Reactive oxygen species and nitric oxide regulate mitochondria-dependent apoptosis and autophagy in evodiamine-treated human cervix carcinoma HeLa cells

The redox environment of the cell is currently thought to be extremely important to control either apoptosis or autophagy. This study reported that reactive oxygen species (ROS) and nitric oxide (NO) generations were induced by evodiamine time-dependently; while they acted in synergy to trigger mitochondria-dependent apoptosis by induction of mitochondrial membrane permeabilization (MMP) through increasing the Bax/Bcl-2 or Bcl-x_L ratio. Autophagy was also stimulated by evodiamine, as demonstrated by the positive autophagosome-specific dye monodansylcadaverine (MDC) staining as well as the expressions of autophagy-related proteins, Beclin 1 and LC3. Pre-treatment with 3-MA, the specific inhibitor for autophagy, dose-dependently decreased cell viability, indicating a survival function of autophagy. Importantly, autophagy was found to be promoted or inhibited by ROS/NO in response to the severity of oxidative stress. These findings could help shed light on the complex regulation of intracellular redox status on the balance of autophagy and apoptosis in anti-cancer therapies.     

Introduction: International conference on forest ecosystems: Ecology,conservation and sustainable management

Abstract

A pollen diagram from a small lake, Kutulahdenlampi, in northern Finland is interpreted in terms of the development of forest vegetation during the Holocene. The abundance of each of the forest taxa is considered independently by means of pollen accumulation rates (PARs), using as the reference material, long term average pollen deposition values monitored by a network of pollen traps. Particular attention is paid to the arrival of spruce and to the species in the original forests that this newcomer replaces. A model of pollen dispersal and deposition developed by Sugita is used to estimate the area around the lake that the pollen assemblage is most clearly reflecting. This relevant source area of pollen (RSAP), for the present day situations is c. 1,500 m. Pollen loadings calculated for a simulated landscape that mimics (i) that of the present day and (ii) for the situation at 8,000 BP (as deduced from the PARs) are compared with the pollen assemblages from the diagram at those points in time, and are seen to be compatible. The advantages of combining PAR and modelling to look at the spatial scale of vegetation reconstructions are discussed.     

Cytoprotective Effects of Nicotinamide Derivatives in Endothelial Cells

Following discovery of NAD⁺-dependent reactions that control gene expression, cytoprotection, and longevity, there has been a renewed therapeutic interest in precursors, such as nicotinamide and its derivatives. We tested 20 analogues of nicotinamide for their ability to protect endothelial cells from peroxynitrite stress and their effect on poly (ADP-ribose) polymerase (PARP) activity. Several nicotinamide derivatives protected endothelial cells from peroxynitrite-induced depletion of cellular NAD⁺ and ATP concentrations, but only some of these compounds inhibited PARP. We conclude that some nicotinamide derivatives provide protection of endothelial cells against peroxynitrite-induced injury independent of inhibition of PARP activity. Preservation of the NAD⁺ pool was a common effect of these compounds.     

Interaction of Tea Catechins with Lipid Bilayers Investigated by a Quartz-Crystal Microbalance Analysis

The quartz-crystal microbalance (QCM) technique was applied to investigate the interaction of tea catechins with lipid bilayers. The association constants obtained from the frequency changes of QCM revealed that (?)epicatechin gallate and (?)epigallocatechin gallate interacted with 1,2-dimyristoyl-sn-glycero-3-phosphocholine ca. 1000 times more strongly than (?)epicatechin and (?)epigallocatechin. The results exhibited good correlation with the strength of biological activity.     

Evaluation of 3-(4′-(4″-fluorophenyl)-6′-phenylpyrimidin-2′-yl)-2-phenylthiazolidin-4-one for in vivo modulation of biomarkers of chemoprevention in the 7,12-dimethylbenz[a]anthracene-induced hamster buccal pouch carcinogenesis

A new bis heterocycle comprising both bioactive 2-aminopyrimidine and thiazolidin-4-one nuclei namely 3-(4′-(4″-fluorophenyl)-6′-phenylpyrimidin-2′-yl)-2-phenylthiazolidin-4-one 3 was synthesized, characterized with the help of melting point, elemental analysis, FT-IR, MS, one-dimensional NMR (¹H, ¹³C) spectra and we evaluated the chemopreventive potential of 3-(4′-(4″-fluorophenyl)-6′-phenylpyrimidin-2′-yl)-2-phenylthiazolidin-4-one based on in vivo inhibitory effects on 7,12-dimethylbenz[a]anthracene (DMBA)-induced hamster buccal pouch carcinogenesis. Administration of 3 effectively suppressed oral carcinogenesis initiated with DMBA as revealed by the reduced incidence of neoplasms. Lipid peroxidation, glutathione (GSH) content, and the activities of glutathione peroxidase (GPx), glutathione S-transferase (GST) were used to biomonitor the chemopreventive potential of 3. Lipid peroxidation was found to be significantly decreased, whereas GSH, GPx, GST, and GGT were elevated in the oral mucosa of tumor-bearing animals. Our data suggest that 3 may exert its chemopreventive effects in the oral mucosa by modulation of lipid peroxidation and enhancing the levels of GSH, GPx, and GST.     

Expression,surface immobilization,and characterization of functional recombinant cannabinoid receptor CB2

Human peripheral cannabinoid receptor CB₂, a G protein-coupled receptor (GPCR) involved in regulation of immune response has become an important target for pharmaceutical drug development. Structural and functional studies on CB₂ may benefit from immobilization of the purified and functional receptor onto a suitable surface at a controlled density and, preferably in a uniform orientation. The goal of this project was to develop a generic strategy for preparation of functional recombinant CB₂ and immobilization at solid interfaces. Expression of CB₂ as a fusion with Rho-tag (peptide composed of the last nine amino acids of rhodopsin) in E. coli was evaluated in terms of protein levels, accessibility of the tag, and activity of the receptor. The structural integrity of CB₂ was tested by ligand binding to the receptor solubilized in detergent micelles, captured on tag-specific monoclonal 1D4 antibody-coated resin. Highly pure and functional CB₂ was obtained by sequential chromatography on a 1D4- and Ni-NTA-resin and its affinity to the 1D4 antibody characterized by surface plasmon resonance (SPR). Either the purified receptor or fusion CB₂ from the crude cell extract was captured onto a 1D4-coated CM4 chip (Biacore) in a quantitative fashion at uniform orientation as demonstrated by the SPR signal. Furthermore, the accessibility of the extracellular surface of immobilized CB₂ and the affinity of interaction with a novel monoclonal antibody NAA-1 was studied by SPR. In summary, we present an integral strategy for purification, surface immobilization, ligand- and antibody binding studies of functional cannabinoid receptor CB₂.     

Enantiopurity Determination of the Enantiomers of the Triple Reuptake Inhibitor Indatraline

The present study describes the development of two approaches for the determination of the enantiopurity of both enantiomers of indatraline. Initially, a method was developed using different chiral solvating agents (CSAs) for diastereomeric discrimination regarding signal separation in ¹H nuclear magnetic resonance (NMR) spectroscopy, revealing MTPA as a promising choice for the differentiation of the indatraline enantiomers. This CSA was also tested for its ideal molar ratio, temperature, and solvent. Optimized conditions could be achieved that made determination of enantiopurity for (1R,3S)‐indatraline up to 98.9% enantiomeric excess (ee) possible. To quantify even higher enantiopurities, a high‐performance liquid chromatography (HPLC) method based on a modified β‐cyclodextrine phase was established. The influence of buffer type, concentration, pH value, percentage and kind of organic modifier, temperature, injection volume as well as sample solvent on chromatographic parameters was investigated. Afterwards, the reliability of the established HPLC method was demonstrated by validation according to the ICH guideline Q2(R1) regarding specificity, accuracy, precision, linearity, and quantitation limit. The developed method proved to be strictly linear within a concentration range of 1.25–1000 μM for the (1R,3S)‐enantiomer and 1.25‐750 μM for its mirror image that enables a reliable determination of enantiopurities up to 99.75% ee for the (1R,3S)‐enantiomer and up to 99.67% ee for the (1S,3R)‐enantiomer. Chirality 25:923–933, 2013. © 2013 Wiley Periodicals, Inc.     

Poly(ADP-ribose) signaling in cell death

Poly(ADP-ribosyl)ation (PARylation) is a reversible protein modification carried out by the concerted actions of poly(ADP-ribose) polymerase (PARP) enzymes and poly(ADP-ribose) (PAR) decomposing enzymes such as PAR glycohydrolase (PARG) and ADP-ribosyl hydrolase 3 (ARH3). Reversible PARylation is a pleiotropic regulator of various cellular functions but uncontrolled PARP activation may also lead to cell death. The cellular demise pathway mediated by PARylation in oxidatively stressed cells has been described almost thirty years ago. However, the underlying molecular mechanisms have only begun to emerge relatively recently. PARylation has been implicated in necroptosis, autophagic cell death but its role in extrinsic and intrinsic apoptosis appears to be less predominant and depends largely on the cellular model used. Currently, three major pathways have been made responsible for PARP-mediated necroptotic cell death: (1) compromised cellular energetics mainly due to depletion of NAD, the substrate of PARPs; (2) PAR mediated translocation of apoptosis inducing factor (AIF) from mitochondria to nucleus (parthanatos) and (3) a mostly elusive crosstalk between PARylation and cell death/survival kinases and phosphatases. Here we review how these PARP-mediated necroptotic pathways are intertwined, how PARylation may contribute to extrinsic and intrinsic apoptosis and discuss recent developments on the role of PARylation in autophagy and autophagic cell death.     

暗腹雪鸡与海兰褐鸡血液生理生化指标的比较

为了解暗腹雪鸡（Tetraogallus himalayensis）正常血液生理生化特性,本文对暗腹雪鸡21项血液生理生化指标进行了测定,并与海兰褐鸡（Gallus gallus domesticus var. Hyline）进行了比较。测定结果,暗腹雪鸡的红细胞纵径及横纵径乘积、平均红细胞容积及平均红细胞血红蛋白含量极显著大于海兰褐鸡（P<0.01）;暗腹雪鸡天冬氨酸氨基转移酶和胆固醇含量均极显著高于海兰褐鸡（P<0.01）。其他各项血液生理生化指标两禽种间之间差异不显著。实验结果表明,暗腹雪鸡血液生理生化指标与家养蛋鸡之间有一定差异,暗腹雪鸡对高原低氧环境有较强的适应性可能与其红细胞体积较大、平均红细胞容积及平均红细胞血红蛋白含量较高有一定关系。     

Intracellular Distribution and Some Properties of β-Glucosidases of a Cellulolytic Pseudomonad

Two distinct forms of β-glucosidase, A and B, were found to occur in the cells of Pseudomonas fluorescens var. cellulosa : A was membrane-bound, while B cytosolic. They differed also from each other in some properties, such as molecular size, kinetic parameters, and susceptibility to various compounds. β-Glucosidase B was partially purified and studied especially of its substrate specificity. The results indicated that it may be an atypical β-glucosidase which possesses a certain character of exo-cellulase.