME (100 mg kg(-1)) but not AG (100 mg kg(-1)) increased mean arterial blood pressure. Neither L-NAME, AG nor the NO donor SIN-1 altered articular oedema induced by zymosan. In conclusion, inhibitors of iNOS decrease pain in zymosan arthritis only when given before the zymosan. This was not due to inhibition of articular PGE(2) release or oedema. NO donors also promoted antinociception in zymosan arthritis without affecting oedema.     

Dual effect of nickel on L-arginine/nitric oxide system in RAW 264.7 macrophages

The immunogenic mechanisms of the potent contact allergen nickel are not completely clear. Nitric oxide (NO) serves as a fundamental signalling and effector molecule in the immune system, but little is known about its possible role in immune reactions elicited by nickel. We investigated the effects of nickel on the L-arginine/inducible NO synthase (iNOS) system in a murine macrophage cell line, RAW 264.7. Both LPS-stimulated and non-stimulated RAW 264.7 cells were incubated in the presence of 0–100 μM nickel sulphate for 24 h. Subsequently, NO production, iNOS protein expression, L-arginine uptake and gene expression of iNOS and cationic amino acid transporter systems (CAT) were measured. We found that 100 μM NiSO₄ increased LPS-induced nitrite production as well as the formation of [³H]-L-citrulline from [³H]-L-arginine in the RAW 264.7 cells. Correspondingly, the expression of iNOS gene and protein was also remarkably enhanced. Nevertheless, nickel had an inhibitory effect on L-arginine transport which disappeared gradually upon LPS-stimulation in parallel with an increase in NO output. LPS was found to significantly amplify CAT-3 as well as CAT-2 mRNA expression, mirroring the increase in L-arginine transport. In the range of 1–10 μM, NiSO₄ did not have any additional effect on CAT mRNA expression, but at 100 μM it was able to enhance CAT-1 and CAT-3 mRNA expression upon LPS stimulation.Our data indicate that nickel interferes with macrophages' L-arginine/NOS system on multiple levels. Considering the potent biological effects of NO, these influences may contribute to nickel toxicity.     

Dual impact of a nitric oxide donor, GEA 3175, in human pulmonary smooth muscle

Nitric oxide (NO) donors could constitute an alternative to inhaled NO as treatment in some patients with pulmonary hypertension. Therefore, the present study investigated the relaxation mechanisms of a novel NO donor, 3-(3-chloro-2-methylphenyl)-5-[[4-methylphenyl)sulphonyl]amino]-)hydroxide (GEA 3175) in segments of human pulmonary arteries and bronchioles, which were mounted in microvascular myographs. GEA 3175 induced concentration-dependent relaxations and was more potent in pulmonary arteries than in bronchioles. A blocker of soluble guanylyl cyclase, 1H-[1,2,4]oxadiazolo[4,3,-a]quinoxalin-1-one (ODQ), and iberiotoxin, a blocker of large-conductance calcium-activated K channels, both reduced relaxations induced by GEA 3175 in pulmonary arteries and bronchioles. Combining of ODQ and iberiotoxin did not produce additional inhibition. GEA 3175 relaxation is mediated through guanylyl cyclase-dependent mechanisms followed by activation of large-conductance calcium-activated K(+) channels. The dilatation of both pulmonary small arteries and airways by GEA 3175 seems advantageous, if it is considered administered as inhalation therapy for pulmonary hypertension.     

Recent developments in nitric oxide donor drugs

During the 1980s, the free radical, nitric oxide (NO), was discovered to be a crucial signalling molecule, with wide-ranging functions in the cardiovascular, nervous and immune systems. Aside from providing a credible explanation for the actions of organic nitrates and sodium nitroprusside that have long been used in the treatment of angina and hypertensive crises respectively, the discovery generated great hopes for new NO-based treatments for a wide variety of ailments. Decades later, however, we are still awaiting novel licensed agents in this arena, despite an enormous research effort to this end. This review explores some of the most promising recent advances in NO donor drug development and addresses the challenges associated with NO as a therapeutic agent.     

Changes in gastric mucosal ulcerogenic responses in rats with adjuvant arthritis: role of nitric oxide

AIM: To examine gastric mucosal ulcerogenic responses to indomethacin and HCl/ethanol in adjuvant arthritic (AA) rats. METHODS AND RESULTS: Arthritis was induced in male Dark Agouti (DA) rats by injection of Freund's complete adjuvant (FCA) into the right hind paw. The gastric ulcerogenic response to indomethacin was markedly worsened in AA rats, depending on the degree of arthritic change. This aggravation of indomethacin-induced gastric lesions in AA rats was significantly prevented by NG-nitro-L-arginine methyl ester (L-NAME) and amino-guanidine as well as dexamethasone. In contrast, the mucosal ulcerogenic response to HCl/ethanol was inhibited in AA rats. The suppression of HCl/ethanol-induced gastric lesions in AA rats was reversed almost totally by L-NAME and aminoguanidine as well as dexamethasone and partly by indomethacin. The expression of inducible nitric oxide synthase (iNOS) mRNA was observed in the stomach of AA rats but not of normal rats. Moreover, the luminal releases of nitric oxide (NO) metabolites as well as prostaglandin (PG) E2 were significantly increased in AA rats. CONCLUSIONS: The gastric mucosal ulcerogenic responses were modified in AA rats, in different manners depending on the irritants; an increase in response to indomethacin and a decrease in response to HCl/ethanol. These changes may both be accounted for by increased production of NO by iNOS, and the latter is also partly related to increased production of PGs.     

Dual effect of nitric oxide on uterine prostaglandin synthesis in a murine model of preterm labour

BACKGROUND AND PURPOSE

Maternal infections are one of the main causes of adverse developmental outcomes including embryonic resorption and preterm labour. In this study a mouse model of inflammation-associated preterm delivery was developed, and used to study the relationship between nitric oxide (NO) and prostaglandins (PGs).

EXPERIMENTAL APPROACH

The murine model of preterm labour was achieved by assaying different doses of bacterial lipopolysaccharides (LPS). Once established, it was used to analyse uterine levels of prostaglandins E₂ and F_2α (by radioimmunoassay), cyclooxygenases (COX) and NOS proteins (by Western blot) and NO synthase (NOS) activity. Effects of inhibitors of COX and NOS on LPS-induced preterm labour were also studied. In vitro assays with a nitric oxide donor (SNAP) were performed to analyse the modulation of prostaglandin production by NO.

KEY RESULTS

Lipopolysaccharide increased uterine NO and PG synthesis and induced preterm delivery. Co-administration of meloxicam, a cyclooxygenase-2 inhibitor, or aminoguanidine, an inducible NOS inhibitor, prevented LPS-induced preterm delivery and blocked the increase in PGs and NO. Notably, the levels of NO were found to determine its effect on PG synthesis; low concentrations of NO reduced PG synthesis whereas high concentrations augmented them.

CONCLUSIONS AND IMPLICATIONS

An infection-associated model of preterm labour showed that preterm delivery can be prevented by decreasing PG or NO production. NO was found to have a dual effect on PG synthesis depending on its concentration. These data contribute to the understanding of the interaction between NO and PGs in pregnancy and parturition, and could help to improve neonatal outcomes.     

一氧化氮供体降压活性研究进展

高血压是一种常见的心血管疾病,目前其发病率逐年上升,并有低龄化的趋势.1980年,Furchgott等发现内皮源性舒张因子(EDRF)具有强大舒张血管作用,后经证实EDRF本质即为一氧化氮(NO).大量研究表明,NO作为信使物质和效应分子,参与体内多种生理、病理过程.     

Blockade of phencyclidine-induced effects by a nitric oxide donor

1. Phencyclidine (PCP) is widely used as an animal model of schizophrenia. The aim of this study was to better understand the role of nitric oxide (NO) in the mechanism of action of PCP and to determine whether positive NO modulators may provide a new approach to the treatment of schizophrenia. 2. The effects of the NO donor, sodium nitroprusside (SNP), were studied in PCP-treated rats. Following drug administration, behavioural changes and the expression of c-fos, a metabolic marker of functional pathways in the brain, were simultaneously monitored. 3. Acute PCP (5 mg kg(-1), i. p.) treatment induced a complex behavioural syndrome, consisting of hyperlocomotion, stereotyped behaviours and ataxia. Treatment with SNP (2 - 6 mg kg(-1), i.p.) by itself produced no effect on any behaviour studied but completely abolished PCP-induced behaviour in a dose- and time-dependent manner. 4. PCP had differential regional effects on c-fos expression in rat brain, suggesting regionally different patterns of neuronal activity. The most prominent immunostaining was observed in the cortical regions. Pre-treatment with SNP blocked PCP-induced c-fos expression at doses similar to those that suppress PCP-induced behavioural effects. 5. These results implicate the NO system in the mechanism of action of PCP. The fact that SNP abolished effects of PCP suggests that drugs targeting the glutamate-NO system may represent a novel approach to the treatment of PCP-induced psychosis and schizophrenia.     

白细胞介素-10对大鼠佐剂性关节炎的治疗作用

目的　研究白细胞介素 10 (IL 10 )对大鼠佐剂性关节炎 (AA)的治疗作用及免疫机制。方法　测量非致炎侧足肿胀度、体重、脏器系数 ,采用淋巴细胞增殖法和滑膜成纤维细胞 (SFCs)增殖法测定细胞增殖能力和IL 1产生 ,应用酶联免疫吸附测定法 (ELISA)检测IL 4水平。结果　IL 10(每鼠 2 ,10 μg·d-1× 9d ,ip)能缓解AA病情 ,使足肿胀度降低 ,体重、胸腺和脾脏系数及脾细胞增殖能力得到改善 ,腹腔巨噬细胞 (PMΦ)和滑膜组织细胞 (SMCs)产生IL 1水平下降 ,抑制SFCs和引流淋巴结细胞 (LNCs)过度增殖 ,使LNCs和SMCs产生IL 4水平提高。结论　IL 10能用于大鼠AA的治疗 ,其治疗机制与IL 10、IL 4和某些其它细胞因子介导的免疫调节相关     

绿原酸对佐剂性关节炎模型大鼠抗炎作用及机制研究

目的探讨绿原酸对佐剂性关节炎模型动物抗炎作用及机制。方法采用大鼠足跖注射Freund's完全佐剂形成佐剂性关节炎(AA),随机分为空白对照组、模型组、阳性对照组(尼美舒利)、绿原酸低剂量组(25 mg·kg-1)、中剂量组(50 mg·kg-1)和高剂量组(100 mg·kg-1)。观察绿原酸对佐剂性关节炎模型大鼠超敏反应与左右足跖宽度、血清肿瘤坏死因子(TNF-α)、白细胞介素-2(IL-2)、循环免疫复合物(CIC)、超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)和丙二醛(MDA)的含量。结果绿原酸中剂量组(50 mg·kg-1)、高剂量组(100 mg·kg-1)能够降低含量,升高SOD和GSH-Px水平,比较差异有统计学意义(P<0.05)。结论绿原酸具有抗佐剂性关节炎(AA)的作用,并通过降低TNF-α、IL-2、CIC和MDA含量及提高抗氧化能力发挥作用。     

红花注射液对佐剂性关节炎大鼠的免疫调节作用

目的：本课题主要研究红花注射液对佐剂性关节炎大鼠（AA）的免疫调节作用。方法：模型制作第15天,给予动物相应药物治疗。治疗20d后,大鼠心脏采血。取全血做TC亚群计数,留取血清用ELISA法测血清IL-1含量;取大鼠腹腔液涂片,做巨噬细胞吞噬功能测定。结果：在红花注射液的作用下,佐剂性关节炎大鼠的巨噬细胞吞噬指数和吞噬百分率、CD4＋T细胞与CD8＋T细胞比值及血清IL-1含量均显著降低。结论：红花注射液可以调节佐剂性关节炎大鼠过于亢进的免疫自稳功能,其治疗效果与雷公藤甲素相仿。     

Inhibitors of nitric oxide synthase in inflammatory arthritis

There is considerable evidence that excessive nitric oxide (NO) synthesized from L-arginine by inducible nitric oxide synthase (iNOS) plays an important pathological role in inflammatory arthritis. Since NO synthesized by constitutive isoforms of NOS has a physiological role, a great deal of activity has been directed at identifying inhibitors of NOS that are selective for the induced isoform. The major chemical areas that have been described so far in the search for such selective iNOS inhibitors and the activity of some of these compounds in animal models of arthritis are reviewed.     

Dual function of nitric oxide in carcinogenesis, reappraisal

Nitric oxide, a unique signalling molecule, is synthesized from L-arginine by a family of isoenzymes called nitric oxide synthase. Function of nitric oxide largely depends on the concentrations of surrounding free radicals and redox state. The local concentration of the nitric oxide defines its anti- or protumorigenic function by interacting with DNA or DNA repair enzymes and tumor suppressor gene, p53, as well. Indoleamine 2,3-dioxygenase activity is induced by interferon-gamma in many human cell types or cancer cells. Indoleamine 2,3- dioxygenase depletes locally available L- tryptophan producing cytotoxic metabolite kynurenine. Nitric oxide regulates the indoleamine 2,3-dioxygenase activity in a dose dependent-manner leading to either enhanced immune response or immune tolerance against tumor tissue. Additionally, the overproduction of nitric oxide may also trigger the cyclooxygenase-lipooxygenase pathways. Nitric oxide synthase expression correlates with cyclooxygenase-2 expression in cancer cells. On the other hand, tetrahydrobiopterin acts as the cofactor for nitric oxide synthase and stabilizes nitric oxide synthase dimers. However, tetrahydrobiopterin acts biphasically; under limited concentrations of tetrahydrobiopterin, nitric oxide generates superoxide radicals. Thus the dual action of nitric oxide and its interaction with the related compounds are of concern for their contribution in the development of carcinogenesis. In this review critical links between the nitric oxide and chronic inflammation associated carcinogenesis are summarized.     

NMI-1182, a gastro-protective cyclo-oxygenase-inhibiting nitric oxide donor

Non-steroidal anti-inflammatory drugs (NSAIDs) are widely used to treat inflammation and to provide pain relief but suffer from a major liability concerning their propensity to cause gastric damage. As nitric oxide (NO) is known to be gastro-protective we have synthesized a NO-donating prodrug of naproxen named NMI-1182. We evaluated two cyclo-oxygenase (COX)-inhibiting nitric oxide donors (CINODs), NMI-1182 and AZD3582, for their ability to be gastroprotective compared to naproxen and for their anti-inflammatory activity. NMI-1182 and AZD3582 were found to produce similar inhibition of COX activity to that produced by naproxen. Both NMI-1182 and AZD3582 produced significantly less gastric lesions after oral administration than naproxen. All three compounds effectively inhibited paw swelling in the rat carrageenan paw edema model. In the carrageenan air pouch model all three compounds significantly reduced PGE₂ levels in the pouch exudate but only NMI-1182 and naproxen inhibited leukocyte influx. These data demonstrate that NMI-1182 has comparable anti-inflammatory activity to naproxen but with a much reduced likelihood to cause gastric damage.     

克班宁透皮贴剂对佐剂性关节炎大鼠的抗炎作用

目的:考察克班宁制备成透皮贴剂后对佐剂性关节炎大鼠的抗炎作用.方法:将60只雄性Wistar大鼠随机分为6组,分别为正常组、模型组、甲氨蝶呤(MTX)组、克班宁贴剂低、中、高剂量组,每组10只.除正常组外,其余大鼠右后足趾皮下注射0.1mL弗氏完全佐剂(CFA)造模,给药组于腹部脱毛区域给予克班宁透皮贴剂,MTX组腹腔...     

Improvement of intestinal absorption of macromolecules by nitric oxide donor

Nitric oxide (NO) is one of the most versatile mediators in mammalian biology. In the present study, we investigated the absorption-enhancing effects of an NO donor, 3-(2-hydroxy-1-methylethyl-2-nitrosohydrazino)-N-methyl-1-propa namine (NOC7), on drugs that are poorly absorbed from the gastrointestinal tract. NOC7 significantly increased the jejunal absorption of fluorescein isothiocyanate dextrans (FDs) of different average molecular weights (4000-20,000). This enhancing effect decreased as the FD molecular weight increased. Another NO donor, S-nitroso-N-acetyl-DL-penicillamine (SNAP), also increased the absorption of FD-4 from the jejunum. The absorption enhancement effect of NOC7 significantly decreased after coadministration with an NO scavenger, 2-(4-carboxyphenyl)-4,4,5, 5-tetramethylimidazole-1-oxyl-3-oxide, sodium salt. Furthermore, the enhancement effect of NOC7 was reversed shortly after cessation of the enhancer treatment. Little damage by NOC7 to the intestinal mucosa was observed in terms of release of lactose dehydrogenase and protein from the intestinal mucosa. NOC7 also increased the absorption of FD-4 by the colon and rectum. The findings suggest that an NO donor can improve the absorption of macromolecules from all regions of the rat intestine with very little mucosal damage and that an NO donor can act as a potent absorption enhancer.     

Synthesis and characterization of S-nitrosoglutathione-oligosaccharide-chitosan as a nitric oxide donor

Objectives: The aim of this work is to synthesize a novel stable and biodegradable nitric oxide (NO) donor polymer based on a chitosan backbone. This polymer needed to be linked to glutathione (GSH), which was nitrosated in a second step. This polymer has been developed as an NO delivery platform that could be further evaluated for an oral delivery in Crohn’s disease.

Methods: The new polymer (named S-nitrosoglutathione-oligosaccharide-chitosan or SNOC) was obtained using a two-step procedure involving the linkage of GSH to chitosan via an amidine reaction followed by a post-nitrosation with NaNO₂. The GSH linkage was assessed using NMR, FTIR and an Ellman’s test, whereas the final NO amount was determined by the Griess and Saville method.

Results: Polymers with different numbers of NO groups were obtained (159.04 ± 64.16 µmol/g of polymer for SNOC G1 and 525.08 ± 151.35 µmol/g of polymer for SNOC G2) depending on the procedure used for production. When tested in an Ussing chamber, SNOC G2 had a sustained release of NO and nitrites for at least 6 h.

Conclusion: We believe that this type of polymer is adapted for the development of various formulations, including microparticles.     

Antiangiogenic effects of S-nitrosocaptopril crystals as a nitric oxide donor

Angiogenesis is the formation of new capillaries from preexisting vessels by migration and proliferation of endothelial cells, which produce a cellular signaling messenger, nitric oxide (NO). The purpose of the present study was to examine the effects of exogenous NO donors on angiogenesis by using a novel crystalline NO donor, S-nitrosocaptopril. The characteristic X-ray diffraction pattern of S-nitrosocaptopril was demonstrated for the first time. On primary capillary endothelial cells pretreated with vascular endothelium growth factor (VEGF), S-nitrosocaptopril (1-500 microM), but not captopril, produced a dose-dependent inhibition of endothelial proliferation. On chick embryos of entire living eggs, gelatin sponges adsorbed with VEGF were implanted on the embryo chorioallantoic membrane to promote vascular growth activity within the sponges. Addition of S-nitrosocaptopril crystals (0.1 mg) to the gelatin sponges markedly reduced vascular density around the sponges, whereas captopril did not inhibit neovascularization. The vascular hemoglobin content surrounding each of the gelatin sponges was determined as a confirmatory test. S-nitrosocaptopril, but not captopril, significantly decreased the hemoglobin content of the embryo tissues immediately surrounding the gelatin sponges. In conclusion, S-nitrosocaptopril exerts an inhibitory effect on angiogenesis. This newly discovered function of S-nitrosocaptopril appears to be governed by distinct structural NO moiety.