Project description:Amyotrophic lateral sclerosis (ALS) is the most common adult-onset paralytic disorder, characterized primarily by a progressive loss of motor neurons in whose degeneration skeletal muscle involvement has been demonstrated. Skeletal muscle is a plastic tissue that responds to insults through proliferation and differentiation of satellite cells. The process of skeletal muscle degeneration and regeneration are finely regulated by signals that regulate satellite cell proliferation and differentiation. It is known that satellite cell differentiation is impaired in ALS, but little is known about the involvement of miRNAs and their role in intercellular communication in skeletal muscle in ALS. Here we demonstrated impaired differentiation of satellite cells derived from ALS mice related to the impairment of myogenic p38MAPK and PKA/pCREB signaling pathways that can be regulated by miR-882 and -134-5p. These miRNAs participate in autocrine signaling in association with miR-26a-5p, that, secreted from WT and captured by ALS myoblasts, enhances ALS-related myoblast differentiation by repressing Smad4-related signals. These findings underscore the urgency of better understanding the role of intercellular communication and the molecular mechanisms underlying ALS pathogenesis and progression. They also suggest that miRNAs could be targeted as potential therapeutic agents to enhance myofiber regeneration.
Project description:We report a genome-wide study of the role of a muscle-specific Igf2/H19 transcriptional enhancer in global histone modifications in primary mouse myocytes. We generated 1.7 billion bases of sequence from DNA purified by the chromatin immunoprecipitation of H3K36me3 in wild type myocytes and myocytes bearing a deletion of the enhancer. Because H3K36me3 is reported to mark transcribed gene regions, we generated these data to develop a list of genes that are directly or indirectly regulated by this muscle enhancer. This study provides additional insight into how this muscle-specific enhancer potentially regulates a transcriptional network in mouse skeletal myocytes.
Project description:Objective: To study the uptake by human embryos of extracellular vesicles secreted by the maternal endometrium, and to investigate their miRNA cargo, in order to describe their role in implantation and early embryo development. Design: Prospective descriptive study. Subjects: Healthy women oocyte donors with confirmed fertility and day 5 human blastocysts. Intervention: Endometrial biopsies were collected from healthy oocyte donors undergoing transvaginal ultrasound-guided cyst aspiration for oocyte retrieval. Main Outcome Measures: Extracellular vesicle were isolated from culture media of primary human endometrial epithelial cells by ultracentrifugation. Concentration and size were analyzed by nanoparticle tracking analysis, their morphology visualized by transmission electron microscopy and extracellular vesicle protein markers expression was determined by western blotting. Vesicles were fluorescently labelled with Bodipy-TR ceramide, and their uptake by human blastocysts was analyzed using confocal microscopy. Analysis of the miRNA cargo of extracellular vesicles was performed using miRNAseq, target genes of the most expressed miRNAs were annotated and functional enrichment analysis was performed. Results: Extracellular vesicle characterization revealed a size within 100-300 nm, and expression of extracellular vesicle protein markers HSP70, TSG101, CD9 and CD81. Fluorescent microscopy showed an efficient extracellular vesicle internalization by human blastocysts within 1-2h, being the fluorescent signal stronger in the hatched area of the embryo. miRNAseq described 149 annotated miRNAs and top 37 most expressed miRNAs targeted 6,592 genes. Functional enrichment analysis of these targeted genes indicated that they participate in several processes related to embryo development, oxygen metabolism, cell cycle, cell differentiation, apoptosis, metabolism, cellular organization or gene expression. Among miRNAs contained in these EVs, hsa-miR-92a-3p, hsa-let-7b-5p, hsa-miR-30a-5p, hsa-miR-24-3p, hsa-miR-21-5p and hsa-let-7a-5p were highly implicated in all these biological processes. Conclusion: Data suggest that extracellular vesicles secreted by human endometrial epithelial cells are internalized by human blastocysts, and transport miRNAs to modulate biological processes related to implantation events and early embryo development. Knowledge of the communication system between human endometrium and embryo via miRNA cargo of these vesicles could describe new biomarkers of implantation success and embryo competence.
Project description:We report gene expression by RNAtag-seq after treatment with 75 different small molecule perturbations in culture of human iPSC-derived cardiac myocytes and genetically matched primary dermal fibroblasts. Perturbations were chosen from the SelleckChem Bioactive Library, among all molecules targeting any kinases or G-protein coupled receptors, and chosen to have as little overlap in annotated targets as possible. Based on these experiments (and others) we show that transcription factors important for cardiac development and cardiac myocyte identity maintenance were frequently up-regulated (i.e., "responsive") after small molecule perturbations of cultured iPSC-derived cardiac myocytes (i.e., responsive). We also show that the set of highly responsive transcription factors in fibroblasts are enriched for barriers to fibroblast reprogramming to iPSC.
Project description:DNA polymerase β (DNA polymerase beta, POLB) belongs to a member of the DNA polymerase X family, mainly involved in various biological metabolic processes such as eukaryotic DNA replication, DNA damage repair, gene recombination and cell cycle regulation. In this study, the muscle development-related gene POLB was screened by selection signature and RNA-seq analysis, and then validated for the proliferation and apoptosis of bovine primary myocytes. It was also found that overexpression of the POLB gene had a pro-apoptosis effect, but interfering with the expression of the gene had no significant effect on cells. Then the analysis of related apoptotic genes revealed that POLB overexpression affected CASP9 gene expression.
Project description:miRNAs are exported to high density lipoproteins (HDL). This study aimed to understand what miRNAs are present in primary human islets from 1 donor