Integration of miRNA and mRNA expression profles reveals microRNA-regulated networks during muscle wasting in cardiac cachexia

Cardiac cachexia (CC) is a common complication of heart failure (HF) associated with muscle wasting and poor patient prognosis. Although different mechanisms have been proposed to explain muscle wasting during CC, its pathogenesis is still not understood. Here, we described an integrative analysis between miRNA and mRNA expression profiles of muscle wasting during CC. Global gene expression profiling identified 1,281 genes and 19 miRNAs differentially expressed in muscle wasting during CC. Several of these deregulated genes are known or putative targets of the altered miRNAs, including miR-29a-3p, miR-29b-3p, miR-210-5p, miR-214, and miR-489. Gene ontology analysis on integrative mRNA/miRNA expression profiling data revealed miRNA interactions affecting genes that regulate extra-cellular matrix (ECM) organization, proteasome protein degradation, citric acid cycle and respiratory electron transport. We further identified 11 miRNAs, including miR-29a-3p and miR-29b-3p, which target 21 transcripts encoding the collagen proteins related to ECM organization. Integrative miRNA and mRNA global expression data allowed us to identify miRNA target genes involved in skeletal muscle wasting in CC. Our functional experiments in C2C12 cells confirmed that miR-29b down-regulates collagen genes and contributes to muscle cell atrophy. Collectively, our results suggest that key ECM-associated miRNAs and their target genes may contribute to CC in HF

Dysregulation of miR-1-3p: An Early Event in Colitis-Associated Dysplasia

Detection of colorectal dysplasia during surveillance colonoscopy remains the best method of determining risk for colitis-associated colorectal cancer (CAC). miRNAs (miRs) show great promise as tissue-specific biomarkers of neoplasia. The goal of this study was to explore the miR expression profile of precancerous dysplastic lesions in the AOM/DSS mouse model and identify early molecular changes associated with CAC. Epithelial cells were laser-microdissected from the colonic mucosa (inflamed versus dysplastic) of mice with AOM/DSS-induced colitis. A miR signature that can distinguish inflamed non-neoplastic mucosa from dysplasia was identified. Bioinformatic analyses led to the discovery of associated miR gene targets and enriched pathways and supported the construction of a network interaction map. miR-1a-3p was one of the miRs with the highest number of predicted targets, including Cdk6. Interestingly, miR-1a-3p and Cdk6 were down- and up-regulated in dysplastic lesions, respectively. Transfection of HCT116 and RKO cells with miR-1a-3p mimics induced apoptosis and cell cycle arrest in G1, suggesting its biological function. A slight reduction in the level of CDK6 transcripts was also observed in cells transfected with miR-1. These data provide novel insight into the early molecular alterations that accompany the development of CAC and identify a miR signature that represents a promising biomarker for the early detection of colitis-associated dysplasia

Decreased miR-497-5p Suppresses IL-6 Induced Atrophy in Muscle Cells

Interleukin-6 (IL-6) is a pro-inflammatory cytokine associated with skeletal muscle wasting in cancer cachexia. The control of gene expression by microRNAs (miRNAs) in muscle wasting involves the regulation of thousands of target transcripts. However, the miRNA-target networks associated with IL6-induced muscle atrophy remain to be characterized. Here, we show that IL-6 promotes the atrophy of C2C12 myotubes and changes the expression of 20 miRNAs (5 up-regulated and 15 down-regulated). Gene Ontology analysis of predicted miRNAs targets revealed post-transcriptional regulation of genes involved in cell differentiation, apoptosis, migration, and catabolic processes. Next, we performed a meta-analysis of miRNA-published data that identified miR-497-5p, a down-regulated miRNAs induced by IL-6, also down-regulated in other muscle-wasting conditions. We used miR-497-5p mimics and inhibitors to explore the function of miR-497-5p in C2C12 myoblasts and myotubes. We found that miR-497-5p can regulate the expression of the cell cycle genes CcnD2 and CcnE1 without affecting the rate of myoblast cellular proliferation. Notably, miR-497-5p mimics induced myotube atrophy and reduced Insr expression. Treatment with miR-497-5p inhibitors did not change the diameter of the myotubes but increased the expression of its target genes Insr and Igf1r. These genes are known to regulate skeletal muscle regeneration and hypertrophy via insulin-like growth factor pathway and were up-regulated in cachectic muscle samples. Our miRNA-regulated network analysis revealed a potential role for miR-497-5p during IL6-induced muscle cell atrophy and suggests that miR-497-5p is likely involved in a compensatory mechanism of muscle atrophy in response to IL-6

Regulation of cardiac microRNAs induced by aerobic exercise training during heart failure

Exercise training (ET) has beneficial effects on the myocardium in heart failure (HF) patients and in animal models of induced cardiac hypertrophy and failure. We hypothesized that if microRNAs (miRNAs) respond to changes following cardiac stress, then myocardial profiling of these miRNAs may reveal cardio-protective mechanisms of aerobic ET in HF. We employed ascending aortic stenosis (AS) inducing HF in Wistar rats. Controls were sham-operated animals. At 18 weeks after surgery, rats with cardiac dysfunction were randomized to 10 weeks of aerobic ET (HF-ET) or to a heart failure sedentary group (HF-S). ET attenuated cardiac remodeling, as well as clinical and pathological signs of HF with maintenance of systolic and diastolic function when compared to HF-S. Global miRNA expression profiling of the cardiac tissue revealed 56 miRNAs differentially regulated in animals in the HF-ET, but only 12 miRNAs were differentially regulated in the HF-S. Out of 23 miRNAs that were differentially regulated in both groups, 17 miRNAs exhibited particularly high increases in expression, including miR-598, miR-429, miR-224, miR-425, and miR-221. From the initial set of deregulated miRNAs, 14 miRNAs with validated targets expressed in cardiac tissue that respond robustly to ET in HF were used to construct a miRNA-mRNA regulatory networks that revealed a set of 203 miRNA-target genes involved in programmed cell death, TGF-β signaling, cellular metabolic processes, cytokine signaling, and cell morphogenesis. Our findings reveal that ET attenuates cardiac abnormalities during HF by regulating cardiac miRNAs with potential role in cardio-protective mechanisms through multiple effects on gene expression.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e tecnológico (CNPq