Ionic mechanisms and Ca2+ dynamics underlying the glucose response of pancreatic β cells: a simulation study

To clarify the mechanisms underlying the pancreatic β-cell response to varying glucose concentrations ([G]), electrophysiological findings were integrated into a mathematical cell model. The Ca(2+) dynamics of the endoplasmic reticulum (ER) were also improved. The model was validated by demonstrating quiescent potential, burst-interburst electrical events accompanied by Ca(2+) transients, and continuous firing of action potentials over [G] ranges of 0-6, 7-18, and >19 mM, respectively. These responses to glucose were completely reversible. The action potential, input impedance, and Ca(2+) transients were in good agreement with experimental measurements. The ionic mechanisms underlying the burst-interburst rhythm were investigated by lead potential analysis, which quantified the contributions of individual current components. This analysis demonstrated that slow potential changes during the interburst period were attributable to modifications of ion channels or transporters by intracellular ions and/or metabolites to different degrees depending on [G]. The predominant role of adenosine triphosphate-sensitive K(+) current in switching on and off the repetitive firing of action potentials at 8 mM [G] was taken over at a higher [G] by Ca(2+)- or Na(+)-dependent currents, which were generated by the plasma membrane Ca(2+) pump, Na(+)/K(+) pump, Na(+)/Ca(2+) exchanger, and TRPM channel. Accumulation and release of Ca(2+) by the ER also had a strong influence on the slow electrical rhythm. We conclude that the present mathematical model is useful for quantifying the role of individual functional components in the whole cell responses based on experimental findings.     

Saccharated colloidal iron enhances lipopolysaccharide-induced nitric oxide production in vivo

We investigated the effects of iron on the production of nitric oxide (NO), inducible NO synthase (iNOS), and plasma cytokines induced by lipopolysaccharide (LPS) in vivo. Male Wistar rats were preloaded with a single intravenous injection of saccharated colloidal iron (Fesin, 70 mg iron/kg body weight) or normal saline as a control, and then given an intraperitoneal injection of LPS (5.0 mg/kg body weight). Rats, preloaded with iron, had evidence of both iron deposition and strong iNOS induction in liver Kupffer cells upon injection of LPS; phagocytic cells in the spleen and lung had similar findings. LPS-induced NO production in iron-preloaded rats was significantly higher than control rats as accessed by NO-hemoglobin levels measured by ESR (electron spin resonance) and NOx (nitrate plus nitrite) levels. Western blot analysis showed that iron preloading significantly enhanced LPS-induced iNOS induction in the liver, but not in the spleen or lung. LPS-induced plasma levels of IL-6, IL-1beta, and TNF-alpha were also significantly higher in iron-preloaded rats as shown by ELISA, but IFN-gamma levels were unchanged. We conclude that colloidal-iron phagocytosed by liver Kupffer cells enhanced LPS-induced NO production in vivo, iNOS induction in the liver, and release of IL-6, IL-1beta, and TNF-alpha.     

Structural basis for allosteric regulation of the monomeric allosteric enzyme human glucokinase

Glucokinase is a monomeric enzyme that displays a low affinity for glucose and a sigmoidal saturation curve for its substrate, two properties that are important for its playing the role of a glucose sensor in pancreas and liver. The molecular basis for these two properties is not well understood. Herein we report the crystal structures of glucokinase in its active and inactive forms, which demonstrate that global conformational change, including domain reorganization, is induced by glucose binding. This suggests that the positive cooperativity of monomeric glucokinase obeys the "mnemonical mechanism" rather than the well-known concerted model. These structures also revealed an allosteric site through which small molecules may modulate the kinetic properties of the enzyme. This finding provided the mechanistic basis for activation of glucokinase as a potential therapeutic approach for treating type 2 diabetes mellitus.     

Range expansion and lineage admixture of the Japanese evergreen tree Machilus thunbergii in central Japan

We investigated the range expansion histories of Machilus thunbergii populations in the Kinki region of central Japan on the basis of nuclear microsatellite data. In the Kinki region, M. thunbergii is typically found in the coastal area, with some fragmented populations inland, around Lake Biwa. Phylogenetic and Bayesian clustering analysis (STRUCTURE analysis) revealed that the inland populations have different genetic components between the west and east sides of Lake Biwa. The population located on the north side of the lake has an admixture of the two genetically differentiated lineages, contributing to an increase in the genetic diversity of the population. Populations around Lake Biwa had lost rare alleles and the F value obtained from STRUCTURE analysis was lower in the coastal populations than in the lake populations. These results suggest that populations around Lake Biwa experienced a bottleneck due to a founder effect during the initial migration to the lake and that glacial refugia of M. thunbergii in the Kinki region existed along the coast.     

Organization and evolution of variable region genes of the human immunoglobulin heavy chain

We have isolated 23 different cosmid clones of the heavy-chain variable region genes (VH) of human immunoglobulin. These clones encompass about 1000 X 10(3) base-pairs of DNA containing 61 VH genes. Characterization of the 23 clones by Southern blot hybridization showed that VH genes belonging to different families were physically linked in many regions. Cluster 71, which was analyzed in detail, comprised seven VH segments arranged in the same orientation with different intervals. This clone contained internal homology regions, each carrying two VH segments of different families. Comparison of the nucleotide sequences of VH segments within each family showed that profiles of accumulation of mutations in framework (FR) and complementarity-determining (CDR) regions were different. CDR had more mutations at amino-acid-substituting positions than at silent positions, whereas FR had the reverse distribution of mutations. Five out of seven VH segments of this cluster were pseudogenes containing various mutations. VH pseudogenes were classified into two distinct groups; one with a few replacement mutations (conserved pseudogenes), and the other with rather extensive mutations (diverged pseudogenes). The possibility that conserved pseudogenes serve as a reservoir of VH segments is discussed.     

健康早产儿24小时脑电图的定量评价

早产儿的生物学行为主要以睡眠为主,因此了解各个睡眠阶段的脑电图模型,对临床客观评价早产儿的大脑发育特点及健康状况具有重要的临床意义。本文旨在探讨健康早产儿不同睡眠阶段脑电图特点,进而为其睡眠阶段的自动判定提供标准。应用多导生理记录仪对24例健康早产儿进行了41次Fp1-C3双电极连续24小时脑电图记录,同时用两台摄像机同步记录眼球运动及肢体运动。依据呼吸、眼球及肢体运动视觉判定睡眠阶段。应用自回两台摄像机同步记录眼球运动最适赤池信息量准则（Min-AIC)、总功率（TP）、δ功率（δP）、θ功率（θP）、α功率（αP）、β功率（βP）及不连续性（Dis）进行分析。结果显示Min-AIC,TP,δP及Dis四个变量依睡眠阶段不同有显著性差异（P＜0．01）。多变量判别分析亦显示联合这四个变量可对睡眠阶段进行判定。联合Min-AIC、TP、δP及Dis四个变量适于对胎龄大于30周的健康早产儿的睡眠阶段进行判定。     

Cloning of cDNA for human T-cell replacing factor (interleukin-5) and comparison with the murine homologue.

We have cloned cDNA for T-cell replacing factor (interleukin-5), which replaces T-cell helper function for normal B cells which secrete immunoglobulin, from human T cell leukemia line, ATL-2, using mouse interleukin-5 cDNA as probe. Total nucleotide sequence of the cDNA (816 base pairs) was determined and compared with that of mouse interleukin-5 cDNA. The cloned cDNA encoded the interleukin-5 precursor of 134 amino acids containing an N-terminal signal sequence. Although the human interleukin-5 precursor is one amino acid longer than the murine homologue, the sizes of the mature proteins appear similar. The nucleotide and amino acid sequence homologies of the coding regions of human and murine interleukin-5 are 77% and 70%, respectively. Human interleukin-5 synthesized by the direction of the cloned cDNA induced immunoglobulin synthesis in human B cells stimulated by Staphylococcus aureus mitogen.     

Roles of Poly(3-Hydroxybutyrate) Depolymerase and 3HB-Oligomer Hydrolase in Bacterial PHB Metabolism

Many poly-3-hydroxybutyrate (PHB)-degrading enzymes have been studied. But biological roles of 3HB-oligomer hydrolases (3HBOHs) and how PHB depolymerases (PHBDPs) and 3HBOHs cooperate in PHB metabolism are not fully elucidated. In this study, several PHBDPs and 3HBOHs from three types of bacteria were purified, and their substrate specificity, kinetic properties, and degradation products were investigated. From the results, PHBDP and 3HBOH seemed to play a role in PHB metabolism in three types of bacteria, as follows: (A) In Ralstonia pickettii T1, an extracellular PHBDP degrades extracellular PHB to various-sized 3HB-oligomers, which an extracellular 3HBOH hydrolyzes to 3HB-monomers. (B) In Acidovorax sp. SA1, an extracellular PHBDP hydrolyzes extracellular PHB to small 3HB-oligomers (dimer and trimer), which an intracellular 3HBOH efficiently degrades to 3HB in the cell. (C) In Ralstonia eutropha H16, an intracellular 3HBOH helps in the degradation of intracellular PHB inclusions by PHBDP.     

Induction of oligodendrocyte differentiation from adult human fibroblast-derived induced pluripotent stem cells

Induced pluripotent stem cells (iPSCs) prepared from somatic cells might become a novel therapeutic tool in regenerative medicine, especially for the central nervous system (CNS). In this study, we attempted to induce O4-positive (O4⁺) oligodendrocytes from adult human fibroblast-derived iPSCs in vitro. We used two adult human iPSC cell lines, 201B7 and 253G1. 201B7 was induced by four-gene transduction (oct4, sox2, klf4, c-myc), and 253G1 was induced by three-gene transduction (oct4, sox2, klf4). We treated these cells with two in vitro oligodendrocyte-directed differentiation protocols that were optimized for human embryonic stem cells. One protocol used platelet-derived growth factor as the major mitogen for oligodendrocyte lineage cells, and the other protocol used epidermal growth factor (EGF) as the mitogen. Although the differentiation efficiency was low (less than 0.01%), we could induce O4⁺ oligodendrocytes from 253G1 cells using the EGF-dependent differentiation protocol. This is the first report of the in vitro induction of oligodendrocytes differentiation from human iPSCs.