Browsing by Subject "smooth muscle cells"

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    Phase II – Biomechanics of Smooth Muscle Cell Differentiation: Experimental Study Using an Innovative In Vitro Mechanical System
    (Office of the Vice Chancellor for Research, 2014-04-11) Collins, Jessica; Sheikh, Zahir; Vipra, Niraj; Yeoh, Joseph
    Smooth muscle cells (SMCs) controls involuntary contractions and express different genotypic and phenotypic traits on specific organs such as blood vessels, bladder and stomach. However, studies have shown different SMC lineages tend to gradually lose specific characteristics due to a static milieu without exerting forces that they would experience naturally when cultured in vitro. The research provided in vivo conditions are mimicked effectively in vitro by applying controlled mechanical loading, SMCs should express their differentiated characteristics. We have validated an innovative mechanical device that simulates the pulsatile stretching SMCs undergo in their in vivo environment. Using the new system and cell and molecular biology techniques, we are evaluating cell differentiation and strain induced alignment when phenotypically modulated SMCs undergo cyclic mechanical loading at 10 and 20 percent strains, for 4, 6, or 8 hours at physiological frequency. We collected proteins after stretch experiments and analyzed via western blot, α-actin, γ-actin, transgelin, and calponin protein expression changes in: coronary SMCs strained 10% and 20% at 4, 6, and 8 hours, bladder SMCs strained 10% at 4, 6, and 8 hours, and BAECs for varying intensities and durations. In order to improvise the machine capability, LabVIEW code is been developed as the user interface providing advantageous of Graphical Approach instead of Cool Muscle Language code. Developed coding provide a complete coverage of acquisition, analysis, reporting, and display features to create modern applications that can scale as system requirements change over time. The next phase of this experiment enable analysis of gene expression using quantitative RT-PCR (qRT-PCR). This facet of research may prove valuable in the analysis of the effect of mechanical stress on maintaining SMC lineage as well as the study of how pathological stretch conditions affect SMC and endothelial cell gene and protein expressions.
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    The Regulation of MicroRNAs by Brahma-Related Gene 1 in Smooth Muscle Cells
    (Office of the Vice Chancellor for Research, 2013-04-05) Chen, Meng; Herring, Paul
    MicroRNAs (miRs) regulate the phenotypic switch of smooth muscle cells (SMCs) that occurs under several pathological conditions such as atherosclerosis. However, little is known about the transcriptional and epigenetic regulation of miR expression in SMCs. To identify miRs that are regulated by the Brahmarelated gene 1 (Brg1)-containing SWI/SNF chromatin remodeling complex we performed a microRNA array screen of RNA isolated from colonic SMCs of mice harboring a smooth muscle-specific knockout of Brg1. Quantitative RT-PCR confirmed changes in expression of several miRs, including miRs-143/145 and miR-133. Expression of dominant negative Brg1 in wild-type SMCs led to decreased expression of miRs-143/145 but not miR-133. The dominant negative Brg1 also blocked the myocardin-mediated induction of miRs-143/145 in 10T1/2 cells. Knockdown of SRF or myocardin decreased expression of miRs-143/145 in SMCs, whereas miR-133 expression was only repressed following SRF knockdown. In Brg1-null SW13 cells, miRs-143/145 but not miR-133 were dramatically induced by myocardin only in the presence of Brg1. Chromatin immunoprecipitation assays revealed that Brg1 is important for myocardin-mediated SRF binding to the miRs-143/145 promoter. Together these data show that Brg1- dependent chromatin remodeling regulates the expression of miRs-143/145 and miR-133 through distinct pathways in SMCs. This implies that chromatin remodeling complexes modulate smooth muscle phenotypes and functions not only through protein coding genes but also non-coding genes.