A long-wavelength fluorescent substrate for continuous fluorometric determination of cellulase activity: resorufin-beta-D-cellobioside

A simple and reliable continuous assay procedure for measurement of cellulase activity from several species using the new substrate resorufin-beta-D-cellobioside (Res-CB) has been developed. The product of enzyme reaction, resorufin, exhibits fluorescence emission at 585 nm with excitation at 571 nm and has a pK(a) of 5.8, which allows continuous measurement of fluorescence turnover at or near physiological pH values. The assay performed using purified cellulase from the microscopic fungus Trichoderma reesei has been shown to give the kinetic parameters K(m) of 112 microM and V(max) of 0.000673 micromol/mL/min. Methods for performing the assay using cellulases isolated from both live Arabidopsis thaliana plant and Aspergillus niger fungal species are presented.     

针刺麻醉复合小剂量瑞芬太尼用于甲状腺良性结节消融术的临床疗效观察

目的:探讨小剂量瑞芬太尼复合针刺麻醉在甲状腺良性结节消融术中的麻醉镇痛效果。方法:选取2015年6月至2016年6月在我院进行甲状腺良性结节射频消融术的患者71例,并将其随机分为对照组(n=35)和复合麻醉组(n=36)。对照组患者接受单纯芬太尼麻醉,复合麻醉组患者接受小剂量瑞芬太尼复合针刺麻醉,观察和比较两组患者麻醉前(T0)、麻醉后(T1)、消融术中(T2)、手术结束(T3)时患者的平均动脉压(MAP)、心率(HR)、血氧饱和度(HPO_2)和VAS评分。结果:复合麻醉组患者T2和T3时间点MAP(95.00±6.09,90.86±3.23)(P0.05)、HR值(65.19±3.52,75.03±6.00)均显著低于对照组(P0.05),HPO_2水平(98.78±1.15,97.81±1.47)均显著高于对照组(P0.05),VAS评分(3.25±1.38,1.69±1.43)均显著低于对照组(P0.05)。结论:电针刺激结合小剂量瑞芬太尼复合麻醉用于甲状腺良性结节消融术可有效镇痛并维持较稳定的血压和心率,效果明显优于单纯芬太尼麻醉。     

Dosimetric effect of intrafraction motion and different localization strategies in prostate SBRT

The aim of this study was to evaluate the dosimetric effect of continuous motion monitoring based localization (Calypso, Varian Medical Systems), gating and intrafraction motion correction in prostate SBRT. Delivered doses were modelled by reconstructing motion inclusive dose distributions for different localization strategies. Actually delivered dose (strategy A) utilized initial Calypso localization, CBCT and additional pre-treatment motion correction by kV-imaging and Calypso, and gating during the irradiation. The effect of gating was investigated by simulating non-gated treatments (strategy B). Additionally, non-gated and single image-guided (CBCT) localization was simulated (strategy C). A total of 308 fractions from 22 patients were reconstructed. The dosimetric effect was evaluated by comparing motion inclusive target and risk organ dose-volume parameters to planned values. Motion induced dose deficits were seen mainly in PTV and CTV to PTV margin regions, whereas CTV dose deficits were small in all strategies: mean ± SD difference in CTVD99% was –0.3 ± 0.4%, −0.4 ± 0.6% and –0.7 ± 1.2% in strategies A, B and C, respectively. Largest dose deficits were seen in individual fractions for strategy C (maximum dose reductions were −29.0% and –7.1% for PTVD95% and CTVD99%, respectively). The benefit of gating was minor, if additional motion correction was applied immediately prior to irradiation. Continuous motion monitoring based localization and motion correction ensured the target coverage and minimized the OAR exposure for every fraction and is recommended to use in prostate SBRT. The study is part of clinical trial NCT02319239.     

Solubility, adsorption, and the thermodynamics of the cutoff effect

For many years, the expression "cutoff effect of anesthesia," has been used to denote the failure of the higher alcohols or paraffins to produce anesthesia. As such, it is used to assess the plausibility of specific models, proposed for anesthesia. However, the uses were shown, in many respects, to be problematic. This article augments the notion of the cutoff to fit for all cases in which only some of the molecules in a homologous series are anesthetics. We find that the location of the cutoff points is affected by three free energy quantities: that of the adsorption of the agent to the anesthetic "site" (f(sl,site)), that of the perturbation of the site (f(ll,site)), and that of the evaporation of the agent from its pure condensed phase (Deltamu degrees (evaporation)). This outcome indicates that the cutoff cannot be attributed to a single parameter. In addition, the analyses that attribute the cutoff to the failure of compounds to obey the much-used Meyer-Overton correlation will have to be amended. This article shows that cutoff results can be used to elucidate the structure of a site.     

A preliminary phylogenetic study of late Palaeozoic spiriferoid brachiopods using cladistic and Bayesian approaches

The brachiopod Superfamily Spiriferoidea diversified greatly and was widely distributed in the late Palaeozoic (Carboniferous–Permian), and yet its phylogeny has been seldom investigated with analytical methods. This is reflected in the current flux of very different classification schemes for this superfamily. This paper provides the first attempt to investigate the phylogenetic relationships of spiriferoid brachiopods through both cladistic and Bayesian analyses involving 24 discrete and continuous characters. The continuous characters, from morphometric data, have been separately discretized using the gap weighting method, and the ‘as such’ option in TNT. Our results highlight the potential significance of continuous characters in reconstructing and elucidating phylogenies, as much as qualitative characters. Building on the outcomes of the analyses, we also briefly evaluate existing classification schemes of Spiriferoidea. We found that none of the existing classifications fully reflect the phylogeny properly; major families within the superfamily, such as Spiriferidae, Choristitidae, and Trigonotretidae, turned out to be polyphyletic. Although this study is considered preliminary, due to the selection of and restriction to certain taxa, combined with the use of a relatively small number of characters, it nevertheless demonstrates that potentially the true phylogenetic relationships of spiriferoid taxa sharply contrast with any of the existing classification schemes. This highlights the need to develop an alternative scheme that takes into account a more comprehensive range of phylogenetic variables.     

Elucidation of T cell signalling models

A potential mechanism that allows T cells to reliably discriminate pMHC ligands involves an interplay between kinetic proofreading, negative feedback and a destruction of this negative feedback. We analyse a detailed model of these mechanisms which involves the TCR, SHP1 and ERK. We discover that the behaviour of pSHP1 negative feedback is of primary importance, and particularly the influence of a kinetic proofreading base negative feedback state on pSHP1 dynamics. The CD8 co-receptor is shown to benefit from a kinetic proofreading locking mechanism and is able to overcome pSHP1 negative influences to sensitise a T cell.     

Optimization of Lipase Purification by Rotating Annular Size-exclusion Chromatography

Crude porcine lipase was purified by continuous rotating annular size-exclusion chromatography. Sephadex G-75 was used as the size-exclusion packing material. Initial studies by this group on a similar unit have been reported [Genest et al. (1998) "Continuous purification of porcine lipase by rotating annular size-exclusion chromatography", Appl. Biochem. Biotechnol. 73, 215-230]. This article presents the results of optimization studies carried out on a modified unit. These modifications resulted in a better performance of the column and a higher throughput. Purification fold values of around 11 were achieved in most runs. The activity recovered was around 99% and the productivity was around 3 mg lipase/mg gel h.     

Efficacy of liposome-encapsulated mepivacaine for infiltrative anesthesia in volunteers

This blinded crossover study evaluated the efficacy and pain sensitivity evoked by a previously reported liposome-encapsulated mepivacaine formulation (). Thirty healthy volunteers received an intraoral injection (1.8?mL), at four different sessions, of the following formulations: 2% mepivacaine with 1:100,000 epinephrine (MVC_2%EPI), 3% mepivacaine (MVC_3%), and 2 and 3% liposome-encapsulated mepivacaine (MVC_2%LUV and MVC_3%LUV). Latency period and duration of anesthesia were assessed by an electrical pulp tester and injection discomfort by a visual analog scale (VAS). Data were analyzed with Tukey-Kramer and Friedman tests (P?<?0.05). No significant difference was found regarding latency period (in minutes) among the formulations (P?>?0.05). The duration of anesthesia after the injection of MVC_3%LUV was higher than the one obtained after the infiltration of MVC_2%LUV and of MVC_3% (P?<?0.05). However, the duration of anesthesia obtained with MVC_3% did not differ from the one obtained with MVC_2%LUV (P?>?0.05). MVC_3%LUV showed lower VAS median values than MVC_2%EPI (P?<?0.05), and there were no significant differences among the others formulations. Liposome-encapsulated 3% mepivacaine showed longer duration of anesthesia, in comparison to the commercial formulation of MVC_3%. MVC_2%LUV was able to produce a similar duration of anesthesia as the 3% commercial formulation, despite the 50% decrease in the anesthetic concentration. Thus, the encapsulation of mepivacaine increased the duration of anesthesia and reduced the injection discomfort caused by vasoconstrictor-associated formulations in healthy volunteers.     

Two continuous coupled assays for ornithine-δ-aminotransferase

We have developed two new continuous coupled assays for ornithine-δ-aminotransferase (OAT) that are more sensitive than previous methods, measure activity in real time, and can be carried out in multiwell plates for convenience and high throughput. The first assay is based on the reduction of Δ¹-pyrroline-5-carboxylate (P5C), generated from ornithine by OAT, using human pyrroline 5-carboxylate reductase 1 (PYCR1), which results in the concomitant oxidation of NADH (nicotinamide adenine dinucleotide, reduced form) to NAD⁺ (nicotinamide adenine dinucleotide, oxidized form). This procedure was found to be three times more sensitive than previous methods and is suitable for the study of small molecules as inhibitors or inactivators of OAT or as a method to determine OAT activity in unknown samples. The second method involves the detection of l-glutamate, produced during the regeneration of the cofactor pyridoxal 5’-phosphate (PLP) of OAT by an unamplified modification of the commercially available Amplex Red l-glutamate detection kit (Life Technologies). This assay is recommended for the determination of the substrate activity of small molecules against OAT; measuring the transformation of l-ornithine at high concentrations by this assay is complicated by the fact that it also acts as a substrate for the l-glutamate oxidase (GluOx) reporter enzyme.     

集群式院外高危心脏病患者远程实时连续监护系统

设计了一种针对院外心血管病患者的远程心电监护系统,包括便携式监护终端和医院监护中心。其特点在于可对身处院外、自由活动的心血管病患者进行远程、实时、连续的监护,同时获得被监护患者的地理位置信息。已完成的整体调试结果表明,本系统不仅能及时发现异常心电图并进行干预,而且还可根据地理位置信息对突发急病的患者进行快速救治。该系统是院内监护系统功能的延伸,有良好的临床应用前景。