Secretogranin II; a Protein Increased in the Myocardium and Circulation in Heart Failure with Cardioprotective Properties

Background: Several beneficial effects have been demonstrated for secretogranin II (SgII) in non-cardiac tissue. As cardiac production of chromogranin A and B, two related proteins, is increased in heart failure (HF), we hypothesized that SgII could play a role in cardiovascular pathophysiology. Methodology/Principal Findings: SgII production was characterized in a post-myocardial infarction heart failure (HF) mouse model, functional properties explored in experimental models, and circulating levels measured in mice and patients with stable HF of moderate severity. SgII mRNA levels were 10.5 fold upregulated in the left ventricle (LV) of animals with myocardial infarction and HF (p<0.001 vs. sham-operated animals). SgII protein levels were also increased in the LV, but not in other organs investigated. SgII was produced in several cell types in the myocardium and cardiomyocyte synthesis of SgII was potently induced by transforming growth factor-beta and norepinephrine stimulation in vitro. Processing of SgII to shorter peptides was enhanced in the failing myocardium due to increased levels of the proteases PC1/3 and PC2 and circulating SgII levels were increased in mice with HF. Examining a pathophysiological role of SgII in the initial phase of post-infarction HF, the SgII fragment secretoneurin reduced myocardial ischemia-reperfusion injury and cardiomyocyte apoptosis by 30% and rapidly increased cardiomyocyte Erk1/2 and Stat3 phosphorylation. SgII levels were also higher in patients with stable, chronic HF compared to age-and gender-matched control subjects: median 0.16 (Q1-3 0.14-0.18) vs. 0.12 (0.10-0.14) nmol/L, p<0.001. Conclusions: We demonstrate increased myocardial SgII production and processing in the LV in animals with myocardial infarction and HF, which could be beneficial as the SgII fragment secretoneurin protects from ischemia-reperfusion injury and cardiomyocyte apoptosis. Circulating SgII levels are also increased in patients with chronic, stable HF and may represent a new cardiac biomarker

"Dynamic aspects of toxin-induced alterations in AMP-activated protein kinase and autophagy in isolated rat hepatocytes" : "Toxin-induced AMPK alterations"

In isolated rat hepatocytes, okadaic acid and cantharidin activate AMP-activated protein kinase (AMPK) by a naringin-sensitive and a naringin-resistant mechanism respectively, probably through inhibition of protein phosphatases PP2A and PP1, respectively. The activating phosphorylation of AMPKá at Thr172 (detected by immunoblotting with a pThr172-specific antibody) induced by a 1-h treatment with okadaic acid was subsequently very stable at all toxin concentrations tested (30-100 nM), persisting even 2 h after drug removal. AMPK phosphorylation induced by cantharidin (30 µM) was also completely stable for 2 h after removal of the toxin. Similarly, in correlation with the stable (Thr172) AMPKá phosphorylation, autophagy was inhibited in an irreversible manner upon incubation with 30 or 50 nM okadaic acid. The stress-activated protein kinases, SAPK/Erk kinase (SEK1) and C-Jun NH2 terminal kinase (JNK), as well as S6 kinase and its substrate, S6, have been suggested as downstream components in a LKB1/AMPK-initiated signalling pathway and are phosphorylated following toxin treatment. However, unlike AMPK, all these phosphoproteins showed a reversal of phosphorylation upon toxin removal, questioning their roles as downstream mediators in an autophagy-regulating pathway. At 30 nM and higher okadaic concentrations, AMPK phosphorylation was evident in immunoblots as at least three distinct and adjacent bands, the uppermost low-mobility form being predominant. This multi-phosphorylated AMPK was clearly enzymatically active in situ, as shown by the effective phosphorylation of its direct substrate, acetyl-CoA carboxylase (ACC) at Ser79. With a shorter incubation time (20 min) as well as at lower okadaic acid concentrations (10-15 nM), the monophosphorylated high-mobility form was the only band present. This monophosphorylated AMPK appeared to be enzymatically inactive in relation to ACC phosphorylation, but still seemed to be able to suppress autophagic sequestration significantly. In addition, monophosphorylated AMPK allowed the phosphorylation of SEK1 (pThr261), JNK (pThr183/Tyr185) and S6K (pThr421/Ser424), but not of S6 (pSer235/236). Thus, mono- and multi- (full) phosphosphorylated AMPK seem to have slightly different substrate preferences. For further examination of the Thr172 AMPK immuno-stained bands, isoelectric focusing (IEF) and 2-D gel electrophoresis were utilized as protein separation methods. Identification by these techniques has been a difficult task, since phosphorylated AMPK has not been detectable. By using a general antibody against the AMPKá subunit, three distinct spots were detected after IEF. Two of these were identified as the á2 isoform

m6A Regulators in Human Adipose Tissue - Depot-Specificity and Correlation With Obesity

Background: N6-methyladenosine (m6A) is one of the most abundant post-transcriptional modifications on mRNA influencing mRNA metabolism. There is emerging evidence for its implication in metabolic disease. No comprehensive analyses on gene expression of m6A regulators in human adipose tissue, especially in paired adipose tissue depots, and its correlation with clinical variables were reported so far. We hypothesized that inter-depot specific gene expression of m6A regulators may differentially correlate with clinical variables related to obesity and fat distribution. Methods: We extracted intra-individually paired gene expression data (omental visceral adipose tissue (OVAT) N=48; subcutaneous adipose tissue (SAT) N=56) of m6A regulators from an existing microarray dataset. We also measured gene expression in another sample set of paired OVAT and SAT (N=46) using RT-qPCR. Finally, we extracted existing gene expression data from peripheral mononuclear blood cells (PBMCs) and single nucleotide polymorphisms (SNPs) in METTL3 and YTHDF3 from genome wide data from the Sorbs population (N=1049). The data were analysed for differential gene expression between OVAT and SAT; and for association with obesity and clinical variables. We further tested for association of SNP markers with gene expression and clinical traits. Results: In adipose tissue we observed that several m6A regulators (WTAP, VIRMA, YTHDC1 and ALKBH5) correlate with obesity and clinical variables. Moreover, we found adipose tissue depot specific gene expression for METTL3, WTAP, VIRMA, FTO and YTHDC1. In PBMCs, we identified ALKBH5 and YTHDF3 correlated with obesity. Genetic markers in METTL3 associate with BMI whilst SNPs in YTHDF3 are associated with its gene expression. Conclusions: Our data show that expression of m6A regulators correlates with obesity, is adipose tissue depot-specific and related to clinical traits. Genetic variation in m6A regulators adds an additional layer of variability to the functional consequences

Circulating MicroRNA-210 Concentrations in Patients with Acute Heart Failure: Data from the Akershus Cardiac Examination 2 Study

Background MicroRNA (miR)-210 expression is induced by acute and chronic hypoxia and provides prognostic information in patients with aortic stenosis and acute coronary syndrome. We hypothesized that circulating miR-210 concentrations could provide diagnostic and prognostic information in patients with acute heart failure (HF). Methods We measured miR-210 concentrations in serum samples on admission from 314 patients hospitalized for acute dyspnea and 9 healthy control subjects. The diagnostic and prognostic properties of miR-210 were tested in patients after adjudication of all diagnoses and with median follow-up of 464 days. Results All patients and control subjects had miR-210 concentrations within the range of detection, and the analytical variation was low as the coefficient of variation of synthetic spike-in RNA was 4%. Circulating miR-210 concentrations were increased in patients with HF compared to healthy control subjects, but miR-210 concentrations did not separate patients with acute HF (n = 143) from patients with non-HF-related dyspnea (n = 171): the area under the curve was 0.50 (95% CI 0.43–0.57). Circulating miR-210 concentrations were associated with mortality (n = 114) after adjustment for clinical risk factors (hazard ratio 1.65 [95% CI 1.03–2.62] per unit miR-210 increase), but this association was attenuated and not significant after adjustment for established cardiac protein biomarkers. Conclusions Circulating miR-210 concentrations are associated with mortality, but do not add to established protein biomarkers for diagnosis or prognosis in patients with acute dyspnea

Circulating MicroRNA-210 Concentrations in Patients with Acute Heart Failure: Data from the Akershus Cardiac Examination 2 Study

Background MicroRNA (miR)-210 expression is induced by acute and chronic hypoxia and provides prognostic information in patients with aortic stenosis and acute coronary syndrome. We hypothesized that circulating miR-210 concentrations could provide diagnostic and prognostic information in patients with acute heart failure (HF). Methods We measured miR-210 concentrations in serum samples on admission from 314 patients hospitalized for acute dyspnea and 9 healthy control subjects. The diagnostic and prognostic properties of miR-210 were tested in patients after adjudication of all diagnoses and with median follow-up of 464 days. Results All patients and control subjects had miR-210 concentrations within the range of detection, and the analytical variation was low as the coefficient of variation of synthetic spike-in RNA was 4%. Circulating miR-210 concentrations were increased in patients with HF compared to healthy control subjects, but miR-210 concentrations did not separate patients with acute HF (n = 143) from patients with non-HF-related dyspnea (n = 171): the area under the curve was 0.50 (95% CI 0.43–0.57). Circulating miR-210 concentrations were associated with mortality (n = 114) after adjustment for clinical risk factors (hazard ratio 1.65 [95% CI 1.03–2.62] per unit miR-210 increase), but this association was attenuated and not significant after adjustment for established cardiac protein biomarkers. Conclusions Circulating miR-210 concentrations are associated with mortality, but do not add to established protein biomarkers for diagnosis or prognosis in patients with acute dyspnea

Prognostic Value of Circulating MicroRNA-210 Levels in Patients with Moderate to Severe Aortic Stenosis

<div><p>Background</p><p>Circulating micro-RNAs have been proposed as a novel class of cardiovascular (CV) biomarkers, but whether they meet analytical requirements and provide additional information to establish risk indices have not been established. miR-210 levels are increased in subjects with low VO₂ max, which is a recognized risk factor in patients with aortic stenosis (AS), and we hypothesized that circulating miR-210 levels may be increased in patients with AS and associated with a poor prognosis.</p><p>Methods</p><p>We measured circulating miR-210 levels by real-time PCR in 57 patients with moderate to severe AS and in 10 age- and gender-matched healthy controls. The merit of miR-210 as a biomarker was assessed according to established criteria, including by comparing miR-210 levels with NT-proBNP and miR-22 levels, which is another miRNA biomarker candidate.</p><p>Results</p><p>All patients and control subjects had miR-210 levels within the range of detection (Cq<35) and the analytical variability was low. Circulating miR-210 levels were 2.0±0.2 [mean±SEM] fold increased in AS patients compared to controls (p = 0.002), whereas miR-22 levels were not differently expressed in the AS patients (0.12±0.06 fold increase, p = 0.45). The increase in miR-210 levels in AS patients was comparable to the increment in NT-proBNP levels: [AUC] 0.82 (95% CI 0.70–0.90) vs. 0.85 (0.75–0.93), respectively, p = 0.71. During a median follow-up of 1287 days, 15 patients (26%) died. There was a significant association between higher circulating levels of miR-210 and increased mortality during follow-up: hazard ratio [supra- vs. inframedian levels] 3.3 (95% CI 1.1–10.5), p = 0.039. Adjusting for other risk indices in multivariate analysis did not attenuate the prognostic merit of circulating miR-210 levels.</p><p>Conclusion</p><p>Circulating miR-210 levels are increased in patients with AS and provide independent prognostic information to established risk indices. Analytical characteristics were also excellent supporting the potential of micro-RNAs as novel CV biomarkers.</p></div

Echocardiographical data in the patients with aortic stenosis and control subjects.

<p>LV, left ventricular; and LVEF, left ventricular ejection fraction.</p><p><b>*</b>n = 53 for E/e’ and n = 52 for E/A ratio.</p