Additional file 1 of miR-30a-5p attenuates hypoxia/reoxygenation-induced cardiomyocyte apoptosis by regulating PTEN protein expression and activating PI3K/Akt signaling pathway

Supplementary Material 1

Serine Protease MP2 Activates Prophenoloxidase in the Melanization Immune Response of <i>Drosophila melanogaster</i>

<div><p>In arthropods, melanization plays a major role in the innate immune response to encapsulate and kill the invasive organisms. It is mediated by a serine protease cascade and is regulated by serpins. The identification of the molecular components of melanization and the regulation of those components are still unclear in <i>Drosophila melanogaster</i>, although some genetic research on the activation of melanization has been reported. Here we report that <i>Drosophila</i> serine protease MP2 directly cleaves both recombinant and native prophenoloxidase-1. Overexpression or repression of MP2 in flies resulted in increased and decreased rates of cleavage, respectively, of prophenoloxidase-1. Moreover, serine protease inhibitor Spn27A formed SDS-stable complexes with MP2, both <i>in vitro</i> and <i>in vivo</i>. The amidase activity of MP2 was inhibited efficiently by Spn27A. Spn27A also prevented MP2 from cleaving prophenoloxidase-1. Taken together, these results indicate that under our experimental conditions MP2 functions as a prophenoloxidase-activating protease, and that this function is inhibited by Spn27A. MP2 and Spn27A thus constitute a regulatory unit in the prophenoloxidase activation cascade in <i>Drosophila</i>. The combination of genetic, molecular genetic and biochemical approaches should allow further advances in our understanding of the prophenoloxidase-activating cascade in insects and indirectly shed further light on protease-cascades in humans and other vertebrates.</p></div

A model for activation of melanization in <i>Drosophila</i> and comparison with <i>M. sexta</i> and <i>T. molitor</i> melanization pathway [10]–[14], [18], [28], [29], [34], [37], [47], [49], [60], [61].

<p><i>Arrows</i> indicate activation of downstream components or steps. <i>Dashed arrows</i> indicate steps that have not been experimentally verified. Regulation of proteases by serpins is indicted.</p

Seed fate of sound and infested acorns of <i>Q. variabilis</i>, <i>Q. aliena,</i> and <i>Q. serrata var. brevipetiolata</i> after being manipulated by Siberian chipmunks.

<p>A, B, and C indicate eaten in situ (EIS); eaten after dispersal (EAD), and cached after dispersal (CAD). Data are expressed as mean ± SE.</p

Activation of purified proMP2_Xa by bovine Factor Xa.

<p>(<b>A</b>) Purified recombinant proMP2_Xa (350 ng) was incubated with Factor Xa (200 ng) at 37°C for 1 h, and the mixtures were separated by 10% SDS-PAGE followed by immunoblot analysis using Anti-His antibodies (diluted 1∶1000). The sizes and positions of molecular weight standards are indicated on the <i>right</i>. Circle, arrow, and diamond indicated proMP2_Xa, catalytic domain of proMP2_Xa, and non-specific cleavage of proMP2_Xa, respectively. (<b>B</b>) Amidase activity assay of activated MP2_Xa using IEAR<i>p</i>NA as a substrate, as described under “Materials and methods”. The bars represent mean ± S.D. (n = 3). <i>Bars</i> labeled with different letters (<i>a</i>, <i>b</i>, and <i>c</i>) are significantly different (analysis of one-way ANOVA followed by Newman-Keuls test, <i>P</i><0.05).</p

SDS-PAGE analysis of recombinant proMP2_Xa, proMP2, Spn27A and PPO1.

<p>The purified recombinant proMP2_Xa (0.7 µg), proMP2 (0.99 µg) and Spn27A (2.6 µg) were treated with SDS sample buffer containing β-mercaptoethanol and separated by 10% SDS-PAGE followed by coomassie brilliant blue staining. Recombinant PPO1 (1.3 µg) was separated by 7.5% SDS-PAGE. The sizes and positions of the molecular mass standards are indicated on the <i>right</i>.</p

The Effects of Clinical Decision Support Systems on Medication Safety: An Overview

<div><p>Background</p><p>The clinical decision support system(CDSS) has potential to improving medication safety. However, the effects of the intervention were conflicting and uncertain. Meanwhile, the reporting and methodological quality of this field were unknown.</p><p>Objective</p><p>The aim of this overview is to evaluate the effects of CDSS on medication safety and to examine the methodological and reporting quality.</p><p>Methods</p><p>PubMed, Embase and Cochrane Library were searched to August 2015. Systematic reviews (SRs) investigating the effects of CDSS on medication safety were included. Outcomes were determined in advance and assessed separately for process of care and patient outcomes. The methodological quality was assessed by Assessment of Multiple Systematic Reviews (AMSTAR) and the reporting quality was examined by Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA).</p><p>Results</p><p>Twenty systematic reviews, consisting of 237 unique randomized controlled trials(RCTs) and 176 non-RCTs were included. Evidence that CDSS significantly impacted process of care was found in 108 out of 143 unique studies of the 16 SRs examining this effect (75%). Only 18 out of 90 unique studies of the 13 SRs reported significantly evidence that CDSS positively impacted patient outcomes (20%). Ratings for the overall scores of AMSTAR resulted in a mean score of 8.3 with a range of scores from 7.5 to 10.5. The reporting quality was varied. Some contents were particularly strong. However, some contents were poor.</p><p>Conclusions</p><p>CDSS reduces medication error by obviously improving process of care and inconsistently improving patient outcomes. Larger samples and longer-term studies are required to ensure more reliable evidence base on the effects of CDSS on patient outcomes. The methodological and reporting quality were varied and some realms need to be improved.</p></div

The results of methodological quality based on AMSTAR.

<p>The results of methodological quality based on AMSTAR.</p

The results of reporting quality assessment.

<p>The results of reporting quality assessment.</p

<i>Drosophila</i> Spn27A binds and inhibits MP2.

<p>SDS-stable complex formation between MP2_Xa and recombinant Spn27A (<b>A</b>) or native Spn27A in <i>Drosophila</i> hemolymph (<b>B</b>). ProMP2_Xa (350 ng) was activated by Factor Xa and then incubated for 30 min at room temperature with 1-fold molar of recombinant Spn27A or 3 µl of hemolymph. In control reactions, proMP2_Xa or Factor Xa was omitted, or proMP2 was used instead. The samples were subjected to SDS-PAGE and immunoblot analysis using mouse anti-His (<i>Left</i>) or rabbit anti-Spn27A (<i>Right</i>) antibodies. The notes used in the figure were: circle, proMP2_Xa; arrow, catalytic domain of proMP2_Xa; diamond, non-specific cleavage of proMP2_Xa; triangles, non-complexed Spn27A; asterisks, Spn27A-MP2_Xa complex. (<b>C</b>) Stoichiometry of inhibition of MP2 by Spn27A. Recombinant Spn27A was incubated with Factor Xa-activated MP2_Xa at different molar ratios for 30 min at room temperature. The residual amidase activity was measured using IEAR<i>p</i>NA as substrate, and plotted as mean ± S.D. (n = 3) against the corresponding molar ratios of Spn27A and MP2_Xa.</p