Verification of growth dependent survival in early life history of Pacific saury <Emphasis Type="Italic">Cololabis saira</Emphasis> using laboratory experiment

To understand the survival processes in Pacific saury during the early life stages, growth histories were compared between individuals that died and those that survived under laboratory conditions. The morphology of Pacific saury drastically changed by 40 day after hatching (DAH) under 20°C, and during this period most individuals reached the juvenile stage (40 mm knob length (KnL)). From hatch to this period, high levels of mortality were observed. The back-calculated growth rates analyzed from otolith microstructure were compared among individuals that died and those that survived this period of metamorphosis. Growth rates of individuals that died were significantly lower since 1 DAH than surviving individuals. Factors contributing to mortality are considered to be the cumulative effects of their delayed growth and their less favorable body conditions (growth dependent survival). The body size (40 mm KnL) that is currently used to estimate the recruitment is concluded to be valid for evaluating the recruitment in the field.     

Production of starch with antioxidative activity by baking starch with organic acids

A starch ingredient with antioxidative activity, as measured by the DPPH method, was produced by baking corn starch with an organic acid; it has been named ANOX sugar (antioxidative sugar). The baking temperature and time were fixed at 170 °C and 60 min, and the organic acid used was selected from preliminary trials of various kinds of acid. The phytic acid ANOX sugar preparation showed the highest antioxidative activity, but the color of the preparation was almost black; we therefore selected L-tartaric acid which had the second highest antioxidative activity. The antioxidative activity of the L-tartaric acid ANOX sugar preparation was stable against temperature, light, and enzyme treatments (α-amylase and glucoamylase). However, the activity was not stable against variations in water content and pH value. The antioxidative activity of ANOX sugar was stabilized by treating with boiled water or nitrogen gas, or by pH adjustment.     

Impact of Platelet Transfusion on Survival of Patients with Intracerebral Hemorrhage after Administration of Anti-Platelet Agents at a Tertiary Emergency Center

This study examined the impact of platelet transfusion (PLT) on the survival of intracerebral hemorrhage (ICH) patients who had been administered anti-platelet agents (APA). This retrospective cohort analysis investigated 432 patients (259 men, 60%) who were newly diagnosed with ICH between January 2006 and June 2011 at the tertiary emergency center of Kitasato University Hospital. Median age on arrival was 67.0 years (range, 40–95 years). ICH was subcortical in 72 patients (16.7%), supratentorial in 233 (53.9%), and infratentorial in 133 (30.8%). PLT was performed in 16 patients (3.7%). Within 90 days after admission to the center, 178 patients (41.2%) had died due to ICH. Before the onset of ICH, 66 patients had been prescribed APA because of atherosclerotic diseases. Multivariate regression analysis indicated APA administration was an independent risk factor for death within 7 days (odds ratio, 5.12; P = 0.006) and within 90 days (hazard ratio, 1.87; P = 0.006) after arrival. Regarding the effect of a PLT in ICH patients with APA, no patient with PLT died. PLT had a survival benefit on patients with ICH, according to our analysis. Further prospective analysis is necessary to confirm the effects of PLT on survival in ICH with APA.     

Multistep ion channel remodeling and lethal arrhythmia precede heart failure in a mouse model of inherited dilated cardiomyopathy

Background

Patients with inherited dilated cardiomyopathy (DCM) frequently die with severe heart failure (HF) or die suddenly with arrhythmias, although these symptoms are not always observed at birth. It remains unclear how and when HF and arrhythmogenic changes develop in these DCM mutation carriers. In order to address this issue, properties of the myocardium and underlying gene expressions were studied using a knock-in mouse model of human inherited DCM caused by a deletion mutation ΔK210 in cardiac troponinT.

Methodology/Principal Findings

By 1 month, DCM mice had already enlarged hearts, but showed no symptoms of HF and a much lower mortality than at 2 months or later. At around 2 months, some would die suddenly with no clear symptoms of HF, whereas at 3 months, many of the survivors showed evident symptoms of HF. In isolated left ventricular myocardium (LV) from 2 month-mice, spontaneous activity frequently occurred and action potential duration (APD) was prolonged. Transient outward (I_to) and ultrarapid delayed rectifier K⁺ (I_Kur) currents were significantly reduced in DCM myocytes. Correspondingly, down-regulation of Kv4.2, Kv1.5 and KChIP2 was evident in mRNA and protein levels. In LVs at 3-months, more frequent spontaneous activity, greater prolongation of APD and further down-regulation in above K⁺ channels were observed. At 1 month, in contrast, infrequent spontaneous activity and down-regulation of Kv4.2, but not Kv1.5 or KChIP2, were observed.

Conclusions/Significance

Our results suggest that at least three steps of electrical remodeling occur in the hearts of DCM model mice, and that the combined down-regulation of Kv4.2, Kv1.5 and KChIP2 prior to the onset of HF may play an important role in the premature sudden death in this DCM model. DCM mice at 1 month or before, on the contrary, are associated with low risk of death in spite of inborn disorder and enlarged heart.     

Discovery of pyrrolo[3,2-c]quinoline-4-one derivatives as novel hedgehog signaling inhibitors

The Hedgehog (Hh) signaling pathway plays a significant role in the regulation of cell growth and differentiation during embryonic development. Since activation of the Hh signaling pathway is implicated in several types of human cancers, inhibitors of this pathway could be promising anticancer agents. Using high throughput screening, thieno[3,2-c]quinoline-4-one derivative 9a was identified as a compound of interest with potent in vitro activity but poor metabolic stability. Our efforts focused on enhancement of in vitro inhibitory activity and metabolic stability, including core ring conversion and side chain optimization. This led to the discovery of pyrrolo[3,2-c]quinoline-4-one derivative 12b, which has a structure distinct from previously reported Hh signaling inhibitors. Compound 12b suppressed stromal Gli1 mRNA expression in a murine model and demonstrated antitumor activity in a murine medulloblastoma allograft model.     

Targeted disruption of the cardiac troponin T gene causes sarcomere disassembly and defects in heartbeat within the early mouse embryo

Cardiac troponin T (cTnT) is a component of the troponin (Tn) complex in cardiac myocytes, and plays a regulatory role in cardiac muscle contraction by anchoring two other Tn components, troponin I (TnI) and troponin C, to tropomyosin (Tm) on the thin filaments. In order to determine the in vivo function of cTnT, we created a null cTnT allele in the mouse TNNT2 locus. In cTnT-deficient (cTnT^−/−) cardiac myocytes, the thick and thin filaments and α-actinin-positive Z-disk-like structures were not assembled into sarcomere, causing early embryonic lethality due to a lack of heartbeats. TnI was dissociated from Tm in the thin filaments without cTnT. In spite of loss of Tn on the thin filaments, the cTnT^−/− cardiac myocytes showed regular Ca²⁺-transients. These findings indicate that cTnT plays a critical role in sarcomere assembly during myofibrillogenesis in the embryonic heart, and also indicate that the membrane excitation and intracellular Ca²⁺ handling systems develop independently of the contractile system. In contrast, heterozygous cTnT^+/− mice had a normal life span with no structural and functional abnormalities in their hearts, suggesting that haploinsufficiency could not be a potential cause of cardiomyopathies, known to be associated with a variety of mutations in the TNNT2 locus.     

Dictyostelium differentiation-inducing factor-3 activates glycogen synthase kinase-3beta and degrades cyclin D1 in mammalian cells

In search of chemical substances applicable for the treatment of cancer and other proliferative disorders, we studied the signal transduction of Dictyostelium differentiation-inducing factors (DIFs) in mammalian cells mainly using HeLa cells. Although DIF-1 and DIF-3 both strongly inhibited cell proliferation by inducing G(0)/G(1) arrest, DIF-3 was more effective than DIF-1. DIF-3 suppressed cyclin D1 expression at both mRNA and protein levels, whereas the overexpression of cyclin D1 overrode DIF-3-induced cell cycle arrest. The DIF-3-induced decrease in the amount of cyclin D1 protein preceded the reduction in the level of cyclin D1 mRNA. The decrease in cyclin D1 protein seemed to be caused by accelerated proteolysis, since it was abrogated by N-acetyl-Leu-Leu-norleucinal, a proteasome inhibitor. DIF-3-induced degradation of cyclin D1 was also prevented by treatment with lithium chloride, an inhibitor of glycogen synthase kinase-3beta (GSK-3beta), suggesting that DIF-3 induced cyclin D1 proteolysis through the activation of GSK-3beta. Indeed, DIF-3 dephosphorylated Ser(9) and phosphorylated tyrosine on GSK-3beta, and it stimulated GSK-3beta activity in an in vitro kinase assay. Moreover, DIF-3 was revealed to induce the nuclear translocation of GSK-3beta by immunofluorescent microscopy and immunoblotting of subcellular protein fractions. These results suggested that DIF-3 activates GSK-3beta to accelerate the proteolysis of cyclin D1 and that this mechanism is involved in the DIF-3-induced G(0)/G(1) arrest in mammalian cells.     

Isonitrile hydratase from Pseudomonas putida N19-2. Cloning,sequencing, gene expression,and identification of its active acid residue

Isonitrile hydratase is a novel enzyme in Pseudomonas putida N19-2 that catalyzes the conversion of isonitriles to N-substituted formamides. Based on N-terminal and internal amino acid sequences, a 535-bp DNA fragment corresponding to a portion of the isonitrile hydratase gene was amplified, which was used as a probe to clone a 6.4-kb DNA fragment containing the whole gene. Sequence analysis of the 6.4-kb fragment revealed that the isonitrile hydratase gene (inhA) was 684 nucleotides long and encoded a protein with a molecular mass of 24,211 Da. Overexpression of inhA in Escherichia coli gave a large amount of soluble isonitrile hydratase exhibiting the same molecular and catalytic properties as the native enzyme from the Pseudomonas strain. The predicted amino acid sequence of inhA showed low similarity to that of an intracellular protease in Pyrococcus horikoshii (PH1704), and an active cysteine residue in the protease was conserved in the isonitrile hydratase at the corresponding position (Cys-101). A mutant enzyme containing Ala instead of Cys-101 did not exhibit isonitrile hydratase activity at all, demonstrating the essential role of this residue in the catalytic function.     

Improvement of pulmonary arterial hypertension,inflammatory response,and epithelium injury by dual activation of cAMP/cGMP pathway in a rat model of monocrotaline-induced pulmonary hypertension

Pulmonary hypertension (PH) is a life-threatening lung disease. PH with concomitant lung diseases, e.g., idiopathic pulmonary fibrosis, is associated with poor prognosis. Development of novel therapeutic vasodilators for treatment of these patients is a key imperative. We evaluated the efficacy of dual activation of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) using an active, small-molecule phosphodiesterase (PDE4)/PDE5 dual inhibitor (Compound A). Compound A increased both cAMP and cGMP levels in WI-38 lung fibroblasts and suppressed the expressions of type-1 collagen α1 chain and fibronectin. Additionally, compound A reduced right ventricular weight/left ventricular weight+septal weight ratio, brain natriuretic peptide expression levels in right ventricle, C─C motif chemokine ligand 2 expression levels in lung, and plasma surfactant protein D. Our data indicate that dual activation of cAMP/cGMP pathways may be a novel treatment strategy for PH.     

p40phox as an alternative organizer to p47phox in Nox2 activation: a new mechanism involving an interaction with p22phox

p40(phox) activated phagocyte NADPH oxidase without p47(phox) in a cell-free system consisting of p67(phox), Rac and cytochrome b(558) relipidated with phosphatidylinositol 3-phosphate. The activation reached to 70% of that by p47(phox). Addition of p47(phox) slightly increased the activation, but not additively. p40(phox) improved the efficiency of p67(phox) in the activation. The C-terminus-truncated p67(phox), p40(phox)(D289A), p40(phox)(R58A), or p40(phox)(W207R) showed an impaired activation. A peptide corresponding to the p22(phox) Pro-rich region suppressed the activation, and far-western blotting revealed its interaction with p40(phox) SH3 domain. Thus, p40(phox) can substitute for p47(phox) in the activation, interacting with p22(phox) and p67(phox) through their specific regions.