Preparation and evaluation of regioselectively substituted amylose derivatives for chiral separations

Six novel regioselectively substituted amylose derivatives with a benzoate at 2‐position and two different phenylcarbamates at 3‐ and 6‐positions were synthesized and their structures were characterized by ¹H nuclear magnetic resonance (NMR) spectroscopy. Their enantioseparation abilities were then examined as chiral stationary phases (CSPs) for high‐performance liquid chromatography (HPLC) after they were coated on 3‐aminopropyl silica gels. Investigations indicated that the substituents at the 3‐ and 6‐positions played an important role in chiral recognition of these amylose 2‐benzoate serial derivatives. The derivatives demonstrated characteristic enantioseparation and some racemates were better resolved on these derivatives than on Chiralpak AD, which is one of the most efficient CSPs, utilizing coated amylose tris(3,5‐dimethylphenylcarbamate) as the chiral selector. Among the derivatives prepared, amylose 2‐benzoate‐3‐(phenylcarbamate/4‐methylphenylcarbamate)‐6‐(3,5‐dimethylphenylcarbamate) exhibited chiral recognition abilities comparable to that of Chiralpak AD and may be useful CSPs in the future. The effect of mobile phase on chiral recognition was also studied. In general, with the decreased concentration of 2‐propanol, better resolutions were obtained with longer retention times. Moreover, when ethanol was used instead of 2‐propanol, poorer resolutions were often achieved. However, in some cases, improved enantioselectivity was achieved with ethanol rather than 2‐propanol as the mobile phase modifier.     

NEIL3-Dependent Regulation of Cardiac Fibroblast Proliferation Prevents Myocardial Rupture

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Fibromodulin Interacts with Collagen Cross-linking Sites and Activates Lysyl Oxidase

The hallmark of fibrotic disorders is a highly cross-linked and dense collagen matrix, a property driven by the oxidative action of lysyl oxidase. Other fibrosis-associated proteins also contribute to the final collagen matrix properties, one of which is fibromodulin. Its interactions with collagen affect collagen cross-linking, packing, and fibril diameter. We investigated the possibility that a specific relationship exists between fibromodulin and lysyl oxidase, potentially imparting a specific collagen matrix phenotype. We mapped the fibromodulin-collagen interaction sites using the collagen II and III Toolkit peptide libraries. Fibromodulin interacted with the peptides containing the known collagen cross-linking sites and the MMP-1 cleavage site in collagens I and II. Interestingly, the interaction sites are closely aligned within the quarter-staggered collagen fibril, suggesting a multivalent interaction between fibromodulin and several collagen helices. Furthermore, we detected an interaction between fibromodulin and lysyl oxidase (a major collagen cross-linking enzyme) and mapped the interaction site to 12 N-terminal amino acids on fibromodulin. This interaction also increases the activity of lysyl oxidase. Together, the data suggest a fibromodulin-modulated collagen cross-linking mechanism where fibromodulin binds to a specific part of the collagen domain and also forms a complex with lysyl oxidase, targeting the enzyme toward specific cross-linking sites.     

Turnover of cell surface-bound capsular polysaccharide in Staphylococcus aureus

The capsular polysaccharide (CPS) of Staphylococcus aureus strain Smith was labelled by growth of bacteria in the presence of radioactive N-acetylglucosamine and was separated from labelled cell wall components by affinity chromatography on wheat germ agglutinin following dissolution of the cells by lysostaphin. The products were partially characterised chemically and immunochemically. Similar labelled components were found in the culture fluid during growth. In a pulse-chase experiment, cell-bound CPS was released continuously into the culture fluid at the same rate as cell wall turnover and there was no evidence of direct excretion of CPS.     

Extracellular vesicles do not contribute to higher circulating levels of soluble LRP1 in idiopathic dilated cardiomyopathy

Idiopathic dilated cardiomyopathy (IDCM) is a frequent cause of heart transplantation. Potentially valuable blood markers are being sought, and low‐density lipoprotein receptor‐related protein 1 (LRP1) has been linked to the underlying molecular basis of the disease. This study compared circulating levels of soluble LRP1 (sLRP1) in IDCM patients and healthy controls and elucidated whether sLRP1 is exported out of the myocardium through extracellular vesicles (EVs) to gain a better understanding of the pathogenesis of the disease. LRP1 α chain expression was analysed in samples collected from the left ventricles of explanted hearts using immunohistochemistry. sLRP1 concentrations were determined in platelet‐free plasma by enzyme‐linked immunosorbent assay. Plasma‐derived EVs were extracted by size‐exclusion chromatography (SEC) and characterized by nanoparticle tracking analysis and cryo‐transmission electron microscopy. The distributions of vesicular (CD9, CD81) and myocardial (caveolin‐3) proteins and LRP1 α chain were assessed in SEC fractions by flow cytometry. LRP1 α chain was preferably localized to blood vessels in IDCM compared to control myocardium. Circulating sLRP1 was increased in IDCM patients. CD9‐ and CD81‐positive fractions enriched with membrane vesicles with the expected size and morphology were isolated from both groups. The LRP1 α chain was not present in these SEC fractions, which were also positive for caveolin‐3. The increase in circulating sLRP1 in IDCM patients may be clinically valuable. Although EVs do not contribute to higher sLRP1 levels in IDCM, a comprehensive analysis of EV content would provide further insights into the search for novel blood markers.     

Structure of the Escherichia coli 0104 polysaccharide and its identity with the capsular K9 polysaccharide

Spontaneous mutants of Rhizobium leguminosarum biovar viciae strain C1204b were selected for their ability to tolerate 0.2 M NaCl, a growth-inhibiting level of salt for the parental strain. Transposon-mediated salt-sensitive mutants of strain C1204b were screened for their inability to grow in 0.08 M NaCl. Quantitation of the free-amino acid pools in the mutants grown in NaCl revealed a dramatic increase in glutamine, serine, glutamate and proline, and to a lesser extent alanine and glycine in the salt-tolerant mutants in comparison with the parental strain exposed to NaCl; but only glutamate and proline increased in the salt-sensitive mutants under NaCl stress. Extracellular polysaccharide levels were quantitated for the salt-tolerant mutants and determined to be approximately two-fold higher than for the parental strain. Although the mutations that occurred in the NaCl-tolerant and NaCl-sensitive strains did not interfere with nodule formation, no nitrogenase activity could be observed in the NaCl tolerant mutants as evaluated by acetylene reduction.     

Role of Asp297 of the AT2 receptor in high-affinity binding to different peptide ligands

To determine how ligand-receptor interaction is affected by the charges of the amino acids at position 2 of the ligands and position 297 of the AT2 receptor, we generated the Asp297Lys mutant of AT2 and a ligand SarAsp²Ile. Asp297Lys mutant lost affinity to Ang II and SarIle however retained partial affinity to ¹²⁵I-CGP42112A. The SarAsp²Ile had high affinity to Asp297Lys (IC₅₀3.5nM) and partial affinity to the AT2 (IC₅₀15nM). Therefore, not only the charge, but also the length of the side arms of the amino acids at position 2 of the ligand and position 297 of the receptor affect their interaction.     

花生四烯酸代谢相关酶和ERK与乳腺癌转移关系

花生四烯酸代谢相关酶环加氧酶(COX)、脂氧合酶(LOX)和活化的胞外信号调节激酶（p-ERK1/2）等与乳腺癌发生发展具有密切关系.为了进一步阐明它们在乳腺癌转移中的作用及其分子机制，应用反转录PCR（RT-PCR）、免疫印迹和免疫组化等方法，分别检测了乳腺癌细胞系、乳腺癌组织、癌旁组织和转移淋巴结组织中COX-2、5-LOX、12R-LOX、cPLA2和p-ERK1/2的表达水平.结果显示，与对照组相比，在具有高转移潜能的乳腺癌LM MCF-7细胞和MDA-MB-231细胞以及转移淋巴结组织中,COX-2、5-LOX、12R-LOX、cPLA2和p-ERK1/2均有较高水平表达.进而发现，p-ERK1/2的特异性抑制剂PD98059可拮抗LM-MCF-7细胞和MDA-MB-231细胞中COX-2、5-LOX、12R-LOX和cPLA2的高表达.上述结果表明，p-ERK1/2可以通过促进花生四烯酸代谢相关酶的表达促进乳腺癌细胞发生转移.     

不同发育时期樟子松（Pinus sylvestris L. var. mongolica Litv.）的电阻抗参数与抗寒性的关系

采用电阻抗图谱（EIS）法和电导（EL）法对不同发育时期的樟子松（Pinus sylvestris L. var. mongolica Litv.）茎和针叶进行了抗寒性测定,试图通过比较两种方法测定抗寒性结果的相关性,找到适合冷冻处理后樟子松抗寒性测定和不经冷冻处理估测抗寒性的EIS参数,完善EIS法测定抗寒性。以8年生樟子松苗为试材,在抗寒锻炼阶段（10月份）和脱锻炼阶段（3月份）分别取样进行EIS和EL测定。结果表明,EIS法胞外电阻率（re）与EL法测定的樟子松抗寒性相关性较高（R2=0.97）,但比EL法求出的抗寒性高。针叶的细胞膜时间恒量（τm）和茎的弛豫时间（τ1）随冷冻温度变化与re表现相似的S曲线,相关分析表明,re（茎和针叶）与τ1（茎）和τm（针叶）的变化有较好的相关性（R2=0.74～0.84）。经Logistic方程拟合,EIS的τm（针叶）和τ1（茎）法与EIS（re）法、EL法测定的樟子松抗寒性相关性也较高（R2=0.88～0.91）,说明针叶τm和茎τ1也可以作为计算抗寒性的参数。另外,8年生樟子松两个发育时期（10月和3月份）未经冷冻的针叶τm与茎的τ2随抗寒性的增强而显著增加,表明不经过冷冻处理样本用τ2（茎）和τm（针叶）估计樟子松抗寒性是很有前途的方法。