Expression Of Myocyte Enhancer Factor 2A Research Articles

Brusatol enhances MEF2A expression to inhibit RCC progression through the Wnt signalling pathway in renal cell carcinoma.

Renal cell carcinoma (RCC) is the most aggressive subtype of kidney tumour with a poor prognosis and an increasing incidence rate worldwide. Brusatol, an essential active ingredient derived from Brucea javanica, exhibits potent antitumour properties. Our study aims to explore a novel treatment strategy for RCC patients. We predicted 37 molecular targets of brusatol based on the structure of brusatol, and MEF2A (Myocyte Enhancer Factor 2A) was selected as our object through bioinformatic analyses. We employed various experimental techniques, including RT-PCR, western blot, CCK8, colony formation, immunofluorescence, wound healing, flow cytometry, Transwell assays and xenograft mouse models, to investigate the impact of MEF2A on RCC. MEF2A expression was found to be reduced in patients with RCC, indicating a close correlation with MEF2A deubiquitylation. Additionally, the protective effects of brusatol on MEF2A were observed. The overexpression of MEF2A inhibits RCC cell proliferation, invasion and migration. In xenograft mice, MEF2A overexpression in RCC cells led to reduced tumour size compared to the control group. The underlying mechanism involves the inhibition of RCC cell proliferation, invasion, migration and epithelial-mesenchymal transition (EMT) through the modulation of Wnt/β-catenin signalling. Altogether, we found that MEF2A overexpression inhibits RCC progression by Wnt/β-catenin signalling, providing novel insight into diagnosis, treatment and prognosis for RCC patients.

Evolutionary conserved circular MEF2A RNAs regulate myogenic differentiation and skeletal muscle development.

Circular RNAs (circRNAs) have been recognized as critical regulators of skeletal muscle development. Myocyte enhancer factor 2A (MEF2A) is an evolutionarily conserved transcriptional factor that regulates myogenesis. However, it remains unclear whether MEF2A produces functional circRNAs. In this study, we identified two evolutionarily conserved circular MEF2A RNAs (circMEF2As), namely circMEF2A1 and circMEF2A2, in chicken and mouse muscle stem cells. Our findings revealed that circMEF2A1 promotes myogenesis by regulating the miR-30a-3p/PPP3CA/NFATC1 axis, whereas circMEF2A2 facilitates myogenic differentiation by targeting the miR-148a-5p/SLIT3/ROBO2/β-catenin signaling pathway. Furthermore, in vivo experiments demonstrated that circMEF2As both promote skeletal muscle growth. We also discovered that the linear MEF2A mRNA-derived MEF2A protein binds to its own promoter region, accelerating the transcription of MEF2A and upregulating the expression of both linear MEF2A and circMEF2As, forming a MEF2A autoregulated positive feedback loop. Moreover, circMEF2As positively regulate the expression of linear MEF2A by adsorbing miR-30a-3p and miR-148a-5p, which directly contribute to the MEF2A autoregulated feedback loop. Importantly, we found that mouse circMEF2As are essential for the myogenic differentiation of C2C12 cells. Collectively, our results demonstrated the evolution, function, and underlying mechanisms of circMEF2As in animal myogenesis, which may provide novel insight for both the farm animal meat industry and human medicine.

HDAC5 inhibition attenuates ventricular remodeling and cardiac dysfunction

BackgroundThis study aimed to investigate the role of histone deacetylase 5 (HDAC5) in ventricular remodeling and explore the therapeutic potential of the HDAC5 inhibitor LMK235.MethodsA transverse aortic constriction (TAC) mouse model and angiotensin II (Ang II)-treated H9C2 cells were used to evaluate the effects of HDAC5 inhibition with LMK235 on ventricular remodeling and cardiac dysfunction. Additionally, the involvement of the extracellular signal-regulated kinase (ERK)/early growth response protein 1 (EGR1) signaling pathway in regulating myocyte enhancer factor 2 A (MEF2A) expression was assessed.ResultsHDAC5 was upregulated in TAC mice and Ang II-treated H9C2 cells, suggesting its involvement in ventricular remodeling and cardiac dysfunction. LMK235 treatment significantly improved cardiac function in TAC mice and attenuated TAC-induced ventricular remodeling and Ang II-induced H9C2 cell hypertrophy. Mechanically, HDAC5 inhibition activated the ERK/EGR1 signaling pathway.ConclusionsOur findings demonstrate that HDAC5 may suppress the activation of ERK/EGR1 signaling to regulate MEF2A expression and therefore participate in cardiac pathophysiology.

Effect of Bovine MEF2A Gene Expression on Proliferation and Apoptosis of Myoblast Cells.

Myocyte enhancer factor 2A (MEF2A) is a member of the myocyte enhancer factor 2 family. MEF2A is widely distributed in various tissues and organs and participates in various physiological processes. This study aimed to investigate the effect of MEF2A expression on the proliferation and apoptosis of bovine myoblasts. CCK8, ELISA, cell cycle, and apoptosis analyses were conducted to assess cell status. In addition, the mRNA expression levels of genes associated with bovine myoblast proliferation and apoptosis were evaluated using RT-qPCR. The results showed that the upregulation of MEF2A mRNA promoted the proliferation rate of myoblasts, shortened the cycle process, and increased the anti-apoptotic rate. Furthermore, the RT-qPCR results showed that the upregulation of MEF2A mRNA significantly increased the cell proliferation factors MyoD1 and IGF1, cell cycle factors CDK2 and CCNA2, and the apoptotic factors Bcl2 and BAD (p < 0.01). These results show that the MEF2A gene can positively regulate myoblast proliferation and anti-apoptosis, providing a basis for the analysis of the regulatory mechanism of the MEF2A gene on bovine growth and development.

Myocyte Enhancer Factor 2A Contributes to the TGF-β1-Mediated Cholangiocyte Epithelial to Mesenchymal Transition and Senescence in Cholestatic Liver Fibrosis.

Cholangiocytes are primary targets in chronic cholestatic liver diseases. Myocyte enhancer factor 2A (MEF2A) is a transcription factor with a crucial role in some fibrogenic diseases. However, whether it contributes to cholestatic liver fibrosis is still obscure. A bile duct-ligated (BDL) mouse model was established to detect MEF2A expression during cholestatic liver fibrosis. In addition, human intrahepatic biliary epithelial cells (HIBECs) were transfected with lentivirus-expressing shMEF2A (LV-shMEF2A) to regulate the expression of MEF2A in vitro. Biomarkers of epithelial to mesenchymal transition (EMT), senescence, and fibrogenesis were evaluated using various assays: Quantitative real-time polymerase chain reaction (qRT-PCR), western blotting, senescence-associated β-galactosidase (SA-β-gal), and immunofluorescence. Furthermore, MEF2A expression and cytoplasm translocation induced by transforming growth factor β1 (TGF-β1) in HIBECs were determined by qRT-PCR, western blotting, and immunofluorescence. The expression of TGF-β1-induced MEF2A, EMT, senescence, and fibrosis markers inhibited by p38 MAPK signaling were evaluated by western blotting. Finally, the peripheral blood from primary biliary cholangitis (PBC) patients and healthy controls (HCs) was collected to analyze expression of MEF2A using Enzyme-linked immunosorbent assay (ELISA). We found that MEF2A expression increased in liver tissues of BDL mice, and positively related to the extent of fibrosis. Silencing MEF2A in HIBECs restrained TGF-β1-induced EMT, senescence, and fibrotic reaction. Moreover, TGF-β1 enhanced the expression of MEF2A and induced its cytoplasm translocation in a concentration- and time-dependent manner, partially through interacting with p38 MAPK. The expression of MEF2A was also higher in the serum of PBC patients than in HCs, and positively correlated with fibrosis degree. Our study demonstrates that MEF2A is a central mediator linking TGF-β1-induced EMT and senescence in HIBECs. We propose it as a novel biomarker of fibrogenesis in cholestatic liver fibrosis. We also suggest inhibiting MEF2A as a potential strategy in treating cholestatic liver fibrosis.

MEF2A Is the Trigger of Resveratrol Exerting Protection on Vascular Endothelial Cell.

Both resveratrol and myocyte enhancer factor 2A (MEF2A) may protect vascular endothelial cell (VEC) through activating the expression of SIRT1. However, the relationship between resveratrol and MEF2A is unclear. We aimed to investigate the deeper mechanism of resveratrol in protecting vascular endothelial cells and whether MEF2A plays a key role in the protective function of resveratrol. Human umbilical vein endothelial cell (HUVEC) was used for in vitro study, and small interfere RNA was used for silencing MEF2A. Silencing MEF2A in the vascular endothelium (VE) of ApoE−/− mice was performed by tail injection with adeno associated virus expressing si-mef2a-shRNA. The results showed that treatment of HUVEC with resveratrol significantly up-regulated MEF2A, and prevented H2O2-induced but not siRNA-induced down-regulation of MEF2A. Under various experimental conditions, the expression of SIRT1 changed with the level of MEF2A. Resveratrol could rescue from cell apoptosis, reduction of cell proliferation and viability induced by H2O2, but could not prevent against that caused by silencing MEF2A with siRNA. Silencing MEF2A in VE of apoE−/− mice decreased the expression of SIRT1, increased the plasma LDL-c, and abrogated the function of resveratrol on reducing triglyceride. Impaired integrity of VE and aggravated atherosclerotic lesion were observed in MEF2A silenced mice through immunofluorescence and oil red O staining, respectively. In conclusion, resveratrol enhances MEF2A expression, and the upregulation of MEF2A is required for the endothelial protective benefits of resveratrol in vitro via activating SIRT1. Our work has also explored the in vivo relevance of this signaling pathway in experimental models of atherosclerosis and lipid dysregulation, setting the stage for more comprehensive phenotyping in vivo and further defining the molecular mechanisms.

Decreased MEF2A Expression Regulated by Its Enhancer Methylation Inhibits Autophagy and May Play an Important Role in the Progression of Alzheimer's Disease.

Alzheimer’s disease (AD) is a neurodegenerative disease characterized by amyloid plaques and neurofibrillary tangles which significantly affects people’s life quality. Recently, AD has been found to be closely related to autophagy. The aim of this study was to identify autophagy-related genes associated with the pathogenesis of AD from multiple types of microarray and sequencing datasets using bioinformatics methods and to investigate their role in the pathogenesis of AD in order to identify novel strategies to prevent and treat AD. Our results showed that the autophagy-related genes were significantly downregulated in AD and correlated with the pathological progression. Furthermore, enrichment analysis showed that these autophagy-related genes were regulated by the transcription factor myocyte enhancer factor 2A (MEF2A), which had been confirmed using si-MEF2A. Moreover, the single-cell sequencing data suggested that MEF2A was highly expressed in microglia. Methylation microarray analysis showed that the methylation level of the enhancer region of MEF2A in AD was significantly increased. In conclusion, our results suggest that AD related to the increased methylation level of MEF2A enhancer reduces the expression of MEF2A and downregulates the expression of autophagy-related genes which are closely associated with AD pathogenesis, thereby inhibiting autophagy.

MEF2A transcriptionally upregulates the expression of ZEB2 and CTNNB1 in colorectal cancer to promote tumor progression

Colorectal cancer (CRC) is one of the leading cancers worldwide, accounting for high morbidity and mortality. The mechanisms governing tumor growth and metastasis in CRC require detailed investigation. The results of the present study indicated that the transcription factor (TF) myocyte enhancer factor 2A (MEF2A) plays a dual role in promoting proliferation and metastasis of CRC by inducing the epithelial-mesenchymal transition (EMT) and activation of WNT/β-catenin signaling. Aberrant expression of MEF2A in CRC clinical specimens was significantly associated with poor prognosis and metastasis. Functionally, MEF2A directly binds to the promoter region to initiate the transcription of ZEB2 and CTNNB1. Simultaneous activation of the expression of EMT-related TFs and Wnt/β-catenin signaling by MEF2A overexpression induced the EMT and increased the frequency of tumor formation and metastasis. The present study identified a new critical oncogene involved in the growth and metastasis of CRC, providing a potential novel therapeutic target for CRC intervention.

The lncRNA MALAT1 participates in regulating coronary slow flow endothelial dysfunction through the miR-181b-5p–MEF2A–ET-1 axis

BackgroundCoronary slow flow (CSF) refers to coronary arteries with no obvious stenosis but have slow coronary flow without effective treatment. The main cause of CSF is endothelial dysfunction. The long non-coding RNA (lncRNA) MALAT1 is involved in regulating endothelial dysfunction, but its role in CSF endothelial dysfunction is still unclear. MethodsWe included 41 CSF patients and 37 controls in the study, who all underwent coronary angiography, echocardiography, and brachial artery flow-mediated dilatation (FMD) examination. Human umbilical vein endothelial cells (HUVECs) stimulated by oxygen–glucose deprivation were used as CSF-induced HUVECs. Plasma endothelin-1 (ET-1) concentrations were determined by enzyme-linked immunosorbent assay (ELISA). The expression levels of MALAT1, miR-181b-5p, myocyte enhancer factor 2A (MEF2A), and ET-1 were measured by qRT-PCR or western blotting. Cell proliferation was determined by 5-ethynyl-2′-deoxyuridine (EdU) and Cell Counting Kit-8 (CCK-8) assays. Apoptosis was examined by flow cytometry. The relationship between miR-181b-5p and MALAT1 or MEF2A was verified by dual-luciferase reporter assay. MEF2A binding directly to the ET-1 promoter region was verified via chromatin immunoprecipitation (ChIP) assay. ResultsMALAT1 and ET-1 were increased, and miR-181b-5p was decreased in the peripheral blood of the CSF patients, and could be used as predictors of CSF. In the CSF-induced HUVECs, MALAT1 was highly expressed, and MALAT1 knockdown improved endothelial function. In contrast, miR-181b-5p was downregulated in the CSF-induced HUVECs, and miR-181b-5p overexpression improved endothelial function. While MEF2A was highly enriched in CSF-induced HUVECs, MEF2A knockdown reduced ET-1 and increased the endothelial function of CSF-induced HUVECs as a transcriptional regulator of ET-1. MALAT1 modulated MEF2A expression positively by sponging miR-181b-5p. ConclusionsEndothelial function is reduced in CSF. MALAT1 participates in regulating CSF endothelial dysfunction through the miR-181b-5p–MEF2A–ET-1 axis, and could provide a new target for CSF treatment.

Long Noncoding RNA Ahit Protects Against Cardiac Hypertrophy Through SUZ12 (Suppressor of Zeste 12 Protein Homolog)-Mediated Downregulation of MEF2A (Myocyte Enhancer Factor 2A).

Long noncoding RNA (lncRNA) can regulate various physiological and pathological processes through multiple molecular mechanisms in cis and in trans. However, the role of lncRNAs in cardiac hypertrophy is yet to be fully elucidated. A mouse lncRNA microarray was used to identify differentially expressed lncRNAs in the mouse hearts following transverse aortic constriction-induced pressure overload comparing to the sham-operated samples. The direct impact of one lncRNA, Ahit, on cardiomyocyte hypertrophy was characterized in neonatal rat cardiomyocytes in response to phenylephrine by targeted knockdown and overexpression. The in vivo function of Ahit was analyzed in mouse hearts by using cardiac-specific adeno-associated virus, serotype 9-short hairpin RNA to knockdown Ahit in combination with transverse aortic constriction. Using catRAPID program, an interaction between Ahit and SUZ12 (suppressor of zeste 12 protein homolog) was predicted and validated by RNA immunoprecipitation and immunoblotting following RNA pull-down. Chromatin immunoprecipitation was performed to determine SUZ12 or H3K27me3 occupancy on the MEF2A (myocyte enhancer factor 2A) promoter. Finally, the expression of human Ahit (leukemia-associated noncoding IGF1R activator RNA 1 [LUNAR1]) in the serum samples from patients of hypertrophic cardiomyopathy was tested by quantitative real-time polymerase chain reaction. A previously unannotated lncRNA, antihypertrophic interrelated transcript (Ahit), was identified to be upregulated in the mouse hearts after transverse aortic constriction. Inhibition of Ahit induced cardiac hypertrophy, both in vitro and in vivo, associated with increased expression of MEF2A, a critical transcriptional factor involved in cardiac hypertrophy. In contrast, overexpression of Ahit significantly attenuated stress-induced cardiac hypertrophy in vitro. Furthermore, Ahit was significantly upregulated in serum samples of patients diagnosed with hypertensive heart disease versus nonhypertrophic hearts (1.46±0.17 fold, P=0.0325). Mechanistically, Ahit directly bound and recruited SUZ12, a core PRC2 (polycomb repressive complex 2) protein, to the promoter of MEF2A, triggering its trimethylation on H3 lysine 27 (H3K27me3) residues and mediating the downregulation of MEF2A, thereby preventing cardiac hypertrophy. Ahit is a lncRNA with a significant role in cardiac hypertrophy regulation through epigenomic modulation. Ahit is a potential therapeutic target of cardiac hypertrophy.

Myocyte enhancer factor 2A delays vascular endothelial cell senescence by activating the PI3K/p-Akt/SIRT1 pathway.

Myocyte enhancer factor 2A (MEF2A) dysfunction is closely related to the occurrence of senile diseases such as cardiocerebrovascular diseases, but the underlying molecular mechanism is unclear. Here, we studied the effects of MEF2A on the senescent phenotype of vascular endothelial cells (VEC) and downstream signaling pathway, and the association between plasma MEF2A levels and coronary artery disease (CAD). Results showed that MEF2A silencing promoted cell senescence and down-regulated PI3K/p-AKT/Sirtuin 1 (SIRT1) expression. MEF2A overexpression delayed cell senescence and up-regulated PI3K/p-AKT/SIRT1. Hydrogen peroxide (H2O2) treatment induced cellular senescence and down-regulated the expression of MEF2A and PI3K/p-AKT/SIRT1. MEF2A overexpression inhibited cellular senescence and the down-regulation of PI3K/p-AKT/SIRT1 induced by H2O2. Further study revealed that MEF2A directly up-regulated the expression of PIK3CA and PIK3CG through MEF2 binding sites in the promoter region. Pearson correlation and logistic regression analysis showed that the plasma level of MEF2A was negatively correlated with CAD, and with age in the controls. These results suggested that MEF2A can directly up-regulate PI3K gene expression, and one of the molecular mechanisms of delaying effect of MEF2A on VEC cell senescence was SIRT1-expression activation through the PI3K/p-Akt pathway. Moreover, the plasma MEF2A levels may be a potential biomarker for CAD risk prediction.

MALAT1 promoted cell proliferation and migration via MALAT1/miR-155/MEF2A pathway in hypoxia of cardiac stem cells.

Accumulating evidence revealed that hypoxia contributed to many human diseases, including ischemic myocardium and heart failure (HF). In recent years, the roles of hypoxia in stem cell survival and cardiac biology have been studied extensively. However, the underlying molecular mechanisms remain to be elucidated. As a leading cause of HF, ischemic heart disease was correlated with hypoxia. In this study, we firstly constructed the hypoxia cell model by CoCl2 in cardiac stem cells (CSCs) and found that hypoxia induced the cell proliferation and migration potential in CSCs. Then, we demonstrated that the expression of metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) was promoted in CoCl2 -induced CSCs hypoxia model. Furthermore, we found that knockdown of MALAT1 inhibited the cell proliferation and migration in CoCl2 -induced CSCs hypoxia model. In addition, we revealed that MALAT1 regulated the microRNA-155 (miR-155) expression in CSCs under both the normal and hypoxia conditions and further, manipulation of the miR-155 expression affected the role of MALAT1 in CoCl2 -induced CSCs hypoxia cell model. We then illustrated that miR-155 regulated the myocyte enhancer factor 2A(MEF2A) expression in CSCs under both the normal and hypoxia conditions and further, changing the expression of MEF2A affected the role of miR-155. Finally, we demonstrated that MALAT1 regulated the MEF2A expression and exerted its role via modulation of the MALAT1/miR-155/MEF2A pathway. Taken together, our study illustrated that MALAT1 promoted the cell proliferation and migration in CoCl2 -induced CSCs hypoxia model, acting mechanistically by promoting MEF2A expression via "sponging" miR-155.

Variants in MEF2A gene in relation with coronary artery disease in Saudi population.

This study investigated the association of variants in myocyte enhancer factor 2A (MEF2A) gene with coronary artery disease (CAD) via case control study on Saudi population. Several studies have indicated a high expression of MEF2A in the human coronary endothelium. The entire (exon 11 putative susceptibility exon) of MEF2A gene was sequenced using direct DNA sequencing method in 120 sporadic patients and 100 controls. Total number of variants were identified and crude odds ratio (OR) with 95% confidence interval (CI) was calculated. In total, three variants were identified, namely, CAG repeats, AGC deletion, and SNP rs: 325400. No significant link was observed between the common (CAG) n polymorphism, AGC deletion, and CAD risk as reported in other populations, but interestingly, rs325400 (G1323T) in Saudis was found to be associated with the CAD with odds ratio 2.0102 (CI = 1.3405-3.0146) and significance of p = 0.00048. None of Saudi subjects (normal as well as diseased) showed 21-bp deletion as reported previously for other populations. In addition, genotype TT of rs325400 is associated with significantly higher levels of LDL-C and lower level of HDL-C. Among the quantitative parameters, lower HDL-C and higher LDL-C was found to be associated with disease. We report that MEF2A gene based on SNP rs325400 (G1323T) can be considered as a susceptibility factor for CAD and presence of T allele makes Saudis at more risk to CAD, while other variants detected in this gene do not have any association in Saudi population.

Inhibition of myocyte-specific enhancer factor 2A improved diabetic cardiac fibrosis partially by regulating endothelial-to-mesenchymal transition.

Cardiac fibrosis is an important pathological process of diabetic cardiomyopathy, the underlying mechanism remains elusive. This study sought to identify whether inhibition of Myocyte enhancer factor 2A (MEF2A) alleviates cardiac fibrosis by partially regulating Endothelial-to-mesenchymal transition (EndMT). We induced type 1 diabetes mellitus using the toxin streptozotocin (STZ) in mice and injected with lentivirus-mediated short-hairpin RNA (shRNA) in myocardium to inhibit MEF2A expression. Protein expression, histological and functional parameters were examined twenty-one weeks post-STZ injection. We found that Diabetes mellitus increased cardiac MEF2A expression, aggravated cardiac dysfunction and myocardial fibrosis through the accumulation of fibroblasts via EndMT. All of these features were abolished by MEF2A inhibition. MEF2A gene silencing by shRNA in cultured human umbilical vein endothelial cells (HUVECs) ameliorated high glucose–induced phenotypic transition and acquisition of mesenchymal markers through interaction with p38MAPK and Smad2. We conclude that inhibition of endothelial cell-derived MEF2A might be beneficial in the prevention of diabetes mellitus-induced cardiac fibrosis by partially inhibiting EndMT through interaction with p38MAPK and Smad2.

Inhibition of MEF2A prevents hyperglycemia-induced extracellular matrix accumulation by blocking Akt and TGF-β1/Smad activation in cardiac fibroblasts

Myocyte enhancer factor 2A (MEF2A) functions in muscle-specific and/or growth factor-related transcription and is involved in cell growth, survival, and apoptosis. To evaluate the role of this transcription factor in cardiac fibroblasts (CFs) in diabetes mellitus, we performed a series of in vitro and in vivo experiments. We used short hairpin RNA (shRNA) to inhibit the expression of MEF2A in CFs in vitro. Inhibition of MEF2A significantly reduced hyperglycemia-induced CF proliferation and migration, myofibroblast differentiation, matrix metalloproteinase (MMP) activities, and collagen production. Furthermore, MEF2A inhibition attenuated HG-induced activation of the mitogen-activated protein kinase (MAPK), Akt, and TGF-β1/Smad signaling pathways. For in vivo analysis in a mouse model, type-1 diabetes was induced by streptozotocinand MEF2A expression was knocked down by myocardial injection with lentivirus carrying shRNA-MEF2A. Cardiac function was assessed by echocardiography. Total collagen deposition was assessed by Masson's trichrome and Picrosirius red staining. Knockdown of MEF2A ameliorated diabetes-induced cardiac dysfunction and collagen deposition. Our study suggests that inhibition of MEF2A could alleviate HG-induced extracellular matrix accumulation by blocking the activation of Akt and TGF-β1/Smad signaling pathway in CFs. Thus, inhibition of MEF2A has therapeutic potential in the treatment of diabetic-induced cardiac remodeling.

(-)-Epicatechin is associated with increased angiogenic and mitochondrial signalling in the hindlimb of rats selectively bred for innate low running capacity.

Alternative approaches to reduce congenital muscle dysfunction are needed in cases where the ability to exercise is limited. (-)-Epicatechin is found in cocoa and may stimulate capillarity and mitochondrial proliferation in skeletal muscle. A total of 21 male rats bred for LCR (low running capacity) from generation 28 were randomized into three groups: vehicle for 30 days (control); (-)-epicatechin for 30 days; and (-)-epicatechin for 30 days followed by 15 days without (-)-epicatechin. Groups 2 and 3 received 1.0 mg of (-)-epicatechin/kg of body mass twice daily, whereas water was given to the control group. The plantaris muscle was harvested for protein and morphometric analyses. In addition, in vitro experiments were conducted to examine the role of (-)-epicatechin on mitochondrial respiratory kinetics at different incubation periods. Treatment for 30 days with (-)-epicatechin increased capillarity (P<0.001) and was associated with increases in protein expression of VEGF (vascular endothelial growth factor)-A with a concomitant decrease in TSP-1 (thrombospondin-1) and its receptor, which remained after 15 days of (-)-epicatechin cessation. Analyses of the p38 MAPK (mitogen-activated protein kinase) signalling pathway indicated an associated increase in phosphorylation of MKK3/6 (MAPK kinase 3/6) and p38 and increased protein expression of MEF2A (myocyte enhancer factor 2A). In addition, we observed significant increases in protein expression of PGC-1α (peroxisome-proliferator-activated receptor γ co-activator 1α), PGC-1β, Tfam and cristae abundance. Interestingly, these increases associated with (-)-epicatechin treatment remained after 15 days of cessation. Lastly, in vitro experiments indicated that acute exposure of LCR muscle to (-)-epicatechin incubation was not sufficient to increase mitochondrial respiration. The results suggest that increases in skeletal muscle capillarity and mitochondrial biogenesis are associated with 30 days of (-)-epicatechin treatment and sustained for 15 days following cessation of treatment. Clinically, the use of this natural compound may have potential application in populations that experience muscle fatigue and are unable to perform endurance exercise.

SUR2a gene is upregulated in skeletal muscle insulin resistance of obese rats: possible correlation with GLUT4 gene

ObjectiveThe aim of this study was to investigate whether SUR2A (sulfonylurea receptor type 2A, ABCC9) MEF2a (myocyte enhancer factor‐2a) and HIF1a (Hypoxia‐inducible factor‐1) can influence glucose uptake in oxidative muscle soleus by regulating GLUT4 expression.MethodsInsulin resistance was induced in male Wistar rats by inducing obesity with monosodium glutamate (MSG). At the age 3 months, half control and MSG‐treated animals started receiving 0.1 mg/kg/day of glimepiride in the drinking water, and distributed in 3 experimental groups: control rats (C), MSG‐obese rats (O) and MSG‐obese rats treated with glimepiride (OG). Animals were submitted to insulin tolerance test (ITT) and samples of soleus muscle were excised for quantification of gene expression by Real Time RT‐PCR and GLUT4 protein.ResultsOG rats presented a 51% increase in SUR2A mRNA content in comparison to O and C rats (p&lt;0.001). O rats showed 16% increase in GLUT4 mRNA content (p&lt;0.01 vs. C), without any increase in GLUT4 protein. After glimepiride treatment, OG rats increased GLUT4 mRNA (21%, p&lt;0.01 vs. C) and protein (131%, p&lt;0.01 vs. C; and 73%, p&lt;0.05 vs. O). No significant differences (p&gt;0.05) in mRNA expression of MEF2a and HIF1a were found.ConclusionWe conclude that the glimepiride increased the GLUT4 expression in soleus of the insulin resistant animals which could be associated to SUR2A genes expression.

Cardiac developmental defects and eccentric right ventricular hypertrophy in cardiomyocyte focal adhesion kinase (FAK) conditional knockout mice

Focal adhesion kinase (FAK) is a nonreceptor tyrosine kinase that plays an important role in integrin-mediated signal transduction. To explore the role and mechanisms of FAK in cardiac development, we inactivated FAK in embryonic cardiomyocytes by crossing the floxed FAK mice with myosin light chain-2a (MLC2a) Cre mice, which expressed Cre as early as embryonic day 9.5 in the heart. The majority of conditional FAK knockout mice generated from MLC2a-Cre (CFKO-2a) died in the embryonic stage with thin ventricular wall and ventricular septal defects. A small fraction of CFKO-2a mice survived to adulthood with spontaneous eccentric right ventricle hypertrophy. Transmission electron microscopy analysis displayed swelling in the rough endoplasmic reticulum in CFKO-2a embryonic cardiomyocytes. We found that decreased cell proliferation, but not increased cell apoptosis or differentiation, is the reason for the thin ventricular wall in CFKO-2a mice. Microarray analysis suggests that myocyte enhancer factor 2a (MEF2a) can be regulated by FAK and that inactivation of FAK in the embryonic heart compromised MEF2a expression. Last, we found that Src, but not PI3K, is important in mediating signal transduction for the regulation of MEF2a by FAK. Together, these results identified the role and mechanisms of FAK in embryonic cardiac development.

Calcineurin-Aα activation enhances the structure and function of regenerating muscles after myotoxic injury

Calcineurin signaling is essential for successful muscle regeneration. Although calcineurin inhibition compromises muscle repair, it is not known whether calcineurin activation can enhance muscle repair after injury. Tibialis anterior (TA) muscles from adult wild-type (WT) and transgenic mice overexpressing the constitutively active calcineurin-A alpha transgene under the control of the mitochondrial creatine kinase promoter (MCK-CnA alpha*) were injected with the myotoxic snake venom Notexin to destroy all muscle fibers. The TA muscle of the contralateral limb served as the uninjured control. Muscle structure was assessed at 5 and 9 days postinjury, and muscle function was tested in situ at 9 days postinjury. Calcineurin stimulation enhanced muscle regeneration and altered levels of myoregulatory factors (MRFs). Recovery of myofiber size and force-producing capacity was hastened in injured muscles of MCK-CnA alpha* mice compared with control. Myogenin levels were greater 5 days postinjury and myocyte enhancer factor 2a (MEF2a) expression was greater 9 days postinjury in muscles of MCK-CnA alpha* mice compared with WT mice. Higher MEF2a expression in regenerating muscles of MCK-CnA alpha* mice 9 days postinjury may be related to an increase of slow fiber genes. Calcineurin activation in uninjured and injured TA muscles slowed muscle contractile properties, reduced fatigability, and enhanced force recovery after 4 min of intermittent maximal stimulation. Therefore, calcineurin activation can confer structural and functional benefits to regenerating skeletal muscles, which may be mediated in part by differential expression of MRFs.

Myocyte-specific enhancer factor 2A is essential for zebrafish posterior somite development

Somite development is governed tightly by genetic factors. In the large-scale mutagenesis screens of zebrafish, no mutations were linked to myocyte enhancer factor 2A (MEF2A) locus. In this study, we find that MEF2A knock-down embryos display a downward tail curvature and have U-shaped posterior somites. Furthermore, we demonstrate that MEF2A is required for Hedgehog signaling. MEF2A inhibition results in induction of apoptosis in the posterior somites. We further find that Hedgehog signaling can negatively regulate MEF2A expression in the somites. Microarray studies reveal a number of genes that are differentially expressed in the MEF2A morphants. Our studies suggest that MEF2A is essential for zebrafish posterior somite development.