氯胺酮抑制内毒素休克大鼠中性粒细胞的CD11b表达和呼吸氧爆发

目的 观察氯胺酮对内毒素休克大鼠中性粒细胞CD11b表达和呼吸氧爆发的影响.方法 腹腔内注射内毒素(10 mg/kg)复制脓毒症模型.30只成年雄性SD大鼠,随机分为5组:生理盐水组;内毒素刺激组;内毒素刺激 氯胺酮(5 mg·kg-1·h-1)治疗组;内毒素刺激 氯胺酮(25 mg·kg-1·h-1)治疗组;内毒素刺激 氯胺酮(50 mg·kg-1·h-1)治疗组.在1、4、6 h使用流式细胞仪检测中性粒细胞CD11b和呼吸氧爆发.结果 与生理盐水组相比内毒素刺激后中性粒细胞的CD11b的表达和呼吸氧爆发明显增加(P＜0.01).与单纯内毒素刺激组相比,使用氯胺酮治疗组CD11b表达和呼吸氧爆发明显减少(P＜0.01).结论 氯胺酮抑制内毒素休克大鼠中性粒细胞的CD11b表达和呼吸氧爆发.     

维拉帕米对人血PMN细胞呼吸暴发和氧自由基的抑制作用

用化学发光法测定：人血ＰＭＮ细胞受ＰＭＡ刺激发生呼吸暴发产生的活性氧；黄嘌呤－黄嘌呤氧化酶体系产生的Ｏ；Ｖｉｔｃ－ＣＵ２＋－酵母多糖产生的·ＯＨ及Ｈ２Ｏ２的释放，同时也观察了维拉帕米对上述体系产生的自由基的影响。结果提示，维拉帕米对ＰＭＮ细胞的呼吸暴发有显著的抑制作用；对Ｏ、Ｈ２Ｏ２有清除作用，呈剂量依赖性，其ＩＣ５０分别为２８．１，２２．３ｎｍｏｌ·Ｌ－１；对·ＯＨ则无作用。     

重组人生长激素对体外人中性粒细胞功能影响

目的观察重组人生长激素(rhGH)对有无脂多糖(LPS)刺激的离体培养中性粒细胞(PMNs)呼吸爆发和粘附功能的影响。方法分离纯化人外周静脉血PMN,加或不加LPS与三个不同剂量(10 ng/ml,100 ng/ml和1000 ng/ml)的rhGH共同孵育3 h,检测不同试验组的PMN呼吸爆发和粘附功能。结果低浓度的rhGH(10 ng/ml)不影响PMNs的呼吸爆发功能和粘附功能,高浓度的的rhGH(100 ng/ml,1000 ng/ml)增强PMN的呼吸爆发和粘附功能,rhGH不影响LPS刺激的PMN的呼吸爆发功能,能增强LPS刺激的PMN的粘附功能。结论一定浓度的rhGH能活化PMNs的呼吸爆发和粘附功能,增加LPS活化的PMNs与内皮细胞的粘附,加重炎症反应时PMNs对组织的损伤作用。     

Investigation of the inhibitory effect of broussochalcone A on respiratory burst in neutrophils

Broussochalcone A, a prenylated chalcone isolated from Broussonetia papyrifera (L.) VENT. (Moraceae), inhibited O₂ consumption in formylmethionyl-leucyl-phenylalanine (fMLP)- and phorbol 12-myristate 13-acetate (PMA)-stimulated rat neutrophils in a concentration-dependent manner with IC₅₀ values of 70.3±4.9 and 63.9±7.1 μM, respectively. Broussochalcone A did not affect the fMLP-induced increase of cellular inositol trisphosphate (IP₃) and [Ca²⁺]_i. However, the enzyme activity of neutrophil cytosolic protein kinase C was effectively suppressed by broussochalcone A. Broussochalcone A had no effect on either [³H]phorbol 12,13-dibutyrate ([³H]PDB) binding to neutrophil cytosolic protein kinase C or on PMA-induced membrane translocation of protein kinase C-β in neutrophils. Broussochalcone A suppressed the enzyme activity of trypsin-treated rat brain protein kinase C in a concentration-dependent manner. In PMA-activated neutrophil particulate NADPH oxidase, broussochalcone A attenuated superoxide anion radical (O₂⁻) generation with an IC₅₀ value of 61.8±5.4 μM. These results show that the inhibitory effect of broussochalcone A on respiratory burst in neutrophils is not mediated by the reduction of phospholipase C activity, but is mediated partly by the suppression of protein kinase C activity through interference with the catalytic region and by the attenuation of O₂⁻ generation from the NADPH oxidase complex.     

Inhibitory effect of plant phenolics on fMLP-induced intracellular calcium rise and chemiluminescence of human polymorphonuclear leukocytes and their chemotactic activity in vitro

Abstract

Context: Polymorphonuclear leukocytes (PMNs) produce oxidants, contributing to systemic oxidative stress. Diets rich in plant polyphenols seem to decrease the risk of oxidative stress-induced disorders including cardiovascular disease.

Objective: The objective of this study was to examine the in vitro effect of each of the 14 polyphenols on PMNs chemotaxis, intracellular calcium response, oxidants production.

Materials and methods: Blood samples and PMNs suspensions were obtained from 60 healthy non-smoking donors and incubated with a selected polyphenol (0.5–10?µM) or a control solvent. We assessed resting and fMLP-dependent changes of intracellular calcium concentration ([Ca²⁺]_i) in PMNs with the Fura-2AM method and measured fMLP-induced luminol enhanced whole blood chemiluminescence (fMLP-LBCL). Polyphenol chemoattractant activity for PMNs was tested with Boyden chambers.

Results: Polyphenols had no effect on resting [Ca²⁺]_i. Unaffected by other compounds, fMLP-dependent increase of [Ca²⁺]_i was inhibited by quercetin and catechol (5?µM) by 32?±?14 and 12?±?10% (p?<?0.04), respectively. Seven of the 14 tested substances (5?µM) influenced fMLP-LBCL by decreasing it. Catechol, quercetin, and gallic acid acted most potently reducing fMLP-LBCL by 49?±?5, 42?±?15, and 28?±?18% (p?<?0.05), respectively. 3,4-Dihydroxyhydrocinnamic, 3,4-dihydroxyphenylacetic, 4-hydroxybenzoic acid, and catechin (5?µM) revealed distinct (p?<?0.02) chemoattractant activity with a chemotactic index of 1.9?±?0.8, 1.8?±?0.7, 1.6?±?0.6, 1.4?±?0.2, respectively.

Conclusion and discussion: Catechol, quercetin, and gallic acid at concentrations commensurate in human plasma strongly suppressed the oxidative response of PMNs. Regarding quercetin and catechol, this could result from an inhibition of [Ca²⁺]_i response.     

Suppression of respiratory burst in human neutrophils by new synthetic pyrrolo-benzylisoquinolines

Reactive oxygen species produced by neutrophils contribute to the pathogenesis of inflammatory diseases. In this study, the inhibition of superoxide anion (O2*-) generation in human neutrophils by new synthetic pyrrolo-benzylisoquinoline derivatives was determined. We found that KW-2, KW-5, and KW-7 (8,9-dimethoxyl-1-(R-phenyl)-5,6-dihydro-pyrrolo[2,1-a]isoquinoline-2,3-dione; where R is 3-chloro, 3-bromo, and 4-methoxy, respectively) were the most effective inhibitors of formyl-L-methionyl-L-leucyl-L-phenylalanine (FMLP)-induced O2*- release in human neutrophils. KW-2, KW-5, and KW-7 displayed no antioxidant or O2*--scavenging ability. The inhibition of O2*- generation was reversed by the protein kinase (PK)A inhibitor, N-(2-((p-bromocinnamyl)amino)ethyl)-5-isoquinolinesulfonamide (H89), but not by the PKG inhibitor (8R,9S,11S)-(-)-2-methyl-9-methoxy-9-methoxycarbonyl-8-methyl-2,3,9,10-tetrahydro-8,11-epoxy-1H,8H,11H-2,7b,11a-triazadibenzo(a,g)cyclocta(cde)trinen-1-one (KT5823), or the soluble guanylate cyclase (sGC) inhibitor, 1H-(1,2,4)oxadiazolo(4,3-a)quinoxalin-1-one (ODQ). KW derivatives increased cellular cyclic AMP concentrations through the inhibition of phosphodiesterase (PDE) activity but not the elevation of adenylate cyclase (AC) activity. These results indicate that inhibition of FMLP-induced respiratory burst in human neutrophils by KW derivatives are cyclic AMP/PKA-dependent and are due to inhibition of PDE. The new chemical skeleton of PDE inhibitors may protect against the progression of inflammation.     

合成鱼腥草素对巨噬细胞呼吸爆发、细胞内游离钙离子浓度及T细胞分泌白细胞介素-2的影响

目的为了进一步分析合成鱼腥草素的免疫调节作用机理 ,研究合成鱼腥草素对于巨噬细胞呼吸爆发、细胞内钙离子浓度以及T细胞分泌白细胞介素水平的影响。方法巨噬细胞分离自大鼠腹腔灌洗液。以 2′ ,7′ 二氯荧光素二乙酯作为荧光指示剂 ,采用流式细胞术检测巨噬细胞的呼吸爆发。以fura 2作为钙离子荧光指示剂采用荧光分光光度法测定细胞内钙离子浓度。利用淋巴细胞分离液采用密度梯度离心法分离外周血T细胞 ,并用尼龙毛柱加以纯化。用ELISA法测定在亚适剂量ConA、白细胞介素 1α(IL 1α)以及白细胞介素 1β(IL 1β)存在的条件下合成鱼腥草素对白细胞介素 2 (IL 2 )分泌的影响。结果合成鱼腥草素可以刺激巨噬细胞呼吸爆发 ,提高细胞内钙离子浓度水平 ,促进T细胞分泌IL 2。结论合成鱼腥草素可能具有激活巨噬细胞和T淋巴细胞的作用 ,从而部分解释合成鱼腥草素的佐剂作用以及治疗感染性疾病的作用机理。     

DDPH对大鼠缺血性脑损伤的保护作用

目的研究DDPH对大鼠缺血性脑损伤的保护作用，并初步探讨其作用机制。方法用线拴法制备大鼠局灶性脑缺血模型，观察DDPH对大鼠脑缺血后神经症状、梗死面积的影响；用大鼠弥漫性不完全性脑缺血模型，观察DDPH对脑缺血后脑组织超氧化物歧化酶（SOD）活性、丙二醛（MDA）含量及组织病理损伤的影响。结果DDPH 10 mg·kg^-1在缺血前30 min ip,局灶性脑缺血模型大鼠在3 h后神经症状明显改善，24 h梗死面积缩小。DDPH使大鼠弥漫性不完全性脑缺血后脑组织内SOD活性增高，MDA含量下降，并明显改善神经细胞的病理性损伤。 结论 DDPH对大鼠缺血性脑损伤有一定保护作用，其机制可能与阻滞钙离子通道、提高SOD活性有关。     

The effects of N-(5-vinyl-1,3-thiazolidin-2-ylidene)phenylamine (5-VTPA) on the changes in free intracellular calcium and the production of reactive oxygen metabolites in human leukocytes

Human leukocytes were exposed to N-(5-vinyl-1,3-thiazolidin-2-ylidene)phenylamine (5-VTPA), a postulated impurity in the case oils that caused the Spanish Toxic Oil Syndrome in 1981. Changes induced by 5-VTPA alone and together with a chemotactic peptide, formyl-methionyl-leucyl-phenylalanine (FMLP), a tumor promoter, phorbol myristate acetate (PMA), or a synthetic diacylglycerol, dioctanoyl-s,n-glycerol (DiC₈) in free intracellular calcium levels ([Ca²⁺]_i) and in the induction of oxidative burst were measured. 5-VTPA elevated dose-dependently [Ca²⁺]_i and induced the production of reactive oxygen metabolites in leukocytes. 5-VTPA also amplified FMLP-induced increase in [Ca²⁺]_i, but was without an effect on FMLP-induced oxidative burst. On the contrary, 5-VTPA amplified dose-dependently PMA- and DiC₈-induced respiratory burst. The present results indicate that 5-VTPA may interfere with transmembrane signalling in human leukocytes. 5-VTPA may elevate [Ca²⁺]_i by acting directly on the membrane, or by acting through Ca²⁺-mobilizing receptors. Moreover, 5-VTPA also clearly amplified responses produced through protein kinase C stimulation. Thus, 5-VTPA may act on human leukocytes by affecting Ca²⁺-metabolism and the activity of protein kinase C.     

Cellular localization of the inhibitory action of abruquinone A against respiratory burst in rat neutrophils

1. The possible mechanisms of action of the inhibitory effect of abruquinone A on the respiratory burst in rat neutrophils in vitro was investigated.
2. Abruquinone A caused an irreversible and a concentration-dependent inhibition of formylmethionyl-leucyl-phenylalanine (fMLP) plus dihydrocytochalasin B (CB)- and phorbol 12-myristate 13-acetate (PMA)-induced superoxide anion (O₂^.−) generation with IC₅₀ values of 0.33±0.05 μg ml⁻¹ and 0.49±0.04 μg ml⁻¹, respectively.
3. Abruquinone A also inhibited O₂ consumption in neutrophils in response to fMLP/CB and PMA. However, abruquinone A did not scavenge the generated O₂^.− in xanthine-xanthine oxidase system and during dihydroxyfumaric acid (DHF) autoxidation.
4. Abruquinone A inhibited both the transient elevation of [Ca²⁺]_i in the absence of [Ca²⁺]_o (IC₅₀ 7.8±0.2 μg ml⁻¹) and the generation of inositol trisphosphate (IP₃) (IC₅₀ 10.6±2.0 μg ml⁻¹) in response to fMLP.
5. Abruquinone A did not affect the enzyme activities of neutrophil cytosolic protein kinase C (PKC) and porcine heart protein kinase A (PKA).
6. Abruquinone A had no effect on intracellular guanosine 3′ : 5′-cyclic monophosphate (cyclic GMP) levels but decreased the adenosine 3′ : 5′-cyclic monophosphate (cyclic AMP) levels.
7. The cellular formation of phosphatidic acid (PA) and phosphatidylethanol (PEt) induced by fMLP/CB was inhibited by abruquinone A with IC₅₀ values of 2.2±0.6 μg ml⁻¹ and 2.5±0.3 μg ml⁻¹, respectively. Abruquinone A did not inhibit the fMLP/CB-induced protein tyrosine phosphorylation but induced additional phosphotyrosine accumulation on proteins of 73–78 kDa in activated neutrophils.
8. Abruquinone A inhibited both the O₂^.− generation in PMA-activated neutrophil particulate NADPH oxidase (IC₅₀ 0.6±0.1 μg ml⁻¹) and the iodonitrotetrazolium violet (INT) reduction in arachidonic acid (AA)-activated cell-free system (IC₅₀ 1.5±0.2 μg ml⁻¹).
9. Collectively, these results indicate that the inhibition of respiratory burst in rat neutrophils by abruquinone A is mediated partly by the blockade of phospholipase C (PLC) and phospholipase D (PLD) pathways, and by suppressing the function of NADPH oxidase through the interruption of electron transport.

     

几种中药有效成分对中性粒细胞呼吸爆发的影响

用化学发光法观察了阿魏酸钠、葛根素、７６４３、麦芽醇、瑞香素、莨菪亭、甘露醇、灯盏花素、６５４－２几种中药有效成分对分叶核粒细胞在酵母聚精刺激下呼吸爆发产生活性氧的抑制作用。结果证明：在药物终浓度为０．００１ｍｍｏｌ·Ｌ－１时，其抑制作用的强度依次为：阿魏酸钠、葛根素＞７６４３、瑞香素、灯盏花素＞６５４－２、莨菪亭。     

Effect of libexin combinatum (libexine and emetine) on phagocytosis by polymorphonuclear granulocytes and alveolar macrophages

Summary The ability of human polymorphonuclear granulocytes and alveolar macrophages from rats to ingestSaccharomyces cerevisiae was tested in vitro in the presence of various concentrations of emetine and libexine. Emetine 4.0 and 40.0 µg/ml caused an increase in the phagocytic index of polymorphonuclear granulocytes and alveolar macrophages. After oral administration of emetine the phagocytosis-enhancing effect was still apparent. Intracellular killing was not influenced by emetine or libexine.     

Activity of Superoxide Dismutase and Catalase in Fenugreek (Trigonella foenum-graecum) in Response to Carbendazim

Fenugreek (Trigonella foenum-graecum) is an annual herb, used as a spice and traditionally as medicine. Fenugreek finds its uses in treating hyperglycemia, hyperlipidemia and disorders of gastro-intestinal and cardiovascular systems. Fenugreek cultivation in India is affected by fungal diseases like root-rot and damping-off and fungicides like carbendazim are used to overcome these infections. Fungicides play both positive and negative role in plants; fungicides protect plants from diseases and also exert oxidative stress simultaneously. This report is on the response of antioxidants, superoxide dismutase and catalase in fenugreek seeds and plants treated to different concentrations of carbendazim.     

N－（4＇－羧苯基）－4－羟基－3，5－二叔丁基苯甲酰胺对十四烷酰佛波醋酸酯刺激大鼠多形核白细胞生成过氧化氢的影响

Ｎ－（４＇－羧苯基）－４－羟基－３，５－二叔丁基苯甲酰胺对十四烷酰佛波醋酸酯刺激大鼠多形核白细胞生成过氧化氢的影响孙士勇，韩锐（中国医学科学院药物研究所，北京１０００５０）新维甲类化合物Ｎ（４＇－羧苯基）－４－羟基－３，５－二叔丁基苯甲酰胺［Ｎ－（４...     

Mechanisms for the induction of oxidative stress in Syrian hamster embryo cells by acrylonitrile.

Chronic administration of acrylonitrile to rats resulted in an increase in the incidence of glial neoplasms of the brain. Recent studies have shown that acrylonitrile induces oxidative stress in rat brain and cultured rat glial cells. Acrylonitrile also induces morphological transformation concomitant with an increase in the formation of oxidized DNA in Syrian Hamster Embryo (SHE) cells in a dose-dependent manner. The mechanism for the induction of oxidative stress in SHE cells remains unresolved. The present study examined the effects of acrylonitrile on enzymatic and nonenzymatic antioxidants in SHE cells. SHE cells were treated with subcytolethal doses of acrylonitrile (0, 25, 50, and 75 microg/ml) for 4, 24, and 48 h. Acrylonitrile (50 microg/ml and 75 microg/ml) increased the amount of reactive oxygen species in SHE cells at all time points. Glutathione (GSH) was depleted and catalase and superoxide dismutase activities were significantly decreased in SHE cells after 4 h of treatment. The inhibition of these antioxidants was temporal, returning to control values or higher after 24 and 48 h. Xanthine oxidase activity was increased following 24 and 48 h treatment with acrylonitrile. 1-aminobenzotriazole, a suicidal P450 enzyme inhibitor, attenuated the effects of acrylonitrile on catalase and xanthine oxidase in SHE cells, suggesting that P450 metabolism is required for acrylonitrile to produce its effects on these enzymes. Additional studies showed that in the absence of metabolic sources acrylonitrile had no effect on either catalase or superoxide dismutase activity. These results suggest that the induction of oxidative stress by acrylonitrile involves a temporal decrease in antioxidants and increase in xanthine oxidase activity that is mediated by oxidative metabolism of acrylonitrile.     

Activity of polymorphonuclear (PMN) leukocytes during bovine herpes virus-1 induced respiratory disease: effect of recombinant bovine interferon alpha I1

Following infection of cattle with bovine herpes virus-1 there is a state of generalized immunosuppression involving various leukocytes including polymorphonuclear (PMN) leukocytes. Since the PMN is considered to be pivotal in recovery from secondary bacterial infections during bovine respiratory disease, investigations were initiated to determine PMN activity in this disease and whether interferon could modulate PMN activity. In this study, the in vivo administration of recombinant interferon alpha-I1 was shown to increase PMN functions as measured by migration/chemotaxis and generation of reactive oxygen species. This augmented activity of PMN appeared to correlate with the reduction of overall clinical disease, that is, number of sick days, lung lesions and weight loss. In the study administration of interferon by the intranasal or intramuscular route were as effective in stimulating PMN function. Based on these studies it was concluded that the reason for improved performance of calves treated with interferon would be due to its immunomodulatory effects on leukocytes. Although interferon did not alter the initial suppression of PMN functions, these functions returned to normal and exceeded normal activities by 7-9 days post-infection, the time when maximal bacterial activity normally is present.     

新生儿呼吸窘迫综合征的护理

目的总结35例新生儿呼吸窘迫综合征患儿的护理体会。方法保暖、营养支持、氧疗和替代疗法的护理,加强呼吸道管理。结果减少合并症的发生,提高早产儿的生存率。结论有效控制体温,正确掌握吸引、气道湿化技术,营养及热量的供给,保持足够的氧浓度是治疗成功的关键。     

超氧化物歧化酶乳酸-羟乙酸共聚物微球的制备及其性质

利用复乳溶剂挥发法制备了超氧化物歧化酶（SOD）的乳酸－羟乙酸共聚物（PLGA）微球，考察了各工艺因素对微球粒径、包封率等的影响，通过扫描电子显微镜（SEM）、差示扫描量热分析（DSC）初步研究了其性质，结果表明，通过调整内水相的体积及浓度，分散相体积及PH值，可得到较高包封率，粒径在20－30μm，形态圆整，表面多孔的SOD微球，DSC表明SOD被有效地包入了PLGA微球中。     

Morphological transformation and oxidative stress induced by cyanide in Syrian hamster embryo (SHE) cells.

Cyanide is a well-established poison known for its rapid lethal action and toxicity. Although long-term mammalian studies examining the carcinogenic potential of cyanide have not been previously reported, cyanide was reported to be positive in Salmonella typhimurium mutagenesis assay and induced aneuploidy in DROSOPHILA: To further evaluate the carcinogenic potential of cyanide, the ability of cyanide to induce morphological transformation in Syrian hamster embryo (SHE) cells was studied. Cyanide induced a dose-dependent increase in morphological transformation in SHE cells following a 7-day continuous treatment. A significant increase in transformation was observed at potassium cyanide doses of 200 microM and greater. Transformation induced by cyanide was inhibited in a dose-related manner by vitamin E, suggesting a role of oxidative stress in the induction of morphological transformation by cyanide. Further, it was shown that 500 microM cyanide induced oxidative DNA damage in SHE cells, evidenced by the formation of 8-hydroxy-2'-deoxyguanosine (50-66% increase over control). The induction of oxidative stress by cyanide involved an early and temporal inhibition of antioxidant enzymes (catalase and superoxide dismutase) as well as an increased production of reactive oxygen species (1.5- to 2.0-fold over control).