Myocyte-specific Enhancer Factor 2C Research Articles

Generation of cardiomyocytes from stem cells cultured on nanofibrous scaffold: Experimental approach for attenuation of myocardial infarction

The current study was constructed to fabricate polyamide based nanofibrous scaffolds (NS) and to define the most promising one for the generation of cardiomyocytes from adipose tissue derived mesenchymal stem cells (ADMSCs). This purpose was extended to assess the potentiality of the generated cardiomyocytes in relieving myocardial infarction (MI) in rats. Production and characterization of NSs were carried out. ADMSCs were cultured on NS and induced to differentiate into cardiomyocytes by specific growth factors. Molecular analysis for myocyte-specific enhancer factor 2 C (MEF2C) and alpha sarcomeric actin (α-SCA) expression was done to confirm the differentiation of ADMSCs into cardiomyocytes for further transplantation into MI induced rats. Implantation of cells in MI afflicted rats boosted heart rate, ST height and PR interval and lessened P duration, RR, QTc and QRS intervals. Also, this type of medication minified serum lactate dehydrogenase (LDH) and creatine kinase-MB (CK-MB) enzymes activity as well as serum and cardiac troponin T (Tn-T) levels and upraised serum and cardiac α-SCA and cardiac connexin 43 (CX 43) levels. Microscopic feature of cardiac tissue sections of rats in the treated groups revealed great renovation in the cardiac microarchitecture. Conclusively, this attempt gains insight into a realistic strategy for recovery of MI through systemic employment of in vitro generated cardiomyocytes.

METTL3 promotes microglial inflammation via MEF2C in spinal cord injury.

Spinal cord injury (SCI) is a significant contributor to disability in contemporary society, resulting in substantial psychological and economic burdens for patients and their family. Microglia-mediated inflammation is an important factor affecting the nerve repair of SCI patients. N6-methyladenosine (m6A) is a prevalent epigenetic modification in mammals, which shows a strong association with inflammation. However, the mechanism of m6A modification regulating microglia-mediated inflammation is still unclear. Here, we observed that METTL3, a m6A methylase, was increased in SCI mice and lipopolysaccharide (LPS)-exposed BV2 cells. Knockdown of METTL3 inhibited the increased expression of iNOS and IL-1β induced by LPS in vitro. Subsequently, MEF2C, myocyte-specific enhancer factor 2C, was decreased in SCI mice and LPS-exposed BV2 cells. Knockdown of MEF2C promoted the expression of iNOS and IL-1β. Sequence analysis showed that there were multiple highly confident m6A modification sites on the MEF2C mRNA. METTL3 antibody could pull down a higher level of MEF2C mRNA than the IgG in RNA binding protein immunoprecipitation assay. Knockdown of METTL3 promoted MEF2C protein expression and MEF2C mRNA expression, accompanied by a reduced m6A modification level on the MEF2C mRNA. Knockdown of MEF2C inhibited the anti-inflammatory effect of METTL3 siRNA. Our results suggest that METTL3 promotes microglia inflammation via regulating MEF2C mRNA m6A modification induced by SCI and LPS treatment.

Mutation of TP53 Confers Ferroptosis Resistance in Lung Cancer Through the FOXM1/MEF2C Axis

Ferroptosis is a highly regulated tumor suppressor process. Loss or mutation of TP53 can cause changes in sensitivity to ferroptosis. Mutations in TP53 may be associated with the malignant or indolent progression of ground glass nodules in early lung cancer, but whether ferroptosis may also be involved in determining this biological process has not yet been determined. Using invivo and invitro gain- and loss-of-function approaches, this study used clinical tissue for mutation analysis and pathological research to show that wild-type TP53 inhibited the expression of forkhead box M1 (FOXM1) by binding to peroxisome proliferator-activated receptor-γ coactivator 1α, maintaining the mitochondrial function and thus affecting the sensitivity to ferroptosis. This function was absent in mutant cells, resulting in overexpression of FOXM1 and ferroptosis resistance. Mechanistically, FOXM1 activated the transcription level of myocyte-specific enhancer factor 2C in the mitogen-activated protein kinase signaling pathway, leading to stress protection when exposed to ferroptosis inducers. This study provides new insights into the mechanism of association between TP53 mutation and ferroptosis tolerance, which can aid a deeper understanding of the role of TP53 in the malignant progression of lung cancer.

Overlap between Central and Peripheral Transcriptomes in Parkinson's Disease but Not Alzheimer's Disease.

Most neurodegenerative disorders take decades to develop, and their early detection is challenged by confounding non-pathological ageing processes. Therefore, the discovery of genes and molecular pathways in both peripheral and brain tissues that are highly predictive of disease evolution is necessary. To find genes that influence Alzheimer’s disease (AD) and Parkinson’s disease (PD) pathogenesis, human RNA-Seq transcriptomic data from Brodmann Area 9 (BA9) of the dorsolateral prefrontal cortex (DLPFC), whole blood (WB), and peripheral blood mononuclear cells (PBMC) were analysed using a combination of differential gene expression and a random forest-based machine learning algorithm. The results suggest that there is little overlap between PD and AD, and the AD brain signature is unique mainly compared to blood-based samples. Moreover, the AD-BA9 was characterised by changes in ‘nervous system development’ with Myocyte-specific enhancer factor 2C (Mef2C), encoding a transcription factor that induces microglia activation, a prominent feature. The peripheral AD transcriptome was associated with alterations in ‘viral process’, and FYN, which has been previously shown to link amyloid-beta and tau, was the prominent feature. However, in the absence of any overlap with the central transcriptome, it is unclear whether peripheral FYN levels reflect AD severity or progression. In PD, central and peripheral signatures are characterised by anomalies in ‘exocytosis’ and specific genes related to the SNARE complex, including Vesicle-associated membrane protein 2 (VAMP2), Syntaxin 1A (STX1A), and p21-activated kinase 1 (PAK1). This is consistent with our current understanding of the physiological role of alpha-synuclein and how alpha-synuclein oligomers compromise vesicle docking and neurotransmission. Overall, the results describe distinct disease-specific pathomechanisms, both within the brain and peripherally, for the two most common neurodegenerative disorders.

WWP1 Deficiency Alleviates Cardiac Remodeling Induced by Simulated Microgravity.

Cardiac muscle is extremely sensitive to changes in loading conditions; the microgravity during space flight can cause cardiac remodeling and function decline. At present, the mechanism of microgravity-induced cardiac remodeling remains to be revealed. WW domain-containing E3 ubiquitin protein ligase 1 (WWP1) is an important activator of pressure overload-induced cardiac remodeling by stabilizing disheveled segment polarity proteins 2 (DVL2) and activating the calcium-calmodulin-dependent protein kinase II (CaMKII)/histone deacetylase 4 (HDAC4)/myocyte-specific enhancer factor 2C (MEF2C) axis. However, the role of WWP1 in cardiac remodeling induced by microgravity is unknown. The purpose of this study was to determine whether WWP1 was also involved in the regulation of cardiac remodeling caused by microgravity. Firstly, we detected the expression of WWP1 and DVL2 in the heart from mice and monkeys after simulated microgravity using western blotting and immunohistochemistry. Secondly, WWP1 knockout (KO) and wild-type (WT) mice were subjected to tail suspension (TS) to simulate microgravity effect. We assessed the cardiac remodeling in morphology and function through a histological analysis and echocardiography. Finally, we detected the phosphorylation levels of CaMKII and HDAC4 in the hearts from WT and WWP1 KO mice after TS. The results revealed the increased expression of WWP1 and DVL2 in the hearts both from mice and monkeys after simulated microgravity. WWP1 deficiency alleviated simulated microgravity-induced cardiac atrophy and function decline. The histological analysis demonstrated WWP1 KO inhibited the decreases in the size of individual cardiomyocytes of mice after tail suspension. WWP1 KO can inhibit the activation of the DVL2/CaMKII/HDAC4 pathway in the hearts of mice induced by simulated microgravity. These results demonstrated WWP1 as a potential therapeutic target for cardiac remodeling and function decline induced by simulated microgravity.

Type I interferon response-related microglial Mef2c deregulation at the onset of Alzheimer's pathology in 5×FAD mice

Alzheimer's disease (AD) is a chronic neurodegenerative disorder with multifactorial etiology. The role of microglia in the pathogenesis of AD has been increasingly recognized in recent years; however, the detailed mechanisms shaping microglial phenotypes in AD-relevant pathological settings remain largely unresolved. Myocyte-specific enhancer factor 2C (Mef2C) is a transcription factor with versatile functions. Recent studies have attributed aging-related microglial changes to type I interferon (IFN-I)-associated Mef2C deregulation. In view of the close relationship between brain aging and AD, it is of great interest to determine microglial Mef2C changes in AD-related conditions. In this study, we have found that suppressed Mef2C nuclear translocation was an early and prominent microglial phenotype in a mouse model of brain amyloidosis (5×FAD mice), which exacerbated with age. Echoing the early Mef2C deregulation and its association with microglial activation, transcriptional data showed elicited IFN-I response in microglia from young 5×FAD mice. Amyloid beta 42 (Aβ42) in its oligomeric forms promoted Mef2C deregulation in microglia on acute organotypic brain slices with augmented microglial activation and synapse elimination via microglial phagocytosis. Importantly, these oligomeric Aβ42-mediated microglial changes were substantially attenuated by blocking IFN-I signaling. The simplest interpretation of the results is that Mef2C, concurring with activated IFN-I signaling, constitutes early microglial changes in AD-related conditions. In addition to the potential contribution of Mef2C deregulation to the development of microglial phenotypes in AD, Mef2C suppression in microglia may serve as a potential mechanistic pathway linking brain aging and AD.

Molecular Mechanisms of Epithelial to Mesenchymal Transition Regulated by ERK5 Signaling.

Extracellular signal-regulated kinase (ERK5) is an essential regulator of cancer progression, tumor relapse, and poor patient survival. Epithelial to mesenchymal transition (EMT) is a complex oncogenic process, which drives cell invasion, stemness, and metastases. Activators of ERK5, including mitogen-activated protein kinase 5 (MEK5), tumor necrosis factor α (TNF-α), and transforming growth factor-β (TGF-β), are known to induce EMT and metastases in breast, lung, colorectal, and other cancers. Several downstream targets of the ERK5 pathway, such as myocyte-specific enhancer factor 2c (MEF2C), activator protein-1 (AP-1), focal adhesion kinase (FAK), and c-Myc, play a critical role in the regulation of EMT transcription factors SNAIL, SLUG, and β-catenin. Moreover, ERK5 activation increases the release of extracellular matrix metalloproteinases (MMPs), facilitating breakdown of the extracellular matrix (ECM) and local tumor invasion. Targeting the ERK5 signaling pathway using small molecule inhibitors, microRNAs, and knockdown approaches decreases EMT, cell invasion, and metastases via several mechanisms. The focus of the current review is to highlight the mechanisms which are known to mediate cancer EMT via ERK5 signaling. Several therapeutic approaches that can be undertaken to target the ERK5 pathway and inhibit or reverse EMT and metastases are discussed.

Myocyte-specific enhancer factor 2c triggers transdifferentiation of adipose tissue-derived stromal cells into spontaneously beating cardiomyocyte-like cells

Cardiomyocyte regeneration is limited in adults. The adipose tissue-derived stromal vascular fraction (Ad-SVF) contains pluripotent stem cells that rarely transdifferentiate into spontaneously beating cardiomyocyte-like cells (beating CMs). However, the characteristics of beating CMs and the factors that regulate the differentiation of Ad-SVF toward the cardiac lineage are unknown. We developed a simple culture protocol under which the adult murine inguinal Ad-SVF reproducibly transdifferentiates into beating CMs without induction. The beating CMs showed the striated ventricular phenotype of cardiomyocytes and synchronised oscillation of the intracellular calcium concentration among cells on day 28 of Ad-SVF primary culture. We also identified beating CM-fated progenitors (CFPs) and performed single-cell transcriptome analysis of these CFPs. Among 491 transcription factors that were differentially expressed (≥ 1.75-fold) in CFPs and the beating CMs, myocyte-specific enhancer 2c (Mef2c) was key. Transduction of Ad-SVF cells with Mef2c using a lentiviral vector yielded CFPs and beating CMs with ~ tenfold higher cardiac troponin T expression, which was abolished by silencing of Mef2c. Thus, we identified the master gene required for transdifferentiation of Ad-SVF into beating CMs. These findings will facilitate the development of novel cardiac regeneration therapies based on gene-modified, cardiac lineage-directed Ad-SVF cells.

Circular RNA circ_DROSHA alleviates the neural damage in a cell model of temporal lobe epilepsy through regulating miR-106b-5p/MEF2C axis

Temporal lobe epilepsy (TLE) is the most prevalent form of acquired epilepsy. Circular RNAs (circRNAs) have recently been highlighted as important regulators in TLE. Nevertheless, the role and mechanism of circRNA Drosha ribonuclease III (circ_DROSHA) in TLE pathogenesis are still unknown. Magnesium-free extracellular solution was used to establish the TLE cell model. The levels of circ_DROSHA, myocyte-specific enhancer factor 2C (MEF2C) and miR-106b-5p were determined by qRT-PCR and western blot. Cell proliferation was detected by the Cell Counting-8 Kit (CCK-8) assay, and cell apoptosis was measured by flow cytometry. Targeted relationships among circ_DROSHA, miR-106b-5p and MEF2C were confirmed by a dual-luciferase reporter or RNA immunoprecipitation (RIP) assay. Our data showed that circ_DROSHA was down-regulated in the serum samples of TLE patients and the TLE cell model. Circ_DROSHA up-regulation alleviated the cytotoxicity of the TLE cell model by enhancing cell proliferation and repressing cell apoptosis. Circ_DROSHA directly bound to miR-106b-5p. Moreover, miR-106b-5p represented a downstream effector of circ_DROSHA function. MEF2C was a direct target of miR-106b-5p, and miR-106b-5p knockdown relieved magnesium-free treatment-induced cell injury by up-regulating MEF2C. Furthermore, circ_DROSHA regulated MEF2C expression via sponging miR-106b-5p. Our study suggested that the enforced expression of circ_DROSHA alleviated the cell damage of the TLE cell model at least in part through the regulation of the miR-106b-5p/MEF2C axis.

O-GlcNAcylation of Mef2c regulates myoblast differentiation

Unlike other types of glycosylation, O-GlcNAcylation is a single glycosylation which occurs exclusively in the nucleus and cytosol. O-GlcNAcylation underlie metabolic diseases, including diabetes and obesity. Furthermore, O-GlcNAcylation affects different oncogenic processes such as osteoblast differentiation, adipogenesis and hematopoiesis. Emerging evidence suggests that skeletal muscle differentiation is also regulated by O-GlcNAcylation, but the detailed molecular mechanism has not been fully elucidated. In this study, we showed that hyper-O-GlcNAcylation reduced the expression of myogenin, a transcription factor critical for terminal muscle development, in C2C12 myoblasts differentiation by O-GlcNAcylation on Thr9 of myocyte-specific enhancer factor 2c. Furthermore, we showed that O-GlcNAcylation on Mef2c inhibited its DNA binding affinity to myogenin promoter. Taken together, we demonstrated that hyper-O-GlcNAcylation attenuates skeletal muscle differentiation by increased O-GlcNAcylation on Mef2c, which downregulates its DNA binding affinity.

Downregulation of lncRNA-11496 in the Brain Contributes to Microglia Apoptosis via Regulation of Mef2c in Chronic T. gondii Infection Mice.

Though it is well known that chronic infections of Toxoplasma gondii (T. gondii) can induce mental and behavioral disorders in the host, little is known about the role of long non-coding RNAs (lncRNAs) in this pathological process. In this study, we employed an advanced lncRNAs and mRNAs integration chip (Affymetrix HTA 2.0) to detect the expression of both lncRNAs and mRNAs in T. gondii Chinese 1 strain infected mouse brain. As a result, for the first time, the downregulation of lncRNA-11496 (NONMMUGO11496) was identified as the responsible factor for this pathological process. We showed that dysregulation of lncRNA-11496 affected proliferation, differentiation and apoptosis of mouse microglia. Furthermore, we proved that Mef2c (Myocyte-specific enhancer factor 2C), a member of the MEF2 subfamily, is the target gene of lncRNA-11496. In a more detailed study, we confirmed that lncRNA-11496 positively regulated the expression of Mef2c by binding to histone deacetylase 2 (HDAC2). Importantly, Mef2c itself could coordinate neuronal differentiation, survival, as well as synapse formation. Thus, our current study provides the first evidence in terms of the modulatory action of lncRNAs in chronic toxoplasmosis in T. gondii infected mouse brain, providing a solid scientific basis for using lncRNA-11496 as a therapeutic target to treat T. gondii induced neurological disorder.

The CaMKII phosphorylation site Thr1604 in the CaV1.2 channel is involved in pathological myocardial hypertrophy in rats

ABSTRACT Residue Thr1604 in the CaV1.2 channel is a Ca2+/calmodulin dependent protein kinase II (CaMKII) phosphorylation site, and its phosphorylation status maintains the basic activity of the channel. However, the role of CaV1.2 phosphorylation at Thr1604 in myocardial hypertrophy is incompletely understood. Isoproterenol (ISO) was used to induce cardiomyocyte hypertrophy, and autocamtide-2-related inhibitory peptide (AIP) was added as a treatment. Rats in a myocardial hypertrophy development model were subcutaneously injected with ISO for two or three weeks. The heart and left ventricle weights, each of which were normalized to the body weight and cross-sectional area of the myocardial cells, were used to describe the degree of hypertrophy. Protein expression levels were detected by western blotting. CaMKII-induced CaV1.2 (Thr1604) phosphorylation (p-CaV1.2) was assayed by coimmunoprecipitation. The results showed that CaMKII, HDAC, MEF2 C, and atrial natriuretic peptide (ANP) expression was increased in the ISO group and downregulated by AIP treatment in vitro. There was no difference in the expression of these proteins between the ISO 2-week group and the ISO 3-week group in vivo. CaV1.2 channel expression did not change, but p-CaV1.2 expression was increased after ISO stimulation and decreased by AIP. In the rat model, p-CaV1.2 levels and CaMKII activity were much higher in the ISO 3-week group than in the ISO 2-week group. CaMKII-induced CaV1.2 channel phosphorylation at residue Thr1604 may be one of the key features of myocardial hypertrophy and disease development.Abbreviations: CaMKII: Ca2+/calmodulin dependent protein kinase II; p-CaMKII: autophosphorylated Ca2+/calmodulin dependent protein kinase II; CaM: calmodulin; AIP: autocamtide-2-related inhibitory peptide; ECC: excitation-contraction coupling; ISO: isoproterenol; BW: body weight; HW: heart weight; LVW: left ventricle weight; HDAC: histone deacetylase; p-HDAC: phosphorylated histone deacetylase; MEF2C: myocyte-specific enhancer factor 2C; ANP: atrial natriuretic peptide; PKC: protein kinase C

Establishing a new human hypertrophic cardiomyopathy-specific model using human embryonic stem cells

Symptom of ventricular hypertrophy caused by cardiac troponin T (TNNT2) mutations is mild, while patients often showed high incidence of sudden cardiac death. The 92nd arginine to glutamine mutation (R92Q) of cTnT was one of the mutant hotspots in hypertrophic cardiomyopathy (HCM). However, there are no such human disease models yet. To solve this problem, we generated TNNT2 R92Q mutant hESC cell lines (heterozygote or homozygote) using TALEN mediated homologous recombination in this study. After directed cardiac differentiation, we found a relative larger cell size in both heterozygous and homozygous TNNT2 R92Q hESC-cardiomyocytes. Expression of atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), and sarcoplasmic reticulum Ca2+-ATPase2a (SERCA2a) were downregulated, while myocyte specific enhancer factor 2c (MEF2c) and the ratio of beta myosin to alpha myosin heavy chain (MYH7/MYH6) were increased in heterozygous TNNT2 R92Q hESC-cardiomyocytes. TNNT2 R92Q mutant cardiomyocytes exhibited efficient responses to heart-related pharmaceutical agents. We also found TNNT2 R92Q heterozygous mutant cardiomyocytes showed increased calcium sensitivity and contractility. Further, engineered heart tissues (EHTs) prepared by combining rat decellularized heart extracellular matrices with heterozygous R92Q mutant cardiomyocytes showed similar drug responses as to HCM patients and increased sensitivity to caspofungin-induced cardiotoxicity. Using RNA-sequencing of TNNT2 R92Q heterozygous mutant cardiomyocytes, we found dysregulation of calcium might participated in the early development of hypertrophy. Our hESC-derived TNNT2 R92Q mutant cardiomyocytes and EHTs are good in vitro human disease models for future disease studies and drug screening.

Key nutritional factors and interactions during larval development of pikeperch (Sander lucioperca)

The effects of 8 nutritional variables (Ca/P, Eicosapentaenoic acid (20:5n-3) + Docosahexaenoic acid (22:6n − 3) (EPA + DHA), Arachidonic acid (20:4n − 6) (ARA), Se, vitamins E, C, D and A) were investigated to identify their respective importance and interactions in pikeperch larval development. In this respect, two modalities (low and high levels) of each variable were tested through a fractional factorial experimental design allowing a reduction from 256 (28) to 16 (28 – 4) experimental units. Survival was significantly higher in larvae fed a high Ca/P diet while larval growth was significantly lower in larvae fed the same diet variant, associated with a higher incidence of kyphosis and pectoral anomalies in these larvae. Lordosis and scoliosis seemed to be mostly affected by dietary long chain polyunsaturated fatty acids (LC-PUFAs). A significant interaction was shown between n-3 LC-PUFA and vitamin C on jaw anomalies, while myocyte-specific enhancer factor 2C (mef2c) gene expression correlated positively with dietary vitamin C increment. Results also demonstrated an effect of the different nutrients and their interactions on the activity levels of digestive enzymatic activities. The results of the present study highlight the importance of the interactions between Ca/P, LC-PUFAs and vitamins C and E, suggesting their essential roles as key nutritional factors influencing pikeperch larval development.

Lipin1 is required for skeletal muscle development by regulating MEF2c and MyoD expression.

Lipin1 is critical for skeletal muscle development. Lipin1 regulates MyoD and myocyte-specific enhancer factor 2C (MEF2c) expression via the protein kinase C (PKC)/histone deacetylase 5-mediated pathway. Inhibition of PKCμ activity suppresses myoblast differentiation by inhibiting MyoD and MEF2c expression. Our previous characterization of global lipin1-deficient (fld) mice demonstrated that lipin1 played a novel role in skeletal muscle (SM) regeneration. The present study using cell type-specific Myf5-cre;Lipin1fl/fl conditional knockout mice (Lipin1Myf5cKO ) shows that lipin1 is a major determinant of SM development. Lipin1 deficiency induced reduced muscle mass and myopathy. Our results from lipin1-deficient myoblasts suggested that lipin1 regulates myoblast differentiation via the protein kinase Cμ (PKCμ)/histone deacetylase 5 (HDAC5)/myocyte-specific enhancer factor 2C (MEF2c):MyoD-mediated pathway. Lipin1 deficiency leads to the suppression of PKC isoform activities, as well as inhibition of the downstream target of PKCμ, class II deacetylase HDAC5 nuclear export, and, consequently, inhibition of MEF2c and MyoD expression in the SM of lipin1Myf5cKO mice. Restoration of diacylglycerol-mediated signalling in lipin1 deficient myoblasts by phorbol 12-myristate 13-acetate transiently activated PKC and HDAC5, and upregulated MEF2c expression. Our findings provide insights into the signalling circuitry that regulates SM development, and have important implications for developing intervention aimed at treating muscular dystrophy.

Novel MEF2C point mutations in Chinese patients with Rett (\u2212like) syndrome or non-syndromic intellectual disability: insights into genotype-phenotype correlation

BackgroundMEF2C (Myocyte-specific enhancer factor 2C) has been associated with neurodevelopmental disorders. This study aimed at delineating the clinical profiles of MEF2C gene mutations.MethodsIn total, 112 Chinese patients with intellectual disability (ID) were recruited, including 44 patients presented with Rett syndrome (RTT) or RTT-like syndrome, and 68 patients with non-syndromic ID. Targeted next-generation sequencing (NGS) was performed. Detailed clinical information was collected.ResultsFive heterozygous MEF2C gene mutations were identified, of which three were novel. The MEF2C mutant rate was 4.5% (5/112) in total, and 6.8% (3/44) in the RTT (−like) cohort. All patients with MEF2C gene mutation presented with cognitive impairment, gross motor delay, speech disorder and autistic features. Four patients had epilepsy, which responded well to antiepileptic drugs. One female was diagnosed with classical RTT, two females with RTT-like syndrome, and two males with non-syndromic ID. Generally, the phenotype of two males with relatively downstream mutations (c.565C > T, p.Arg 189*; c.766C > T, p.Arg 256*) was milder than that of three females with upstream mutations (c.48C > G, p.Asn16Lys; c.334G > T, p.Glu112* and c.403-1G > T).ConclusionsOur findings expanded the current understanding of the consequences of MEF2C dysfunctions, especially MEF2C point mutations. MEF2C mutations are associated with a broad clinical spectrum, ranged from classical RTT to non-syndromic ID. Through our study, it can be inferred that there is correlation between the phenotype and MEF2C-genotype, the mutation site. Overall, the MEF2C gene mutational analysis should be performed in ID cohort, especially in patients with features overlapped with RTT.

Comprehensive analysis of multi Ewing sarcoma microarray datasets identifies several prognosis biomarkers.

Ewing sarcoma (ES) is a common primary malignancy in children and adolescents. Progression of treatment methods hasn't contributed a lot to the imrovement of prognosis. To identify potential prognostic biomarkers, a meta-analysis pipeline of multi-gene expression datasets for ES from the Gene Expression Omnibus (GEO) was performed. Three datasets were screened and differential expression genes (DEGs) in ES samples compared with normal tissues were identified through limma package and subjected to network analysis. As a result, 1,470 DEGs were obtained which were mainly involved in biological processes associated with immune response and transcription regulation. Network analysis obtained 22 core genes with high network degree and fold change. Kaplan-Meier analysis based on ES datasets from The Cancer Genome Atlas identified five genes, including glycogen phosphorylase, muscle-associated, myocyte-specific enhancer factor 2C, tripartite motif containing 63, budding uninhibited by benzimidazoses1 and Ras GTPase-activating protein 1, whose altered expression profiles are significantly associated with survival. Changes of their expression values were further confirmed through RT-qPCR in ES cell and normal cell lines. Those genes may be considered as potential prognostic biomarkers of ES and should be helpful for its early diagnosis and treatment.

MicroRNA-139-5p elevates skeletal myogenic differentiation of human adult dental pulp stem cells through Wnt/β-catenin signaling pathway.

The aim of the present study was to identify a microRNA (miRNA or miR)-based biomarker and therapeutic target for skeletal myogenic differentiation of human adult dental pulp stem cells (ADSCs). miRNA expression was measured using reverse transcription-quantitative polymerase chain reaction; cell viability assay and lactate dehydrogenase (LDH) activity levels were measured using MTT and LDH activity kits, respectively. Apoptosis assay and caspase-3/9 activity levels were measured using flow cytometry and caspase-3/9 activity kits, respectively. Western blot analysis and immunofluorescence microscopy measured the protein expression of myocyte-specific enhancer factor 2C, myogenic differentiation 1, myosin heavy chain, Wnt and β-catenin. Overexpression of miR-139-5p promoted cell growth and induced skeletal myogenic differentiation of ADSCs. Downregulation of miR-139-5p inhibited cell growth and reduced skeletal myogenic differentiation of ADSCs. Overexpression of miR-139-5p induced Wnt/β-catenin signaling pathway and Wnt/β-catenin signaling pathway was suppressed by anti-miR-139-5p in ADSCs. Wnt inhibitor reduced the effect of miR-139-5p on skeletal myogenic differentiation of ADSCs. In conclusion, miR-139-5p elevates skeletal myogenic differentiation of human ADSCs through the Wnt/β-catenin signaling pathway.

The phosphodiesterase-4 inhibitor roflumilast reverts proteolysis in skeletal muscle cells of patients with COPD cachexia.

Peripheral muscle weakness and mass loss are characteristic features in severe chronic obstructive pulmonary disease (COPD). We hypothesized that the phosphodiesterase (PDE)-4 inhibitor roflumilast-induced cAMP may ameliorate proteolysis and metabolism in skeletal muscles of COPD patients with severe muscle wasting. In myogenic precursor cells (isolated from muscle biopsies and cultured up to obtain differentiated myotubes) from 10 severe COPD patients and 10 healthy controls, which were treated with 1 μM roflumilast N-oxide (RNO) for three time cohorts (1, 6, and 24 h), genes of antioxidant defense and oxidative stress marker, myogenesis and muscle metabolism, proteolysis (tyrosine release assay) and ubiquitin-proteasome system markers, autophagy, and myosin isoforms were analyzed using RT-PCR and immunoblotting. In COPD patients at 6 h RNO treatment, myotube tyrosine release, total protein ubiquitination, and tripartite motif-containing protein 32 levels were significantly lower than healthy controls, whereas at 24 h RNO treatment, myotube myosin heavy chain ( MyHC) -I and MyHC-IIx expression levels were upregulated in both patients and controls. In the 6-h RNO cohort, in patients and controls, myotube expression of nuclear factor (erythroid-derived 2)-like 2 ( NRF2) and its downstream antioxidants sirtuin-1, FGF-inducible 14, and insulin-like growth factor-1 was upregulated, whereas that of myocyte-specific enhancer factor 2C, myogenic differentiation, myogenin, myostatin, atrogin-1, and muscle RING-finger protein-1 was downregulated. In myotubes of severe COPD patients with cachexia, roflumilast-induced cAMP signaling exerts beneficial effects by targeting muscle protein breakdown (tyrosine release), along with reduced expression of proteolytic markers of the ubiquitin-proteasome system and that of myostatin. In both patients and controls, roflumilast also favored antioxidant defense through upregulation of the NRF2 pathway and that of the histone deacetylase sirtuin-1, whereas it improved the expression of slow- and fast-twitch myosin isoforms. These findings show that muscle dysfunction and wasting may be targeted by roflumilast-induced cAMP signaling in COPD. These results have potential therapeutic implications, as this PDE-4 inhibitor is currently available for the treatment of systemic inflammation and exacerbations in patients with severe COPD. NEW & NOTEWORTHY In myotubes of cachectic chronic obstructive pulmonary disease (COPD) patients, cAMP signaling exerted beneficial effects by targeting muscle proteolysis and reducing gene expression of proteolytic markers of the ubiquitin-proteasome system and that of myostatin. In myotubes of patients and controls, roflumilast also favored antioxidant defense through upregulation of the nuclear factor (erythroid-derived 2)-like 2 pathway, of sirtuin-1, and of gene expression of slow- and fast-twitch isoforms. These findings have potential clinical implications for the treatment of muscle wasting in patients with COPD and cachexia.

Podocyte-specific Rac1 deficiency ameliorates podocyte damage and proteinuria in STZ-induced diabetic nephropathy in mice

Activation of Ras-related C3 botulinum toxin substrate 1 (Rac1) has been implicated in diverse kidney diseases, yet its in vivo significance in diabetic nephropathy (DN) is largely unknown. In the present study, we demonstrated a podocyte-specific Rac1-deficient mouse strain and showed that specific inhibition of Rac1 was able to attenuate diabetic podocyte injury and proteinuria by the blockade of Rac1/PAK1/p38/β-catenin signaling cascade, which reinstated the integrity of podocyte slit diaphragms (SD), rectified the effacement of foot processes (FPs), and prevented the dedifferentiation of podocytes. In vitro, we showed Rac1/PAK1 physically bound to β-catenin and had a direct phosphorylation modification on its C-terminal Ser675, leading to less ubiquitylated β-catenin, namely more stabilized β-catenin, and its nuclear migration under high-glucose conditions; further, p38 activation might be responsible for β-catenin nuclear accumulation via potentiating myocyte-specific enhancer factor 2C (MEF2c) phosphorylation. These findings provided evidence for a potential renoprotective and therapeutic strategy of cell-specific Rac1 deficiency for DN and other proteinuric diseases.