In this study, we aimed to systematically profile global RNA N6-methyladenosine (m⁶A) modification patterns in a mouse model of diabetic cardiomyopathy (DCM). Patterns of m⁶A in DCM and normal hearts were analyzed via m⁶A-specific methylated RNA immunoprecipitation followed by high-throughput sequencing (MeRIP-seq) and RNA sequencing (RNA-seq). m⁶A-related mRNAs were validated by quantitative real-time PCR analysis of input and m⁶A immunoprecipitated RNA samples from DCM and normal hearts. A total of 973 new m⁶A peaks were detected in DCM samples and 984 differentially methylated sites were selected for further study, including 295 hypermethylated and 689 hypomethylated m⁶A sites (fold change (FC) > 1.5, P < 0.05). Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) Pathway analyses indicated that unique m⁶A-modified transcripts in DCM were closely linked to cardiac fibrosis, myocardial hypertrophy, and myocardial energy metabolism. Total m⁶A levels were higher in DCM, while levels of the fat mass and obesity-associated (FTO) protein were downregulated. Overexpression of FTO in DCM model mice improved cardiac function by reducing myocardial fibrosis and myocyte hypertrophy. Overall, m⁶A modification patterns were altered in DCM, and modification of epitranscriptomic processes, such as m⁶A, is a potentially interesting therapeutic approach.

Background: The P38 mitogen-activated protein kinase (MAPK) pathway plays an essential role in CVB3-induced diseases. We previously demonstrated microRNA-21 has potential inhibitory effect on the MAP2K3 which locates upstream of P38 MAPK and was upregulated in mouse hearts upon CVB3 infection. However, the effect and underlying mechanism of miRNA-21 on CVB3 infection remain unclear. Methods: We detected continuous changes of cellular miRNA-21 and P38 MAPK proteins expression profiling post CVB3 infection in vitro within 12 h. P38 MAPK signaling was inhibited by the specific inhibitor, small interfering RNA and miRNA-21 mimic in vitro, CVB3 replication, cell apoptosis rate and proliferation were detected. Viral load in the mice heart, cardiomyocyte apoptosis rate and histological of the heart were also detected in the mice model of viral myocarditis pretreated with miRNA-21-lentivirus. Results: We observed significant upregulation of miRNA-21 expression followed by suppression of the MAP2K3/P38 MAPK signaling in CVB3-infected Hela cells. The inactivation of the MAP2K3/P38 MAPK signaling by P38 MAPK specific inhibitor, small interfering RNA against MAP2K3, or miRNA-21 overexpression significantly inhibited viral progeny release from CVB3-infected cells. Mechanistically, when compared with control miRNA, miRNA-21 showed no effect on capsid protein VP1 expression and viral load within host cells, while significantly reversing CVB3-induced caspase-3 activation and cell apoptosis rate, further promoting proliferation of infected cells, which indicates the inhibitory effect of miRNA-21 on CVB3 progeny release. In the in vivo study, when compared with control miRNA, miRNA-21 pretreatment remarkably inactivated the MAP2K3/P38 MAPK signaling in mice and protected them against CVB3 infection as evidenced by significantly alleviated cell apoptosis rate, reduced viral titers, necrosis in the heart as well as by remarkably prolonged survival time. Conclusions: miRNA-21 were reverse correlated with P38 MAPK activation post CVB3 infection, miRNA-21 overexpression significantly inhibited viral progeny release and decreased myocytes apoptosis rate in vitro and in vivo, suggesting that miRNA-21 may serve as a potential therapeutic agent against CVB3 infection through targeting the MAP2K3/P38 MAPK signaling.

MicroRNAs (miRNAs) are important post-transcriptional regulators during adipocyte adipogenesis. MiR-206-3p, a tissue-specific miRNA, is absent in white adipocytes. In this study, we examined the roles of mmu-miR-206-3p in the adipogenic differentiation of 3T3-L1 preadipocytes. The miR-206-3p expression has shown an apparent decreasing trend after induction, and sustained low expression throughout the differentiation of 3T3-L1 cells. miR-206-3p blocked the adipogenic differentiation of 3T3-L1 cells by attenuating c-Met expression; the inhibition effect of miR-206 to the adipogenic differentiation can be counteracted by restoring c-Met expression. In addition, miR-206-3p decreased the phosphorylation of Akt, which is the downstream effector of c-Met in the PI3K/Akt signaling pathway. These data indicate that miR-206-3p inhibits adipocyte adipogenesis through silencing c-Met and subsequently inactivating the PI3K/Akt signaling pathway.

Folic acid (FA) is an antioxidant that can reduce reactive oxygen species generation and can blunt cardiac dysfunction during ischemia. We hypothesized that FA supplementation prevents cardiac fibrosis and cardiac dysfunction induced by obesity. Six-week-old C57BL6/J mice were fed a high-fat diet (HFD), normal diet (ND), or an HFD supplemented with folic acid (FAD) for 14 weeks. Cardiac function was measured using a transthoracic echocardiographic exam. Phenotypic analysis included measurements of body and heart weight, blood glucose and tissue homocysteine (Hcy) content, and heart oxidative stress status. HFD consumption elevated fasting blood glucose levels and caused obesity and heart enlargement. FA supplementation in HFD-fed mice resulted in reduced fasting blood glucose, heart weight, and heart tissue Hcy content. We also observed a significant cardiac systolic dysfunction when mice were subjected to HFD feeding as indicated by a reduction in the left ventricular ejection fraction and fractional shortening. However, FAD treatment improved cardiac function. FA supplementation protected against cardiac fibrosis induced by HFD. In addition, HFD increased malondialdehyde concentration of the heart tissue and reduced the levels of antioxidant enzyme, glutathione, and catalase. HFD consumption induced myocardial oxidant stress with amelioration by FA treatment. FA supplementation significantly lowers blood glucose levels and heart tissue Hcy content and reverses cardiac dysfunction induced by HFD in mice. These functional improvements of the heart may be mediated by the alleviation of oxidative stress and myocardial fibrosis.

Wilms’ tumor is also called nephroblastoma and is the most common pediatric renal cancer. Several genetic and epigenetic factors have been found to account for the development of Wilms’ tumor. MiRNAs play important roles in this tumorigenic process. In the present study, we aimed to investigate the role of miR-140-5p in nephroblastoma by identifying its targets, as well as its underlying molecular mechanism of action. The miRNA expression profile of nephroblastoma samples was investigated and the targets of miR-140-5p were predicted and validated using the miRNA luciferase reporter method. Moreover, the roles of miR-140-5p in regulating nephroblastoma cell proliferation, migration and cell cycle were analyzed by the CCK8, migration and flow cytometry assays, respectively. The downstream protein of the direct target of miR-140-5p was also identified. miR-140-5p was downregulated in Wilms’ tumor tissues, whereas in the nephroblastoma cell lines G401 and WT-CLS1 that exhibited high levels of miRNA-140-5p, inhibition of cellular proliferation and metastasis were noted as well as cell cycle arrest at the G1/S phase. TGFBRI and IGF1R were identified as direct target genes for miRNA-140-5p. In addition, SMAD2/3 and p-AKT were regulated by TGFBRI and IGF1R separately and participated in the miRNA-140-5p regulatory network. Ectopic expression of TGFBR1 and IGF-1R could abrogate the inhibitory effect of miR-140-5p. We demonstrated that miRNA-140-5p participates in the progression of Wilms’ tumor by targeting the TGFBRI/SMAD2/3 and the IGF-1R/AKT signaling pathways.