DNA molecule manipulation by motor proteins for analysis at the single-molecule level

Massively parallel and individual DNA manipulation for analysis has been demonstrated by designing a fully self-assembled molecular system using motor proteins. DNA molecules were immobilized by trapping in a polyacrylamide gel replica, and were digested by a restriction enzyme, XhoI, for DNA analysis. One end of the λDNA was modified with biotin and the other end was modified with digoxin molecules by fragment labeling and ligation methods. The digoxin-functionalized end was immobilized on a glass surface coated with anti-digoxigenin antibody. The biotinylated end was freely suspended and experienced Brownian motion in a buffer solution. The free end was attached to a biotinylated microtubule via avidin–biotin biding and the DNA was stretched by a kinesin-based gliding assay. A stretched DNA molecule was fixed between the gel and coverslip to observe the cleavage of the DNA by the enzyme, which was supplied through the gel network structure. This simple process flow from DNA manipulation to analysis offers a new method of performing molecular surgery at the single-molecule scale. Figure DNA molecule manipulation by motor proteins for analysis at the single-molecule level     

1H and 13C NMR assignments for four triterpenoid saponins from Albizziae cortex

Four triterpenoid saponins were isolated from Albizziae cortex, and a complete assignment of their (1)H and (13)C NMR spectra was carried out using 1D and 2D NMR ((1)H-(1)H COSY, HSQC, HMBC, and HSQC-TOCSY) methods. Their (1)H NMR assignments were reported for the first time and some of their (13)C NMR spectral data reported in literature were corrected.     

A new quantitative analysis of significant timing differences between externally cued and self-initiated motor tasks in an fMRI study

It is generally accepted that the temporal resolution of blood oxygenation level dependent functional MRI is limited due to the inherent latency and longevity of the haemodynamic response. However, in this study we introduce a technique for measurement of timing differences from within the same brain region in two (or more) separate tasks that allows accurate determination of cortical timing differences 200 ms. Our technique, based on a novel use of linear regression analysis, is shown to yield accurate results both in simulated and experimental data. We show that cortical timing differences measured using fMRI are consistent with published electrophysiological results. Measurement of timing differences using this technique could prove a useful strategy for identifying neural network components in a wide range of cognitive paradigms.     

Metabolomics comparison of chemical components and metabolic regulations in different parts of Eucommia ulmoides Oliv

Eucommia ulmoides Oliv. (EU) is an herb with variety of pharmacological activities. As the demand for EU as a medical source increases, its limited availability is becoming an urgent problem. The bark of EU—Eucommia cortex (EC)—was the main medicinal part of EU, whose yield has been too low to meet market requirements. The EU leaves—Eucommiae folium (EF)—obtained easily from the trees, traditionally considered less valuable than the EU bark. This study compared chemical components and metabolic regulations in different parts of EU using metabolomics to explore the potential for alternative medical sources of EU. The results demonstrated that the composition of EC differs slightly from the composition of EF, and there are some differences in the content of individual ingredients between EC and EF. Combined pharmacodynamic/metabolomics analysis showed that both EC and EF could improve the bone metabolism markers and trabecular microarchitecture of ovariectomized rats mainly by intervening in amino acid metabolism. Our data suggest that EF is a potential alternative medicinal option when EU resources are scarce, and this study provides a mechanistic framework for the benefits of EF in osteoporosis disease.     

A serial functional connectivity MRI study in healthy individuals assessing the variability of connectivity measures: reduced interhemispheric connectivity in the motor network during continuous performance

To date, little data is available on the reproducibility of functional connectivity MRI (fcMRI) studies. Here, we tested the variability and reproducibility of both the functional connectivity itself and different statistical methods to analyze this phenomenon. In the main part of our study, we repeatedly examined two healthy subjects in 10 sessions over 6 months with fcMRI. Cortical areas involved in motor function were examined under two different cognitive states: during continuous performance (CP) of a flexion/extension task of the fingers of the right hand and while subjects were at rest. Connectivity to left primary motor cortex (lSM1) was calculated by correlation analysis. The resulting correlation coefficients were transformed to z-scores of the standard normal distribution. For each subject, multisession statistical analyses were carried out with the z-score maps of the resting state (RS) and the CP experiments. First, voxel based t tests between the two groups of fcMRI experiments were performed. Second, ROI analyses were carried out for contralateral right SM1 and for supplementary motor area (SMA). For both ROI, mean and maximum z-score were calculated for each experiment. Also, the fraction of significantly (P<.05) correlated voxels (FCV) in each ROI was calculated. To evaluate the differences between the RS and the CP condition, paired t tests were performed for the mean and maximum z-scores, and Wilcoxon signed ranks tests for matched pairs were carried out for the FCV. All statistical methods and connectivity measures under investigation yielded a distinct loss in left–right SM1 connectivity under the CP condition. For SMA, interindividual differences were apparent. We therefore repeated the fcMRI experiments and the ROI analyses in a group of seven healthy subjects (including the two subjects of the main study). In this substudy, we were able to verify the reduction of left–right SM1 connectivity during unilateral performance. Still, the direction of SMA to lSM1 connectivity change during the CP condition remained undefined as four subjects showed a connectivity increase and three showed a decrease. In summary, we were able to demonstrate a distinct reduction in left–right SM1 synchrony in the CP condition compared to the RS both in the longitudinal and in the multisubject study. This effect was reproducible with all statistical methods and all measures of connectivity under investigation. We conclude that despite intra- and interindividual variability, serial and cross-sectional assessment of functional connectivity reveals stable and reliable results.     

Origins of intersubject variability of blood oxygenation level dependent and arterial spin labeling fMRI: implications for quantification of brain activity

Accurate localization of brain activity using blood oxygenation level-dependent (BOLD) functional magnetic resonance imaging (fMRI) has been challenged because of the large BOLD signal within distal veins. Arterial spin labeling (ASL) techniques offer greater sensitivity to the microvasculature but possess low temporal resolution and limited brain coverage. In this study, we show that the physiological origins of BOLD and ASL depend on whether percent change or statistical significance is being considered. For BOLD and ASL fMRI data collected during a simple unilateral hand movement task, we found that in the area of the contralateral motor cortex the centre of gravity (CoG) of the intersubject coefficient of variation (CV) of BOLD fMRI was near the brain surface for percent change in signal, whereas the CoG of the intersubject CV for Z-score was in close proximity of sites of brain activity for both BOLD and ASL. These findings suggest that intersubject variability of BOLD percent change is vascular in origin, whereas the origin of inter-subject variability of Z-score is neuronal for both BOLD and ASL. For longer duration tasks (12 s or greater), however, there was a significant correlation between BOLD and ASL percent change, which was not evident for short duration tasks (6 s). These findings suggest that analyses directly comparing percent change in BOLD signal between pre-defined regions of interest using short duration stimuli, as for example in event-related designs, may be heavily weighted by large-vessel responses rather than neuronal responses.     

分光光度法测定车用甲醇汽油中的甲醇

建立了可见分光光度法测定甲醇汽油中甲醇含量的方法.实验结果表明,当甲醇含量在0-2.0mL/L时,吸光度与浓度能很好地遵循朗伯-比耳定律,相关系数为0.9934,该方法的平均加标回收率为96.9％,精密度实验M15和M20甲醇汽油的变异系数CV分别为1.1％和1.0％.     

In vivo temporal EPR study using a region-selected intensity determination method to estimate cerebral reducing ability in rats treated with olanzapine

Region-selected intensity determination (RSID) is a method for obtaining the temporal changes in electron paramagnetic resonance (EPR) signal intensity from a target region, without the use of complicated procedures employed in the conventional imaging methods. An in vivo 700-MHz radio frequency EPR spectrometer equipped with a bridged loop-gap resonator was used with the RSID method to estimate intracerebral reducing ability in the rat following acute administration of olanzapine (OZP) or haloperidol (HPD). To this end, temporal changes in EPR signal intensity of target regions (the striatum and the prefrontal cortex) of rats which had received a blood-brain-barrier-permeable nitroxide radical (3-hydroxymethyl-2,2,5,5-tetramethylpyrrolidine-1-oxyl) via an intravenous route were observed. The half-lives of EPR signal intensity in both regions of OZP- or HPD-treated rats were significantly longer than in control animals. This indicated that reducing abilities of the striatum and cerebral cortex decreased in the rats to which either OZP or HPD had been acutely administered.     

猴子大脑皮层组织的拉曼光谱研究

猴子的大脑皮层主要由灰质和神经元组成,是大脑中调控行动和思维的区域。选取在生命成长过程中具有代表性的3个年龄段(幼年、成年、老年)猴子的大脑皮层组织,首次测得其拉曼光谱。实验结果显示,3个不同年龄段的猴子大脑皮层的拉曼光谱最明显的差异表现在1 000～1 400 cm~(-1)和2 800～3 000cm~(-1)范围内。随着年龄的增长,归属于脂类的拉曼峰1 313和2 885 cm~(-1)的强度越来越强,而归属于蛋白的拉曼峰1 338和2 932 cm~(-1)的强度却逐渐减弱,并且在老年猴子大脑皮层的拉曼谱中出现了2个归属于脂类的拉曼峰1 296和2 850 cm,I_(1313)/I_(1338)和I_(2885)/I_(2932)的变化趋势很好地描述了这种差异。研究表明,随着年龄的不断增长,猴子大脑皮层中脂类的含量逐渐增加,这样可以很好地说明大脑皮层中脂类的聚积是导致衰老的一个重要原因,为进一步研究人类和动物的生命活动规律、疾病的形成机理提供了很有价值的参考。     

利用基于~1H NMR的代谢组学分析研究重离子辐射对大鼠额叶皮质区的影响

太空辐射尤其是重离子辐射可造成DNA的破坏、细胞死亡、以及一些癌症的发生,是人类深空探索进程中急需克服的难题. 本文通过重离子加速器产生¹²C⁶⁺重离子束对大鼠头部进行一定剂量的辐射,模拟空间重离子辐射对中枢神经系统(CNS)的生物学效应. 采用基于¹H NMR的代谢组学方法对辐射大鼠大脑额叶皮质区进行了测定分析,结合数据的统计分析和检验,发现了包括一些重要CNS神经递质在内的代谢物含量发生明显变化. 这些代谢物主要为：牛磺酸、乳酸、谷氨酸、4-氨基丁酸、以及磷酸胆碱等. 结合差异蛋白质组结果分析,包括4-氨基丁酸、谷氨酸、乳酸、牛磺酸等在内的代谢物参与的主要生物途径,如神经递质的合成途径,以及神经递质受体介导的信号途径可能受重离子辐射的负面影响. 这些发现将为进一步阐明重离子辐射效应的分子机制提供有利信息,从而为从生物学途径探寻有效重离子辐射防护措施提供依据.