Studies on pituitary melanotrophs reveal the novel GABAB antagonist CGP 35-348 to be the first such compound effective on endocrine cells.

One obstacle to understanding the action and physiological significance of the responsiveness of various endocrine cells to gamma-aminobutyric acid (GABA) has been that previously available substances, all active as GABAB antagonists in the nervous system, are ineffective on endocrine cells. The introduction of a potent new member of this class, CGP 35-348, of very different chemical structure, encouraged us to examine its effect on endocrine cells. For this purpose, we studied melanotroph secretion from pituitary neurointermediate lobes. We found that CGP 35-348, in contrast to previously available members of this class, suppressed completely, in rat and toad, secretory responses to baclofen, the classic GABAB agonist. Analysis, in toad, showed CGP 35-348 did not affect responses to GABAA agonists (muscimol; isoguvacine), dopamine, or neuropeptide Y. When tested against GABA, the physiological ligand present in the innervation of melanotrophs (along with dopamine and neuropeptide Y), CGP 35-348 completely suppressed the secretory response, which, in toad, is purely inhibitory and unaffected by bicuculline, the specific GABAA antagonist. In addition, CGP 35-348 unmasked a stimulant effect that bicuculline blocked. In CGP 35-348, we thus have a new tool with which to analyse responses to GABA and their physiological involvement in endocrine cells.     

Suramin, a novel antitumor compound

Suramin, a polyanionic compound originally synthesized for use as an antiparasitic agent, has recently entered clinical trials for the treatment of a variety of human cancers refractory to conventional modalities of therapy. This is based on suramin's ability to bind and to inactivate growth factor and enzyme systems critical to cellular homeostasis and proliferation. In addition, this compound possesses adrenocorticolytic properties in vivo and exerts significant cytostatic and cytocidal effects against a variety of human tumor cell lines in vitro. Pilot studies using suramin have thus far been conducted in adrenocortical carcinoma, prostate cancer refractory to conventional hormonal manipulation and nodular lymphomas.     

Studies on compound I formation of the lignin peroxidase from Phanerochaete chrysosporium

Ligninase, isolated from the wood-destroying fungus Phanerochaete chrysosporium, catalyzes the oxidation of lignin and lignin-related compounds. Ligninase reacts with H2O2 to form the classical peroxidase intermediates Compounds I and II. We have determined the activation energy of ligninase Compound I formation to be 5.9 kcal/mol. The effect of pH and ionic strength on the rate of ligninase Compound I formation was studied. In contrast to all other peroxidases, no pH effect was observed. This is despite homology of active-site amino acids residues (Tien, M., and Tu, C.-P. D. (1987) Nature 326, 520-523) which are proposed to affect the pH profile of Compound I formation. Ligninase Compound I formation can also be supported by organic peroxides. The second-order rate constants with the organic peroxides are lower, suggesting that H2O2 is the preferred substrate.     

Studies on repository compound stability in DMSO under various conditions

The chemical stability of repository compounds is affected by various environmental conditions during long-term storage. Studies were carried out to evaluate the effects of the following potential causes of instability of compounds in DMSO at a 10-mM concentration: water, oxygen, freeze/thaw cycles, and storage container material. A set of compounds was selected for the study based on structural diversity and functional group representation. Compound concentration was determined with liquid chromatography/ultraviolet spectroscopy/mass spectrometry (LC/UV/MS) analysis relative to an internal standard added to each sample. An accelerated study was conducted, and results demonstrate that most compounds are stable for 15 weeks at 40 degrees C. Water is more important in causing compound loss than oxygen. The freeze/thaw cycle study was done with freezing at -15 degrees C and thawing under nitrogen atmosphere at 25 degrees C. Two methods were used to redissolve compounds after thawing: agitation and repeated aspiration/dispense. The results indicate no significant compound loss after 11 freeze/thaw cycles. Compound recovery was also measured from glass and polypropylene containers for 5 months at room temperature, and no significant difference was found for these 2 types of containers.     

Apoptotic cell death by the novel natural compound, cinobufotalin

Cinobufotalin (CB), one of the bufadienolides prepared from toad venom, was investigated for its cytotoxicity, and the underneath mechanism involved. We primarily utilized DNA fragmentation assay and microscopic observation to assess the effect of various doses of CB in human lymphoma U937 cells. Following that, we investigated other parameters involved in cell death mechanism such as reactive oxygen species (ROS), mitochondrial membrane potential (MMP), and apoptotic proteins activation. HeLa cells were concomitantly used to generalize the data observed. Our results show that CB caused significant DNA fragmentation, decrease of MMP, and an increase in the intracellular Ca(2+) ion and ROS production. In addition, CB induced upregulation of Fas protein, proteolytic activation of cytochrome c, caspase-2, -3, -8 and -9 together with the activation of Bid and Bax. Our findings were further validated using either Fas/FasL antagonist or pan-caspase inhibitor to significantly inhibit CB-induced DNA fragmentation. In our study, we suggest that CB induces caspase dependent cell death in U937 cells, and that Fas plays a role in CB-induced apoptosis. Altogether, our data provides novel insights of the mechanism of action of CB and its potential as a future chemotherapeutic agent.     

后基因组时代的结核新疫苗研究

利用蛋白质组技术、生物信息学的DNA芯片等技术筛选适当的保护性抗原,重点在亚单位疫苗的DNA疫苗方面构建预防性和治疗性的疫苗,并配合使用亚单位疫苗,DNA疫苗和减毒活疫苗是值得重视的方向。     

杜仲叶中总黄酮物质超声提取的研究

采用醇提法、超声提取法探讨了从杜仲叶中提取总黄酮类物质的最佳工艺。试验结果表明：通过单因素试验和正交试验,超声提取效果最好。其最佳条件是：用浓度为400%、按料液比1：60的比例的乙醇浸泡24h,再用超声提取45min,其杜仲叶中总黄酮类物质的提出率可达25．43％。     

月腺大戟根中有效成分乙素和丙素的结构研究

本文主要报道从月腺大戟(Euphorbia ebracteolata Hayata)根中分离出的对结核菌有抑制作用的晶Ⅵ(乙素)和晶Ⅶ(丙素)的化学结构。经化学反应及光谱鉴定,确定晶Ⅵ为2,4-二羟基-6-甲氧基-3-甲基-苯乙酮,晶Ⅶ为2-羟基-6-甲氧基-3-甲基-1-苯乙酮-4-6-葡萄糖甙。乙素为首次分离的天然产物,丙素为新的化合物。该植物提取物对结核病有明显疗效。     

The effect of a novel anti-adherent compound on the adherence of oral streptococci to hydroxyapatite

The effect of the copolymer M239, 144 with and without chlorhexidine on the adherence of oral streptococci to saliva-coated hydroxyapatite was investigated. At 1% w/v M239, 144 reduced the adherence of Streptococcus sanguis NCTC 7863 by 94%. It had a moderate effect on the adherence of other Strep. sanguis strains and a Streptococcus gordonii strain but had no effect on the adherence of Streptococcus oralis or Streptococcus mutans. Chlorhexidine did not influence the anti-adhesive properties of 1% w/v M239, 144.     

Studies on the formation of germ cells in a compound ascidian Botryllus,primigenus oka

Gametogenesis of a compound ascidian Botryllus primigenus was studied histologically. On either side of the zooid (stage 9), in the gonadal space between the epidermis and the atrial epithelium, either a single testis or a complex of an egg follicle and a testis can be formed. The egg follicle consists of a single ovum (occasionally two ova) and its accessory cells and is connected with the atrial epithelium by the follicle stalk. The egg follicle is always accompanied by the brood pouch, a diverticulum of the atrial cavity. The testis is equipped with a vestigial spermiduct and is attached to the atrial epithelium. Buds of stage 8 comprise, besides the developing testes and, egg follicles, loose aggregations of hemoblasts and oocytes of early developmental stages, which are already accompanied by primary follicular cells. Both the oocytes and the primary follicular cells seem to arise from the hemoblasts. The young oocytes are isolated in the gonadal space of the buds nnd are transferred to buds of the succeeding generations until they finally mature. In the bud of stage 3, a compact mass of cells appears, attaching to tbe inner vesicle on either side of the body. It is derived from the hemoblasts lodged there in the preceding generation and presumably also from the circulating hemoblasts. When the cell mass receives a large oocyte derived from the preceding generation, part of the cell mass differentiates into egg envelopes, forming an egg follicle, and a follicle stalk and the remainder into a testis. When the cell mass receives no oocyte, it differentiate as a whole into a testis. In the egg follicle thus formed the outer and inner follicular cells increase in number by mitotic division. Subsequently, initial test cells are derived from the inner follicle by migration across the developing chorion; then they increas2 in number by mitosis. In the testis, meiosis and spermiogenesis take place.     

A novel dicyclodextrinyl ditelluride compound with antioxidant activity.

Reactive oxygen species (ROS) primarily arise from products of normal metabolic activities and are thought to be the etiology of many diseases. A novel dicyclodextrinyl ditelluride (2-TeCD) compound was designed to be a functional mimic of the glutathione peroxidase that normally removes ROS. 2-TeCD exhibited highly catalytic efficiency and good water solubility. Antioxidant activity was studied by using ferrous sulfate/ascorbate-induced mitochondria damage model system. 2-TeCD protected the mitochondria against oxidative damage in a dose-dependent manner and exhibited also great antioxidant ability in comparison with 2-phenyl-1,2-benziososelenazol-3(2H)-one. The mimic may result in better clinical therapies for the treatment of ROS-mediated diseases.     

A novel dicyclodextrinyl diselenide compound with glutathione peroxidase activity

A 6A,6A'-dicyclohexylamine-6B,6B'-diselenide-bis-beta-cyclodextrin (6-CySeCD) was designed and synthesized to imitate the antioxidant enzyme glutathione peroxidase (GPX). In this novel GPX model, beta-cyclodextrin provided a hydrophobic environment for substrate binding within its cavity, and a cyclohexylamine group was incorporated into cyclodextrin in proximity to the catalytic selenium in order to increase the stability of the nucleophilic intermediate selenolate. 6-CySeCD exhibits better GPX activity than 6,6'-diselenide-bis-cyclodextrin (6-SeCD) and 2-phenyl-1,2-benzoisoselenazol-3(2H)-one (Ebselen) in the reduction of H(2)O(2), tert-butyl hydroperoxide and cumenyl hydroperoxide by glutathione, respectively. A ping-pong mechanism was observed in steady-state kinetic studies on 6-CySeCD-catalyzed reactions. The enzymatic properties showed that there are two major factors for improving the catalytic efficiency of GPX mimics. First, the substrate-binding site should match the size and shape of the substrate and second, incorporation of an imido-group increases the stability of selenolate in the catalytic cycle. More efficient antioxidant ability compared with 6-SeCD and Ebselen was also seen in the ferrous sulfate/ascorbate-induced mitochondria damage system, and this implies its prospective therapeutic application.     

Studies on the kinetic mechanism and the phosphoryl-enzyme compound of the Escherichia coli acetate kinase reaction.

The Escherichia coli acetate kinase was further examined to determine whether a phosphoryl-enzyme intermediate (or compound) was kinetically discernible or chemically essential in the catalysis of that reaction. The extent of phosphoryl-enzyme formation as monitored by gel filtration of the enzyme-phosphoryl donor substrate complex (E-P) was found to be quite variable. Isolated E-P could be reduced with sodium borohydride and the extent of formation of α-amino-δ-hydroxyvalerate paralleled the observed degree of phosphorylation of the acetate kinase. Incubation with 1 m neutral hydroxylamine of the enzyme, which had been preincubated with acetyl-P for 30–60 min, led to parallel loss of the enzyme's ability to catalyze the net reaction and the ADP ATP exchange reaction. The percentage of activity loss depended upon the concentration of acetyl-P with which the enzyme was preincubated. These observations point toward a requirement for an unmodified active site carboxyl group which may be phosphorylated during the reaction course. The theoretical basis for a new and simple alternative substrate protocol for segregating sequential and ping-pong mechanisms is presented. When applied to acetate kinase, the findings clearly rule out a ping-pong kinetic mechanism. This was also confirmed by initial rate measurements with propionyl-P and ADP, and the weight of the evidence now clearly favors a modified random substrate addition pathway as presented herein. The data do not in any way exclude E-P formation as a requirement for catalysis. Finally, the rates of cold inactivation at 0 °C and reactivation at 29 °C were determined in order to minimize the possible influence of this phenomenon on the kinetic studies.