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The rising incidences of atherosclerosis have necessitated efforts to identify novel targets for therapeutic interventions. In the present study, we observed increased expression of the mechanosensitive calcium channel Piezo1 transcript in mouse and human atherosclerotic plaques, correlating with infiltration of PIEZO1-expressing macrophages. In vitro administration of Yoda1, a specific agonist for PIEZO1, led to increased foam cell apoptosis and enhanced phagocytosis by macrophages. Mechanistically, PIEZO1 activation resulted in intracellular F-actin rearrangement, elevated mitochondrial ROS levels and induction of mitochondrial fragmentation upon PIEZO1 activation, as well as increased expression of anti-inflammatory genes. In vivo, ApoE−/− mice treated with Yoda1 exhibited regression of atherosclerosis, enhanced stability of advanced lesions, reduced plaque size and necrotic core, increased collagen content, and reduced expression levels of inflammatory markers. Our findings propose PIEZO1 as a novel and potential therapeutic target in atherosclerosis. [ABSTRACT FROM AUTHOR]

Background: To avoid the overuse of antibiotics, non-steroidal anti-inflammatory drugs (NSAIDs), acting via cyclooxygenase (COX) inhibition, have been used to reduce pain and as an alternative treatment for uncomplicated urinary tract infections (UTIs). However, clinical studies evaluating NSAIDs versus antibiotics have reported an increased risk of acute pyelonephritis. Therefore, we hypothesized that COX inhibition could compromise the innate immune response and contribute to complications in patients with uncomplicated UTI. Results: We here demonstrate that in particular COX-2 inhibition led to decreased expression of the antimicrobial peptides psoriasin and human β-defensin-2 in human uroepithelial cells. Psoriasin expression was altered in neutrophils and macrophages. COX-2 inhibition also had impact on the inflammasome mediated IL-1β expression in response to uroepithelial E. coli infection. Further, COX-2 inhibition downregulated free radicals and the epithelial barrier protein claudin 1, favoring infectivity. In addition, conditioned media from COX-2 inhibited uroepithelial cells infected with E. coli failed to activate macrophages. Conclusions: Taken together, our data suggests an adverse innate immune effect of COX-2 inhibition on uroepithelial cells during UTI. [ABSTRACT FROM AUTHOR]

Background and Purpose: Tyrosine kinase inhibitors (TKI) used to treat chronic myeloid leukaemia (CML) have been associated with cardiovascular side effects, including reports of calcific aortic valve stenosis. The aim of this study was to establish the effects of first and second generation TKIs in aortic valve stenosis and to determine the associated molecular mechanisms.Experimental Approach: Hyperlipidemic APOE*3Leiden.CETP transgenic mice were treated with nilotinib, imatinib or vehicle. Human valvular interstitial cells (VICs) were isolated and studied in vitro. Gene expression analysis was perfromed in aortic valves from 64 patients undergoing aortic valve replacement surgery.Key Results: Nilotinib increased murine aortic valve thickness. Nilotinib, but not imatinib, promoted calcification and osteogenic activation and decreased autophagy in human VICs. Differential tyrosine kinase expression was detected between healthy and calcified valve tissue. Transcriptomic target identification revealed that the discoidin domain receptor DDR2, which is preferentially inhibited by nilotinib, was predominantly expressed in human aortic valves but markedly downregulated in calcified valve tissue. Nilotinib and selective DDR2 targeting in VICs induced a similar osteogenic activation, which was blunted by increasing the DDR2 ligand, collagen.Conclusions and Implications: These findings suggest that inhibition of DDR2 by nilotinib promoted aortic valve thickening and VIC calcification, with possible translational implications for cardiovascular surveillance and possible personalized medicine in CML patients. [ABSTRACT FROM AUTHOR]

Easily accessible biomarkers are needed to diagnose cardiovascular disease precisely – particularly, to distinguish between disease subtypes that are encountered in clinical practice. Per the hypothesis that plasma miRNA is valuable for this purpose, we performed complete transcriptional profiling of an miRNA discovery-set in 14 samples: three patients with ST-elevated acute myocardial infarction (STEMI) at baseline and after three months of follow-up, four with stable ischaemic heart disease (stable-IHD) and four healthy age-matched volunteers. Our aim was to determine whether we could distinguish patients with unstable plaques from stable patients following a STEMI event. After analysing miRNA profiles, we conducted a validation study comparing three-month STEMI (n=40) with stable-IHD (n=35), which confirmed that miR-486-3P differentiates patients with three-month STEMI from those with stable-IHD (P=0.019). [ABSTRACT FROM AUTHOR]

Introduction: An assay to determine glucocorticoid (GC) responsiveness in humans could be used to monitor GC non-responsiveness in states of GC insufficiency and could provide a tool to adapt GC treatment to individual patients. We propose an ex vivo assay to test GC responsiveness in peripheral leukocytes. The assay was evaluated in a human experimental model of surgery-induced inflammation. Patients and methods: Changes in expression of the GC-regulated genes GILZ, IL1R2, FKBP5, and HLA-DR and glucocorticoid receptor alpha (GRα) were determined by reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) in peripheral leukocytes from surgical patients and healthy blood donors (total n=60) in response to low (1 nM) and high (1 μM) dexamethasone (DEX). The final selection of a suitable endogenous control gene was based on the studies of stability during DEX treatment and inflammation. Correlations between pre- and postoperative GC-induced gene expression, the postoperative systemic inflammatory and metabolic response (CRP, IL-6, white blood cell count, cytokines, resistin, free fatty acids, glucose, insulin, and adiponectin), and the clinical outcome were analyzed. The length of stay in the intensive care unit (ICU-LOS), the length of stay in the hospital, and postoperative complications were used to measure clinical outcome. Results: When the blood donors were compared to the patients, there were no significant differences in the regulation of the genes in response to DEX, except for GRα. Preoperative, but not postoperative, gene regulation of GILZ and GRα was negatively correlated to ICU-LOS (P<0.05 and P<0.01, respectively). Preoperative GILZ and FKBP5 gene regulation was negatively correlated to postoperative systemic TNFα and MIP-1α levels. Conclusion: We suggest that this assay could be used to determine GC responsiveness. An alteration in preoperative GC responsiveness may be related to a patient's ability to recover from surgically induced inflammatory stress. [ABSTRACT FROM AUTHOR]

Pefilipin 2 (PLIN2) is the most abunda lipid droplet (LD)-associated protein in nonadipose tissue, and its expression correlates with intracellular lipid accumulation. Here we identified a missens polymorphism, Ser251Pro, that has major effect on protein structure and function, along with an influence on human plasma triglyceride concentration. The evolutionarily conserved Ser251Pro polymorphism was identified with the ClustalW program. Structure modeling using 3D-JigSaw and the Chimera package revealed that the Pro251 allele disrupts a predicted α-helix in PIXq2 Analyses of macrophages from individuals carrying Ser251Pro variants and human embryonic kidney 29 (HEK293) cells stably transfected with either of the alleles demonstrated that the Pro251 variant caus increased lipid accumulation and decreased fipolysis Analysis of LD size distribution in stably transfected cells showed that the minor Pro251 allele resulted in an increased number of small LDs per cell and increased perilipin 3 protein expression levels as compared with cells carrying the major Ser251 allele. Genotyping of 2113 individuals indicated that the Pro251 variant is associated with decreased plasma triglyceride an very low-density lipoprotein concentrations. Altogether, these data provide the first evidence of a polymorphis in PLIN2 that affects PLIN2 function and may influence the development of metabolic and cardiovascular diseases. [ABSTRACT FROM AUTHOR]

Interprofessional education (IPE) involving undergraduate health professionals is expected to promote collaboration in their later careers. The role of IPE between doctors and biomedical scientists has not been explored at the undergraduate level. Our aim was to introduce IPE sessions for medical and biomedical students in order to identify the benefits and barriers to these groups learning together. Medical and biomedical students together discussed laboratory results, relevant literature, and ideas for developing new diagnostic tools. The programme was evaluated with questionnaires and interviews. While there was general support for the idea of IPE, medical and biomedical students responded differently. Biomedical students were more critical, wanted more explicit learning objectives and felt that their professional role was often misunderstood. The medical students were more enthusiastic but regarded the way the biomedical students communicated concerns about their perceived role as a barrier to effective interprofessional learning. We conclude that stereotyping, which can impede effective collaborations between doctors and biomedical scientists, is already present at the undergraduate level and may be a barrier to IPE. Effective learning opportunities should be supported at the curriculum level and be designed to specifically enable a broad appreciation of each other's future professional roles. [ABSTRACT FROM AUTHOR]

An inquiry-based laboratory course was created in an effort to increase student responsibility in learning and to improve teaching in areas related to molecular medicine. Authentic medical cases with both scientific and clinical aspects formed the basis of a project-oriented course that also included student laboratory work focused on the disease-related proteins. Students used basic biochemical techniques to develop and test hypotheses relating their results to the clinical findings. The course also included patient demonstrations to personalize students' knowledge of case presentations, lectures on basic biochemical principles relevant to the molecular basis of the cases, and seminars by invited guests with expertise in translational medicine. Students developed proposals for future research as part of the final examination. An inquiry matrix was used to evaluate the degree of learning responsibility taken during the course. By allowing for openness in how to explore the case including choice of methods and interpretation of unexpected results, students gained confidence in their ability to solve problems, formulate and test hypotheses, and collaborate with both clinical and non-clinical professionals. [ABSTRACT FROM AUTHOR]

Activators of peroxisome proliferator-activated receptor (PPAR)γ have been studied intensively for their insulin-sensitizing properties and antidiabetic effects. Recently, a specific PPARδ activator (GW501516) was reported to attenuate plasma glucose and insulin levels when administered to genetically obese ob/ob mice. This study was performed to determine whether specific activation of PPARδ has direct effects on insulin action in skeletal muscle. Specific activation of PPARδ using two pharmacological agonists (GW501516 and GW0742) increased glucose uptake independently of insulin in differentiated C2C12 myotubes. In cultured primary human skeletal myotubes, GW501516 increased glucose uptake independently of insulin and enhanced subsequent insulin stimulation. PPARδ agonists increased the respective phosphorylation and expression of AMP-activated protein kinase 1.9-fold (P < 0.05) and 1.8-fold (P < 0.05), of extracellular signal-regulated kinase ½ mitogen-activated protein kinase (MAPK) 2.2-fold (P < 0.05) and 1.7-fold (P < 0.05), and of p38 MAPK 1.2-fold (P < 0.05) and 1.4-fold (P < 0.05). Basal and insulin-stimulated protein kinase B/Akt was unaltered in cells preexposed to PPARδ agonists. Preincubation of myotubes with the p38 MAPK inhibitor SB203580 reduced insulin- and PPARδ-mediated increase in glucose uptake, whereas the mitogen-activated protein kinase kinase inhibitor PD98059 was without effect. PPARδ agonists reduced mRNA expression of PPARδ, sterol regulatory element binding protein (SREBP)-la, and SREBP-1c (P < 0.05). In contrast, mRNA expression of PPARγ, PPARγ coactivator 1, GLUT1, and GLUT4 was unaltered. Our results provide evidence to suggest that PPARδ agonists increase glucose metabolism and promote gene regulatory responses in cultured human skeletal muscle. Moreover, we provide biological validation of PPARδ as a potential target for antidiabetic therapy. Diabetes 54:1157-1163, 2005 [ABSTRACT FROM AUTHOR]

Highly active antiretroviral therapy (HAART) has improved the prognosis of human immunodeficiency virus (HIV)-infected patients but is associated with severe adverse events, such as lipodystrophy and insulin resistance. Rosiglitazone did not increase subcutaneous fat in patients with HAART-associated lipodystrophy (HAL) in a randomized, double-blind, placebo-controlled trial, although it attenuated insulin resistance and decreased liver fat content. The aim of this study was to examine effects of rosiglitazone on gene expression in subcutaneous adipose tissue in 30 patients with HAL. The mRNA concentrations in subcutaneous adipose tissue were measured using real-time PCR. Twenty-four-week treatment with rosiglitazone (8 mg/day) compared with placebo significantly increased the expression of adiponectin, peroxisome proliferator-activated receptor-γ (PPARγ), and PPARγ coactivator 1 and decreased IL-6 expression. Expression of other genes involved in lipogenesis, fatty acid metabolism, or glucose transport, such as acyl-CoA synthase, adipocyte lipid-binding protein, CD45, fatty acid transport protein-1 and -4, GLUT1, GLUT4, keratinocyte lipid-binding protein, lipoprotein lipase, PPARδ, and sterol regulatory element-binding protein 1c, remained unchanged. Rosiglitazone also significantly increased serum adiponectin concentration. The change in serum adiponectin concentration was inversely correlated with the change in fasting serum insulin concentration and liver fat content. In conclusion, rosiglitazone induced significant changes in gene expression in subcutaneous adipose tissue and ameliorated insulin resistance in patients with HAL. Increased expression of adiponectin might have mediated most of the favorable insulin-sensitizing effects of rosiglitazone in these patients. [ABSTRACT FROM AUTHOR]

Insulin-like growth factor-binding protein 6 (IGFBP6), an extracellular protein with preferential affinity for insulin-like growth factor (IGF) II, belongs to a family of binding proteins with at least six members. We have characterized the genomic structure and the chromosomal location of the human IGFBP6, which is present in the human genome as a single-copy gene spanning 4.7 kb. It consists of four exons, encoding the translated regions, with sizes of 334, 146, 120, and 123 bp, while the intervening introns are 2661, 182, and 844 bp. Three mRNA cap sites were localized 101, 100, and 96 bp upstream of the ATG translation start codon as determined by S1 nuclease analysis. The proximal 5′-flanking region does not have any TATA or CAAT consensus sequences. The IGFBP6 was localized to Chr 12 by analysis of somatic cell hybrids and regionalized to 12q13 by fluorescence DNA in situ hybridization. [ABSTRACT FROM AUTHOR]

Insulin-like growth-factor-binding proteins (IGFBPs) constitute a family of structurally related proteins that specifically bind insulin-like growth factors and modulate their functions. In this study, the chromosomal localization was determined for the gene encoding IGFBP4, i.e. inhibitory-IGFBP. A polymerase chain reaction (PCR) fragment corresponding to the previously published cDNA sequence was used to isolate overlapping cosmid clones. By fluorescent in situ hybridization to metaphase chromosomes, the IGFBP4 gene was then localized to chromosomal region 17q21-21.1. This result was in agreement with PCR analysis of a panel of somatic cell hybrids. [ABSTRACT FROM AUTHOR]

Autophagy serves as a cell survival mechanism which becomes dysregulated under pathological conditions and aging. Aortic valve thickening and calcification causing left ventricular outflow obstruction is known as calcific aortic valve stenosis (CAVS). CAVS is a chronic and progressive disease which increases in incidence and severity with age. Currently, no medical treatment exists for CAVS, and the role of autophagy in the disease remains largely unexplored. To further understand the role of autophagy in the progression of CAVS, we analyzed expression of key autophagy genes in healthy, thickened, and calcified valve tissue from 55 patients, and compared them with nine patients without significant CAVS, undergoing surgery for aortic regurgitation (AR). This revealed a upregulation in autophagy exclusively in the calcified tissue of CAVS patients. This difference in autophagy between CAVS and AR was explored by LC3 lipidation in valvular interstitial cells (VICs), revealing an upregulation in autophagic flux in CAVS patients. Inhibition of autophagy by bafilomycin-A1 led to a decrease in VIC survival. Finally, treatment of VICs with high phosphate led to an increase in autophagic activity. In conclusion, our data suggests that autophagy is upregulated in the calcified tissue of CAVS, serving as a compensatory and pro-survival mechanism. [ABSTRACT FROM AUTHOR]

Background: Takotsubo cardiomyopathy (TCM), also known as “broken heart syndrome”, is a type of heart failure characterized by transient ventricular dysfunction in the absence of obstructive coronary lesions. Although associated with increased levels of catecholamines, pathophysiological mechanisms are unknown. Relapses and family heritability indicate a genetic predisposition. Several small studies have investigated associations between three different loci; the β1-adrenic receptor (ADRB1), G-protein-coupled receptor kinase 5 (GRK5), Bcl-associated athanogene 3 (BAG3) and TCM but no consensus has been reached. Methods: Participants were recruited using the Swedish Coronary Angiography and Angioplasty Register (SCAAR). TCM patients without coronary artery disease (CAD)(n = 258) were identified and age- and sex-matched subjects with (n = 164) and without (n = 243) CAD were selected as controls. DNA was isolated from saliva and genotyped for candidate single nucleotide polymorphisms in the ADRB1, GRK5 and BAG3 genes. Allele frequencies and Odds Ratios (OR) with 95% Confidence Intervals (CI) for the investigated polymorphisms were compared, respectively calculated for TCM patients and controls. Results: There were no differences in allele frequencies between TCM patients and controls. OR (CI) for TCM patients having at least one minor allele using controls as reference were 1.07 (0.75–1.55) for ADRB1, 0.45 (0.11–1.85) for GRK5 and 1.27 (0.74–2.19) for BAG3. Conclusion: By genotyping a large takotsubo cohort, we demonstrate a lack of association between candidate SNPs in the ADRB1, GRK5 and BAG3 genes, earlier suggested to contribute to TCM. Our result indicates a need to expand the search for new genetic candidates contributing to TCM. [ABSTRACT FROM AUTHOR]

Context:Nonfasting (postprandial) triglyceride concentrations have emerged as a clinically significant cardiovascular disease risk factor that results from accumulation of remnant triglyceride-rich lipoproteins (TRLs) in the circulation. The remnant TRLs are cleared from the circulation by hepatic uptake, but the specific mechanisms involved are unclear. The syndecan-1 heparan sulfate proteoglycan (HSPG) pathway is important for the hepatic clearance of remnant TRLs in mice, but its relevance in humans is unclear. Objective:We sought to determine whether polymorphisms of the genes responsible for HSPG assembly and disassembly contribute to atherogenic dyslipoproteinemias in humans. Patients And Design:We performed an oral fat load in 68 healthy subjects. Lipoproteins (chylomicrons and very low density lipoproteins 1 and 2) were isolated from blood, and the area under curve and incremental area under curve for postprandial variables were calculated. Single nucleotide polymorphisms in genes encoding syndecan-1 and enzymes involved in the synthesis or degradation of HSPG were genotyped in the study subjects. Results:Our results indicate that the genetic variation rs2281279 in SULF2 associates with postprandial clearance of remnant TRLs and triglyceride levels in healthy subjects. Furthermore, the SNP rs2281279 in SULF2 associates with hepatic SULF2 mRNA levels. Conclusions:In humans, mild but clinically relevant postprandial hyperlipidemia due to reduced hepatic clearance of remnant TRLs may result from genetic polymorphisms that affect hepatic HSPG. [ABSTRACT FROM AUTHOR]