尼日利亚螫毛果(Cnestis ferruginca)中一种具有清除自由基的活性化合物的分离鉴定

用二苯基苦基苯肼自由基薄层实验法（DPPH-TLC-ASSAY）,从尼日利亚螫毛果（Cnestis ferruginca)植物的甲醇提取物中发现了一种具有很强的清除自由基活性的物质;用柱色谱及反相制备柱色谱（RP-LPPLC）,制备出了该活性物质的纯品;用质谱（HR ESI-MS）,核磁共振氢谱（^1H0,碳谱（^13C,DEPT)。紫外光谱及化学方法对该物质进行了结构鉴定,鉴定结果为对苯酚基-6-O-反式咖啡酸基-β-D-吡喃葡萄糖甙。     

魔芋葡甘露寡糖的制备、化学结构及对胰岛NO自由基释放量的影响

魔芋精粉经 β 甘露聚糖酶酶解成寡糖后 ,用活性炭柱进行分离纯化 ,以不同浓度 (5 % ,10 % ,2 0 % )的乙醇洗脱 .研究不同洗脱组分对链脲佐菌素 (STZ)诱导糖尿病模型的胰岛NO自由基释放量的影响 .发现 1mg ml以 5 %乙醇洗脱的寡糖可以使胰岛培养液中的NO自由基释放量平均下降2 5 4 % (P <0 0 5 ) ,0 1mg ml以 5 %乙醇洗脱的寡糖使NO自由基水平下降 2 0 % (P <0 0 5 ) .结果表明 ,5 %乙醇洗脱的魔芋寡糖对保护胰岛免受链脲佐菌素 (STZ)的破坏有一定的作用 .用凝胶色谱、红外光谱、元素分析、核磁共振光谱、质谱等方法初步分析了 5 %乙醇洗脱的魔芋寡糖的化学结构 .发现该糖是一种四糖 ,分子量为 6 6 6 .其推测性结构式为 :β D Man(1→ 4 ) β D Man(1→ 4 ) β D Glc(1→ 4 )α D Man ,β D Man(1→ 4 ) β D Glc(1→ 4 ) β D Man(1→ 4 )α D Man或 β D Glc(1→ 4 ) β D Man(1→4 ) β D Man(1→ 4 )α D Man .     

Immunological Monitoring of Fenton Fragmentation of Fibrinogen

Fibrinogen is transformed into insoluble “neofibe” by reaction with up to IOOpM Cu(II) and 1.5 mM ascorbate. The soluble peptides which are released during the reaction can be monitored by amino acid analysis and by measuring released keto-carbonyl (with DNPH). Immunologic characterization of the soluble peptides. with anibodies directed against fibrino-peptide A (FPA) clearly show the release of this epitope. optimally at 50 pM Cu(II). Anti-FPB gives no evidence for the release of that epitope. However, N-terminal amino acid analyses reveals the presence of 3 peptides terminating in ALA (alpha chain FPA). GLU (beta chain FPB) and SER/ASP (unknown). The release of fibrinopeptides is interpreted within the context of a general mechanism for OH'-induced peptide chain cleavage via intermediate Schiff-base hydrolysis.     

The involvement of ethanol in the free radical reaction of 6-hydroxydopamine

ESR spin trapping technique was used to detect and analyze free radical formation. When 6-hydroxydomine (6-OHDA) was incubated alone or in the presence of a free radical generating system (H₂O₂ and FeSO₄), hydroxyl free radicals were observed in a concentration-dependent manner. Glutathione was found to be the most effective scavenger of the ESR signal when compared with vitamin E or Mannitol. The addition of ethanol resulted in the formation of the pure hydroxyethyl free radicals. The amount of hydroxyethyl free radicals in the system was dependent upon the concentration of ethanol and the formation of hydroxyethyl free radicals correlated well with the extent of lipid peroxidation and the loss of enzymic activity of the membrane-bound (Na⁺, K⁺)-ATPase. We suggest that in the biological system ethanol may potentiate the neurotoxicity of 6-OHDA with the formation of hydroxyethyl free radicals, which are longer-lived and far more damaging to membranes that the hydroxyl radicals. These data lead us to further hypothesize that the neuronal degeneration caused by 6-OHDA and other compounds that generate free radicals could be potentiated in the presence of ethanol.     

Free radical damage to cultured porcine aortic endothelial cells and lung fibroblasts: Modulation by culture conditions

Summary Culture conditions modulating cell damage from xanthine plus xanthine oxidase-derived partially reduced oxygen species were studied. Porcine thoracic aorta endothelial cells and porcine lung fibroblasts were maintained in monolayer culture. Cells were prelabeled with⁵¹Cr before xanthine plus xanthine oxidase exposure. Endothelial cells showed 30 to 100% more lysis than fibroblasts and thus seemed more sensitive to this oxidant stress. The effect of cell culture age, as indicated by population doubling level (PDL), was examined. Response of low PDL endothelial cells and fibroblasts subjected to oxidant stress was compared with the response of PDL 15 cells. Both low PDL endothelial cells and fibroblasts responded differently to the lytic effect of xanthine oxidase-derived free radicals than did higher PDL cells. Specific activities of the antioxidant enzymes catalase, managanese superoxide dismutase, copper-zinc superoxide dismutase, glutathione peroxidase, and glucose-6-phosphate dehydrogenase were measured in both low and high PDL fibroblasts and endothelial cells. Antioxidant enzyme specific activities could only partially explain the differences in response to oxidant stress between fibroblasts and endothelial cells and between low and high PDL cells. Cell culture medium composition modulated the rate of production, and relative proportions of xanthine plus xanthine oxidase-derived partially reduced species of oxygen, i.e. superoxide, hydrogen peroxide, and hydroxyl radical. Serum content of medium was important in modulating free radical generation; superoxide production rates decreased 32%, H₂O₂ became undetectable, and hydroxyl radical generation decreased 54% in the presence of 10% serum. The medium protein and iron content also modulated free radical generation. The data suggest that cell culture media constituents, cell type, and cell culture age greatly affect in vitro response of cells subjected to oxidant stress. Research supported by American Lung Association Fellowship Training Grant and Research Training Grant, the R. J. Reynolds Corporation, and National Institutes of Health Grants HL29784 and 1 HL 23805.     

内皮衍生舒张因子对缺血（缺氧）,再灌注（复氧）心肌的保护作用

血管内皮产生的内皮衍生舒张因子（ｅｎｄｏｔｈｅｌｉｕｍ－ｄｅｒｉｖｅｄ ｒｅｌａｘｉｎｇ ｆａｃｔｏｒ,ＥＤＲＦ）即一氧化氮（ｎｉｔｒｉｃ ｏｘｉｄｅ,ＮＯ）本工作分别在大鼠Ｌａｎｇｅｎｄｏｒｆｆ离体心脏灌流模型和培养的大鼠心肌细胞上观察了ＮＯ、ＮＯ的前体物质Ｌ－精氨酸（Ｌ－Ａｒｇ）、ＮＯ的前体物质Ｌ－精氨酸（Ｌ－Ａｒｇ）、ＮＯ的合成阻断剂Ｌ－硝基精氨酸（Ｌ－ＮＮＡ）对心肌缺血（缺氧）再灌注（复氧     

Oxidation and nitration of ribonuclease and lysozyme by peroxynitrite and myeloperoxidase

In spite of the many studies on protein modifications by reactive species, knowledge about the products resulting from the oxidation of protein-aromatic residues, including protein-derived radicals and their stable products, remains limited. Here, we compared the oxidative modifications promoted by peroxynitrite and myeloperoxidase/hydrogen peroxide/nitrite in two model proteins, ribonuclease (6Tyr) and lysozyme (3Tyr/6Trp). The formation of protein-derived radicals and products was higher at pH 5.4 and 7.4 for myeloperoxidase and peroxynitrite, respectively. The main product was 3-nitro-Tyr for both proteins and oxidants. Lysozyme rendered similar yields of nitro-Trp, particularly when oxidized by peroxynitrite. Hydroxylated and dimerized products of Trp and Tyr were also produced, but in lower yields. Localization of the main modified residues indicates that peroxynitrite decomposes to radicals within the proteins behaving less specifically than myeloperoxidase. Nitrogen dioxide is emphasized as an important protein modifier.     

改良乳腺癌根治术后切口感染的病原学特征、影响因素及其凝血纤溶功能研究

摘要 目的：探讨改良乳腺癌根治术后切口感染的病原学特征、影响因素及其对凝血纤溶功能的影响。方法：选取我院于2016年6月~2020年9月期间收治的390例行改良乳腺癌根治术的乳腺癌患者,分析改良乳腺癌根治术后切口感染的病原学特征、影响因素及术后切口感染对凝血纤溶功能的影响。结果：390例行改良乳腺癌根治术的乳腺癌患者,术后发生切口感染28例,术后切口感染率为7.18%（28/390）,将未发生术后切口感染的患者纳为未感染组（n=362）,发生的纳为感染组（n=28）。28例发生感染的患者共分离培养病原菌36株,其中革兰阳性菌14株,占比38.89%（14/36）,以金黄色葡萄球菌、粪肠球菌为主。革兰阴性菌21株,占比58.33%（21/36）,以大肠埃希菌、铜绿假单胞菌为主。改良根治术后乳腺癌患者切口感染的影响因素包括手术时间、术后住院时间、合并基础疾病、引流时间、年龄、白蛋白（P<0.05）。多因素Logistic回归分析发现：合并基础疾病、年龄≥60岁、白蛋白＜35 g/L、手术时间≥120 min均是改良乳腺癌根治术后切口感染的影响因素（P＜0.05）。两组术后30 d凝血酶原时间（PT）、活化部分凝血活酶时间（APTT）、凝血酶时间（TT）升高,且未感染组高于感染组（P<0.05）,纤维蛋白原（FIB）降低,且未感染组低于感染组（P<0.05）。结论：改良乳腺癌根治术后切口感染较为常见,致病菌以革兰阴性菌为主,年龄、合并基础疾病、白蛋白、手术时间均是其影响因素,同时术后切口感染的发生可影响凝血纤溶功能的恢复,临床医生应积极采取措施预防术后切口感染的发生,从而保证手术治疗效果。     

Rescuing activity of oxygen-damaged pyruvate formate-lyase by a spare part protein

Pyruvate formate-lyase (PFL) is a glycyl radical enzyme (GRE) that converts pyruvate and coenzyme A into acetyl-CoA and formate in a reaction that is crucial to the primary metabolism of many anaerobic bacteria. The glycyl radical cofactor, which is posttranslationally installed by a radical S-adenosyl-L-methionine (SAM) activase, is a simple and effective catalyst, but is also susceptible to oxidative damage in microaerobic environments. Such damage occurs at the glycyl radical cofactor, resulting in cleaved PFL (cPFL). Bacteria have evolved a spare part protein termed YfiD that can be used to repair cPFL. Previously, we obtained a structure of YfiD by NMR spectroscopy and found that the N-terminus of YfiD was disordered and that the C-terminus of YfiD duplicates the structure of the C-terminus of PFL, including a β-strand that is not removed by the oxygen-induced cleavage. We also showed that cPFL is highly susceptible to proteolysis, suggesting that YfiD rescue of cPFL competes with protein degradation. Here, we probe the mechanism by which YfiD can bind and restore activity to cPFL through enzymatic and spectroscopic studies. Our data show that the disordered N-terminal region of YfiD is important for YfiD glycyl radical installation but not for catalysis, and that the duplicate β-strand does not need to be cleaved from cPFL for YfiD to bind. In fact, truncation of this PFL region prevents YfiD rescue. Collectively our data suggest the molecular mechanisms by which YfiD activation is precluded both when PFL is not damaged and when it is highly damaged.     

The specific activation of TRPC4 by Gi protein subtype

The classical type of transient receptor potential channel (TRPC) is a molecular candidate for Ca²⁺-permeable cation channels in mammalian cells. Especially, TRPC4 has the similar properties to Ca²⁺-permeable nonselective cation channels (NSCCs) activated by muscarinic stimulation in visceral smooth muscles. In visceral smooth muscles, NSCCs activated by muscarinic stimulation were blocked by anti-Gαi/o antibodies. However, there is still no report which Gα proteins are involved in the activation process of TRPC4. Among Gα proteins, only Gαi protein can activate TRPC4 channel. The activation effect of Gαi was specific for TRPC4 because Gαi has no activation effect on TRPC5, TRPC6 and TRPV6. Coexpression with muscarinic receptor M2 induced TRPC4 current activation by muscarinic stimulation with carbachol, which was inhibited by pertussis toxin. These results suggest that Gαi is involved specifically in the activation of TRPC4.