Caryophyllene Sesquiterpenes from a Chaetomium globosum Endophyte of the Canadian Medicinal Plant Empetrum nigrum.

Punctaporonins T (1) and U (2), new caryophyllene sesquiterpenes, were isolated with three known punctaporonins, A (3), B (4), and C (5), from the endophytic fungus Chaetomium globosum (TC2-041). The structures and relative configurations of punctaporonins T and U were elucidated based on a combination of HRESIMS, 1D/2D NMR spectroscopic analysis, and X-ray diffraction analysis, while their absolute configuration is presumed to be consistent with the co-isolated 3-5 on biogenetic arguments. Compound 1 showed weak inhibitory activity against both Mycobacterium tuberculosis and Staphylococcus aureus.

Tolypocaibols: Antibacterial Lipopeptaibols from a Tolypocladium sp. Endophyte of the Marine Macroalga Spongomorpha arcta.

Two new lipopeptaibols, tolypocaibols A (1) and B (2), and the mixed NRPS-polyketide-shikimate natural product maximiscin [(P/M)-3)] were isolated from a Tolypocladium sp. fungal endophyte of the marine alga Spongomorpha arcta. Analysis of NMR and mass spectrometry data revealed the amino acid sequences of the lipopeptaibols, which both comprise 11 residues with a valinol C-terminus and a decanoyl acyl chain at the N-terminus. The configuration of the amino acids was determined by Marfey's analysis. Tolypocaibols A (1) and B (2) showed moderate, selective inhibition against Gram-positive and acid-fast bacterial strains, while maximiscin [(P/M)-3)] showed moderate, broad-spectrum antibiotic activity.

Pandemic practice: Horror fans and morbidly curious individuals are more psychologically resilient during the COVID-19 pandemic.

One explanation for why people engage in frightening fictional experiences is that these experiences can act as simulations of actual experiences from which individuals can gather information and model possible worlds. Conducted during the COVID-19 pandemic, this study (n = 310) tested whether past and current engagement with thematically relevant media fictions, including horror and pandemic films, was associated with greater preparedness for and psychological resilience toward the pandemic. Since morbid curiosity has previously been associated with horror media use during the COVID-19 pandemic, we also tested whether trait morbid curiosity was associated with pandemic preparedness and psychological resilience during the COVID-19 pandemic. We found that fans of horror films exhibited greater resilience during the pandemic and that fans of "prepper" genres (alien-invasion, apocalyptic, and zombie films) exhibited both greater resilience and preparedness. We also found that trait morbid curiosity was associated with positive resilience and interest in pandemic films during the pandemic. Taken together, these results are consistent with the hypothesis that exposure to frightening fictions allow audiences to practice effective coping strategies that can be beneficial in real-world situations.

ß-arrestin-biased agonism of ß-adrenergic receptor regulates Dicer-mediated microRNA maturation to promote cardioprotective signaling.

RATIONALE: MicroRNAs (miRs) are small, non-coding RNAs that function to post-transcriptionally regulate target genes. First transcribed as primary miR transcripts (pri-miRs), they are enzymatically processed by Drosha into premature miRs (pre-miRs) and further cleaved by Dicer into mature miRs. Initially discovered to desensitize ß-adrenergic receptor (ßAR) signaling, ß-arrestins are now well-appreciated to modulate multiple pathways independent of G protein signaling, a concept known as biased signaling. Using the ß-arrestin-biased ßAR ligand carvedilol, we previously showed that ß-arrestin1 (not ß-arrestin2)-biased ß1AR (not ß2AR) cardioprotective signaling stimulates Drosha-mediated processing of six miRs by forming a multi-protein nuclear complex, which includes ß-arrestin1, the Drosha microprocessor complex and a single-stranded RNA binding protein hnRNPA1. OBJECTIVE: Here, we investigate whether ß-arrestin-mediated ßAR signaling induced by carvedilol could regulate Dicer-mediated miR maturation in the cytoplasm and whether this novel mechanism promotes cardioprotective signaling. METHODS AND RESULTS: In mouse hearts, carvedilol indeed upregulates three mature miRs, but not their pre-miRs and pri-miRs, in a ß-arrestin 1- or 2-dependent manner. Interestingly, carvedilol-mediated activation of miR-466g or miR-532-5p, and miR-674 is dependent on ß2ARs and ß1ARs, respectively. Mechanistically, ß-arrestin 1 or 2 regulates maturation of three newly identified ßAR/ß-arrestin-responsive miRs (ß-miRs) by associating with the Dicer maturation RNase III enzyme on three pre-miRs of ß-miRs. Myocardial cell approaches uncover that despite their distinct roles in different cell types, ß-miRs act as gatekeepers of cardiac cell functions by repressing deleterious targets. CONCLUSIONS: Our findings indicate a novel role for ßAR-mediated ß-arrestin signaling activated by carvedilol in Dicer-mediated miR maturation, which may be linked to its protective mechanisms.

Determining the mode of action of anti-mycobacterial C17 diyne natural products using expression profiling: evidence for fatty acid biosynthesis inhibition.

BACKGROUND: The treatment of microbial infections is becoming increasingly challenging because of limited therapeutic options and the growing number of pathogenic strains that are resistant to current antibiotics. There is an urgent need to identify molecules with novel modes of action to facilitate the development of new and more effective therapeutic agents. The anti-mycobacterial activity of the C17 diyne natural products falcarinol and panaxydol has been described previously; however, their mode of action remains largely undetermined in microbes. Gene expression profiling was therefore used to determine the transcriptomic response of Mycobacterium smegmatis upon treatment with falcarinol and panaxydol to better characterize the mode of action of these C17 diynes. RESULTS: Our analyses identified 704 and 907 transcripts that were differentially expressed in M. smegmatis after treatment with falcarinol and panaxydol respectively. Principal component analysis suggested that the C17 diynes exhibit a mode of action that is distinct to commonly used antimycobacterial drugs. Functional enrichment analysis and pathway enrichment analysis revealed that cell processes such as ectoine biosynthesis and cyclopropane-fatty-acyl-phospholipid synthesis were responsive to falcarinol and panaxydol treatment at the transcriptome level in M. smegmatis. The modes of action of the two C17 diynes were also predicted through Prediction of Activity Spectra of Substances (PASS). Based upon convergence of these three independent analyses, we hypothesize that the C17 diynes inhibit fatty acid biosynthesis, specifically phospholipid synthesis, in mycobacteria. CONCLUSION: Based on transcriptomic responses, it is suggested that the C17 diynes act differently than other anti-mycobacterial compounds in M. smegmatis, and do so by inhibiting phospholipid biosynthesis.

ß-arrestin1-biased ß1-adrenergic receptor signaling regulates microRNA processing.

RATIONALE: MicroRNAs (miRs) are small, noncoding RNAs that function to post-transcriptionally regulate gene expression. First transcribed as long primary miR transcripts (pri-miRs), they are enzymatically processed in the nucleus by Drosha into hairpin intermediate miRs (pre-miRs) and further processed in the cytoplasm by Dicer into mature miRs where they regulate cellular processes after activation by a variety of signals such as those stimulated by ß-adrenergic receptors (ßARs). Initially discovered to desensitize ßAR signaling, ß-arrestins are now appreciated to transduce multiple effector pathways independent of G-protein-mediated second messenger accumulation, a concept known as biased signaling. We previously showed that the ß-arrestin-biased ßAR agonist, carvedilol, activates cellular pathways in the heart. OBJECTIVE: Here, we tested whether carvedilol could activate ß-arrestin-mediated miR maturation, thereby providing a novel potential mechanism for its cardioprotective effects. METHODS AND RESULTS: In human cells and mouse hearts, carvedilol upregulates a subset of mature and pre-miRs, but not their pri-miRs, in ß1AR-, G-protein-coupled receptor kinase 5/6-, and ß-arrestin1-dependent manner. Mechanistically, ß-arrestin1 regulates miR processing by forming a nuclear complex with hnRNPA1 and Drosha on pri-miRs. CONCLUSIONS: Our findings indicate a novel function for ß1AR-mediated ß-arrestin1 signaling activated by carvedilol in miR biogenesis, which may be linked, in part, to its mechanism for cell survival.

δPKC interaction with the d subunit of F1Fo ATP synthase impairs energetics and exacerbates ischemia/reperfusion injury in isolated rat hearts.

Previously, we demonstrated protection against hypoxic injury in neonatal cardiac myocytes and reduced release of cardiac troponin I from perfused rat hearts by a novel peptide inhibitor [NH2-YGRKKRRQRRRMLATRALSLIGKRAISTSVCAGRKLALKTIDWVSFDYKDDDDK-] of the delta protein kinase C (δPKC) interaction with the "d" subunit of mitochondrial F1Fo ATP synthase (dF1Fo). This peptide was developed in our laboratory and contains: an HIV-Tat protein transduction domain; a mitochondrial targeting motif; the δPKC-dF1Fo inhibitor sequence; and a FLAG epitope. In the present study the δPKC-dF1Fo inhibitor attenuated co-immunoprecipitation of δPKC with dF1Fo, improved recovery of contractility, diminished levels of tissue t-carbonyls and 4-hydroxy-2-nonenal (HNE), and reduced 2,3,5-triphenyltetrazolium chloride-monitored infarct size following simulated global ischemia/reperfusion (IR) exposures. Perfusion of hearts with this peptide prior to IR enhanced ATP levels 2.1-fold, improved ADP (state 3)- and FCCP (maximal)-stimulated respiration in mitochondrial oxygen consumption assays, and attenuated Ca(++)-induced mitochondrial swelling following ischemic injury. Mitochondrial membrane potential (assessed by JC-1) was also improved 1.6-fold by the inhibitor in hearts subsequently exposed to IR injury. Brief IR exposures did not cause mitochondrial loss of cytochrome c in the presence or absence of the inhibitor. Additionally, the inhibitor did not modify accumulation of the autophagy marker LC3II after brief IR injury. Our results support the potential for this first-in-class peptide as a translational agent for combating cardiac IR injury.

Dibenz[b,f]oxepin and Antimycobacterial Chalcone Constituents of Empetrum nigrum.

Two new dibenz[b,f]oxepins, empetroxepins A and B (1 and 2), and seven known compounds (3-9) were isolated from an extract of the Canadian medicinal plant Empetrum nigrum that significantly inhibited the growth of Mycobacterium tuberculosis H37Ra. The structures of 1 and 2 were established through analysis of NMR and MS data. The antimycobacterial activity of the plant extract was attributed primarily to the presence of two chalcone derivatives (6 and 7) that exhibited selective antimycobacterial activity (IC50 values of 23.8 and 32.8 µM, respectively) in comparison to mammalian (HEK 293) cells (IC50 values of 109 and 249 µM, respectively).

Optimisation of the microplate resazurin assay for screening and bioassay-guided fractionation of phytochemical extracts against Mycobacterium tuberculosis.

INTRODUCTION: Because of increased resistance to current drugs, there is an urgent need to discover new anti-mycobacterial compounds for the development of novel anti-tuberculosis drugs. The microplate resazurin assay (MRA) is commonly used to evaluate natural products and synthetic compounds for anti-mycobacterial activity. However, the assay can be problematic and unreliable when screening methanolic phytochemical extracts. OBJECTIVE: To optimise the MRA for the screening and bioassay-guided fractionation of phytochemical extracts using Mycobacterium tuberculosis H37Ra. METHODS: The effects of varying assay duration, resazurin solution composition, solvent (dimethyl sulphoxide - DMSO) concentration and type of microtitre plate used on the results and reliability of the MRA were investigated. The optimal bioassay protocol was applied to methanolic extracts of medicinal plants that have been reported to possess anti-mycobacterial activity. RESULTS: The variables investigated were found to have significant effects on the results obtained with the MRA. A standardised procedure that can reliably quantify anti-mycobacterial activity of phytochemical extracts in as little as 48 h was identified. The optimised MRA uses 2% aqueous DMSO, with an indicator solution of 62.5 µg/mL resazurin in 5% aqueous Tween 80 over 96 h incubation. CONCLUSION: The study has identified an optimal procedure for the MRA when used with M. tuberculosis H37Ra that gives rapid, reliable and consistent results. The assay procedure has been used successfully for the screening and bioassay-guided fractionation of anti-mycobacterial compounds from methanol extracts of Canadian medicinal plants.

Quantitative Measurement of Dissection Resistance in Intimal and Medial Layers of Human Coronary Arteries.

The left anterior descending (LAD) coronary artery is the most frequently involved vessel in coronary artery dissection, a cause of acute coronary syndrome or sudden cardiac death. The biomechanical mechanisms underlying arterial dissection are not well understood. This study investigated the dissection properties of LAD specimens harvested from explanted hearts at the time of cardiac transplantation, from patients with primary dilated cardiomyopathy (n=12). Using a previously validated approach uniquely modified for these human LAD specimens, we quantified the local energy release rate, G, within different arterial layers during experimental dissection events (tissue tearing). Results show that the mean values of G during arterial dissection within the intima and within the media in human LADs are 20.7±16.5 J/m2 and 10.3±5.0 J/m2, respectively. The difference in dissection resistance between tearing events occurring within the intima and within the media is statistically significant. Our data fall in the same order of magnitude as most previous measurements of adhesive strength in other human arteries, with the differences in measured values of G within the layers most likely due to histologically observed differences in the structure and composition of arterial layers.

Isolation and bioassay screening of medicinal plant endophytes from eastern Canada.

Eighty-one distinct fungal endophytes were isolated from 12 traditionally used medicinal plants from New Brunswick, Canada. This is the first report of endophytes from 8 of the 12 host plants. One hundred and sixty-two crude extracts derived from the mycelia and spent fermentation broths of liquid cultures of each endophyte were screened for antibacterial and antifungal activity. Twenty-two extracts were active against Staphylococcus aureus while 30 were active against Pseudomonas aeruginosa. Twelve crude extracts were found to be active against Candida albicans.

Modulation of the protein kinase Cdelta interaction with the "d" subunit of F1F0-ATP synthase in neonatal cardiac myocytes: development of cell-permeable, mitochondrially targeted inhibitor and facilitator peptides.

The F(1)F(0)-ATP synthase provides approximately 90% of cardiac ATP, yet little is known regarding its regulation under normal or pathological conditions. Previously, we demonstrated that protein kinase Cdelta (PKCdelta) inhibits F(1)F(0) activity via an interaction with the "d" subunit of F(1)F(0)-ATP synthase (dF(1)F(0)) in neonatal cardiac myocytes (NCMs) (Nguyen, T., Ogbi, M., and Johnson, J. A. (2008) J. Biol. Chem. 283, 29831-29840). We have now identified a dF(1)F(0)-derived peptide (NH(2)-(2)AGRKLALKTIDWVSF(16)-COOH) that inhibits PKCdelta binding to dF(1)F(0) in overlay assays. We have also identified a second dF(1)F(0)-derived peptide (NH(2)-(111)RVREYEKQLEKIKNMI(126)-COOH) that facilitates PKCdelta binding to dF(1)F(0). Incubation of NCMs with versions of these peptides containing HIV-Tat protein transduction and mammalian mitochondrial targeting sequences resulted in their delivery into mitochondria. Preincubation of NCMs, with 10 nm extracellular concentrations of the mitochondrially targeted PKCdelta-dF(1)F(0) interaction inhibitor, decreased 100 nm 4beta-phorbol 12-myristate 13-acetate (4beta-PMA)-induced co-immunoprecipitation of PKCdelta with dF(1)F(0) by 50 +/- 15% and abolished the 30 nm 4beta-PMA-induced inhibition of F(1)F(0)-ATPase activity. A scrambled sequence (inactive) peptide, which contained HIV-Tat and mitochondrial targeting sequences, was without effect. In contrast, the cell-permeable, mitochondrially targeted PKCdelta-dF(1)F(0) facilitator peptide by itself induced the PKCdelta-dF(1)F(0) co-immunoprecipitation and inhibited F(1)F(0)-ATPase activity. In in vitro PKC add-back experiments, the PKCdelta-F(1)F(0) inhibitor blocked PKCdelta-mediated inhibition of F(1)F(0)-ATPase activity, whereas the facilitator induced inhibition. We have developed the first cell-permeable, mitochondrially targeted modulators of the PKCdelta-dF(1)F(0) interaction in NCMs. These novel peptides will improve our understanding of cardiac F(1)F(0) regulation and may have potential as therapeutics to attenuate cardiac injury.

Attenuation of the hypoxia-induced protein kinase Cdelta interaction with the 'd' subunit of F1Fo-ATP synthase in neonatal cardiac myocytes: implications for energy preservation and survival.

The F1Fo-ATP synthase provides most of the heart's energy, yet events that alter its function during injury are poorly understood. Recently, we described a potent inhibitory effect on F1Fo-ATP synthase function mediated by the interaction of PKCdelta (protein kinase Cdelta) with dF1Fo ('d' subunit of the F1Fo-ATPase/ATP synthase). We have now developed novel peptide modulators which facilitate or inhibit the PKCdelta-dF1Fo interaction. These peptides include HIV-Tat (transactivator of transcription) protein transduction and mammalian mitochondrial-targeting sequences. Pre-incubation of NCMs (neonatal cardiac myocyte) with 10 nM extracellular concentrations of the mitochondrial-targeted PKCdelta-dF1Fo interaction inhibitor decreased Hx (hypoxia)-induced co-IP (co-immunoprecipitation) of PKCdelta with dF1Fo by 40+/-9%, abolished Hx-induced inhibition of F1Fo-ATPase activity, attenuated Hx-induced losses in F1Fo-derived ATP and protected against Hx- and reperfusion-induced cell death. A scrambled-sequence (inactive) peptide, which contained HIV-Tat and mitochondrial-targeting sequences, was without effect. In contrast, the cell-permeant mitochondrial-targeted PKCdelta-dF1Fo facilitator peptide, which we have shown previously to induce the PKCdelta-dF1Fo co-IP, was found to inhibit F1Fo-ATPase activity to an extent similar to that caused by Hx alone. The PKCdelta-dF1Fo facilitator peptide also decreased ATP levels by 72+/-18% under hypoxic conditions in the presence of glycolytic inhibition. None of the PKCdelta-dF1Fo modulatory peptides altered the inner mitochondrial membrane potential. Our studies provide the first evidence that disruption of the PKCdelta-dF1Fo interaction using cell-permeant mitochondrial-targeted peptides attenuates cardiac injury resulting from prolonged oxygen deprivation.

Electrophysiological ventricular substrate of stroke: a prospective cohort study in the Atherosclerosis Risk in Communities (ARIC) study.

OBJECTIVES: The goal of the study was to determine an association of cardiac ventricular substrate with thrombotic stroke (TS), cardioembolic stroke (ES) and intracerebral haemorrhage (ICH). DESIGN: Prospective cohort study. SETTING: The Atherosclerosis Risk in Communities (ARIC) study in 1987-1989 enrolled adults (45-64 years), selected as a probability sample from four US communities (Minneapolis, Minnesota; Washington, Maryland; Forsyth, North Carolina; Jackson, Mississippi). Visit 2 was in 1990-1992, visit 3 in 1993-1995, visit 4 in 1996-1998 and visit 5 in 2011-2013. PARTICIPANTS: ARIC participants with analysable ECGs and no history of stroke were included (n=14 479; age 54±6 y; 55% female; 24% black). Ventricular substrate was characterised by cardiac memory, spatial QRS-T angle (QRS-Ta), sum absolute QRST integral (SAIQRST), spatial ventricular gradient magnitude (SVGmag), premature ventricular contractions (PVCs) and tachycardia-dependent intermittent bundle branch block (TD-IBBB) on 12-lead ECG at visits 1-5. OUTCOME: Adjudicated TS included a first definite or probable thrombotic cerebral infarction, ES-a first definite or probable non-carotid cardioembolic brain infarction. Definite ICH was included if it was the only stroke event. RESULTS: Over a median 24.5 years follow-up, there were 899 TS, 400 ES and 120 ICH events. Cox proportional hazard risk models were adjusted for demographics, cardiovascular disease, risk factors, atrial fibrillation, atrial substrate and left ventricular hypertrophy. After adjustment, PVCs (HR 1.72; 95% CI 1.02 to 2.92), QRS-Ta (HR 1.15; 95% CI 1.03 to 1.28), SAIQRST (HR 1.20; 95% CI 1.07 to 1.34) and time-updated SVGmag (HR 1.19; 95% CI 1.08 to 1.32) associated with ES. Similarly, PVCs (HR 1.53; 95% CI 1.03 to 2.26), QRS-Ta (HR 1.08; 95% CI 1.01 to 1.16), SAIQRST (HR 1.07; 95% CI 1.01 to 1.14) and time-updated SVGmag (HR 1.11; 95% CI 1.04 to 1.19) associated with TS. TD-IBBB (HR 3.28; 95% CI 1.03 to 10.46) and time-updated SVGmag (HR 1.23; 95% CI 1.03 to 1.47) were associated with ICH. CONCLUSIONS: PVC burden (reflected by cardiac memory) is associated with ischaemic stroke. Transient cardiac memory (likely through TD-IBBB) precedes ICH.

Antimycobacterial screening of traditional medicinal plants using the microplate resazurin assay.

Multidrug-resistant Mycobacterium tuberculosis strains have rapidly become a global health concern. North American First Nations communities have used traditional medicines for generations to treat many pulmonary infections. In this study, we evaluated the antimycobacterial activity of 5 medicinal plants traditionally used as general therapeutics for pulmonary illnesses and specifically as treatments for tuberculosis. Aqueous extracts of Aralia nudicaulis, Symplocarpus foetidus, Heracleum maximum, Juniperus communis, and Acorus calamus were screened for antimycobacterial activity against Bacillus Calmette-Guérin, Mycobacterium avium, and M. tuberculosis H37Ra using the colorimetric microplate resazurin assay. Extracts of Acorus calamus and H. maximum root demonstrated significant antimycobacterial activity comparable to that of the rifampin control (2 microg/mL). Evaluation of the cytotoxicity of these 2 extracts using the MTT assay also showed that the extracts were less toxic to 3 human cell lines than was the DMSO positive control. This study demonstrates that aqueous extracts of the roots of H. maximum and Acorus calamus possess strong in vitro antimycobacterial activity, validates traditional knowledge, and provides potential for the development of urgently needed novel antituberculous therapeutics.

Evidence of Inverse Hall-Petch Behavior and Low Friction and Wear in High Entropy Alloys.

We present evidence of inverse Hall-Petch behavior for a single-phase high entropy alloy (CoCrFeMnNi) in ultra-high vacuum and show that it is associated with low friction coefficients (~0.3). Grain size measurements by STEM validate a recently proposed dynamic amorphization model that accurately predicts grain size-dependent shear strength in the inverse Hall-Petch regime. Wear rates in the initially soft (coarse grained) material were shown to be remarkably low (~10-6 mm3/N-m), the lowest for any HEA tested in an inert environment where oxidation and the formation of mixed metal-oxide films is mitigated. The combined high wear resistance and low friction are linked to the formation of an ultra-nanocrystalline near-surface layer. The dynamic amorphization model was also used to predict an average high angle grain boundary energy (0.87 J/m2). This value was used to explain cavitation-induced nanoporosity found in the highly deformed surface layer, a phenomenon that has been linked to superplasticity.

Assessing the Structure of the Five Factor Model of Personality (IPIP-NEO-120) in the Public Domain.

Assessment of individual differences in personality traits is arguably one of the hallmarks of psychological research. Testing the structural validity of trait measurements is paramount in this endeavor. In the current study, we investigated 30 facet traits in one of the accessible and comprehensive public-domain Five Factor Model (FFM) personality inventories, IPIP-NEO-120 (Johnson, 2014), using one of the largest US samples to date (N = 320,128). We present structural loadings for all trait facets organized into respective FFM-trait domain (Neuroticism, Extraversion, Openness, Agreeableness, and Conscientiousness). Both hierarchical second-order and bi-factor models showed tolerable model fit indices, using confirmatory factor analysis in a structural equation modeling (SEM) framework. Some facet traits were substantially more representative than others for their respective trait domain, which facilitate further discussions on FFM-construct content. We conclude that IPIP-NEO is sufficiently structurally robust for future use, for the benefit of research and practice in personality assessment.

Protein precoating of elastomeric tissue-engineering scaffolds increased cellularity, enhanced extracellular matrix protein production, and differentially regulated the phenotypes of circulating endothelial progenitor cells.

BACKGROUND: Optimal cell sources and scaffold-cell interactions remain unanswered questions for tissue engineering of heart valves. We assessed the effect of different protein precoatings on a single scaffold type (elastomeric poly (glycerol sebacate)) with a single cell source (endothelial progenitor cells). METHODS AND RESULTS: Elastomeric poly (glycerol sebacate) scaffolds were precoated with laminin, fibronectin, fibrin, collagen types I/III, or elastin. Characterized ovine peripheral blood endothelial progenitor cells were seeded onto scaffolds for 3 days followed by 14 days incubation. Endothelial progenitor cells were CD31+, vWF+, and alpha-SMA- before seeding confirmed by immunohistochemistry and immunoblotting. Both precoated and uncoated scaffolds demonstrated surface expression of CD31+ and vWF+, alpha-SMA+ cells and were found in the "interstitium" of the scaffold. Protein precoating of elastomeric poly (glycerol sebacate) scaffolds revealed significantly increased cellularity and altered the phenotypes of endothelial progenitor cells, which resulted in changes in cellular behavior and extracellular matrix production. Moreover, mechanical flexure testing demonstrated decreased effective stiffness of the seeded scaffolds compared with unseeded controls. CONCLUSIONS: Scaffold precoating with extracellular matrix proteins can allow more precise "engineering" of cellular behavior in the development of tissue engineering of heart valves constructs by altering extracellular matrix production and cell phenotype.