Claudia Bang

The miRNA-212/132 family regulates both cardiac hypertrophy and cardiomyocyte autophagy

Pathological growth of cardiomyocytes (hypertrophy) is a major determinant for the development of heart failure, one of the leading medical causes of mortality worldwide. Here we show that the microRNA (miRNA)-212/132 family regulates cardiac hypertrophy and autophagy in cardiomyocytes. Hypertrophic stimuli upregulate cardiomyocyte expression of miR-212 and miR-132, which are both necessary and sufficient to drive the hypertrophic growth of cardiomyocytes. MiR-212/132 null mice are protected from pressure-overload-induced heart failure, whereas cardiomyocyte-specific overexpression of the miR-212/132 family leads to pathological cardiac hypertrophy, heart failure and death in mice. Both miR-212 and miR-132 directly target the anti-hypertrophic and pro-autophagic FoxO3 transcription factor and overexpression of these miRNAs leads to hyperactivation of pro-hypertrophic calcineurin/NFAT signalling and an impaired autophagic response upon starvation. Pharmacological inhibition of miR-132 by antagomir injection rescues cardiac hypertrophy and heart failure in mice, offering a possible therapeutic approach for cardiac failure.

Diagnostic and prognostic impact of six circulating microRNAs in acute coronary syndrome

Circulating microRNAs may have diagnostic potential in acute coronary syndrome (ACS). Previous studies, however, were based on low patient numbers and could not assess the relation of microRNAs to clinical characteristics and their potential prognostic value. We thus assessed the diagnostic and prognostic value of cardiomyocyte-enriched microRNAs in the context of clinical variables and a sensitive myonecrosis biomarker in a larger ACS cohort. MiR-1, miR-133a, miR-133b, miR-208a, miR-208b, and miR-499 concentrations were measured by quantitative reverse transcription PCR in plasma samples obtained on admission from 444 patients with ACS. High-sensitivity troponin T (hsTnT) was measured by immunoassay. Patients were followed for 6 months regarding all-cause mortality. In a multiple linear regression analysis that included clinical variables and hsTnT, miR-1, miR-133a, miR-133b, and miR-208b were independently associated with hsTnT levels (all P<0.001). Patients with myocardial infarction presented with higher levels of miR-1, miR-133a, and miR-208b compared with patients with unstable angina. However, all six investigated microRNAs showed a large overlap between patients with unstable angina or myocardial infarction. MiR-133a and miR-208b levels were significantly associated with the risk of death in univariate and age- and gender-adjusted analyses. Both microRNAs lost their independent association with outcome upon further adjustment for hsTnT. The present study tempers speculations about the potential usefulness of cardiomyocyte-enriched microRNAs as diagnostic or prognostic markers in ACS.

Circulating MicroRNAs as Biomarkers and Potential Paracrine Mediators of Cardiovascular Disease

MicroRNAs (miRNAs) comprise a class of small, noncoding RNAs that control expression of complementary target mRNAs. Dysregulation of intracellular miRNA expression has been described in various diseases, including a number of cardiovascular conditions. Functional studies have shown a role for miRNAs in cardiac fibrosis, hypertrophy, angiogenesis, and heart failure.1,–,3 These findings suggest a new therapeutic entry point for cardiac disease and illustrate the broad therapeutic potential of miRNA modulation. Initial reports have detected circulating extracellular miRNAs in the serum/plasma of patients with cancer.4 Despite the existence of RNases, miRNAs remain stable in serum and other body fluids. One explanation is the inclusion of miRNAs into lipid or lipoprotein complexes such as exosomes5 or microvesicles.6 Subsequently, altered concentrations of miRNAs have been found in patients with various cardiovascular diseases. Here, we review the current knowledge about circulating miRNAs during coronary artery disease (CAD), myocardial infarction, and heart failure (Table). A further new and exciting function of circulating miRNAs in the cardiovascular system may be their potential to serve as paracrine signaling molecules.7 The detection of circulating miRNAs in serum/plasma8,9 suggests that miRNAs may fulfill biological functions outside the cell and serve as potential biomarkers for diseases. Circulating miRNAs are protected from RNase-dependent degradation by several mechanisms, including their inclusion in microvesicles, exosomes, and apoptotic bodies as well as through the formation of protein-miRNA complexes resistant to degradation (Figure). In addition, circulating miRNAs are quite stable even after multiple freeze-thaw cycles.8 Recent studies have shown that miRNAs are actively secreted in microvesicles or exosomes from different cell types.6,10 Despite the current knowledge about the existence of circulating miRNAs and the intercellular transfer of miRNAs from donor …

Exosomes: New players in cell–cell communication

Exosomes are small membrane vesicles of endosomal origin, which are secreted from a variety of cell types. During the 1980s exosomes were first described as organelles to remove cell debris and unwanted molecules. The discovery that exosomes contain proteins, messenger and microRNAs suggests a role as mediators in cell-to-cell communication. Exosomes can be transported between different cells and influence physiological pathways in the recipient cells. In the present review, we will summarize the biological function of exosomes and their involvement in physiological and pathological processes. Moreover, the potential clinical application of exosomes as biomarkers and therapeutic tools will be discussed.

Novel techniques and targets in cardiovascular microRNA research

MicroRNAs (miRNAs) are highly conserved, tiny (∼22 nucleotides) non-coding RNAs that have emerged as potent regulators of mRNA translation. miRNAs exhibit fine-tuning of the control of proteins involved in cell signalling (AE) pathways and in vital cellular and developmental processes. miRNAs are expressed in cardiovascular tissues, and multiple functional aspects of miRNAs underscore their key role in cardiovascular (patho)physiology. The development and increasing use of novel molecular biology tools have contributed to the recent success in miRNA research. In the present review, we discuss current updates on important and novel miRNA techniques, including: (i) miRNA screening tools; (ii) bioanalytical target prediction tools; (iii) target validation tools; and (iv) manipulative miRNA expression tools. We also present an update about recently identified miRNA targets that play a key role in cardiovascular development and disorders.

Osteopontin is indispensible for AP1-mediated angiotensin II-related miR-21 transcription during cardiac fibrosis

Aims Osteopontin (OPN) is a multifunctional cytokine critically involved in cardiac fibrosis. However, the underlying mechanisms are unresolved. Non-coding RNAs are powerful regulators of gene expression and thus might mediate this process. Methods and results OPN and miR-21 were significantly increased in cardiac biopsies of patients with myocardial fibrosis. Ang II infusion via osmotic minipumps led to specific miRNA regulations with miR-21 being strongly induced in wild-type (WT) but not OPN knockout (KO) mice. This was associated with enhanced cardiac collagen content, myofibroblast activation, ERK-MAP kinase as well as AKT signalling pathway activation and a reduced expression of Phosphatase and Tensin Homologue (PTEN) as well as SMAD7 in WT but not OPN KO mice. In contrast, cardiotropic AAV9-mediated overexpression of OPN in vivo further enhanced cardiac fibrosis. In vitro, Ang II induced expression of miR-21 in WT cardiac fibroblasts, while miR-21 levels were unchanged in OPN KO fibroblasts. As pri-miR-21 was also increased by Ang II, we studied potential involved upstream regulators; Electrophoretic Mobility Shift and Chromatin Immunoprecipitation analyses confirmed activation of the miR-21 upstream-transcription factor AP-1 by Ang II. Recombinant OPN directly activated miR-21, enhanced fibrosis, and activated the phosphoinositide 3-kinase pathway. Locked nucleic acid-mediated miR-21 silencing ameliorated cardiac fibrosis development in vivo. Conclusion In cardiac fibrosis related to Ang II, miR-21 is transcriptionally activated and targets PTEN/SMAD7 resulting in increased fibroblast survival. OPN KO animals are protected from miR-21 increase and fibrosis development due to impaired AP-1 activation and fibroblast activation.

Impairment of Wound Healing in Patients With Type 2 Diabetes Mellitus Influences Circulating MicroRNA Patterns via Inflammatory Cytokines

Objective—MicroRNAs (miRNA/miR) are stably present in body fluids and are increasingly explored as disease biomarkers. Here, we investigated influence of impaired wound healing on the plasma miRNA signature and their functional importance in patients with type 2 diabetes mellitus. Approach and Results—miRNA array profiling identified 41 miRNAs significantly deregulated in diabetic controls when compared with patients with diabetes mellitus–associated peripheral arterial disease and chronic wounds. Quantitative real-time polymerase chain reaction validation confirmed decrease in circulating miR-191 and miR-200b levels in type 2 diabetic versus healthy controls. This was reverted in diabetic subjects with associated peripheral arterial disease and chronic wounds, who also exhibited higher circulating C-reactive protein and proinflammatory cytokine levels compared with diabetic controls. miR-191 and miR-200b were significantly correlated with C-reactive protein or cytokine levels in patients with diabetes mellitus. Indeed, proinflammatory stress increased endothelial- or platelet-derived secretion of miR-191 or miR-200b. In addition, dermal cells took up endothelial-derived miR-191 leading to downregulation of the miR-191 target zonula occludens-1. Altered miR-191 expression influenced angiogenesis and migratory capacities of diabetic dermal endothelial cells or fibroblasts, respectively, partly via its target zonula occludens-1. Conclusions—This study reports that (1) inflammation underlying nonhealing wounds in patients with type 2 diabetes mellitus influences plasma miRNA concentrations and (2) miR-191 modulates cellular migration and angiogenesis via paracrine regulation of zonula occludens-1 to delay the tissue repair process.

Cardiovascular importance of the microRNA-23/27/24 family.

Please cite this paper as: Bang C, Fiedler J, Thum T. Cardiovascular importance of the microRNA‐23/27/24 family. Microcirculation19: 208–214, 2012.

TWIST1 regulates the activity of ubiquitin proteasome system via the miR-199/214 cluster in human end-stage dilated cardiomyopathy.

BACKGROUND The transcription factor TWIST1 has been described to regulate the microRNA (miR)-199/214 cluster. Genetic disruption of TWIST1 resulted in a cachectic phenotype and early death of the knock-out mice. This might be connected to the activity of the ubiquitin-proteasome-system (UPS), as miR-199a has been suggested to regulate the ubiquitin E2 ligases Ube2i and Ube2g1. METHODS Cardiac tissue from explanted hearts of 42 patients with dilated cardiomyopathy and 20 healthy donor hearts were analysed for protein expression of TWIST1 and its inhibitors Id-1, MuRF-1 and MAFbx, the expression of miR-199a, -199b and -214, as well as the activity of the UPS by using specific fluorogenic substrates. RESULTS TWIST1 was repressed in patients with dilated cardiomyopathy by 43% (p=0.003), while Id1 expression was unchanged. This was paralleled by a reduced expression of miR-199a by 38 ± 9% (p=0.053), miR-199b by 36 ± 13% (p=0.019) and miR-214 by 41 ± 11% (p=0.0158) compared to donor hearts. An increased peptidylglutamyl-peptide-hydrolysing activity (p<0.0001) was observed in the UPS, while the chymotrypsin-like and trypsin-like activities were unchanged. The protein levels of the rate limiting ubiquitin E3-ligases MuRF-1 and MAFbx were up-regulated (p=0.005 and p=0.0156, respectively). Mechanistically silencing of TWIST1 using siRNA in primary rat cardiomyocytes led to a down-regulation of the miR-199/214 cluster and to a subsequent up-regulation of Ube2i. CONCLUSION The TWIST1/miR-199/214 axis is down-regulated in dilated cardiomyopathy, which is likely to play a role in the increased activity of the UPS. This may contribute to the loss of cardiac mass during dilatation of the heart.

Circulating long-non coding RNAs as biomarkers of left ventricular diastolic function and remodelling in patients with well-controlled type 2 diabetes

Contractile dysfunction is underdiagnosed in early stages of diabetic cardiomyopathy. We evaluated the potential of circulating long non-coding RNAs (lncRNAs) as biomarkers of subclinical cardiac abnormalities in type 2 diabetes. Forty-eight men with well-controlled type 2 diabetes and 12 healthy age-matched volunteers were enrolled in the study. Left ventricular (LV) parameters were measured by magnetic resonance imaging. A panel of lncRNAs was quantified in serum by RT-qPCR. No differences in expression levels of lncRNAs were observed between type 2 diabetes patients and healthy volunteers. In patients with type 2 diabetes, long intergenic non-coding RNA predicting cardiac remodeling (LIPCAR) was inversely associated with diastolic function, measured as E/A peak flow (P < 0.050 for all linear models). LIPCAR was positively associated with grade I diastolic dysfunction (P < 0.050 for all logistic models). Myocardial infarction-associated transcript (MIAT) and smooth muscle and endothelial cell-enriched migration/differentiation-associated long noncoding RNA (SENCR) were directly associated with LV mass to LV end-diastolic volume ratio, a marker of cardiac remodelling (P < 0.050 for all linear models). These findings were validated in a sample of 30 patients with well-controlled type 2 diabetes. LncRNAs are independent predictors of diastolic function and remodelling in patients with type 2 diabetes.