TSP-1 inhibited late EPCs tubule formation on Matrigel.

<p>(A) Late EPCs were plated on Matrigel with TSP-1 at different concentrations for 8 hrs (Scale bar = 200 µm, 100×magnification). (B) Quantification of total tube length per high power field (HPF, 100×magnification) was presented as mean±S.D. of three independent experiments (#p<0.05 versus no TSP-1 intervention, *p<0.01 versus no TSP-1 intervention).</p

Clinical characteristics and serum TSP-1 level in all patients.

<p>LVEF indicates left ventricular ejection fraction, previous MI indicates previous myocardial infarction,eGFR indicates estimated glomerular filtration rate, TC indicates total cholesterol, TG indicates total triglyceride, LDL-C indicates low-density lipoprotein-cholesterol, HDL-C indicates high-density lipoprotein-cholesterol.</p>*<p>p<0.01 compared with control, #p<0.05 compared with control,$<0.05 compared with Collateral group I.</p

TSP-1 inhibits VEGF induced VEGFR2 phosphorylation through CD47.

<p>(A) Late EPCs were treated with VEGF (25 ng/ml) for 0, 5, 10, 15 min respectively. Total protein was extracted and the expression of FLK-1, phospho-VEGFR2 (Tyr1175) was determined by Western blot. (B)Seventy-two hrs after transfection of CD47-specific siRNA or control siRNA, the expression of CD47 in late EPCs was determined by western blotting analysis in three independent experiments (#p<0.05 versus control siRNA). (C)Seventy-two hrs after transfection of CD47-specific siRNA or control siRNA, late EPCs were treated with TSP-1(2 µg/ml) for 30 min and then VEGF(25 ng/ml) for 5 min. Total protein was extracted and the expression of FLK-1, phospho-VEGFR2 (Tyr1175) was determined by Western blot analysis. (D) Quantification of VEGFR2 phosphorylation normalized to VEGFR2 in three independent experiments(#p<0.05 versus control siRNA, *p<0.01 versus control siRNA).</p

Flow cytometric analysis of CD34+ cells and morphological and immunophenotypical characterization of early and late EPCs.

<p>(A) Flow cytometric analysis of CD34 expression after isolation by anti-CD34 microbeads. Shown are representative data from 3 independent experiments using cells isolated from different cord blood with similar results. Isotype controls are used. (B) Early EPCs cultured for 7 days and late EPCs cultured for 14 days (Scale bar = 100 µm, 200×magnification). (C)Early EPCs are shown to uptake DiI-Ac-LDL(red) (Scale bar = 100 µm, 200×magnification). Immunocytochemistry of VEGFR2(red),CD31 (red), and DAPI(blue) was demonstrated in early EPCs (Scale bar = 50 µm,400×magnification). (D) Immunocytochemistry of VEGFR2(red), vWF(green), CD31(red),and DAPI(blue) was demonstrated in late EPCs(Scale bar = 50 µm,400×magnification). Shown are representative data from 3 independent experiments using early EPCs isolated from different cord blood and 3 independent experiments using late EPCs isolated from different cord blood.</p

CD47 antibody attenuated TSP-1′s inhibition on angiogenesis of late EPCs.

<p>(A) Late EPCs were plated on Matrigel and treated with anti-CD47 antibody (2.5 µg/ml), anti-integrin β1 antibody (2 µg/ml) or control IgG for 30 min and then treated with TSP-1(2 µg/ml) as described (Scale bar = 400 µm, 50×magnification). (B) Total tube length per HPF (100×magnification) was measured. Values are presented as the mean±S.D. of three independent experiments (#p<0.05 versus control siRNA with TSP-1 stimulation)</p

TSP-1 inhibited early EPCs incorporation into tube-like structure.

<p>(A) Early EPCs pretreated with TSP-1 at different concentrations for 1 hr were labeled with a DiI fluorescent marker(red) and coplated with HUVECs(transparent) to form tubule structures on Matrigel(Scale bar = 200 µm,100×magnification). (B–D) Early EPCs pretreated with TSP-1 at different concentrations for 1 hr(B), 6 hrs(C), 12 hrs(D) were coplated with HUVECs on Matrigel. Quantifications of incorporated EPCs per field were presented as mean±S.D of three independent experiments (#p<0.05 versus no TSP-1 intervention).</p

Time Course of Current of Injury Is Related to Acute Stability of Active-Fixation Pacing Leads in Rabbits

<div><p>Background</p><p>Magnitude of current of injury (COI) consequent to pacemaker lead fixation is recognized as a predictor of acute lead stability. It is unclear whether dynamic monitoring of COI after lead fixation provides additional information beyond a single assessment performed at the time of fixation.</p> <p>Objectives</p><p>This study was aimed to test the hypothesis that the time course of COI is related to acute lead stability.</p> <p>Methods and Results</p><p>Active fixation leads with fixed screw were anchored to either Langendorff-perfused rabbit hearts endocardially or in vivo hearts epicardially in manners of contact the helix with no rotation, half rotation and full rotation, respectively. Intracardiac electrogram (EGM) was monitored dynamically from onset to resolution of COI, and magnitudes of intrinsic R wave and COI, including ST-segment elevation, ST/R and intracardiac EGM duration (IED), were measured. A digital force gauge was applied to assess lead stability. In vitro, COI in contacted leads was significantly smaller than those in half rotated (<i>p</i><0.05) and fully rotated leads (<i>p</i><0.05), and presented most precipitous recovery to baseline (1.5±1.1 min, <i>p</i><0.05). Half-rotated and fully rotated leads manifested the same magnitude of COI right after placement. However, the time course of COI was significantly longer in fully rotated leads than that in half rotated leads (26.5±2.8 min <i>vs.</i> 5.6±2.0 min, <i>p</i><0.05). Similar findings were observed in vivo. The time course of COI was significantly correlated with the force needed to detach the lead from myocardium (<i>r</i> = 0. 72, n = 48, <i>p</i><0.001).</p> <p>Conclusions</p><p>Time course of COI is related to acute lead stability in rabbits. One might be misled by a single assessment of COI magnitude right after lead placement, whereas persistence of COI is likely to be a useful indicator of adequate lead stability.</p> </div

Correlation between COI time course and acute Lead Stability.

<p>COI time course from onset to resolution was significantly correlated with the force in order to detach the lead from myocardium (r = 0.72, n = 48, p<0.001).</p

Comparison of Intracardiac EGM variables derived from isolated rabbit hearts.

<p>Panel A. Intrinsic R wave amplitude showed no significant dissimilarity regarding to different lead positionings. Panel B. Contacted leads presented the smallest magnitude of ST-segment elevation with the most rapid decline, followed by half rotated leads, while fully rotated leads showed the biggest COI amplitude and the slowest recovery. Note that there was no difference between half and fully rotated leads at 0 min. Panel C and D depicted the same findings as Panel B in the value of ST/R and IED, respectively. *: <i>p</i><0.05 (compared with half and fully rotated leads); ^†: <i>p</i><0.05 and ^‡: <i>p</i><0.01 (compared with fully rotated leads).</p

Comparison of Intracardiac EGM variables derived from in vivo hearts.

<p>Panel A, B and C showed the similar findings observed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0057727#pone-0057727-g003" target="_blank">Figure 3</a>. Panel D. No difference in IED between half and fully rotated leads at any time point. *: <i>p</i><0.05 and ^†: <i>p</i><0.01 (compared with half and fully rotated leads); ^‡: <i>p</i><0.05 and: §: <i>p</i><0.01 (compared with fully rotated leads).</p