Highly Coordinated Gene Regulation in Mouse Skeletal Muscle Regeneration

Mammalian skeletal muscles are capable of regeneration after injury. Quiescent satellite cells are activated to reenter the cell cycle and to differentiate for repair, recapitulating features of myogenesis during embryonic development. To understand better the molecular mechanism involved in this process in vivo, we employed high density cDNA microarrays for gene expression profiling in mouse tibialis anterior muscles after a cardiotoxin injection. Among 16,267 gene elements surveyed, 3,532 elements showed at least a 2.5-fold change at one or more time points during a 14-day time course. Hierarchical cluster analysis and semiquantitative reverse transcription-PCR showed induction of genes important for cell cycle control and DNA replication during the early phase of muscle regeneration. Subsequently, genes for myogenic regulatory factors, a group of imprinted genes and genes with functions to inhibit cell cycle progression and promote myogenic differentiation, were induced when myogenic stem cells started to differentiate. Induction of a majority of these genes, including E2f1 and E2f2, was abolished in muscles lacking satellite cell activity after gamma radiation. Regeneration was severely compromised in E2f1 null mice but not affected in E2f2 null mice. This study identifies novel genes potentially important for muscle regeneration and reveals highly coordinated myogenic cell proliferation and differentiation programs in adult skeletal muscle regeneration in vivo

MicroRNA expression distinguishes SCLC from NSCLC lung tumor cells and suggests a possible pathological relationship between SCLCs and NSCLCs

<p>Abstract</p> <p>Background</p> <p>Recent studies have shown that microRNAs (miRNAs) play roles in tumorigenesis and are reliable classifiers of certain cancer types and subtypes. However, the role of miRNAs in the pathogenesis and diagnosis of small cell carcinoma (SCLC), the majority of which represent the most aggressive lung tumors, has not been investigated.</p> <p>Methods</p> <p>In order to explore miRNA involvement in the pathogenesis of small cell lung carcinoma (SCLC) and the potential role of miRNAs in SCLC diagnosis, we compared the miRNA expression profile of a set of SCLC cell lines to that of a set of non-small cell lung cancer (NSCLC) cell lines and normal immortalized human bronchial epithelial cells (HBECs) using microarray analysis.</p> <p>Results</p> <p>Our results show that miRNA profiles reliably distinguish SCLC cell lines from NSCLC and HBEC cell lines. Further analysis of the miRNA expression profile of the two subtypes of lung cancer cell lines indicates that the expression levels of the majority of the miRNAs that are differentially expressed in SCLC cells relative to NSCLC cells and HBECs show a progressive trend from HBECs to NSCLC cells to SCLC cells.</p> <p>Conclusions</p> <p>The distinctive miRNA expression signature of SCLCs relative to NSCLCs and HBECs suggests that miRNA profiles have the potential to serve as a diagnostic marker of SCLC lung tumors. The progressive trend of miRNA profile changes from HBECs to NSCLCs to SCLCs suggests a possible pathological relationship between SCLCs and NSCLCs, and suggests that the increasing dysregulation of miRNA expression may play a role in lung tumor progression. The specific role of these miRNAs in lung tumor pathogenesis and differentiation need to be investigated further in future studies.</p

The Complete Exosome Workflow Solution: From Isolation to Characterization of RNA Cargo

Exosomes are small (30–150 nm) vesicles containing unique RNA and protein cargo, secreted by all cell types in culture. They are also found in abundance in body fluids including blood, saliva, and urine. At the moment, the mechanism of exosome formation, the makeup of the cargo, biological pathways, and resulting functions are incompletely understood. One of their most intriguing roles is intercellular communication—exosomes function as the messengers, delivering various effector or signaling macromolecules between specific cells. There is an exponentially growing need to dissect structure and the function of exosomes and utilize them for development of minimally invasive diagnostics and therapeutics. Critical to further our understanding of exosomes is the development of reagents, tools, and protocols for their isolation, characterization, and analysis of their RNA and protein contents. Here we describe a complete exosome workflow solution, starting from fast and efficient extraction of exosomes from cell culture media and serum to isolation of RNA followed by characterization of exosomal RNA content using qRT-PCR and next-generation sequencing techniques. Effectiveness of this workflow is exemplified by analysis of the RNA content of exosomes derived from HeLa cell culture media and human serum, using Ion Torrent PGM as a sequencing platform