小剂量洋地黄对急性心肌梗死PCI术后合并心力衰竭患者心率变异性的影响

目的:探讨早期应用小剂量洋地黄类药物对急性心肌梗死(Acute myocardial infarction,AMI)行经皮冠状动脉介入治疗(Percutaneous coronary intervention,PCI)术后合并心力衰竭患者心率变异性(Heart rate variability,HRV)的影响。方法:入选32例在发病24小时内接受PCI治疗且合并心力衰竭的AMI患者,再灌注后随机分为洋地黄组(西地兰0.2 mg,n=17)和对照组(生理盐水20 m L,n=15)。在用药前、用药后30分钟、用药后3小时、用药后6小时、用药后12小时、用药后24小时进行5分钟HRV分析。结果:1洋地黄组的心率在用药6小时后显著小于对照组(P0.05);2洋地黄组SDNN在用药后3小时-6小时显著大于对照组(P0.05),两组RMSSD比较无显著统计学差别(P0.05);3洋地黄组LFnorm在用药后3小时-6小时显著大于对照组(P0.05);用药3小时后,洋地黄组HFnorm显著大于对照组(P0.05),LF/HF显著小于对照组(P0.05)。结论:小剂量洋地黄可以显著降低AMI PCI术后合并心力衰竭患者的心率、逆转迷走神经与交感神经活性的失衡状态,改善HRV。     

Activation of conventional and novel protein kinase C isozymes by different diacylglycerol molecular species

A variety of diacylglycerol (DG) molecular species are produced in stimulated cells. Conventional (α, βII and γ) and novel (δ, ε, η and θ) protein kinase C (PKC) isoforms are known to be activated by DG. However, a comprehensive analysis has not been performed. In this study, we analyzed activation of the PKC isozymes in the presence of 2–2000 mmol% 16:0/16:0-, 16:0/18:1-, 18:1/18:1-, 18:0/20:4- or 18:0/22:6-DG species. PKCα activity was strongly increased by DG and exhibited less of a preference for 18:0/22:6-DG at 2 mmol%. PKCβII activity was moderately increased by DG and did not have significant preference for DG species. PKCγ activity was moderately increased by DG and exhibited a moderate preference for 18:0/22:6-DG at 2 mmol%. PKCδ activity was moderately increased by DG and exhibited a preference for 18:0/22:6-DG at 20 and 200 mmol%. PKCε activity moderately increased by DG and showed a moderate preference for 18:0/22:6-DG at 2000 mmol%. PKCη was not markedly activated by DG. PKCθ activity was the most strongly increased by DG and exhibited a preference for 18:0/22:6-DG at 2 and 20 mmol% DG. These results indicate that conventional and novel PKCs have different sensitivities and dependences on DG and a distinct preference for shorter and saturated fatty acid-containing and longer and polyunsaturated fatty acid-containing DG species, respectively. This differential regulation would be important for their physiological functions.     

New Cardenolide Glycosides from the Seeds of Digitalis purpurea and Their Cytotoxic Activity

A chemical investigation of Digitalis purpurea seeds led to the isolation of three new cardenolide glycosides (1, 8 and 11), together with 12 known cardenolide glycosides (2–7, 9, 10 and 12–15). The structures of 1, 8 and 11 were determined by 1D and 2D NMR spectroscopic analyses and the results of an acid or enzymatic hydrolysis. The cytotoxic activity of the isolated compounds (1–15) against HL-60 leukemia cells was examined. Compounds 2, 9, 11 and 12 showed potent cytotoxicity against HL-60 cells with respective 50% inhibition concentration (IC₅₀) values of 0.060, 0.069, 0.038, and 0.034 µM. Compounds 2, 9 and 11 also exhibited potent cytotoxic activity against HepG2 human liver cancer cells with respective IC₅₀ values of 0.38, 0.79, and 0.71 µM. An investigation of the structure-activity relationship showed that the cytotoxic activity was reduced by the introduction of a hydroxy group at C-16 of the digitoxigenin aglycone, methylation of the C-3' hydroxy group at the fucopyranosyl moiety, and acetylation of the C-3' hydroxy group at the digitoxopyranoyl moiety.     

Design,synthesis and biological evaluation of novel human monoamine oxidase B inhibitors based on a fragment in an X-ray crystal structure

Herein we report our efforts of developing reversible selective hMAO-B inhibitors based on isatin, a fragment in an X-ray crystal structure. Five different scaffolds were designed and many compounds were synthesized. Among them, compound A3 demonstrated very high potency and isoform selectivity against hMAO-B, 11 and 13 times more potent (IC₅₀?=?3?nM) and 23.64 and 6.8 times more selective than the marked drugs, selegiline and safinamide. However, the endeavors to modify the polar 3-one group of isatin, that is in a hydrophobic environment in the binding site of hMAO-B, to small nonpolar hydrophobic groups did not bring about improved hMAO-B inhibitors, which may challenge our understanding of molecular interactions and molecular recognition in biological systems.     

Why three Rho proteins? RhoA, RhoB, RhoC, and cell motility

Higher vertebrates have 3 Rho GTPases, RhoA, RhoB, and RhoC, which share 85% amino acid sequence identity. Here, we compare and contrast the roles of RhoA, B, and C in the regulation of the cytoskeleton and cell motility. Despite their similarity, some regulators and effectors show preferential interaction with RhoA, B, or C, and the three proteins show differences in function in cells. RhoA plays a key role in the regulation of actomyosin contractility. RhoB, which is localized primarily on endosomes, has been shown to regulate cytokine trafficking and cell survival, while RhoC may be more important in cell locomotion. In cancer cells, the expression and activity of RhoA, B, and C is altered in different ways. Together, this evidence suggests that although the 3 isoforms of Rho are structurally highly homologous, they have different cellular functions.     

Carbonic anhydrase activators: Activation of the human cytosolic isozyme III and membrane-associated isoform IV with amino acids and amines

An activation study of the human carbonic anhydrase (hCA, EC 4.2.1.1) isoforms hCA III (cytosolic) and IV (membrane-associated) with a series of natural and non-natural amino acids and aromatic/heterocyclic amines is reported. hCA III was efficiently activated by d-His, serotonin, pyridyl-alkylamines, and aminoethyl-piperazine/morpholine (K_As of 91nM–1.12 μM), whereas the best hCA IV activators were 4-amino-phenylalanine, serotonin, and 4-(2-aminoethyl)-morpholine (K_As of 79 nM–3.14 μM). Precise steric and electronic requirements are needed to be present in the molecules of effective CA III/IV activators, in order to assure an adequate fit within the enzyme active site for the formation of the enzyme-activator complex, and for efficient proton transfer processes between the active site and the reaction medium. The activation profiles of CA III and IV are distinct from those of all other mammalian CA isoforms investigated so far for their interaction with amino acids and amines.     

Comparative genomics analysis of metallothioneins in twelve Drosophila species

Metallothioneins (MTs) are a superfamily of Cys-rich polypeptides that bind heavy metal ions, both for physiological and detoxification purposes. They are present in all organisms, but their origin is probably polyphyletic, so that MT evolutionary studies are rather scarce. We present a thorough search and analysis of the MT coding sequences in the 12 Drosophila genomes completely sequenced, taking as reference the features reported for D. melanogaster, where four isogenes (MtnA to MtnD) are known and deeply characterized. We include a fifth isoform in this study, named MtnE, and recently annotated. The MTs polymorphism pattern is essentially the same for the 12 Drosophila species. Invariably, a MtnA form and an MtnB-cluster, comprising the MtnB-to-MtnE forms in tandem array, are observed. The whole set of genes are kept in the same synteny element (Muller E), but implicated in rearrangement events (mainly inversions), encompassing all or some of the isogenes. Gene exon/intron architecture, and cDNA and protein sequences appear highly conserved through Drosophila speciation, concordantly with an essential function for MT isoforms in flies, even for those previously considered as minor products. Data presented here will be comprehensively analyzed to provide a valuable guide for future MT evolutionary, structure and function studies.     

人血红素加氧酶同工酶的分离纯化初报

首次报道了人肝脏血红素加氧酶（hemeoxggenase,HO）的同工酶的分离纯化,并初步探讨了它们的性质．采用DEAE-SephadexA-25和羟基磷灰石柱层析法从人肝脏分离纯化H0的同工酶,酶活性检测、SDS-PAGE分析结果显示,人肝脏微粒体含HO的同工酶,按洗脱先后顺序分别得到分子量为30000和36000的HO-1和HO-2．酶促反应中需相同辅酶参与,其中酶活性HO-1明显高于HO-2,两者之比为2.4:1,从分子量和酶的催化活性分析发现,HO-1属诱导型的传统HO.HO-2为新发现的非诱导型HO的同工酶．     

不同因子对毛地黄叶离体培养强心苷含量的影响

对毛地黄幼叶离体培养中,影响毛地黄强心苷含量的因子进行了研究。培养基的组成成分对毛地黄强心苷的合成十分重要,ＭＳ＋２,４－Ｄ１．０ｍｇ／Ｌ＋ＢＡ１．５ｍｇ／Ｌ＋活性炭０．５％有明显促进作用;外源植物激素是合成毛地黄强心苷必需的,在ＭＳ＋２,４－Ｄ１．０ｍｇ／Ｌ基本培养基中附加ＢＡ,ＢＡ的浓度在０．２－１．５ｍｇ／Ｌ范围内,浓度越大,强心苷含量越高;不同继代培养细胞的时期对毛地黄强心苷的合成起重要作用,随着继代的增加,毛地黄强心苷的含量呈渐降趋势;细胞的生长后期对毛地黄强心苷的合成效果最佳;不同光质的光照培养对毛地黄强心苷的积累也有影响,蓝光明显提高强心苷的合成量。     

Brain expression of the calcineurin inhibitor RCAN1 (Adapt78)

RCAN1 (Adapt78) is an endogenous inhibitor of calcineurin, an important intracellular phosphatase that mediates many cellular responses to calcium. RCAN1 is expressed in multiple organs, especially heart, skeletal muscle and brain. In brain, it is thought to be important due to its strong expression, developmental regulation, abundance of target protein (calcineurin), and putative links to multiple brain-related disorders. Surprisingly, however, few studies have examined RCAN1 protein expression here. This has led to some confusion in the field over the exact nature and cell-type expression of isoform 4, the more studied of the two major RCAN1 protein isoforms, in brain. Here we characterize RCAN1 brain isoforms in more detail by assessing their size and distribution under conditions of calcium elevation, a hallmark of the isoform 4 response, and using rodent models to allow for more expanded analyses. We find that the 25-29 kDa version of this protein, reported in many non-brain studies, is indeed also present in neurons, and most observable after calcium induction. We also observe that expression of isoform 4 is not specific to neurons, as both microglia and astrocyte cells in culture exhibit a strong induction of isoform 4 protein following calcium stress that is not observable in non-stressed tissue sections. Isoform 1 expression is also observable in a primary glial cell-type (rat microglia). Finally, our observations confirm previous reports of low or non-detectable constitutive isoform expression in non-stressed glia, and of a larger sized, RCAN1 antibody-interacting species. These studies extend and complement previous studies on RCAN isoforms toward better understanding the role of RCAN1 in brain function and as a potential new target for treating calcineurin-related brain disorders.     

Evolution of tubulin isoforms during the sporophytic development of Allomyces arbuscula

Abstract Tubulins extracted from the sporophytic developmental stages of Allomyces arbuscula have been characterised by one- and two-dimensional SDS-PAGE gels immunoblotted with monoclonal antibodies as α-, acetylated α- ( M _r 57 kDa both) and β- ( M _r 55 kDa) isoforms. The zoosporangial isoforms could be characterised only when precautions were taken to inhibit the strong tubulin proteolytic activity at this stage. The zoospores and zoosporangia contained greater amounts of the acetylated α- and β-isoforms than the mycelium, while the non acetylated α-isoform was present in greater quantity in the mycelium than in the zoospores or zoosporangia.     

豌豆卷须肌动蛋白Ⅱ类异型体cDNA克隆的序列分析

分析１５个豌豆卷须肌动蛋白ｃＤＮＡ 克隆的限制性内切酶图谱,发现在豌豆卷须中至少存在三类肌动蛋白异型体,分别命名为Ⅰ类（ＰＥＡｃ Ⅰ）、Ⅱ类（ＰＥＡｃ Ⅱ）和Ⅲ类（ＰＥＡｃ Ⅲ）异型体．三类异型体的克隆数目分别为１０、４和１个,表明三类异型体在豌豆卷须中的表达是不同的,很可能具有组织或发育阶段的特异性．对Ⅱ类异型体的三个ｃＤＮＡ 克隆ＰＥＡｃ３、ＰＥＡｃ９和ＰＥＡｃ１１进行了全序列测定,所测定的序列已被ＧｅｎＢａｎｋ 数据库所接受．测序结果表明,ＰＥＡｃ３、ＰＥＡｃ９和ＰＥＡｃ１１的序列长度分别为１５５０、１６８０和１０９１个核苷酸,其中编码区长１１３４个核苷酸,编码的氨基酸长度为３７７（ＰＥＡｃ１１缺少编码氨基端前９６个氨基酸的核苷酸序列）．三个克隆的核苷酸序列完全相同,差别仅在于３′非翻译区的长度不同,即ｐｏｌｙ（Ａ）的加入位点不同．这说明它们可能是由同一基因转录而来,但转录后的加工过程不同．豌豆卷须中肌动蛋白基因ｐｏｌｙ（Ａ）加入位点的使用,可能与组织或发育的特异性表达有关．此外,豌豆卷须肌动蛋白三类异型体之间的核苷酸序列同源性为８０％ ,氨基酸序列同源性为９４％ ．     

Elevated Variance in Heart Rate During Slow-Wave Sleep After Late-Night Physical Activity

This study investigates the effect of mild physical activity before bedtime on the sleep pattern and heart rate during the night. Nine healthy subjects underwent a habituation night, a reference night, and a physical induction night. The physical induction night did not alter the sleep pattern. Physical activity before bedtime resulted in higher heart rate variance during slow-wave sleep. The low-frequency/high-frequency component (LF/HF) ratio during slow-wave sleep in the physical induction night was significantly higher than during the reference night. Increased mean heart rate and higher LF/HF ratio are related to decreased parasympathetic dominance. Exercise up to 1?h before bedtime thus seems to modify the quality of sleep. (Author correspondence: Daniel.Berckmans@biw.kuleuven.be)     

Regulation of vascular function by RCAN1 (ADAPT78)

RCAN1 (Adapt78) functions mainly, if not exclusively, as a regulator of calcineurin, a phosphatase that mediates many cellular responses to calcium. Identification of this regulatory activity has led to a surge of interest in RCAN1, since calcineurin is involved in many cellular and tissue functions, and its abnormal expression is associated with multiple pathologies. Recent studies have implicated RCAN1 as a regulator of angiogenesis. To more fully investigate the role of RCAN1 in vascular function, we first extended previous studies by assessing RCAN1 response in cultured endothelial cells to various vascular agonists. Strong induction of isoform 4 but not isoform 1 was observed in human umbilical vein- and bovine pulmonary aortic-endothelial cells in response to VEGF, thrombin, and ATP but not other agonists. Inductions were both calcium and calcineurin dependent, with the relative effect of each agonist cell-type dependent. Ectopic RCAN1 expression also inhibited calcineurin signaling in the HUVEC cells. Based on these strong RCAN1 responses and a lack of RCAN1-associated vascular studies beyond angiogenesis, we investigated the potential role of RCAN1 in vascular tone using whole mounted mesenteric artery. RCAN1 knockout mice exhibited an attenuated mesenteric vasoconstriction to phenylephrine as compared with wild-type. Overall contractility was unaffected, suggesting that this component of smooth muscle action is similar in the two mouse strains. Constriction in the knockout artery appeared to be potentiated by the addition of the nitric oxide synthase (NOS) inhibitor l-NAME, suggesting that elevated nitric oxide (NO) production occurs in the knockout vasculature and contributes to the weakened vasoconstriction. Our results reveal a newly identified vascular role for RCAN1, and a potential new target for treating vascular- and calcineurin-related disorders.     

Structure and property based design, synthesis and biological evaluation of γ-lactam based HDAC inhibitors

Histone deacetylases (HDACs) are involved in post-translational modification and gene expression. Cancer cells recruited amounts of HDACs for their survival by epi-genetic down regulation of tumor suppressor genes. HDACs have been the promising targets for treatment of cancer, and many HDAC inhibitors have been investigated nowadays. In previous study, we synthesized δ-lactam core HDAC inhibitors which showed potent HDAC inhibitory activities as well as cancer cell growth inhibitory activities. Through QSAR study of the δ-lactam based inhibitors, the smaller core is suggested as more active than larger one because it fits better in narrow hydrophobic tunnel of the active pocket of HDAC enzyme. The smaller γ-lactam core HDAC inhibitors were designed and synthesized for biological and property optimization. Phenyl, naphthyl and thiophenyl groups were introduced as the cap groups. Hydrophobic and bulky cap groups increase potency of HDAC inhibition because of hydrophobic interaction between HDAC and inhibitors. In overall, γ-lactam based HDAC inhibitors showed more potent than δ-lactam analogues.