Quantitative mechanical evaluation and analysis of Drosophila embryos through the stages of embryogenesis

The fruit fly Drosophila embryo is one of the most important model organisms in genetics and developmental biology research. To better understand the biomechanical properties involved in Drosophila embryo research, this work presents a mechanical characterization of living Drosophila embryos through the stages of embryogenesis. Measurements of the mechanical forces of Drosophila embryos are implemented using a novel, in situ, and minimally invasive force sensing tool with a resolution in the range of microN. The measurements offer an essential understanding of penetration force profiles during the microinjection of Drosophila embryos. Sequentially quantitative evaluation and analysis of the mechanical properties, such as Young's modulus, stiffness, and mechanical impedance of living Drosophila embryos are performed by extracting the force measurements throughout the stages of embryogenesis. Experimental results illustrate the changing mechanical properties of Drosophila embryos during development, and thus mathematical models are proposed. The evaluation provides a critical step toward better understanding of the biomechanical properties of Drosophila embryos during embryogenesis, and could contribute to more efficient and significant genetic and embryonic development research on Drosophila.     

Fabrication and characterization of poly(lactic acid)/acetyl tributyl citrate/carbon black as conductive polymer composites

By using acetyl tributyl citrate (ATBC) as the plasticizer of poly(lactic acid) (PLA) and carbon black (CB) as conductive filler, electrically conductive polymer composites (CPC) with different CB and ATBC contents were prepared. FTIR revealed that the interaction existed between PLA/ATBC matrix and CB filler and ATBC could improve this interaction. The rheology showed that ATBC could obviously decrease the shear viscosity and improve the fluidity of the composites but just the reverse for CB. With the increasing of CB contents, the enforcement effect, storage modulus, and glass-transition temperature increased but the elongation at break decreased. PLA/ATBC/CB composites exhibited the low electrical percolation thresholds of 0.516, 1.20, 2.46, and 2.74 vol % CB at 30, 20, 10, and 0 wt % ATBC. The conductivity of the composite containing 3.98 vol % CB and 30 wt % ATBC reached 1.60 S/cm. Scanning electron microscopy revealed that the addition of ATBC facilitated the dispersion of the CB in the PLA matrix. Water vapor permeability (WVP) showed that, at the same CB contents, the more ATBC contents there were, the less the values of WVP were.     

Conformational behavior of giant DNA through binding with Ag+ and metallization

The conformational behavior of a long single-chain double-stranded DNA in solutions of free silver ions and silver nanoparticles generated via the reduction of AgNO3 by NaBH4 was monitored by fluorescence and electron microscopies and UV spectroscopy. The interaction of monovalent silver ions with DNA induces shrinking of a DNA-coiled polymer chain as a result of a decrease in the DNA persistence length through the complexation of Ag+ with DNA bases. In contrast, the reduction of silver ions by NaBH4 in DNA solutions triggers DNA compaction: a DNA transition from elongated coil state into a compact state. This transition is continuous, unlike the all-or-none discrete DNA compaction that is commonly seen with multications. It is suggested that the collapse of DNA is accompanied by growth aggregation of silver nanoparticles generated on the DNA template.     

Proteomic profiling of nonenzymatically glycated proteins in human plasma and erythrocyte membranes

Nonenzymatic glycation of peptides and proteins by d-glucose has important implications in the pathogenesis of diabetes mellitus, particularly in the development of diabetic complications. In this work, we report the first proteomics-based characterization of nonenzymatically glycated proteins in human plasma and erythrocyte membranes from individuals with normal glucose tolerance, impaired glucose tolerance, and type 2 diabetes mellitus. Phenylboronate affinity chromatography was used to enrich glycated proteins and glycated tryptic peptides from both human plasma and erythrocyte membranes. The enriched peptides were subsequently analyzed by liquid chromatography coupled with electron transfer dissociation-tandem mass spectrometry, resulting in the confident identification of 76 and 31 proteins from human plasma and erythrocyte membranes, respectively. Although most of the glycated proteins could be identified in samples from individuals with normal glucose tolerance, slightly higher numbers of glycated proteins and more glycation sites were identified in samples from individuals with impaired glucose tolerance and type 2 diabetes mellitus.     

SirT1 gain of function increases energy efficiency and prevents diabetes in mice

In yeast, worms, and flies, an extra copy of the gene encoding the Sirtuin Sir2 increases metabolic efficiency, as does administration of polyphenols like resveratrol, thought to act through Sirtuins. But evidence that Sirtuin gain of function results in increased metabolic efficiency in mammals is limited. We generated transgenic mice with moderate overexpression of SirT1, designed to mimic the Sirtuin gain of function that improves metabolism in C. elegans. These mice exhibit normal insulin sensitivity but decreased food intake and locomotor activity, resulting in decreased energy expenditure. However, in various models of insulin resistance and diabetes, SirT1 transgenics display improved glucose tolerance due to decreased hepatic glucose production and increased adiponectin levels, without changes in body weight or composition. We conclude that SirT1 gain of function primes the organism for metabolic adaptation to insulin resistance, increasing hepatic insulin sensitivity and decreasing whole-body energy requirements. These findings have important implications for Sirtuin-based therapies in humans.     

转基因哺乳动物细胞自合成多不饱和脂肪酸

亚油酸、亚麻酸是哺乳动物体内的必需脂肪酸,但哺乳动物由于缺乏△12和ω-3脂肪酸脱氢酶而自身不能合成．△12和ω-3脂肪酸脱氢酶存在于真菌、植物和一些低等动物中．为了实现哺乳动物细胞亚油酸的自身合成,克隆了线虫编码△12脂肪酸脱氢酶的FAT-2基因eDNA序列,通过优化密码子,构建真核表达载体,稳定转染细胞,经抗生素筛选获得稳定整合FAT-2基因的CHO细胞．PCR和RNA印迹（Northern blot）验证了基因的整合和表达．气相色谱分析细胞的脂肪酸含量表明,FAT-2基因的表达显著提高了转基因细胞中亚油酸的含量,亚油酸含量为阴性对照细胞的2．4倍．研究结果表明,低等动物△12脂肪酸脱氢酶可以重建哺乳动物多不饱和脂肪酸合成途径,并利用细胞中的油酸合成亚油酸．上述研究为进一步利用转基因技术促进农业动物合成多不饱和脂肪酸从而提高食品营养价值奠定基础．     

Tetrahydro anthranilic acid as a surrogate for anthranilic acid: application to the discovery of potent niacin receptor agonists

The design, synthesis, and biological activity of a series of cycloalkene acid-based niacin receptor agonists are described. This led to the discovery that tetrahydro anthranilic acid is an excellent surrogate for anthranilic acid. Several compounds were identified that were potent against the niacin receptor, had enhanced cytochrome P450 selectivity against subtypes CYP2C8 and CYP2C9, and improved oral exposure in mice.     

Discovery of piperidine-aryl urea-based stearoyl-CoA desaturase 1 inhibitors

A series of structurally novel stearoyl-CoA desaturase1 (SCD1) inhibitors has been identified via molecular scaffold manipulation. Preliminary structure–activity relationship (SAR) studies led to the discovery of potent, and orally bioavailable piperidine-aryl urea-based SCD1 inhibitors. 4-(2-Chlorophenoxy)-N-[3-(methyl carbamoyl)phenyl]piperidine-1-carboxamide 4c exhibited robust in vivo activity with dose-dependent desaturation index lowering effects.     

Cell-cycle synchronization of fibroblasts derived from transgenic cloned cattle ear skin: effects of serum starvation, roscovitine and contact inhibition

The purpose of the present study was to evaluate the effects of serum-starvation, contact-inhibition and roscovitine treatments on cell-cycle synchronization at the G0/G1 stage of ear skin fibroblasts isolated from transgenic cloned cattle. The developmental competence of re-cloned embryos was also examined. Our results showed that the proportion of G0/G1 cells from the serum-starved group at 3, 4 or 5 days was significantly higher compared with 1 or 2 days only (91.5, 91.7 and 93.5% versus 90.1 and 88.8%, respectively, p < 0.05); whilst there was no statistical difference among cells at 3, 4 or 5 days. For roscovitine-treated cells, the proportion of G0/G1 cells at 2, 3, 4 or 5 days was significantly higher than those treated for 1 day only (91.1, 90.1, 89.4 and 91.3% versus 86.51%, respectively, p < 0.05). The proportion of contact-inhibited G0/G1 cells rose significantly with treatment time, but was similar at 3, 4 and 5 days (89.4, 90.4, 91.4, 91.6 and 92.1%, respectively, p < 0.05). The efficiency of obtaining G0/G1 phase cells was lower when roscovitine treatment was employed to synchronize the cell cycle compared with the serum-starvation and contact-inhibition methods (89.7 versus 91.1% and 91.0%, p < 0.05). Moreover, obvious differences were observed in the rate of fused couplets and blastocysts (89.88 +/- 2.70 versus 87.40 +/- 5.13; 44.10 +/- 8.62 versus 58.38 +/- 13.28, respectively, p < 0.05), when nuclear transfer embryos were reconstructed using donors cells that had been serum starved or contact inhibited for 3 days. Our data indicate that 3 day treatment is feasible for harvesting sufficient G0/G1 cells to produce re-cloned transgenic bovine embryos, regardless of whether serum-starvation, contact-inhibition or roscovitine treatments are used as the synchronization methods.     

The COP9 signalosome: more than a protease

The COP9 signalosome (CSN) is a conserved protein complex that functions in the ubiquitin-proteasome pathway. After two decades of research, we now know that the CSN is a multi-subunit protease that regulates the activity of cullin-RING ligase (CRL) families of ubiquitin E3 complexes. The CSN is rapidly emerging as a key player in the DNA-damage response, cell-cycle control and gene expression. The independent functions of CSN5 (also known as JAB1) add to the complexity of the CSN machinery. Here, we provide an updated view of the structure, functions and regulation of this protein complex.