Effect of insulin detemir, compared to human insulin, on 3T3-L1 adipogenesis

Insulin detemir (DET) represents a myristic acid (MA)-coupled insulin derivative with protracted action due to reversible albumin binding. As compared to human insulin (HI), DET provokes no or only minor body weight gain in vivo. Therefore, we compared DET's and HI's adipogenic effects. 3T3-L1 preadipocytes were differentiated with 5 nmol/l HI, 5 nmol/l DET (=DET(equimolar)), or 20 nmol/l DET (=DET(equipotent); equipotent in terms of the reported metabolic potency in vitro). Due to differentiation-suppressive effects, albumin was excluded from the studies. During the induction period, only HI allowed clonal expansion. Moreover, HI induced a 200-fold increase in specific glycerol-3-phosphate dehydrogenase activity, whereas DET(equimolar) and DET(equipotent) were markedly less adipogenic (P相似文献   

Hepatoprotective effect and mechanistic insights of deoxyelephantopin,a phyto-sesquiterpene lactone,against fulminant hepatitis

Deoxyelephantopin (DET) is an abundant sesquiterpene lactone isolated from an anecdotally hepatoprotective phytomedicine, Elephantopus scaber. Our objective in this study was to provide scientific evidence for the in vivo efficacy and the underlying mechanisms of action of DET in lipopolysaccharide/d-galactosamine (LPS/D-GalN)-induced fulminant hepatitis. We investigated both the protective effect of pretreatment with DET (10 mg/kg body weight, Pre-DET10) prior to administration of LPS/D-GalN and the therapeutic effect of treatment with 10 mg/kg DET (Post-DET10) or the hepatoprotective drug silymarin (Post-SM10) following the administration of LPS/D-GalN. Our data showed that Pre-DET10 prevented LPS/D-GalN-induced infiltration of F4/80 monocytes/macrophages and an increase of nitrotyrosine and cyclooxygenase-2 protein in liver tissues. Further, Post-DET10 and Psot-SM10 treatments protected against liver cell apoptosis. All three treatments suppressed serum aminotransferase activities, tumor necrosis factor-alpha and interleukin-6 levels, and serum and hepatic matrix metalloproteinase-9 activity. The Pre-DET10 or Post-DET10 and Post-SM10 treatments in combination with inhibition of heme oxygenase-1 expression ultimately decreased protection of mice from LPS/D-GalN-induced mortality, with decreased survival from 75% and 62.5% to 50%, respectively. Results obtained from serial liver scintigraphy with ^99mTc-diisopropyl iminodiacetic acid (DISIDA) on single-photon emission computed tomography analysis showed that both liver uptake and excretion times of DISIDA were significantly delayed in LPS/D-GalN-treated animals and were effectively recovered by DET and silymarin treatment. This report demonstrates that DET functions in the modulating multiple molecular targets or signaling pathways that counteract inflammation during the progression of fulminant hepatitis and may serve as a novel lead compound for future development of anti-inflammatory or hepatoprotective agents.     

Mammalian DET1 regulates Cul4A activity and forms stable complexes with E2 ubiquitin-conjugating enzymes

DET1 (de-etiolated 1) is an essential negative regulator of plant light responses, and it is a component of the Arabidopsis thaliana CDD complex containing DDB1 and COP10 ubiquitin E2 variant. Human DET1 has recently been isolated as one of the DDB1- and Cul4A-associated factors, along with an array of WD40-containing substrate receptors of the Cul4A-DDB1 ubiquitin ligase. However, DET1 differs from conventional substrate receptors of cullin E3 ligases in both biochemical behavior and activity. Here we report that mammalian DET1 forms stable DDD-E2 complexes, consisting of DDB1, DDA1 (DET1, DDB1 associated 1), and a member of the UBE2E group of canonical ubiquitin-conjugating enzymes. DDD-E2 complexes interact with multiple ubiquitin E3 ligases. We show that the E2 component cannot maintain the ubiquitin thioester linkage once bound to the DDD core, rendering mammalian DDD-E2 equivalent to the Arabidopsis CDD complex. While free UBE2E-3 is active and able to enhance UbcH5/Cul4A activity, the DDD core specifically inhibits Cul4A-dependent polyubiquitin chain assembly in vitro. Overexpression of DET1 inhibits UV-induced CDT1 degradation in cultured cells. These findings demonstrate that the conserved DET1 complex modulates Cul4A functions by a novel mechanism.     

Characterization of polyamines having agonist, antagonist, and inverse agonist effects at the polyamine recognition site of the NMDA receptor

The endogenous polyamines spermine and spermidine increase the binding of [3H]MK-801 to NMDA receptors. This effect is antagonized by diethylenetriamine (DET). We report here that spermine increases the rates of both association and dissociation of binding of [3H]MK-801, suggesting that it increases the accessibility of the binding site for MK-801 within the ion channel of the receptor complex. 1,10-Diaminodecane (DA10) inhibited the binding of [3H]MK-801. This effect was due to a decrease in the rate of association with no change in the rate of dissociation of [3H]MK-801. The effect of DA10 was not mediated by an action of DA10 at the binding sites for glutamate, glycine, Mg2+, or Zn2+, and was attenuated by DET. This suggests that DA10 acts at the polyamine recognition site. In hippocampal neurons the NMDA-elicited current was decreased by DA10, an effect opposite to that of spermine. The effects of spermine and DA10 were selectively blocked by DET. It is concluded that DA10 acts as a negative allosteric modulator or inverse agonist at the polyamine recognition site of the NMDA receptor.     

DGT/DET与CID技术联用获取环境微界面元素异质性分布特征：进展与展望

环境中广泛存在的微界面是物质循环和能量交换的门户，显著影响元素(如S、Fe和P)的迁移转化过程与生态效应。而微界面元素分布具有高度的时空异质性，因此开发和应用原位且高分辨率的表征技术手段显得尤为必要。薄膜扩散梯度(DGT)和薄膜扩散平衡(DET)技术是研究微界面元素和化合物分布及其生化过程的利器。近30年的大量实验证据表明，DGT/DET通过扩散-吸附/平衡作用原位被动采样结合后续高分辨化学分析手段，比如一维/二维膜物理切割-化学仪器分析、激光剥蚀-电感耦合等离子体质谱(LA-ICP-MS)以及计算机/比色密度成像计量法(CID),可以获取元素的高分辨浓度分布和生物有效性信息以及多元素间耦合作用的动态过程。在上述3种后续分析方法中，CID手段是最为便捷、快速和廉价的，空间分辨率与LA-ICP-MS分析相当，纵横方向均可达几十至几百微米，推广应用潜力巨大。本文在对DGT/DET和CID技术概述的基础上，总结了技术联用的思路，并依据获取二维高分辨图像的步骤差异对技术联用的进展进行了分类介绍，一类是原位即时显色成像，比如以AgI为吸附膜的DGT与CID方法联用测定硫化物，另一类是预处理后显色...     

Effects of diethyldithiocarbamate on the acute toxicity and the acetylcholinesterase inhibition by methylparathion in mice]

Acute toxicity of methylparathion male mice is reduced by DET in dose-range from 50 to 300 mg/kg. Mean survival time in LD100 range of methylparathion is significantly prolonged. Inhibition of AChE in vivo is reduced, too. The observed dose-effect-relations show an approx. 10-fold increase in equi-effective doses after DET-pretreatment. DET has no protective effect both in vivo and in vitro, if methylparathion is added to the brain homogenate of mice. As probable mechanism of action a change in methylparathion-metabolism induced by DET in the system of mixed functional oxidases is discussed.     

Deoxyelephantopin impedes mammary adenocarcinoma cell motility by inhibiting calpain-mediated adhesion dynamics and inducing reactive oxygen species and aggresome formation

We previously showed that deoxyelephantopin (DET), a plant sesquiterpene lactone, exhibits more profound suppression than paclitaxel (PTX) of lung metastasis of mammary adenocarcinoma TS/A cells in mice. Proteomics studies suggest that DET affects actin cytoskeletal protein networks and downregulates calpain-mediated proteolysis of several actin-associated proteins, whereas PTX mainly interferes with microtubule proteins. Here, DET was observed to significantly deregulate adhesion formation in TS/A cells, probably through inhibition of m-calpain activity. Epithelial growth factor (EGF)-mediated activation of Rho GTPase Rac1 and formation of lamellipodia in TS/A cells were remarkably suppressed by DET treatment. Further, DET impaired vesicular trafficking of EGF and induced protein carbonylation and formation of centrosomal aggregates in TS/A cells. DET-induced reactive oxygen species were observed to be the upstream stimulus for the formation of centrosomal ubiquitinated protein aggregates that might subsequently restrict cancer cell motility. PTX, however, caused dramatic morphological changes, interfered with microtubule networking, and moderately inhibited calpain-mediated cytoskeletal and focal adhesion protein cleavage in TS/A cells. This study provides novel mechanistic insights into the pharmacological action of DET against metastatic mammary cell migration and suggests that modulation of oxidative stress might be a potential strategy for treatment of metastatic breast cancer.     

Evidence for FUS6 as a component of the nuclear-localized COP9 complex in Arabidopsis.

The pleiotropic CONSTITUTIVE PHOTOMORPHOGENIC (COP), DEETIOLATED (DET), and FUSCA (FUS) loci are essential regulatory genes involved in the light control of seedling developmental patterns in Arabidopsis. Although COP1, DET1, COP9, and FUS6 (also called COP11) have been cloned, their biochemical activities and interactions remain elusive. We have recently suggested that multiple pleiotropic COP, DET, and FUS genes may encode subunits of a large regulatory complex. In this study, we generated specific antibodies against Arabidopsis FUS6 and show that accumulation of both COP9 and FUS6 is coordinated in the pleiotropic cop, det, and fus mutant backgrounds and in wild-type plants throughout development. Both COP9 and FUS6 cofractionated into identical high molecular mass fractions in an analytical gel filtration assay, and neither was found in its monomeric form. Moreover, antibodies raised against either COP9 or FUS6 selectively coimmunoprecipitated both proteins. We have also developed an Arabidopsis protoplast immunolocalization assay and demonstrated that the COP9 complex is localized in the nucleus and that its nuclear localization is not affected by light conditions or tissue types. The integrated genetic and biochemical results strongly support the conclusion that both COP9 and FUS6 are components of the nuclear-localized COP9 complex. Therefore, we have provided the strongest evidence for the conclusion that at least some of the pleiotropic COP, DET, and FUS loci act in the same signaling pathway.     

Climacostol inhibits <Emphasis Type="Italic">Tetrahymena</Emphasis> motility and mitochondrial respiration

Climacostol is a resorcinol derivative that is produced by the ciliate Climacostomum virens. Exposure to purified climacostol results in lethal damage to the predatory ciliate Dileptus margaritifer and several other ciliates. To elucidate the mechanism of climacostol toxic action, we have investigated the effects of this compound on the swimming behavior of Tetrahymena thermophila and the respiratory system of isolated rat liver mitochondria. When added to living T. thermophila cells, climacostol markedly increased the turning frequency that was accompanied by a decrease in swimming velocity and subsequently by cell death. Observations by DIC and fluorescence microscopy showed morphological alterations in climacostol treated T. thermophila, indicating that climacostol might exert cytotoxic action on this organism. In the experiment with isolated rat liver mitochondria, climacostol was found to inhibit the NAD-linked respiration, but had no apparent effect on succinate-linked respiration. This finding indicates that climacostol specifically inhibits respiratory chain complex I in mitochondria. The combination of results suggest that the inhibition of mitochondrial respiration may be the cytotoxic mechanism of climacostol’s defenses against predatory protozoa.     

Modulation of positive inotropy and metabolic coronary dilatation by myocardial alpha-adrenoceptors

Cardiac hyperactivity and its consequent metabolically induced coronary vasodilation (MCD) were studied in isolated, perfused, electrically paced rat hearts. The alpha-adrenoceptor agonists, phenylephrine and methoxamine, produced a concentration-dependent inhibition of the inotropic responses to noradrenaline, dobutamine, isoprenaline, tyramine, and glucagon, while relatively potentiating their MCD reactions. This inhibition was unrelated to the alpha-agonists' known inotropic action and was not affected by catecholamine depletion of the heart. Withdrawal of the alpha-agonists or administration of the alpha-adrenoceptor antagonists phentolamine, phenoxybenzamine, or prazosin returned the inotropic and MCD reactions to normal. Neither the MCD response to electrically induced tachycardia nor the inotropic reactions produced by calcium chloride were affected by alpha-adrenoceptor agonists or antagonists. Alone, alpha-adrenoceptor antagonists were shown to potentiate the inotropic responses to noradrenaline and isoprenaline while the MCD was relatively diminished. The responses to glucagon were unaltered by alpha-antagonists. We postulate that myocardial reactivity to sympathetic stimulation can be modulated through alpha-adrenoceptors by the inhibition of processes that mediate cardiostimulation at post-beta-adrenoceptor sites, together with facilitation of those leading up to MCD. Accordingly, this modulation would act to prevent ischaemic damage to the heart by acting to limit the inotropic responses to increasing sympathetic stimulation while maximizing the blood supply to the myocardium.     

The effects of Sp7/Osterix gene silencing in the chondroprogenitor cell line, ATDC5

Ubiquitin is a small polypeptide and ubiquitination is the post-translational modification by ubiquitin protein, resulting in degradation of target proteins by the 26S proteasome complex. Here, we found that E3 ubiquitin ligase SINAT5, an Arabidopsis homologue of the Drosophila SINA RING-finger protein, interacts directly with LHY, a component of the circadian oscillator, and DET1, a negative regulator of light-regulated gene expression. We also found that SINAT5 has E3 ubiquitination activity for LHY but not for DET1. Interestingly, LHY ubiquitination by SINAT5 was inhibited by DET1. Late flowering of sinat5 mutants indicates that flowering time can be controlled by DET1 through regulation of LHY stability by SINAT5.     

Kallikrein-thrombolytic and hypotensive action in cats--preliminary results.

An intravenous injection of kallikrein produced hypotensive and thrombolytic effects in anesthetized cats, using the blood superfused tendon technique. The thrombolytic action of kallikrein was mediated by an unstable substance. The generation of this substance was abolished by either acetylsalicylic acid (ASA) or aprotonin and enhanced by captopril. The hypotensive action of kallikrein was only partially inhibited by ASA. It is proposed that both these pharmacological effects of kallikrein are mediated by bradykinin which in turn releases prostacyclin from the endothelium. However, in contrast to the thrombolytic effect of kallikrein which is totally mediated by prostacyclin the hypotensive action of kallikrein depends not only on prostacyclin but also on another endothelium-derived vasorelaxant, e.g. EDRF.     

ABI3 affects plastid differentiation in dark-grown Arabidopsis seedlings

The Arabidopsis ABSCISIC ACID-INSENSITIVE3 (ABI3) protein has been identified previously as a crucial regulator of late seed development. Here, we show that dark-grown abi3 plants, or abi3 plants returned to the dark after germination in the light, developed and maintained an etioplast with a prominent prolamellar body at developmental stages in which the wild type did not. Overexpression of ABI3 led to the preservation of the plastid ultrastructure that was present at the onset of darkness. These observations suggest that ABI3 plays a role in plastid differentiation pathways in vegetative tissues. Furthermore, the analysis of deetiolated (det1) abi3 double mutants revealed that DET1 and ABI3 impinge on a multitude of common processes. During seed maturation, ABI3 required DET1 to achieve its full expression. Mature det1 abi3 seeds were found to be in a highly germinative state, indicating that germination is controlled by both DET1 and ABI3. During plastid differentiation in leaves of dark-grown plants, DET1 is required for the action of ABI3 as it is during seed development. Together, the results suggest that ABI3 is at least partly regulated by light.     

Penalized loss functions for Bayesian model comparison

The deviance information criterion (DIC) is widely used for Bayesian model comparison, despite the lack of a clear theoretical foundation. DIC is shown to be an approximation to a penalized loss function based on the deviance, with a penalty derived from a cross-validation argument. This approximation is valid only when the effective number of parameters in the model is much smaller than the number of independent observations. In disease mapping, a typical application of DIC, this assumption does not hold and DIC under-penalizes more complex models. Another deviance-based loss function, derived from the same decision-theoretic framework, is applied to mixture models, which have previously been considered an unsuitable application for DIC.     

Synergy of in vitro Thrombolytic Action of Combinations of Recombinant Staphylokinase and Single-Chain Urokinase-Type Plasminogen Activator

Kinetics of lysis of human plasma clots immersed in plasma were studied in vitro at 37°C under the influence of recombinant staphylokinase, single-chain urokinase-type plasminogen activator (scu-PA), and their simultaneous and consecutive combinations. Staphylokinase and scu-PA caused concentration- and time-dependent lysis of the clots; 32 nM staphylokinase and 75 nM scu-PA separately caused 50% lysis in 4 h. At these equally effective concentrations staphylokinase in 4 h induced a significantly lesser exhaustion of the plasma plasminogen, ₂-antiplasmin, and fibrinogen than scu-PA. Combinations of staphylokinase (<30 nM) and scu-PA (<75 nM) rendered synergic thrombolytic action on the clots. The synergy of thrombolytic action was more pronounced on the simultaneous addition of the two agents than on their consecutive addition, scu-PA 30 min after staphylokinase. In 4 h after the addition, staphylokinase (25 nM) or scu-PA (15 nM) induced 24% and 2% lysis, respectively, whereas the simultaneous and consecutive combination of the same concentrations of these agents induced 58% and 50% lysis, respectively. The simultaneous combination of 15 nM staphylokinase and 15 nM scu-PA resulted in maximal 3.8-fold increase in the thrombolytic effect as compared to the expected total effect of the individual agents. Synergic combinations of the two agents caused lesser exhaustion of plasma plasminogen, ₂-antiplasmin, and fibrinogen as compared with the expected total effect of these agents used separately. Thus, simultaneous and consecutive combinations of staphylokinase and scu-PA in a relatively narrow range of their concentrations possessed synergistic fibrinselective thrombolytic action on the plasma clot in vitro.     

DET1, a negative regulator of light-mediated development and gene expression in arabidopsis,encodes a novel nuclear-localized protein

The mechanisms by which plants integrate light signals to modify endogenous developmental programs are largely unknown. One candidate for a signal transduction component that may integrate light with developmental pathways is the Arabidopsis DET1 gene product. Here we report the positional cloning of the DET1 locus and show that DET1 is a unique nuclear-localized protein. An analysis of a number of det1 mutants indicates that mutants with partial DET1 activity develop as light-grown plants in the dark. det1 null mutants share this phenotype, but also display severe defects in temporal and spatial regulation of gene expression. These results suggest that DET1 acts in the nucleus to control the cell type-specific expression of light-regulated promoters.     

Fabrication and optimal design of differential electromagnetic transducer for implantable middle ear hearing device

A differential electromagnetic transducer (DET), with similar frequency characteristics to those of a normal middle ear, is designed and implemented for use in an implantable middle ear hearing device (IMEHD). To optimize the characteristics of the DET that depend on the electromagnetically forced vibration, a theoretical analysis is conducted to design the vibrating part. The electromagnetic force of the DET is simulated according to the design parameters of the coil size using a finite element analysis (FEA). As a result, the maximal vibration force is achieved when the optimal length and thickness of the cylindrical coil is 70% of the length of the magnets and 56% of their radius. The vibration characteristics of the DET are then simulated when applying the maximal force. The optimally designed DET is implemented using MEMS technology and vibration experiments carried out with the fabricated DET in an unloaded state. The vibrating displacement of the DET is about 200 nm within a range between 0.1 and 1.5 kHz when a current of 1 mA(rms) is applied to the coil. To investigate the usefulness of the DET, in vitro and in vivo experiments are conducted using the ossicular chain of a cadaver and guinea pig, and the results verify that the implemented DET performs well as a transducer for an IMEHD.