Tetanus toxin selectively impairs anti-tumoral but not anti-microbial macrophage-mediated effector functions

Abstract The present study was designed to establish the susceptibility of macrophage-mediated effector functions to tetanus toxin (TT). Using the murine macrophage cell line, GG2EE, generated in vitro by v- raf /v- myc oncogenes, we have previously provided evidence that TT selectively inhibits interferon gamma (IFN-γ), but not basal, lysozyme activity. Here we show that while neither phagocytic nor candidacidal activities are affected by TT treatment, antitumoral activity is significantly impaired after exposure to TT. This phenomenon, which is dose-dependent, is fully ascribed to the holotoxin, as heat inactivated TT, C or A-B fragments result ineffective. Furthermore, C but not A-B fragment competes with TT in abrogating its inhibitory effects. Overall, these data indicate that TT is not a broad-spectrum, down-regulating signal on macrophage-mediated functions, thus implying that its toxic action is exerted on specific molecular targets.     

烷化溶血磷脂ET—18—OCH3对K562细胞作用的研究

为研究烷化溶血磷脂ET－18－OCH3（ALP）的抗白血病效果。本文以K562细胞为研究对象,通过台蓝拒染法测定ALP作用后K562细胞的生长抑制率和生长曲线;甲基纤维素半固体培养法测定克隆原细胞的存尖率;流式细胞仪检测K562细胞P210蛋白表达;TR－PCR半定量法测定细胞的bcr-abl mRNA;采用流式细胞仪进行DNA 及是民镜观察细胞形态学改变。结果显示,K562细胞经ALP处理后细胞生长明显受抑制,呈作用时间和剂量的依赖性,IC50为31.6(24h),22.3(48h),14.8(72h)μg/ml;细胞增殖速度显著降低,克隆原细胞存活曲线呈指数型,而正常对照组细胞的CFU－GM则未受影响;ATP还可使KT562细胞P210及bcr-abl mRNA水平下调,并有诱导细胞凋亡的作用,说明ALP对K562细胞生长具有明显抑制作用,并有诱导细胞凋亡的作用,提示ALP具有一定的抗白血病效应。     

Exogenous Glycine Betaine Reduces Drought Damage by Mediating Osmotic Adjustment and Enhancing Antioxidant Defense in <i>Phoebe hunanensis</i>

Drought stress negatively impacts growth and physiological processes in plants. The foliar application of glycine betaine (GB) is an effective and low-cost approach to improve the drought tolerance of trees. This study examined the effect of exogenously applied GB on the cell membrane permeability, osmotic adjustment, and antioxidant enzyme activities of Phoebe hunanensis Hand.-Mazz under drought stress. Two levels (0 and 800 mL) of water irrigation were tested under different applied GB concentrations (0, 50, 100, and 200 mM). Drought stress decreased the relative water content by 58.5% while increased the electric conductivity, malondialdehyde, proline, soluble proteins, soluble sugars, and antioxidant enzyme activities (superoxide dismutase, catalase, peroxidase) by up to 62.9%, 42.4%, 87.0%, 19.1%, 60.5%, 68.3%, 71.7%, and 83.8%, respectively, on the 25^th day. The foliar application of GB, especially at 100 mM, increased the relative water content of P. hunanensis leaves under drought stress. The concentration of GB from 50 to 100 mM effectively alleviated the improvement of cell membrane permeability and inhibited the accumulation of membrane lipid peroxidation products. Under drought stress, the concentrations of proline, soluble proteins, and soluble sugars in the leaves of P. hunanensis increased as the applied GB concentration was increased and the water stress time was prolonged. Exogenously applied GB decreased oxidative stress and improved antioxidant enzyme activities as compared with treatments without GB application. Furthermore, the physiological and biochemical indexes of P. hunanensis showed a certain dose effect on exogenous GB concentration. These results suggest that GB helps maintain the drought tolerance of P. hunanensis.     

Synthesis and biological activity of FGLamide allatostatin analogs with Phe3 residue modifications

A FGLamide allatostatin neuropeptide mimic ( H17 ) is a potential insect growth regulator which inhibits the production of juvenile hormone by the corpora allata. To find more evidence to reveal the structure–activity relationships of the Phe³ residue in the C‐terminal conserved pentapeptide and search for novel analogs with high activity, a series of Phe³ residue‐modified analogs were designed and synthesized using H17 as the lead compound. Bioassay using juvenile hormone (JH) production by corpora allata of the cockroach Diploptera punctata indicated that analogs 4 , 11 , and 13 showed strong ability to inhibit JH production in vitro, with IC₅₀ of 38.5, 22.5, and 26 nM, respectively. As well, the activity of analog 2 (IC₅₀: 89.5 nM) proved roughly equivalent to that of H17 . Based on the primary structure–activity relationships of Phe³ residue, we suggest that for analogs containing six‐membered aromatic rings, removing the methylene group of Phe³ or an o‐halogen or p‐halogen‐substituted benzene ring could increase the ability to inhibit biosynthesis of JH. This study will be useful for the design of new allatostatin analogs for insect management. Copyright © 2016 European Peptide Society and John Wiley & Sons, Ltd.     

The synthesis and characterization of a series of cobalt(II) β-ketoaminato complexes and their cytotoxic activity towards human tumor cell lines

A series of square planar cobalt(II) compounds bearing tetradentate β-ketoaminato ligands with variation in the number of ―CF₃ ligand substituents has been prepared and structurally and spectroscopically characterized. The fluorinated β-ketoamine ligands were prepared utilizing a multistep reaction sequence employing a silylenol protecting group. An additional tetrahedral cobalt compound bearing two bidentate β-ketoaminato ligands was also prepared and characterized.Cytotoxic activity of the cobalt-containing complexes was evaluated using six human cell lines; including two different prostate cancer cell lines (PC-3 and VCaP), acute monocytic leukemia (THP-1), astrocytoma (U-373 MG), hepatocellular carcinoma (HepG2), and neuroblastoma (SH-SY5Y) cells. The cobalt compounds are more active than their corresponding ligands. The activity is cell type specific; the cobalt compounds exhibit strong activity against human prostate cancer and monocytic leukemia cells but weak or no activity against neuroblastoma, astrocytoma, and liver carcinoma cells. Activity generally increases with a greater number of ―CF₃ substituents, and square planar complexes exhibit greater activity than the tetrahedral derivative. The mechanisms of activity against human PC-3 prostate cancer cells involve caspase-3 and two different mitogen-activated protein kinases. The addition of a thiol antioxidant reduced cytotoxicity, suggesting the possible involvement of reactive oxygen species. These cobalt complexes may represent a novel class of cytotoxic drugs selective towards certain types of tumors.     

两种多糖对日本沼虾酚氧化酶活力的影响

在饲料中分别添加 1‰的云芝多糖和虫草多糖 ,对日本沼虾进行投喂 ,通过连续测定日本沼虾血清中的酚氧化酶活力 ,研究两种多糖对日本沼虾免疫机能的作用。结果表明 ,在平均水温为 2 3 8℃± 3 8℃时 ,云芝组、虫草组分别较对照组提高了 34 52 %、84 62 % ;在平均水温为 1 0 8± 2 4℃时 ,云芝组和虫草组分别较对照组提高了30 0 2 %、5 1 2 % ,因而云芝多糖和虫草多糖能明显地增强日本沼虾血清的酚氧化酶活力。另经实验表明 ,在王雷所确立的酚氧化酶活力的检测条件下 ,其PO活力为 8 3units/mL ,而在Horowitz所确立的检测条件下 ,其PO活力为2 90units/mL ,并且后者检测方法重复性好 ,能较好地反映日本沼虾的免疫状态。     

蛇足石杉内生真菌Shiraia sp. Slf14中赖氨酸脱羧酶基因的克隆、表达及功能

【目的】阿尔茨海默症治疗药物石杉碱甲(Huperzine A,Hup A)的生物合成途径起始于赖氨酸脱羧酶(Lysine decarboxylase,LDC)。本研究克隆及表达了来源于产Hup A的植物内生真菌的LDC基因,并研究了其功能。【方法】采用RT-PCR扩增法,从一株产Hup A的蛇足石杉内生真菌Shiraia sp.Slf14获得LDC基因,构建表达质粒p ET-22b-LDC与p ET-32a-LDC,转化感受态细胞E.coli BL21,加入IPTG至终浓度为1×10~(–3) mol/L,于24°C、200 r/min培养8 h,诱导表达LDC蛋白质;通过Ni~(2+)金属亲和层析纯化重组LDC并建立酶促反应体系,利用TLC检测了LDC催化活性。利用生物信息学软件分析了LDC的理化性质及蛋白质的空间结构。【结果】成功克隆并异源表达出重组蛋白LDC与Trx-LDC,经SDS-PAGE电泳鉴定分子量分别为24.4 k Da和42.7 k Da,与预计大小相符。TLC结果表明LDC与Trx-LDC均具有赖氨酸脱羧酶活性。【结论】本研究从产Hup A的蛇足石杉内生真菌Shiraia sp.Slf14中成功克隆到LDC基因并进行了异源表达,检测到了其催化活性,为丰富LDC分子信息及阐明内生真菌中Hup A生物合成机制提供参考数据。     

Evaluation of Phytochemical Compositions and Biological Properties of Achillea gypsicola at Different Phenological Stages

Phytochemicals, which are commonly found at different levels in many medicinal plants, are natural strong antioxidants used in traditional medicine. In this research, determination of differences of phytochemical compositions and biological properties were aimed as periodically (pre‐, full and post flowering) and daily (6 am, 1 pm and 8 pm) in Achillea gypsicola Hub.‐Mor . The volatile oils belonging to A. gypsicola were obtained by hydrodistillation and analyzed by gas chromatography‐flame ionization detection (GC‐FID) and gas chromatography‐mass spectrometry (GC/MS). The antimicrobial activities of the volatile oils were determined with disc diffusion method. The microdilution method was used to determine minimum inhibitory concentration (MIC). Total phenolic and flavonoid contents were determined by spectrophotometric methods and antioxidant capacities were evaluated by 2,2‐diphenyl‐1‐picrylhydrazyl (DPPH) free radical, reducing power (RP) and metal chelating activity (MCA) assay. In addition, the phenolic acid and flavonoid compositions were evaluated by reversed phase‐high‐performance liquid chromatography (RP‐HPLC). This study presented a comprehensive report for the first time on evaluation of the phytochemical composition and the biological properties of A. gypsicola at different phenological stages. Thirty‐two compounds, containing the major component as camphor, 1,8‐cineole and borneol, were detected. Designated harvest time for the highest yield of volatile oils was found to be at full flowering stage‐1 pm. It has been observed that the volatile oil composition changes periodically and even daily. Also, in this research, menthol and menthone were found as the composition of volatile oil in Achillea species for the first time. Full flowering stage was found as the richest period in terms of phenolic acid and flavonoid compositions of A. gypsicola for the first time. The species examined in this research showed a high antioxidant and antimicrobial activity in comparison to other studies with Achillea species. The volatile oils exhibited high performances with range of inhibition zones (8.3–42.3 mm) and minimum inhibitory concentration values (2.25—144 μg/ml). Besides, a high correlation between antioxidant activity and phenolic content of A. gypsicola was found. These results suggest that A. gypsicola can be used as a safe source in the cosmetic, food and pharmaceutical industries.     

Pyrethroids as Promising Marine Antifoulants: Laboratory and Field Studies

Due to the regulations and bans regarding the use of traditional toxic chemicals against marine fouling organisms and the practical impediments to the commercialization of natural product antifoulants, there is an urgent need for compounds that are antifouling-active, environmentally friendly, and have a potential for commercial application. In this study, a series of common, commercially available pyrethroid products, which are generally used as environmentally safe insecticides, was evaluated for antifouling activity in the laboratory using an anti-settlement test with cyprids of the barnacle Balanus albicostatus and also in a field experiment. Laboratory assay showed that all eleven pyrethroids (namely, rich d-trans-allethrin, Es-biothrin, rich d-prallethrin, S-prallethrin, tetramethrin, rich d-tetramethrin, phenothrin, cyphenothrin, permethrin, cypermethrin, and high active cypermethrin) were able to inhibit barnacle settlement (EC₅₀ range of 0.0316 to 87.00 μg/ml) without significant toxicity. Analysis of structure–activity relationships suggested that the cyano group at the α-carbon position had a significant influence on the expression of antifouling activity in pyrethroids. In the field, the antifouling activity of pyrethroids was further confirmed, with the most potent pyrethroids being cypermethrin and high active cypermethrin, which displayed efficiency comparable with that of tributyltin. In summary, our investigation indicated that these pyrethroids have a great and practical commercial potential as antifouling agents.