mRNA PGC-1α levels in blood samples reliably correlates with its myocardial expression: study in patients undergoing cardiac surgery

et al.[Objective]: Peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) is a transcriptional coactivator that has been proposed to play a protective role in mouse models of cardiac ischemia and heart failure, suggesting that PGC-1α could be relevant as a prognostic marker. Our previous studies showed that the estimation of peripheral mRNA PGC-1α expression was feasible and that its induction correlated with the extent of myocardial necrosis and left ventricular remodeling in patients with myocardial infarction. In this study, we sought to determine if the myocardial and peripheral expressions of PGC-1α are well correlated and to analyze the variability of PGC-1α expression depending on the prevalence of some metabolic disorders. [Methods]: This was a cohort of 35 consecutive stable heart failure patients with severe aortic stenosis who underwent an elective aortic valve replacement surgery. mRNA PGC-1α expression was simultaneously determined from myocardial biopsy specimens and blood samples obtained during surgery by quantitative PCR, and a correlation between samples was made using the Kappa index. Patients were divided into two groups according to the detection of baseline expression levels of PGC-1α in blood samples, and comparisons between both groups were made by chi-square test or unpaired Student’s t-test as appropriate. [Results]: Based on myocardial biopsies, we found that mRNA PGC-1α expression in blood samples showed a statistically significant correlation with myocardial expression (Kappa index 0.66, p<0.001). The presence of higher systemic PGC-1α expression was associated with a greater expression of some target genes such as silent information regulator 2 homolog-1 (x-fold expression in blood samples: 4.43±5.22 vs. 1.09±0.14, p=0.044) and better antioxidant status in these patients (concentration of Trolox: 0.40±0.05 vs. 0.34±0.65, p=0.006). [Conclusions]: Most patients with higher peripheral expression also had increased myocardial expression, so we conclude that the non-invasive estimation of mRNA PGC-1α expression from blood samples provides a good approach of the constitutive status of the mitochondrial protection system regulated by PGC-1α and that this could be used as prognostic indicator in cardiovascular disease.Grant from Sociedad Valenciana de Cardiología, 2013 to Óscar Fabregat-Andrés.Peer Reviewe

List of deregulated miRNAs determined by TaqMan Human miRNA Low-Density Array between gastric cancer (n = 13) and normal gastric (n = 12) tissues (FDR adjusted p-value <0.01 and fold change >2).

<p>List of deregulated miRNAs determined by TaqMan Human miRNA Low-Density Array between gastric cancer (n = 13) and normal gastric (n = 12) tissues (FDR adjusted p-value <0.01 and fold change >2).</p

Receiver operating characteristic (ROC) curves of differentially expressed miRNAs in plasma between GC patients and healthy controls.

<p>ROC curves of miR-375 (<b>a</b>); miR-148a-3p (<b>b</b>); and miR-223 (<b>c</b>); the combination of miR-375 and miR-148a-3p (<b>d</b>).</p

Expression levels of candidate miRNA in tissue and plasma samples.

<p>The boxplots for miR-148a-3p (<b>a</b>); miR-204-5p (<b>b</b>); miR-223-3p (<b>c</b>); miR-375 (<b>d</b>); miR-135a-5p (<b>e</b>) and miR-155-5p (<b>f</b>) represent the results of qRT-PCR comparing GC samples with healthy controls. qRT-PCR data are represented as log2 2-(deltaCt) values. The red and blue colors shows expression levels of candidate miRNAs in tissue and plasma samples, respectively. *—FDR adjusted p < 0.05 by Mann-Whitney U test in tissue. **- FDR adjusted p < 0.05 by Mann-Whitney U test in plasma.</p

Network of candidate miRNAs and their putative target genes.

<p>Network includes the individual miRNAs (red circles) and four types of their predicted mRNA target genes (hexagons), obtained from miRTarBase and miRecords databases. The pink color represents target genes which are regulated by a single miRNA. The orange and green colors indicate target genes regulated simultaneously by two or three distinct miRNAs, respectively. GC-associated target genes retrieved from DisGeNet database are represented by blue hexagons. The databases included in the regulatory interaction networks are identified by the color of the connecting arrows: miRTarBase (blue) and miRecords (red).</p

Clinical characteristics of the gastric cancer patients and healthy controls.

<p><i>H</i>. <i>pylori—Helicobacter pylori;</i> GC–gastric cancer.</p><p>Clinical characteristics of the gastric cancer patients and healthy controls.</p

Expression levels of <i>BCL2</i> and <i>DNMT3B</i> in GC tissue and correlation analysis with their putatively targeting miRNAs.

<p>(<b>a</b>) Expression levels of <i>BCL2</i> and <i>DNMT3B</i> was analyzed using qRT-PCR. The data are represented as log2 2-(deltaCt) values; Pearson correlation analysis: (<b>b</b>) between relative expression levels of <i>DNMT3B</i> and relative expression levels miR-375; (<b>c</b>) between relative expression levels of <i>DNMT3B</i> and relative expression levels miR-148a-3p; (<b>d</b>) between relative expression levels of <i>BCL2</i> and relative expression levels miR-148a-3p; (<b>e</b>) between relative expression levels of <i>BCL2</i> and relative expression levels miR-204-5p; (<b>f</b>) between relative expression levels of <i>BCL2</i> and relative expression levels miR-375 in gastric tissue samples. P value below 0.05 was considered significant.</p

Enrichment analysis of GC-associated genes in miR-148a-3p, miR-204-5p, miR-223-3p and miR-375 target genes by using hypergeometrical distribution.

<p>GC–gastric cancer.</p><p>Enrichment analysis of GC-associated genes in miR-148a-3p, miR-204-5p, miR-223-3p and miR-375 target genes by using hypergeometrical distribution.</p

Analysis of Deregulated microRNAs and Their Target Genes in Gastric Cancer

<div><p>Background</p><p>MicroRNAs (miRNAs) are widely studied non-coding RNAs that modulate gene expression. MiRNAs are deregulated in different tumors including gastric cancer (GC) and have potential diagnostic and prognostic implications. The aim of our study was to determine miRNA profile in GC tissues, followed by evaluation of deregulated miRNAs in plasma of GC patients. Using available databases and bioinformatics methods we also aimed to evaluate potential target genes of confirmed differentially expressed miRNA and validate these findings in GC tissues.</p><p>Methods</p><p>The study included 51 GC patients and 51 controls. Initially, we screened miRNA expression profile in 13 tissue samples of GC and 12 normal gastric tissues with TaqMan low density array (TLDA). In the second stage, differentially expressed miRNAs were validated in a replication cohort using qRT-PCR in tissue and plasma samples. Subsequently, we analyzed potential target genes of deregulated miRNAs using bioinformatics approach, determined their expression in GC tissues and performed correlation analysis with targeting miRNAs.</p><p>Results</p><p>Profiling with TLDA revealed 15 deregulated miRNAs in GC tissues compared to normal gastric mucosa. Replication analysis confirmed that miR-148a-3p, miR-204-5p, miR-223-3p and miR-375 were consistently deregulated in GC tissues. Analysis of GC patients’ plasma samples showed significant down-regulation of miR-148a-3p, miR-375 and up-regulation of miR-223-3p compared to healthy subjects. Further, using bioinformatic tools we identified targets of replicated miRNAs and performed disease-associated gene enrichment analysis. Ultimately, we evaluated potential target gene <i>BCL2</i> and <i>DNMT3B</i> expression by qRT-PCR in GC tissue, which correlated with targeting miRNA expression.</p><p>Conclusions</p><p>Our study revealed miRNA profile in GC tissues and showed that miR-148a-3p, miR-223-3p and miR-375 are deregulated in GC plasma samples, but these circulating miRNAs showed relatively weak diagnostic performance as sole biomarkers. Target gene analysis demonstrated that <i>BCL2</i> and <i>DNMT3B</i> expression in GC tissue correlated with their targeting miRNA expression.</p></div

The volcano plot of aberrantly expressed miRNAs detected in TLDA.

<p>The green color represents significantly (FDR adjusted p < 0.01) differentially expressed miRNAs with fold change > 2. The red color indicates significantly differentially expressed miRNAs with fold change < 2.</p