Histidine77, glutamic acid81, glutamic acid123, threonine126, asparagine194, and tryptophan197 of the human emopamil binding protein are required for in vivo sterol delta 8-delta 7 isomerization

The human emopamil binding protein (hEBP) exhibits sterol Delta8-Delta7 isomerase activity (EC 5.3.3.5) upon heterologous expression in a sterol Delta8-Delta7 isomerization-deficient erg2-3 yeast strain. Ala scanning mutagenesis was used to identify residues in the four putative transmembrane alpha-helices of hEBP that are required for catalytic activity. Isomerization was assayed in vivo by spectrophotometric quantification of Delta5,7-sterols. Out of 64 Ala mutants of hEBP only H77A-, E81A-, E123A-, T126A-, N194A-, and W197A-expressing yeast strains contained 10% or less of wild-type (wt) Delta5,7-sterols. All substitutions of these six residues with functionally or structurally similar amino acid residues failed to fully restore catalytic activity. Mutants E81D, T126S, N194Q, and W197F, but not H77N and E123D, still bound the enzyme inhibitor 3H-ifenprodil. Changed equilibrium and kinetic binding properties of the mutant enzymes confirmed our previous suggestion that residues required for catalytic activity are also involved in inhibitor binding [Moebius et al. (1996) Biochemistry 35, 16871-16878]. His77, Glu81, Glu123, Thr126, Asn194, and Trp197 are localized in the cytoplasmic halves of the transmembrane segments 2-4 and are proposed to line the catalytic cleft. Ala mutants of Trp102, Tyr105, Asp109, Arg111, and Tyr112 in a conserved cytoplasmic domain (WKEYXKGDSRY) between transmembrane segments 2 and 3 contained less than 10% of wt Delta5,7-sterols, implying that this region also could be functionally important. The in vivo complementation of enzyme-deficient yeast strains with mutated cDNAs is a simple and sensitive method to rapidly analyze the functional consequences of mutations in sterol modifying enzymes.     

Quantification of regional left ventricular dyssynchrony by magnetic resonance imaging

Cardiac resynchronization therapy is an established treatment in patients with symptomatic heart failure and intraventricular conduction delay. Electrical dyssynchrony is typically adopted to represent myocardial activation dyssynchrony, which should be compensated by cardiac resynchronization therapy. One third of the patients, however, does not respond to the therapy. Therefore, imaging modalities aimed at the mechanical dyssynchrony estimation have been recently proposed to improve patient selection criteria. This paper presents a novel fully automated method for regional mechanical left ventricular dyssynchrony quantification in short-axis magnetic resonance imaging. The endocardial movement is described by time-displacement curves with respect to an automatically determined reference point. Different methods are proposed for time-displacement curve analysis aimed at the regional contraction timing estimation. These methods were evaluated in two groups of subjects with (nine patients) and without (six patients) left bundle branch block. The contraction timing standard deviation showed a significant increase for left bundle branch block patients with all the methods. A novel method based on phase spectrum analysis may be however preferred due to a better specificity (99.7%) and sensitivity (99.0%). In conclusion, this method provides a valuable prognostic indicator for heart failure patients with dyssynchronous ventricular contraction and it opens new possibilities for regional timing analysis.     

孔子论死亡

孔子有限的几句关于死亡的话语其实透露出孔子对如何生的思考。＂未知生,焉知死？＂这句话告诉我们中国哲学和西方哲学一样,肇始于学会如何看待死亡。＂朝闻道,夕死可矣＂反映了君子肩负传承道德的人生使命;仁则是君子在传道途中不畏惧死亡的根本原因;孔子追求的是一种有意识的生和死。在某种意义上,死亡是一个人生命的完成。     

Simultaneous determination of the micro-, meso-, and macropore size fractions of porous polymers by a combined use of Fourier transform near-infrared diffuse reflection spectroscopy and multivariate techniques

Fourier transform near-infrared (FT-NIR) diffuse reflection spectroscopy was used in combination with principal component analysis and partial least-squares regression to simultaneously determine the physical and the chemical parameters of a porous poly(p-methylstyrene-co-1,2-bis(p-vinylphenyl)ethane) (MS/BVPE) monolithic polymer. Chemical variations during the synthesis of the polymer material can alter the pore volume and pore area distributions within the polymer scaffold. Furthermore, mid-infrared and near-infrared (NIR) spectroscopic chemical imaging was implemented as a tool to assess the uniformity of the samples. The presented study summarizes the comparative results derived from the spectral FT-NIR data combined with chemometric techniques. The relevance of the interrelation of physical and chemical parameters is highlighted whereas the amount of MS/BVPE (%, v/v) and the quantity (%) of micropores (diameter, d < 6 nm), mesopores (6 nm < d < 50 nm), and macropores (50 nm < d < 200 nm) could be determined with one measurement. For comparison of the quantitative data, the standard error of prediction (SEP) was used. The SEP for determining the MS/BVPE amount in the samples showed 0.35%, for pore volume quantiles 1.42-8.44%, and for pore area quantiles 0.38-1.45%, respectively. The implication of these results is that FT-NIR spectroscopy is a suitable technique for the screening of samples with varying physicochemical properties and to quantitatively determine the parameters simultaneously within a few seconds.