Saponin 1 Induces Apoptosis and Suppresses NF-κB-Mediated Survival Signaling in Glioblastoma Multiforme (GBM)

Saponin 1 is a triterpeniod saponin extracted from Anemone taipaiensis, a traditional Chinese medicine against rheumatism and phlebitis. It has also been shown to exhibit significant anti-tumor activity against human leukemia (HL-60 cells) and human hepatocellular carcinoma (Hep-G2 cells). Herein we investigated the effect of saponin 1 in human glioblastoma multiforme (GBM) U251MG and U87MG cells. Saponin 1 induced significant growth inhibition in both glioblastoma cell lines, with a 50% inhibitory concentration at 24 h of 7.4 µg/ml in U251MG cells and 8.6 µg/ml in U87MG cells, respectively. Nuclear fluorescent staining and electron microscopy showed that saponin 1 caused characteristic apoptotic morphological changes in the GBM cell lines. Saponin 1-induced apoptosis was also verified by DNA ladder electrophoresis and flow cytometry. Additionally, immunocytochemistry and western blotting analyses revealed a time-dependent decrease in the expression and nuclear location of NF-κB following saponin 1 treatment. Western blotting data indicated a significant decreased expression of inhibitors of apoptosis (IAP) family members,(e.g., survivin and XIAP) by saponin 1. Moreover, saponin 1 caused a decrease in the Bcl-2/Bax ratio and initiated apoptosis by activating caspase-9 and caspase-3 in the GBM cell lines. These findings indicate that saponin 1 inhibits cell growth of GBM cells at least partially by inducing apoptosis and inhibiting survival signaling mediated by NF-κB. In addition, in vivo study also demonstrated an obvious inhibition of saponin 1 treatment on the tumor growth of U251MG and U87MG cells-produced xenograft tumors in nude mice. Given the minimal toxicities of saponin 1 in non-neoplastic astrocytes, our results suggest that saponin 1 exhibits significant in vitro and in vivo anti-tumor efficacy and merits further investigation as a potential therapeutic agent for GBM.     

Automated and Accurate Detection of Soma Location and Surface Morphology in Large-Scale 3D Neuron Images

Automated and accurate localization and morphometry of somas in 3D neuron images is essential for quantitative studies of neural networks in the brain. However, previous methods are limited in obtaining the location and surface morphology of somas with variable size and uneven staining in large-scale 3D neuron images. In this work, we proposed a method for automated soma locating in large-scale 3D neuron images that contain relatively sparse soma distributions. This method involves three steps: (i) deblocking the image with overlap between adjacent sub-stacks; (ii) locating the somas in each small sub-stack using multi-scale morphological close and adaptive thresholds; and (iii) fusion of the repeatedly located somas in all sub-stacks. We also describe a new method for the accurate detection of the surface morphology of somas containing hollowness; this was achieved by improving the classical Rayburst Sampling with a new gradient-based criteria. Three 3D neuron image stacks of different sizes were used to quantitatively validate our methods. For the soma localization algorithm, the average recall and precision were greater than 93% and 96%, respectively. For the soma surface detection algorithm, the overlap of the volumes created by automatic detection of soma surfaces and manually segmenting soma volumes was more than 84% for 89% of all correctly detected somas. Our method for locating somas can reveal the soma distributions in large-scale neural networks more efficiently. The method for soma surface detection will serve as a valuable tool for systematic studies of neuron types based on neuron structure.     

Binding of chara Myosin globular tail domain to phospholipid vesicles

Binding of Chara myosin globular tail domain to phospholipid vesicles was investigated quantitatively. It was found that the globular tail domain binds to vesicles made from acidic phospholipids but not to those made from neutral phospholipids. This binding was weakened at high KCl concentration, suggesting that the binding is electrostatic by nature. The dissociation constant for the binding of the globular tail domain to 20% phosphatidylserine vesicles (similar to endoplasmic reticulum in acidic phospholipid contents) at 150 mM KCl was 273 nM. The free energy change due to this binding calculated from the dissociation constant was -37.3 kJ mol(-1). Thus the bond between the globular tail domain and membrane phospholipids would not be broken when the motor domain of Chara myosin moves along the actin filament using the energy of ATP hydrolysis (DeltaG degrees ' = -30.5 kJ mol(-1)). Our results suggested that direct binding of Chara myosin to the endoplasmic reticulum membrane through the globular tail domain could work satisfactorily in Chara cytoplasmic streaming. We also suggest a possible regulatory mechanism of cytoplasmic streaming including phosphorylation-dependent dissociation of the globular tail domain from the endoplasmic reticulum membrane.     

西鄂尔多斯4种荒漠植物光合作用特征与差异性

植物对光能的高效吸收、传递和转换机理是光合作用的核心。为了厘清西鄂尔多斯地区4种荒漠植物光合生理生态适应性和生境适宜性,运用LI-6800光合作用测定系统对四合木(Tetraena mongolica)、霸王(Sarcozygium xanthoxylon)、沙冬青(Ammopiptanthus mongolicus)和白刺(Nitraria tangutorum)进行光合作用日变化进行了测定。研究结果表明:(1) 4种荒漠植物的光合作用日变化的净光合速率(Pn)、蒸腾速率(Tr)、气孔导度(Gs)和胞间CO_2浓度(Ci)均表现出明显的分异;(2)净光合速率均呈"双峰"曲线的变化趋势,4种荒漠植物的净光合速率(Pn)依次排序为四合木(4.37μmol m~(-2) s~(-1))霸王(3.58μmol m~(-2) s~(-1))沙冬青(2.63μmol m~(-2) s~(-1))和白刺(2.26μmol m~(-2) s~(-1))。说明四合木和近缘种霸王比其他二种荒漠植物具有较强的光合生理生态适应性与生境适宜性;(3)从影响光合作用的有关生理生态因子来看,净光合速率与气孔导度与蒸腾速率、水分利用效率呈现明显的正相关性,与微气象因子大气相对湿度(RH)的相关性不明显;(4)水分因子是限制4种荒漠植物生长的重要因素之一。该研究以期为我国西北荒漠区珍稀濒危植物的保护提供案例借鉴与理论依据。     

Acetaminophen-induced antinociception via central 5-HT(2A) receptors.

Acetaminophen is one of the most widely used analgesic drugs. Although the mechanism of analgesic action of acetaminophen is still not known, the involvement of the central serotonin (5-hydroxytryptamine: 5-HT) system is one possibility. In the present study, we examined the antinociceptive effect of acute and chronic intraperitoneally (i.p.) administered acetaminophen by tail flick latency measurements in the rat. A significantly increased tail flick latency was observed in acute and 15-day acetaminophen-treated rats, but not in 30-day acetaminophen-treated rats, at a dose of 400 mg/kg/day. To investigate the plasticity of receptors at postsynaptic membrane, we conducted a series of experiments by radioligand binding method on frontal cortex and brainstem membrane. The technique involved radioligand binding with [phenyl-4-³H]spiperone and ketanserin for studying 5-HT_2A receptor characteristics. A significant decrease in the maximum number of 5-HT_2A binding sites (B_max) was demonstrated in all treatment groups with acetaminophen 300 and 400 mg/kg on frontal cortex membrane, whereas the value of the dissociation equilibrium constant (K_d) remained unchanged. The down-regulation of 5-HT_2A binding sites in frontal cortex was of a lesser magnitude after 30 days of treatment and the tail flick latency was as in the control animals. These results suggest that down-regulation of 5-HT_2A receptor in response to 5-HT release is a major step in the mechanism underlying analgesia produced by this agent. On the contrary, chronic use of acetaminophen may result in 5-HT depletion, which in turn produces re-adaptation of postsynaptic 5-HT_2A receptors. These data provide further evidence for a central 5-HT-dependent antinociceptive effect of acetaminophen.     

One-electron oxidation-reduction properties of hepatic NADH-cytochrome b5 reductase

The one-electron oxidation-reduction properties of flavin in hepatic NADH-cytochrome b5 reductase were investigated by optical absorption spectroscopy, electron paramagnetic resonance (EPR), and potentiometric titration. An intermediate with a peak at 375 nm previously described by Iyanagi (1977) [ Iyanagi , T. (1977) Biochemistry 16, 2725-2730] was confirmed to be a red anionic semiquinone. The NAD+-bound reduced enzyme was oxidized by cytochrome b5 via the semiquinone intermediate. This indicates that electron transfer from flavin to cytochrome b5 proceeds in two successive one-electron steps. Autoxidation of the NAD+-bound reduced enzyme was slower than that of the NAD+-free reduced enzyme and was accompanied by the appearance of an EPR signal. Midpoint redox potentials of the consecutive one-electron-transfer steps in the presence of excess NAD+ were Em,1 = -88 mV and Em,2 = 147 mV at pH 7.0. This corresponds to a semiquinone formation constant of 8. The values of Em,1 and Em,2 were also studied as a function of pH. A mechanism for electron transfer from NADH to cytochrome b5 is discussed on the basis of the one-electron redox potentials of the enzyme and is compared with the electron-transfer mechanism of NADPH-cytochrome P-450 reductase.     

Regulatory functions of allosteric enzymes in far-from-equilibrium systems

A theoretical study is made on catalytic activities of allosteric enzymes in non-equilibrium systems. It is demonstrated that the amount of chemical flow catalyzed by allosteric enzymes in systems maintained far-from-equilibrium can be qualitatively different from that familiar in the near-equilibrium situations. To be more specific, a study is made of a system containing two chemical species, S and P, and an allosteric enzyme, E, which catalyzes the reaction of the interconversion between them. This system interacts with its environment in a way characterized by a set of controlled parameters. By this interaction the system is maintained far-from-equilibrium. More than one steady state is possible for a certain range of controlled parameters. For continuous changes of the controlled parameters, discontinuous transitions between the multiple steady states can occur. This new aspect of the enzyme kinetics of allosteric proteins may play a role in the regulation of metabolic flows within living cells.     

Paleoecologic and stratigraphic significance of the freshwater algae <Emphasis Type="Italic">Pediastrum</Emphasis> in the Upper Cretaceous (Turonian) marine deposits,north Western Desert,Egypt

Palynological investigations of samples collected from the Abu Roash Formation, Faghur Hj5-1 well, north Western Desert, Egypt show a low diversity in palynomorph assemblage. This assemblage is mainly dominated by a clear proliferation of Pediastrum (and other allied algal forms, e.g., Scenedesmus and Botryococcus) which today lives exclusively in freshwater. Such a prominent record within the current marine deposits could be considered a good biostratigraphic datum in the Upper Cretaceous (Turonian) period rather than an ecologic event in the north Western Desert, Egypt. The influence of freshwater input on the studied deposits is proven by the presence of heavy minerals including kyanite, zircon, staurolite, and amphiboles encountered in siliciclastic sediments. Most of these minerals are sub-rounded to rounded so they were derived probably from sedimentary rocks. In the studied succession, the presence of Pediastrum reflects sea level fall (i.e., lowstand systems tracts). The occurrence of Pediastrum and other algae in such marine deposits reflects the predominant deposition of fluviatile sediments related to the discharge of rivers into shelf seas.     

Down-regulation of IL-8 expression in human airway epithelial cells through helper-dependent adenoviral-mediated RNA interference

Interleukin (IL)-8 is a potent neutrophil chemotactic factor and a crucial mediator in neutrophil-dependent inflammation.Various cell types produce IL-8, either in response to external stimuli such as cytokines or bacterial infection, or after malignant transformation. Anti-IL-8 strategies have been considered for anti-inflammatory therapy. In this paper we demonstrate that the RNA interference technique can be used to efficiently down-regulate IL-8 protein expression in airway epithelial cells. We used a helper-dependent adenoviral vector to express a small hairpin (sh)RNA targeting human IL-8 in cultured airway epithelial cells (IB3-1, Cftr; C38, Cftr-corrected) stimulated with TNF-α, IL-1β or heat-inactivated Burkholderia cenocepacia. Stimulated IL-8 expression in IB3-1 and C38 cells was significantly reduced by shRNA expression. The shRNA targeting IL-8 had no effect on the activation of NF-κB, or on the protein levels of IκB or IL-6, suggesting that this anti-IL-8 strategy was highly specific, and therefore may offer potential for the treatment of inflammatory diseases.     

Induction of drug-metabolizing enzymes and transporters in human bronchial epithelial cells by beclomethasone dipropionate

Inhaled steroids are the most potent anti-inflammatory therapy commonly used in bronchial asthma. There are, however, a small number of asthmatic patients who do not respond to inhaled steroid-treatment. The stimulation of metabolism and excretion of inhaled drugs at bronchial tissues might lead to a decrease in the effect of the drugs, although the molecular mechanism of this resistance is unclear. In this study, we found that beclomethasone dipropionate (BDP) stimulated the expression of mRNAs for uridine 5'-diphosphate glucuronosyl transferase 2B4 and 2B11, and transporters such as multidrug resistance P-glycoprotein, multidrug resistance-associated protein 1 and 2 in cultured bronchial epithelial cells. It is possible that the individual differences of expression of drug metabolizing enzymes and transporters and their enhancement with BDP are implicated in the individual differences of reactivity over steroid medical treatment.