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Non-vitamin K antagonist oral anticoagulants (NOACs) represent a paradigm shift in the treatment of non-valvular atrial fibrillation (AF) with major practice guidelines around the world recommending NOACs over vitamin K antagonist oral anticoagulants for initial treatment of AF for stroke prevention. Here we describe the evidence collated and the process followed for the successful inclusion of NOACs into the 21st WHO Model List of Essential Medicines (EML). Individual NOACs have been reported to be non-inferior or superior to warfarin in preventing stroke and systemic embolism in eligible AF patients with a reduction in the risk of stroke and systemic embolism and a lower risk of major bleeding in patients with non-valvular AF compared with warfarin in both RCTs and real-world data. The successful inclusion of NOACs in the WHO EML is an important step forward in the global fight against cardiovascular morbidity and mortality, especially in low- and middle-income countries, where the burden of disease is high and limited access to diagnosis and treatment translates into a higher burden of morbidity, mortality, and economic costs., Competing Interests: MDH has received support from the American Heart Association, Verily, and AstraZeneca for work unrelated to this research. MDH has received salary support from the American Medical Association for his role as an associate editor for JAMA Cardiology. The George Institute for Global Health has a patent and license and has received investment funding with intent to commercialize fixed-dose combination therapy through its social enterprise business, George Medicines. LN received speakers fees from Daiichi Sankyo unrelated to this research. EJZ received speakers fees from Novartis, Pfizer and Bayer unrelated to this research. The author alone is responsible for the views expressed in this article, which do not necessarily represent the views, decisions or policies of the institution., (Copyright: © 2020 The Author(s).)

High adipose tissue (AT) accumulation in the body increases the risk for many metabolic and chronic diseases. This work investigated the capacity of the flavonoid (-)-epicatechin to prevent undesirable modifications of AT in mice fed a high-fat diet. Studies were focused on thoracic aorta perivascular AT (taPVAT), which is involved in the control of blood vessel tone, among other functions. Male C57BL/6J mice were fed for 15 weeks a high-fat diet with or without added (-)-epicatechin (20 mg per kg body weight per d). In high-fat diet fed mice, (-)-epicatechin supplementation: (i) prevented the expansion of taPVAT, (ii) attenuated the whitening of taPVAT (according to the adipocyte morphology, diameter, and uncoupling-protein 1 (UCP-1) levels) and (iii) blunted the increase in plasma glucose and cholesterol. The observed taPVAT modifications were not associated with alterations in the aorta wall thickness, aorta tumor necrosis factor-alpha (TNF-α) and NADPH-oxidase 2 (NOX2) expression, and endothelial nitric oxide synthase (eNOS) phosphorylation levels. In summary, our results indicate (-)-epicatechin as a relevant bioactive protecting from the slow and silent development of metabolic and chronic diseases as they are associated with excessive fat intake.

Background: Women differ from men in their left ventricular (LV) structure, function and remodeling with age and diseases. The LV assist device (LVAD) unloads the LV and reversely remodels the heart. We sought to define the effects of sex on longitudinal reverse remodeling after LVAD implantation., Methods and Results: Cardiac structure and function were assessed by serial echocardiograms. Mixed effect regression models were constructed to assess the independent contribution of sex to longitudinal changes in cardiac structure and function. A total of 355 consecutive patients with advanced heart failure received continuous flow LVADs between 2006 and 2016. The average age was 56 ± 13 years, 73% were men, and 67% were black. Early (within 3 months) after LVAD implantation, women had a greater reduction in LV dimensions and a greater increase in LV ejection fraction compared with men. These differences were independent of age, body surface area, device type, or ischemic etiology of heart failure. At long-term follow-up, LV dimensions increased slightly over time in women compared with men, but overall, earlier changes were maintained., Conclusion: Women had significantly more favorable longitudinal changes in cardiac structure and function in response to LV unloading compared with men. Understanding the cause of sex difference in reverse remodeling after LVAD may help to devise novel therapeutic strategies for women with advanced heart failure., (Copyright © 2020 Elsevier Inc. All rights reserved.)

The expression of the extracellular sulfatase SULF2 has been associated with increased hepatocellular carcinoma (HCC) growth and poor patient survival. However, the molecular mechanisms underlying SULF2-associated tumor growth remain unclear. To address this gap, here we developed a transgenic mouse overexpressing Sulf2 in hepatocytes under the control of the transthyretin promoter. In this model, Sulf2 overexpression potentiated diethylnitrosamine-induced HCC. Further analysis indicated that the transcription factor GLI family zinc finger 1 (GLI1) mediates Sulf2 expression during HCC development. A cross of the Sulf2 -overexpressing with Gli1 -knockout mice revealed that Gli1 inactivation impairs SULF2-induced HCC. Transcriptomic analysis revealed that Sulf2 overexpression is associated with signal transducer and activator of transcription 3 (STAT3)-specific gene signatures. Interestingly, the Gli1 knockout abrogated SULF2-mediated induction of several STAT3 target genes, including suppressor of cytokine signaling 2/3 ( Socs2/3 ); Pim-1 proto-oncogene, Ser/Thr kinase ( Pim1 ); and Fms-related tyrosine kinase 4 ( Flt4 ). Human orthologs were similarly regulated by SULF2, dependent on intact GLI1 and STAT3 functions in HCC cells. SULF2 overexpression promoted a GLI1-STAT3 interaction and increased GLI1 and STAT3 enrichment at the promoters of their target genes. Interestingly, the SULF2 overexpression resulted in GLI1 enrichment at select STAT3 consensus sites, and vice versa. siRNA-mediated STAT3 or GLI1 knockdown reduced promoter binding of GLI1 and STAT3, respectively. Finally, chromatin-capture PCR confirmed long-range co-regulation of SOCS2 and FLT3 through changes in promoter conformation. These findings define a mechanism whereby SULF2 drives HCC by stimulating formation of a GLI1-STAT3 transcriptional complex., (© 2020 Carr et al.)

Prognostic models have been developed to help make decisions in the treatment of pulmonary embolism (PE). Among them, the Pulmonary Embolism Severity Index (PESI) and simplified PESI (sPESI), however they have not been validated in our setting. The objective was to evaluate PESI and sPESI scores ability to predict in-hospital mortality in patients with PE in Argentina. We analyzed a database of 75 academic centers in Argentina that included consecutive patients with PE from 2016 to 2017. The scores were prospectively calculated, and in-hospital and 30 days mortality were assessed. The validation of the models was assessed through discrimination using the area under the ROC curve (AUC), and calibration with the Hosmer-Lemeshow (HL) test. The cohort included 684 patients. In-hospital mortality was 12% and at 30 days an additional 3.2% mortality was registered. The AUC (95% CI) for in-hospital mortality was 0.75 (0.69-0.81) for PESI and 0.77 (0.71-0.82) for sPESI (p = 0.2 between scores). AUC of 30-day mortality 0.75 (0.68-0.8) and 0.78 (0.74-0.83) for PESI and sPESI (p = 0.2 between scores). Both models presented good calibration. The PESI and sPESI risk scores demonstrated similar performance and good accuracy in predicting hospital and 30-day mortality. Both scores can be established as simple prediction tools for PE patients in Argentina.

Dissemination of methicillin-resistant-Staphylococcus aureus/(MRSA) is a worldwide concern both in hospitals [healthcare-associated-(HA)-MRSA] and communities [community-associated-(CA)-MRSA]. Knowledge on when and where MRSA colonization is acquired and what clones are involved is necessary, to focus efforts for prevention of hospital-acquired MRSA-infections., Methods: A prospective/longitudinal cohort study was performed in eight Argentina hospitals (Cordoba/ October-December/2014). Surveillance cultures for MRSA (nose-throat-inguinal) were obtained on admission and at discharge. MRSA strains were genetically typed as CA-MRSA G and HA-MRSA G genotypes., Results: Overall, 1419 patients were screened and 534 stayed at hospital for ≥3 days. S. aureus admission prevalence was 30.9% and 4.2% for MRSA. Overall MRSA acquisition rate was 2.3/1000 patient-days-at-risk with a MRSA acquisition prevalence of 1.96% (95%CI: 1.0%-3.4%); 3.2% of patients were discharged back to community with MRSA. CA-MRSA G accounted for 84.6% of imported, 100.0% of hospital-acquired and 94% of discharged MRSA strains. Most imported and acquired MRSA strains belonged to two major epidemic CA-MRSA clones spread in Argentina: PFGEtypeI-ST5-IVa-t311-PVL + and PFGEtypeN/ST30-IVc-t019-PVL + ., Conclusions: CA-MRSA clones, particularly ST5-IV-PVL + and ST30-IV-PVL + , with main reservoir in the community, not only enter but also are truly acquired within hospital, causing healthcare-associated-hospital-onset infections, having a transmission capacity greater or similar than HA-MRSA G . This information is essential to develop appropriate MRSA infection prevention-control programs, considering hospital and community., (Copyright © 2019 The British Infection Association. Published by Elsevier Ltd. All rights reserved.)

Due to the coronavirus disease 2019 (COVID-19) pandemic, a wide number of compounds are under scrutiny regarding their antiviral activity, one of them being hydroxychloroquine. Cardiac aspects of the use of chloroquine and hydroxychloroquine are reviewed in this manuscript. A non-systematic review of the medical literature was performed. Information about their safety and efficacy as antimalarials, antivirals, as well as in the long-term treatment of rheumatic diseases was collected. We found an anti-inflammatory effect with reduction of longterm cardiovascular events, a very infrequent heart disease due to a lysosomal effect of the drug, and at the hemodynamic level hypotension, tachycardia, and QT interval prolongation, exacerbated when combined with azithromycin. However, the rate of adverse cardiac events of hydroxychloroquine (and chloroquine) was low.

Background/objectives: Interplay between the Hedgehog (HH) and epidermal growth factor receptor (EGFR) pathways modulating the outcome of their signaling activity have been reported in various cancers including pancreatic ductal adenocarcinoma (PDAC). Therefore, simultaneous targeting of these pathways may be clinically beneficial. This Phase I study combined HH and EGFR inhibition in metastatic PDAC patients., Methods: Combined effects of HH and EGFR inhibition using Vismodegib and Erlotinib with or without gemcitabine in metastatic solid tumors were assessed by CT. Another cohort of patients with metastatic PDAC was evaluated by FDG-PET and tumor biopsies-derived biomarkers., Results: Treatment was well tolerated with the maximum tolerated dose cohort experiencing no grade 4 toxicities though 25% experienced grade 3 adverse effects. Recommended phase II dose of Vismodegib and Erlotinib were each 150 mg daily. No tumor responses were observed although 16 patients achieved stable disease for 2-7 cycles. Paired biopsy analysis before and after first cycle of therapy in PDAC patients showed reduced GLI1 mRNA, phospho-GLI1 and associated HH target genes in all cases. However, only half of the cases showed reduced levels of desmoplasia or changes in fibroblast markers. Most patients had decreased phospho-EGFR levels., Conclusions: Vismodegib and Erlotinib combination was well-tolerated although overall outcome in patients with metastatic PDAC was not significantly impacted by combination treatment. Biomarker analysis suggests direct targets inhibition without significantly affecting the stromal compartment. These findings conflict with pre-clinical mouse models, and thus warrant further investigation into how upstream inhibition of these pathways is circumvented in PDAC., Competing Interests: Declaration of competing interest The authors declared no conflicts with this submission., (Copyright © 2019. Published by Elsevier B.V.)

The COVID-19 pandemic has led to measures of social isolation, labor restrictions, a strong information campaign and the suspension of scheduled medical activities. The aim of this study was to describe the impact of these measures on the number of hospitalizations in Cardiovascular Intensive Care Units, with the hypothesis that the social behavior generated by this emergency promotes a decreased demand for medical care, even when severe cardiovascular disease is involved. We compared the number of admissions in March-April 2010-2019 versus March-April 2020, based on a prospective study including six institutions (three public and three private) that use Epi-Cardio® as a multicenter registry of cardiovascular care unit discharge. Altogether, we included 6839 patients discharged during the 11-year study period (2010-2020). The average number of patient admissions on March-April 2010-19 was 595 (CI 95%: 507-683) and decreased to 348 in 2020 (fall of 46.8%, p < 0.001). The reasons for hospitalization were classified into 11 groups and a statistically significant reduction was seen in 10 of these groups: cardiovascular surgery 72.3%, electrophysiological interventions 67.8%, non-ST acute coronary syndromes 52.6%, angioplasties 47.6%, arrhythmias 48.7%, heart failure 46%, atrial fibrillation 35.7%, ST elevation myocardial infarction 34.7%, non cardiac chest pain 31.8%, others 51.6%. Although with low prevalence, hypertensive crisis increased in 89%. The abrupt decrease observed in the number of admissions due to critical pathologies may be considered an "adverse effect" related to the measures adopted, with potentially severe consequences. This trend could be reversed by improving public communication and policy adjustment.

Autophagy has been shown to be a key cellular event controlling tumor growth in different neoplasms including hepatocellular carcinoma (HCC). Although this biological role of autophagy has been clearly established, the mechanism underlying its regulation remains elusive. Here, we demonstrate a role of sulfatase 2 (SULF2), a 6-O-endosulfatase modulating various growth factors and cytokine-related signaling pathways controlling tumor cell proliferation and survival, in the regulation of autophagy in HCC cells. SULF2 increased autophagosome formation, shown by increased LC3B-II protein and green fluorescent protein-LC3 puncta. Increased fusion between autophagosomes and lysosomes/lysosomal enzymes, higher expression of lysosomal membrane protein, and an increase in autolysosomes were also shown by western blot, immunofluorescence, and electron microscopy of SULF2-expressing cells, indicating enhanced autophagic flux. In contrast, RNA-interference silencing of SULF2 in Huh7 cells induced lysosomal membrane permeabilization with diffuse cytosolic staining of cathepsin D and punctate staining of galectin-3. Analysis of the mechanism showed that inhibition of lysosome-associated protein transmembrane 4 beta (LAPTM4B), a gene induced by SULF2, resulted in decreased autophagosome formation, decreased fusion between autophagosomes and lysosomes, and increased lysosomal membrane permeabilization. Interestingly, down-regulation of LAPTM4B also phenocopies the knockdown of SULF2, significantly reducing cell viability and colony formation. Conclusion: Our results demonstrate a role for SULF2 in the regulation of autophagic flux that is mediated through LAPTM4B induction in HCC cells, and provide a foundation for future translational efforts targeting autophagy in liver malignancies., (© 2019 The Authors. Hepatology Communications published by Wiley Periodicals, Inc., on behalf of the American Association for the Study of Liver Diseases.)

Objective: Despite advances in the identification of epigenetic alterations in pancreatic cancer, their biological roles in the pathobiology of this dismal neoplasm remain elusive. Here, we aimed to characterise the functional significance of histone lysine methyltransferases (KMTs) and demethylases (KDMs) in pancreatic tumourigenesis., Design: DNA methylation sequencing and gene expression microarrays were employed to investigate CpG methylation and expression patterns of KMTs and KDMs in pancreatic cancer tissues versus normal tissues. Gene expression was assessed in five cohorts of patients by reverse transcription quantitative-PCR. Molecular analysis and functional assays were conducted in genetically modified cell lines. Cellular metabolic rates were measured using an XF24-3 Analyzer, while quantitative evaluation of lipids was performed by liquid chromatography-mass spectrometry (LC-MS) analysis. Subcutaneous xenograft mouse models were used to evaluate pancreatic tumour growth in vivo., Results: We define a new antitumorous function of the histone lysine (K)-specific methyltransferase 2D (KMT2D) in pancreatic cancer. KMT2D is transcriptionally repressed in human pancreatic tumours through DNA methylation. Clinically, lower levels of this methyltransferase associate with poor prognosis and significant weight alterations. RNAi-based genetic inactivation of KMT2D promotes tumour growth and results in loss of H3K4me3 mark. In addition, KMT2D inhibition increases aerobic glycolysis and alters the lipidomic profiles of pancreatic cancer cells. Further analysis of this phenomenon identified the glucose transporter SLC2A3 as a mediator of KMT2D-induced changes in cellular, metabolic and proliferative rates., Conclusion: Together our findings define a new tumour suppressor function of KMT2D through the regulation of glucose/fatty acid metabolism in pancreatic cancer., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2019. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: In two interim analyses of this trial, patients with advanced heart failure who were treated with a fully magnetically levitated centrifugal-flow left ventricular assist device were less likely to have pump thrombosis or nondisabling stroke than were patients treated with a mechanical-bearing axial-flow left ventricular assist device., Methods: We randomly assigned patients with advanced heart failure to receive either the centrifugal-flow pump or the axial-flow pump irrespective of the intended goal of use (bridge to transplantation or destination therapy). The composite primary end point was survival at 2 years free of disabling stroke or reoperation to replace or remove a malfunctioning device. The principal secondary end point was pump replacement at 2 years., Results: This final analysis included 1028 enrolled patients: 516 in the centrifugal-flow pump group and 512 in the axial-flow pump group. In the analysis of the primary end point, 397 patients (76.9%) in the centrifugal-flow pump group, as compared with 332 (64.8%) in the axial-flow pump group, remained alive and free of disabling stroke or reoperation to replace or remove a malfunctioning device at 2 years (relative risk, 0.84; 95% confidence interval [CI], 0.78 to 0.91; P<0.001 for superiority). Pump replacement was less common in the centrifugal-flow pump group than in the axial-flow pump group (12 patients [2.3%] vs. 57 patients [11.3%]; relative risk, 0.21; 95% CI, 0.11 to 0.38; P<0.001). The numbers of events per patient-year for stroke of any severity, major bleeding, and gastrointestinal hemorrhage were lower in the centrifugal-flow pump group than in the axial-flow pump group., Conclusions: Among patients with advanced heart failure, a fully magnetically levitated centrifugal-flow left ventricular assist device was associated with less frequent need for pump replacement than an axial-flow device and was superior with respect to survival free of disabling stroke or reoperation to replace or remove a malfunctioning device. (Funded by Abbott; MOMENTUM 3 ClinicalTrials.gov number, NCT02224755.)., (Copyright © 2019 Massachusetts Medical Society.)

Klebsiella pneumoniae (Kp) is a globally disseminated opportunistic pathogen that can cause life-threatening infections. It has been found as the culprit of many infection outbreaks in hospital environments, being particularly aggressive towards newborns and adults under intensive care. Many Kp strains produce extended-spectrum β-lactamases, enzymes that promote resistance against antibiotics used to fight these infections. The presence of other resistance determinants leading to multidrug-resistance also limit therapeutic options, and the use of 'last-resort' drugs, such as polymyxins, is not uncommon. The global emergence and spread of resistant strains underline the need for novel antimicrobials against Kp and related bacterial pathogens. To tackle this great challenge, we generated multiple layers of 'omics' data related to Kp and prioritized proteins that could serve as attractive targets for antimicrobial development. Genomics, transcriptomics, structuromic and metabolic information were integrated in order to prioritize candidate targets, and this data compendium is freely available as a web server. Twenty-nine proteins with desirable characteristics from a drug development perspective were shortlisted, which participate in important processes such as lipid synthesis, cofactor production, and core metabolism. Collectively, our results point towards novel targets for the control of Kp and related bacterial pathogens.

The World Health Organization (WHO) estimates that 40% of tuberculosis (TB) cases are not diagnosed and treated correctly. Even though there are several diagnostic tests available in the market, rapid, easy, inexpensive detection, and drug susceptibility testing (DST) of Mycobacterium tuberculosis is still of critical importance specially in low and middle-income countries with high incidence of the disease. In this work, we have developed a microscopy-based methodology using the reporter mycobacteriophage mCherry bomb ϕ for detection of Mycobacterium spp. and phenotypic determination of rifampicin resistance within just days from sputum sample collection. Fluoromycobacteriophage methodology is compatible with regularly used protocols in clinical laboratories for TB diagnosis and paraformaldehyde fixation after infection reduces biohazard risks with sample analysis by fluorescence microscopy. We have also set up conditions for discrimination between M. tuberculosis complex (MTBC) and non-tuberculous mycobacteria (NTM) strains by addition of p -nitrobenzoic acid (PNB) during the assay. Using clinical isolates of pre-XDR and XDR-TB strains from this study, we tested mCherry bomb Φ for extended DST and we compared the antibiotic resistance profile with those predicted by whole genome sequencing. Our results emphasize the utility of a phenotypic test for M. tuberculosis extended DST. The many attributes of mCherry bomb Φ suggests this could be a useful component of clinical microbiological laboratories for TB diagnosis and since only viable cells are detected this could be a useful tool for monitoring patient response to treatment.

Available genomic data for pathogens has created new opportunities for drug discovery and development to fight them, including new resistant and multiresistant strains. In particular structural data must be integrated with both, gene information and experimental results. In this sense, there is a lack of an online resource that allows genome wide-based data consolidation from diverse sources together with thorough bioinformatic analysis that allows easy filtering and scoring for fast target selection for drug discovery. Here, we present Target-Pathogen database (http://target.sbg.qb.fcen.uba.ar/patho), designed and developed as an online resource that allows the integration and weighting of protein information such as: function, metabolic role, off-targeting, structural properties including druggability, essentiality and omic experiments, to facilitate the identification and prioritization of candidate drug targets in pathogens. We include in the database 10 genomes of some of the most relevant microorganisms for human health (Mycobacterium tuberculosis, Mycobacterium leprae, Klebsiella pneumoniae, Plasmodium vivax, Toxoplasma gondii, Leishmania major, Wolbachia bancrofti, Trypanosoma brucei, Shigella dysenteriae and Schistosoma Smanosoni) and show its applicability. New genomes can be uploaded upon request., (© The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.)

The morbidity of bone fractures and defects is steadily increasing due to changes in the age pyramid. As such, novel biomaterials that are able to promote the healing and regeneration of injured bones are needed to overcome the limitations of auto-, allo-, and xenografts, while providing a ready-to-use product that may help to minimize surgical invasiveness and duration. In this regard, recombinant biomaterials, such as elastin-like recombinamers (ELRs), are very promising as their design can be tailored by genetic engineering, thus allowing scalable production and batch-to-batch consistency, among others. Furthermore, they can self-assemble into physically crosslinked hydrogels above a certain transition temperature, in this case body temperature, but are injectable below this temperature, thereby markedly reducing surgical invasiveness. In this study, we have developed two bioactive hydrogel-forming ELRs, one including the osteogenic and osteoinductive bone morphogenetic protein-2 (BMP-2) and the other the Arg-Gly-Asp (RGD) cell adhesion motif. The combination of these two novel ELRs results in a BMP-2-loaded extracellular matrix-like hydrogel. Moreover, elastase-sensitive domains were included in both ELR molecules, thereby conferring biodegradation as a result of enzymatic cleavage and avoiding the need for scaffold removal after bone regeneration. Both ELRs and their combination showed excellent cytocompatibility, and the culture of cells on RGD-containing ELRs resulted in optimal cell adhesion. In addition, hydrogels based on a mixture of both ELRs were implanted in a pilot study involving a femoral bone injury model in New Zealand white rabbits, showing complete regeneration in six out of seven cases, with the other showing partial closure of the defect. Moreover, bone neoformation was confirmed using different techniques, such as radiography, computed tomography, and histology. This hydrogel system therefore displays significant potential in the regeneration of bone defects, promoting self-regeneration by the surrounding tissue with no involvement of stem cells or osteogenic factors other than BMP-2, which is released in a controlled manner by elastase-mediated cleavage from the ELR backbone.

The aim of this study is to evaluate the effects of the inoculation of the saltmarsh plant (Halimione portulacoides) with Pseudomonas sp. SPN31 nah+ combined with exposure to 2-methylnaphthalene (2-MtN) on the plant rhizosphere and endosphere bacterial communities as well as on plant health. To achieve this goal, microcosm experiments were set up. Denaturing gradient gel electrophoresis (DGGE) profiles and statistical analysis showed that rhizosphere and endosphere bacterial communities had distinct responses to plant inoculation and/or exposure to 2-MtN. PCR-sequencing analysis of nah genes encoding for 2-MtN degrading enzymes suggested the presence of Pseudomonas sp. SPN31 nah+ in the endosphere of H. portulacoides with 2-MtN contamination. Moreover, a significant effect in the photosynthetic performance of inoculated plants was detected. To conclude, despite the potential beneficial effect of plant inoculation with Pseudomonas sp. SPN31 nah+ endophytic bacteria may have on plant health, no significant effect on the removal of MtN was detected for the level of contamination used in the study.

Existing antiangiogenic approaches to treat metastatic hepatocellular carcinoma (HCC) are weakly effectual, prompting further study of tumor angiogenesis in this disease setting. Here, we report a novel role for sulfatase 2 (SULF2) in driving HCC angiogenesis. Sulf2-deficient mice (Sulf2 KO) exhibited resistance to diethylnitrosamine-induced HCC and did not develop metastases like wild-type mice (Sulf2 WT). The smaller and less numerous tumors formed in Sulf2 KO mice exhibited a markedly lower microvascular density. In human HCC cells, SULF2 overexpression increased endothelial proliferation, adhesion, chemotaxis, and tube formation in a paracrine fashion. Mechanistic analyses identified the extracellular matrix protein periostin (POSTN), a ligand of αvβ3/5 integrins, as an effector protein in SULF2-induced angiogenesis. POSTN silencing in HCC cells attenuated SULF2-induced angiogenesis and tumor growth in vivo The TGFβ1/SMAD pathway was identified as a critical signaling axis between SULF2 and upregulation of POSTN transcription. In clinical HCC specimens, elevated levels of SULF2 correlated with increased microvascular density, POSTN levels, and relatively poorer patient survival. Together, our findings define an important axis controlling angiogenesis in HCC and a mechanistic foundation for rational drug development. Cancer Res; 77(3); 632-45. ©2016 AACR., (©2016 American Association for Cancer Research.)

Purpose: Genomic testing has increased the quantity of information available to oncologists. Unfortunately, many identified sequence alterations are variants of unknown significance (VUSs), which thus limit the clinician's ability to use these findings to inform treatment. We applied a combination of in silico prediction and molecular modeling tools and laboratory techniques to rapidly define actionable VUSs., Materials and Methods: Exome sequencing was conducted on 308 tumors from various origins. Most single nucleotide alterations within gene coding regions were VUSs. These VUSs were filtered to identify a subset of therapeutically targetable genes that were predicted with in silico tools to be altered in function by their variant sequence. A subset of receptor tyrosine kinase VUSs was characterized by laboratory comparison of each VUS versus its wild-type counterpart in terms of expression and signaling activity., Results: The study identified 4,327 point mutations of which 3,833 were VUSs. Filtering for mutations in genes that were therapeutically targetable and predicted to affect protein function reduced these to 522VUSs of interest, including a large number of kinases. Ten receptortyrosine kinase VUSs were selected to explore in the laboratory. Of these, seven were found to be functionally altered. Three VUSs (FGFR2 F276C, FGFR4 R78H, and KDR G539R) showed increased basal or ligand-stimulated ERK phosphorylation compared with their wild-type counterparts, which suggests that they support transformation. Treatment of a patient who carried FGFR2 F276C with an FGFR inhibitor resulted in significant and sustained tumor response with clinical benefit., Conclusion: The findings demonstrate the feasibility of rapid identification of the biologic relevance of somatic mutations, which thus advances clinicians' ability to make informed treatment decisions., Competing Interests: AUTHORS’ DISCLOSURES OF POTENTIAL CONFLICTS OF INTEREST The following represents disclosure information provided by authors of this manuscript. All relationships are considered compensated. Relationships are self-held unless noted. I = Immediate Family Member, Inst = My Institution. Relation-ships may not relate to the subject matter of this manuscript. For more information about ASCO’s conflict of interest policy, please refer to www.asco.org/rwc or po.ascopubs.org/site/ifc. Jan B. Egan No relationship to disclose David L. Marks Stock and Other Ownership Interests: WebMD, Cardinal Health (I), Prothena (I), Perrigo (I) Tara L. Hogenson No relationship to disclose Anne M. Vrabel No relationship to disclose Ashley N. Sigafoos No relationship to disclose Ezequiel J. Tolosa No relationship to disclose Ryan M. Carr No relationship to disclose Stephanie L. Safgren No relationship to disclose Elisa Enriquez Hesles No relationship to disclose Luciana L. Almada No relationship to disclose Paola A. Romecin-Duran No relationship to disclose Eriko Iguchi No relationship to disclose Aryan Ala’Aldeen No relationship to disclose Jean-Pierre A. Kocher No relationship to disclose Gavin R. Oliver No relationship to disclose Naresh Prodduturi No relationship to disclose David W. Mead No relationship to disclose Asif Hossain No relationship to disclose Norine E. Huneke No relationship to disclose Colleen M. Tagtow No relationship to disclose Sikander Ailawadhi Consulting or Advisory Role: Amgen, Takeda Pharmaceuticals, Novartis Research Funding: Pharmacyclics (Inst) Travel, Accommodations, Expenses: Amgen, Takeda Pharmaceuticals, Novartis Stephen M. Ansell Honoraria: WebMD, Research To Practice Research Funding: Bristol-Myers Squibb (Inst), Celldex Therapeutics (Inst), Seattle Genetics (Inst), Merck (Inst), Affimed (Inst), Trillium Therapeutics (Inst) Michaela S. Banck No relationship to disclose Alan H. BryceNo relationship to disclose Estrella M. Carballido No relationship to disclose Asher A. Chanan-Khan No relationship to disclose Kelly K. Curtis Employment: INC Research Stock and Other Ownership Interests: INC Research Travel, Accommodations, Expenses: INC Research Ernesto ResnikNo relationship to disclose Chelsea D. Gawryletz No relationship to disclose Ronald S. Go Speakers’ Bureau: OnLive, Takeda Pharmaceuticals Thorvardur R. Halfdanarson Consulting or Advisory Role: Lexicon (Inst), Ipsen (Inst), Merrimack (Inst) Research Funding: Esanex (Inst), Ipsen (Inst), Boston Biomedical (Inst) Thai H. Ho No relationship to disclose Richard W. Joseph Consulting or Advisory Role: Bristol-Myers Squibb, Nektar, Genoptix, Eisai, Bristol-Myers Squibb Research Funding: Merck, Bristol-Meyers Squibb, Roche (Inst), Genentech (Inst), X4P Pharmaceuticals (Inst), Amgen (Inst) Prashant Kapoor Consulting or Advisory Role: Sanofi (Inst) Research Funding: Amgen (Inst), Takeda Pharmaceuticals (Inst) Aaron S. Mansfield Consulting or Advisory Role: TrovaGene (Inst), Genentech (Inst), Bristol-Myers Squibb (Inst) Travel, Accommodations, Expenses: Bristol-Myers Squibb Nathalie Meurice No relationship to disclose Amulya A. Nageswara Rao No relationship to disclose Grzegorz S. Nowakowski Consulting or Advisory Role: Celgene (Inst), MorphoSys (Inst), Genentech (Inst) Research Funding: Celgene (Inst) Animesh Pardanani No relationship to disclose Sameer A. Parikh Research Funding: Pharmacyclics (Inst) John C. Cheville No relationship to disclose Andrew L. Feldman Consulting or Advisory Role: Infinity Pharmaceuticals Patents, Royalties, Other Intellectual Property: Inventor of technology for which the Mayo Clinic holds an unlicensed patent or has submitted a patent application (Inst) Ramesh K. Ramanathan Honoraria: Taiho Pharmaceutical, Cerulean Pharma, Pharmacyclics, Insys Therapeutics, Novocure Consulting or Advisory Role: Cerulean Pharma, Novocure, INSYS Therapeutics, Pharmacyclics Research Funding: AbbVie (Inst), Celgene (Inst), Merck (Inst), Schering Plough (Inst), Merrimack (Inst), Boston Biomedical (Inst), BERG (Inst), Superlab Far East (Inst) Steven I. Robinson Travel, Accommodations, Expenses: Tricon Pharmaceuticals Raoul Tibes Research Funding: Novartis (Inst), Merck (Inst), Seattle Genetics (Inst), Boehringer Ingelheim (Inst), Astellas Pharma (Inst), Astex Pharmaceuticals (Inst)Heidi D. Finnes Jennifer B. McCormick No relationship to disclose Robert R. McWilliams Consulting or Advisory Role: Merrimack (Inst) Research Funding: PRISM BioLab (Inst), Genentech (Inst), Novartis (Inst), NewLink Genetics (Inst), Eli Lilly (Inst), Aduro Biotech (Inst), Pfizer (Inst), Sanofi (Inst) Travel, Accommodations, Expenses: AstraZeneca Aminah Jatoi Research Funding: Entera Health, Boston Biologics Mrinal M. Patnaik No relationship to disclose Alvin C. Silva No relationship to disclose Eric D. Wieben Patents, Royalties, Other Intellectual Property: Champions Oncology–licensed IP-targeted therapy with abiraterone acetate ($0 received), WuXi AppTec–licensed IP-breast cancer xenografts ($12,737 to Mayo Clinic; Inst), Affymetrix-licensed IP on several variants in drug processing enzymes ($0 in past 2 years), Life Technologies–licensed IP on several variants in drug processing enzymes ($0 in past 2 years) Tammy M. McAllister No relationship to disclose Kandelaria M. Rumilla No relationship to disclose Sarah E. Kerr Research Funding: Abbott Molecular Konstantinos N. Lazaridis No relationship to disclose Gianrico Farrugia No relationship to disclose A. Keith Stewart Consulting or Advisory Role: Celgene, Bristol-Myers Squibb, Janssen Pharmaceuticals, Roche, Amgen Karl J. Clark Patents, Royalties, Other Intellectual Property: Inventor on patents associated with the Sleeping Beauty transposon system Eileen J. Kennedy Patents, Royalties, Other Intellectual Property: Patent on unrelated work (US patent 9,458,189) for “ligation of stapled polypeptides” issued October 4, 2016 Eric W. Klee Patents, Royalties, Other Intellectual Property: Royalties on software development Mitesh J. Borad Stock and Other Ownership Interests: GlaxoSmithKline, Gilead Sciences Consulting or Advisory Role: G1 Therapeutics, TD2, Fujifilm (Inst), Agios (Inst), INSYS Therapeutics (Inst), Novartis (Inst), ArQule (Inst), Celegene (Inst), Inspyr Therapeutics, Halozyme Therapeutics (Inst) Research Funding: Boston Biomedical (Inst), miRNA Therapeutics (Inst), Senhwa Biosciences (Inst), Medimmune (Inst), Bioline (Inst), Agios (Inst), Halozyme Therapeutics (Inst), Celgene (Inst), Threshold Pharmaceuticals (Inst), Toray (Inst), Dicerna Pharmaceuticals (Inst), Sillajen (Inst), Eisai (Inst), Taiho Pharmaceuticals (Inst), EMD Serono (Inst), Isis Pharmaceuticals (Inst), Incyte (Inst), Sun BioPharma (Inst), ARIAD Pharmaceuticals (Inst), ImClone Systems (Inst) Travel, Accommodations, Expenses: ArQule, Celgene, AstraZeneca Martin E. Fernandez-Zapico No relationship to disclose

Although the differentiation of oncogenically transformed basal progenitor cells is one of the key steps in prostate tumorigenesis, the mechanisms mediating this cellular process are still largely unknown. Here we demonstrate that an expanded p63 + and CK5 + basal/progenitor cell population, induced by the concomitant activation of oncogenic Kras(G12D) and androgen receptor (AR) signaling, underwent cell differentiation in vivo The differentiation process led to suppression of p63-expressing cells with a decreased number of CK5 + basal cells but an increase of CK8 + luminal tumorigenic cells and revealed a hierarchal lineage pattern consisting of p63 + /CK5 + progenitor, CK5 + /CK8 + transitional progenitor, and CK8 + differentiated luminal cells. Further analysis of the phenotype showed that Kras-AR axis-induced tumorigenesis was mediated by Gli transcription factors. Combined blocking of the activators of this family of proteins (Gli1 and Gli2) inhibited the proliferation of p63 + and CK5 + basal/progenitor cells and development of tumors. Finally, we identified that Gli1 and Gli2 exhibited different functions in the regulation of p63 expression or proliferation of p63 + cells in Kras-AR driven tumors. Gli2, but not Gli1, transcriptionally regulated the expression levels of p63 and prostate sphere formation. Our study provides evidence of a novel mechanism mediating pathological dysregulation of basal/progenitor cells through the differential activation of the Gli transcription factors. Also, these findings define Gli proteins as new downstream mediators of the Kras-AR axis in prostate carcinogenesis and open a potential therapeutic avenue of targeting prostate cancer progression by inhibiting Gli signaling., (© 2016 by The American Society for Biochemistry and Molecular Biology, Inc.)

Crude glycerin, a potential energy source for ruminant animals, has been evaluated, mainly, in diets with high starch content. However, a limit number of studies have evaluated the inclusion of crude glycerin in low starch diets. This study aimed to evaluate the effects of the association of crude glycerin with corn grain or citrus pulp on carcass traits and meat quality of Nellore bulls (n=30, 402±31 kg initial weight). The treatment consisted of: CON=control, without crude glycerin; CG10=10% of crude glycerin and corn grain; CG15=15% of crude glycerin and corn grain; CP10=10% of crude glycerin and citrus pulp; CP15=15% of crude glycerin and citrus pulp. The performance parameters and carcass traits were not affected by treatments (P>0.05). The inclusion of crude glycerin decreased yellow color intensity and increased fatty acids pentadecanoic and heptadecenoic in meat (P<0.05), without affecting neither the concentration of polyunsaturated fatty acids nor the relationship of saturated and unsaturated fatty acids. The association of crude glycerin with corn or citrus pulp has no adverse effects on carcass characteristics and meat quality.

Our aim was to understand the involvement of Sonic hedgehog (Shh) morphogen in the oriented distribution of neural crest cells (NCCs) toward the optic vesicle and to look for potential disorders of this guiding mechanism after ethanol exposure. In vitro directional analysis showed the chemotactic response of NCCs up Shh gradients and to notochord co-cultures (Shh source) or to their conditioned medium, a response inhibited by anti-Shh antibody, receptor inhibitor cyclopamine and anti-Smo morpholino (MO). Expression of the Ptch-Smo receptor complex on in vitro NCCs was also shown. In whole embryos, the expression of Shh mRNA and protein was seen in the ocular region, and of Ptch, Smo and Gli/Sufu system on cephalic NCCs. Anti-Smo MO or Ptch-mutated plasmid (Ptch1(Δloop2)) impaired cephalic NCC migration/distribution, with fewer cells invading the optic region and with higher cell density at the homolateral mesencephalic level. Beads embedded with cyclopamine (Smo-blocking) or Shh (ectopic signal) supported the role of Shh as an in vivo guide molecule for cephalic NCCs. Ethanol exposure perturbed in vitro and in vivo NCC migration. Early stage embryos treated with ethanol, in a model reproducing Fetal Alcohol Syndrome, showed later disruptions of craniofacial development associated with abnormal in situ expression of Shh morphogen. The results show the Shh/Ptch/Smo-dependent migration of NCCs toward the optic vesicle, with the support of specific inactivation with genetic and pharmacological tools. They also help to understand mechanisms of accurate distribution of embryonic cells and of their perturbation by a commonly consumed teratogen, and demonstrate, in addition to its other known developmental functions, a new biological activity of cellular guidance for Shh., (Copyright © 2016 Elsevier GmbH. All rights reserved.)

In this study, we evaluated the effects of total corn replacement with crude glycerin on carcass characteristics and meat quality of feedlot lambs fed high-concentrate diets with low starch. Forty non-castrated Santa Ines lambs (23.5 ± 3.56 kg BW) were assigned to a randomized complete block design with five dietary treatments: 0%, 7.5%, 15%, 22.5%, or 30% crude glycerin, replacing corn. Animals were slaughtered at a BW of 38 kg after 72 ± 20 days. The addition of up to 30% crude glycerin reduced carcass weight and yield (P ≤ 0.02). Odd-chain fatty acids, oleic, palmitoleic, total unsaturated, and monounsaturated fatty acids were increased (P ≤ 0.01) while CLA tended to increase in glycerin-fed lambs (P = 0.06). Crude glycerin decreased stearic, palmitic, transvaccenic, total saturated fatty acids, and atherogenicity index (P < 0.01). High concentrations of crude glycerin in low-starch diets reduced carcass weights, nevertheless improved meat quality by increasing unsaturated and odd-chain fatty acid contents., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Although estuarine microphytobenthos (MPB) is frequently exposed to excessive light and temperature conditions, little is known on their interactive effects on MPB primary productivity. Laboratory and in situ experiments were combined to investigate the short-term joint effects of high light (HL) and high temperature (37 °C versus 27 °C) on the operating efficiency of photoprotective processes [vertical migration versus non-photochemical quenching (NPQ)] exhibited by natural benthic diatom communities from two intertidal flats in France (FR) and Portugal (PT). A clear latitudinal pattern was observed, with PT biofilms being more resistant to HL stress, regardless the effect of temperature, and displaying a lower relative contribution of vertical migration to photoprotection and a stronger NPQ in situ. However, higher temperature leads to comparable effects, with photoinhibition increasing to about three times (i.e. from 3% to 10% and from 8% to 22% in PT and FR sites respectively). By using a number of methodological novelties in MPB research (lipid peroxidation quantification, Lhcx proteins immunodetection), this study brings a physiological basis to the previously reported depression of MPB photosynthetic productivity in summer. They emphasize the joint role of temperature and light in limiting, at least transiently (i.e. during emersion), MPB photosynthetic activity in situ., (© 2014 Society for Applied Microbiology and John Wiley & Sons Ltd.)

Mice with heterozygous loss of the tumor suppressor Patched1 (Ptch) develop rhabdomyosarcoma (RMS)-like tumors. However, Ptch transcripts are consistently overexpressed in these tumors. We have recently shown that the upregulated transcripts are derived from the mutated Ptch allele thus leading to the hypothesis that the wild-type allele is repressed during RMS development. Here we describe epigenetic changes taking place at the Ptch locus during RMS development. We showed a lower degree of DNA-methylation in methylation-sensitive CpG regions of the Ptch promoter in RMS compared to normal muscle from heterozygous Ptch animals. In agreement with these results, treatment of heterozygous Ptch mice with the DNA demethylating agent 5-aza-2-deoxycytidine (5-aza-dC) between embryonic days E9.5-E11.5 significantly accelerated RMS formation. Since Ptch promoter methylation occurs after/around E13.5, the window for RMS initiation during embryogenesis, these results provide additional evidence that Ptch promoter hypomethylation may contribute to RMS formation. We have also demonstrated increased trimethylation of histone H3 lysine 4 (H3K4me3) and preferential binding of Gli1, a known Ptch activator, to the mutant locus in RMS. Together, these findings support an alternative model for RMS formation in heterozygous Ptch mice including loss of methylation and concomitant occupancy by activating histone marks of mutant Ptch.

The effect of ethanol in modulating the intensity and duration of the perceived sourness induced by citric acid was studied. Magnitude Estimation-Converging Limits method was applied to rate the sourness of seven solutions (3-70 mM) of citric acid in aqueous solution presented alone and mixed with 8% V/V or 15% V/V ethanol. Dynamic sourness ratings of 5, 15, and 45 mM citric acid alone and mixed with the same two ethanol levels were assessed by the Time Intensity Method (TI). Results were consistent with both methods. Sourness changed with citric acid concentration and ethanol levels. From TI measurements, a similar interactive pattern was obtained for parameters as duration, area under the curve, peak and average intensity.

Light-response curves (LCs) of chlorophyll fluorescence are widely used in plant physiology. Most commonly, LCs are generated sequentially, exposing the same sample to a sequence of distinct actinic light intensities. These measurements are not independent, as the response to each new light level is affected by the light exposure history experienced during previous steps of the LC, an issue particularly relevant in the case of the popular rapid light curves. In this work, we demonstrate the proof of concept of a new method for the rapid generation of LCs from nonsequential, temporally independent fluorescence measurements. The method is based on the combined use of sample illumination with digitally controlled, spatially separated beams of actinic light and a fluorescence imaging system. It allows the generation of a whole LC, including a large number of actinic light steps and adequate replication, within the time required for a single measurement (and therefore named "single-pulse light curve"). This method is illustrated for the generation of LCs of photosystem II quantum yield, relative electron transport rate, and nonphotochemical quenching on intact plant leaves exhibiting distinct light responses. This approach makes it also possible to easily characterize the integrated dynamic light response of a sample by combining the measurement of LCs (actinic light intensity is varied while measuring time is fixed) with induction/relaxation kinetics (actinic light intensity is fixed and the response is followed over time), describing both how the response to light varies with time and how the response kinetics varies with light intensity.

Purpose: Gastroesophageal reflux disease (GERD) is associated with allograft dysfunction after lung transplantation (LTX). We attempted to identify outcomes in LTX recipients with clinical evidence of GERD., Methods: Retrospective review of 162 LTX recipients at our institution between January 1994 and June 2006 was performed. GERD was confirmed in symptomatic patients by esophagogastroduodenoscopy (EGD) and/or esophagography. Occurrence of biopsy-proven obliterative bronchiolitis (OB) and bronchiolitis obliterans syndrome (BOS) were analyzed. Kaplan-Meier analysis of survival and Cox proportional hazard analysis of risk factors were performed., Results: GERD was diagnosed in 21 (13%) of patients, usually following LTX (71%). There was no difference in mean survival (1603 +/- 300 vs. 1422 +/- 131 days; log rank P > 0.05), or development of OB (5% vs. 6%, respectively; P > 0.05) in patients with GERD compared with patients without GERD. However, there was correlation between GERD and BOS (P = 0.01)., Conclusions: Symptomatic GERD is increased following LTX. Patients with symptomatic GERD demonstrated an increased incidence of BOS, but survival was not affected in this study. More sensitive and specific diagnostic tools should be implemented in all LTX recipients to investigate the impact of symptomatic and silent GERD and thus improve outcomes after LTX.

Heart failure is associated with abnormalities in betaAR cascade regulation, calcium cycling, expression of inflammatory mediators and apoptosis. Adenoviral mediated gene transfer of betaARKct has beneficial indirect effects on these pathologic processes upon the left ventricular myocardium. The concomitant biochemical changes that occur in the right ventricle have not been well characterized. Sprague-Dawley rats underwent aortic banding and were followed by echocardiography. After a decrease in fractional shortening of 25% from baseline, intracoronary injection of adenoviral-betaARKct (n=14) or adenoviral-beta-galactosidase (control, n=13) was performed. Rats were randomly euthanized on post-operative day 7, 14 or 21. Protein analysis including RV myocardial levels of betaARKct, betaARK1, SERCA(2a), inflammatory tissue mediators (IL-1, IL-6 and TNF-alpha), apoptotic markers (bax and bak), and MAP kinases (jnk, p38 and erk) was performed. ANOVA was employed for group comparison. Adenoviral-betaARKct treated animals showed increased expression of betaARKct and decreased levels of betaARK1 compared with controls. This treatment group also demonstrated normalization of SERCA(2a) expression and decreased levels of the inflammatory markers IL-1, IL-6 and TNF-alpha. The pro-apoptotic markers bax and bak were similarly improved. Ventricular levels of the MAP kinase jnk were increased. Differences were most significant 7 days after gene transfer, but the majority of these changes persisted at 21 days. These results suggest that attenuation of the pathologic mechanisms of beta adrenergic receptor desensitization, SERCA(2a) expression, inflammation and apoptosis, not only occur in the left ventricle but also in the right ventricular myocardium after intracoronary gene transfer of betaARKct during heart failure.

Objective: We hypothesized that intracoronary adenoviral-mediated delivery of betaARKct would improve heart failure associated pathophysiologic abnormalities related to exercise capacity, cardiac contractility, systemic inflammation and volume overload., Methods: After aortic banding, a cohort of Sprague-Dawley rats was followed by echocardiography. When an absolute decline of 25% in fractional shortening was detected, animals were randomized to intracoronary delivery of Ad.ssARKct (n=14), Ad.beta-Gal (n=13), or followed without any other further intervention (n=18). Assessment of exercise tolerance and hemodynamic profile and measurement of markers of systemic inflammation and volume overload was performed at 7, 14, and 21 days after gene delivery. Data were analyzed using ANOVA., Results: Animals receiving Ad.ssARKct showed improved exercise tolerance compared to Ad.Gal-treated animals at 14 days (507+/-26 s vs. 408+/-19 s, P=0.01) and 21 days (526+/-55 s vs. 323+/-19 s, P<0.001) following injection. Animals receiving Ad.ssARKct demonstrated improved +dP/dtmax (mean+/-SD, 5,581+/-960 mmHg/s vs. 3,134+/-438 mmHg/s, P<0.01) and -dP/dtmax (mean+/-SD, -3,494+/-1,269 mmHg/s vs. -1,925+/-638 mmHg/s, P<0.01) compared to Ad.Gal-treated animals at 7 days. These differences were observed up to 21 days following injection. Serum levels of IL-1, IL-6, and TNF-alpha, as well as ANP were also decreased in animals receiving Ad.betaARKct., Conclusions: Genetic modulation of heart failure using the betaARKct gene was associated with improved exercise capacity and cardiac function as well as amelioration in heart failure-associated profiles of systemic inflammation and volume overload.

Objective: To examine fat biopsy samples from lean insulin-sensitive and obese insulin-resistant nondiabetic individuals for evidence of endoplasmic reticulum (ER) stress., Research Design and Methods: Subcutaneous fat biopsies were obtained from the upper thighs of six lean and six obese nondiabetic subjects. Fat homogenates were used for proteomic (two-dimensional gel and MALDI-TOF/TOF), Western blot, and RT-PCR analysis., Results: Proteomic analysis revealed 19 differentially upregulated proteins in fat of obese subjects. Three of these proteins were the ER stress-related unfolded protein response (UPR) proteins calreticulin, protein disulfide-isomerase A3, and glutathione-S-transferase P. Western blotting revealed upregulation of several other UPR stress-related proteins, including calnexin, a membrane-bound chaperone, and phospho c-jun NH(2)-terminal kinase (JNK)-1, a downstream effector protein of ER stress. RT-PCR analysis revealed upregulation of the spliced form of X-box binding protein-1s, a potent transcription factor and part of the proximal ER stress sensor inositol-requiring enzyme-1 pathway., Conclusions: These findings represent the first demonstration of UPR activation in subcutaneous adipose tissue of obese human subjects. As JNK can inhibit insulin action and activate proinflammatory pathways, ER stress activation of JNK may be a link between obesity, insulin resistance, and inflammation.

Objectives: In a rat model of pressure overload hypertrophy, we studied the effects of intracoronary delivery of mesenchymal stem cells on hemodynamic performance, exercise capacity, systemic inflammation, and left ventricular reverse remodeling., Methods: Sprague-Dawley rats underwent aortic banding and were followed up by echocardiographic scanning. After a decrease in fractional shortening of 25% from baseline, animals were randomized to intracoronary injection of mesenchymal stem cells (MSC group; n = 28) or phosphate-buffered saline solution (control group; n = 20). Hemodynamic and echocardiographic assessment, swim testing to exhaustion, and measurement of inflammatory markers were performed before the rats were humanely killed on postoperative day 7, 14, 21, or 28., Results: Injection of mesenchymal stem cells improved systolic function in the MSC group compared with the control group (mean +/- standard deviation: maximum dP/dt 3048 +/- 230 mm Hg/s vs 2169 +/- 97 mm Hg/s at 21 days and 3573 +/- 741 mm Hg/s vs 1363 +/- 322 mm Hg/s at 28 days: P < .001). Time to exhaustion was similarly increased in the MSC group compared with controls (487 +/- 35 seconds vs 306 +/- 27 seconds at 28 days; P < .01). Serum levels of interleukins 1 and 6, tumor necrosis factor-alpha, and brain natriuretic peptide-32 were significantly decreased in animals treated with mesenchymal stem cells. Stem cell transplantation improved left ventricular fractional shortening at 21 and 28 days. Left ventricular end-systolic and end-diastolic diameters were also improved at 28 days., Conclusions: In this model of pressure overload hypertrophy, intracoronary delivery of mesenchymal stem cells during heart failure was associated with an improvement in hemodynamic performance, maximal exercise tolerance, systemic inflammation, and left ventricular reverse remodeling. This study suggests a potential role of this treatment strategy for the management of hypertrophic heart failure resulting from pressure overload.