Engineering of functional smooth muscle cells from gastrointestinal myofibroblast

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 DESCRIPTION (provided by applicant): Dysfunctions of smooth muscle cells (SMCs) are associated with hypertrophy, hyperplasia and fibrosis found in SM diseases including gastrointestinal (GI) bowel obstruction and Hirschprung's disease. Defective SMCs are dedifferentiated cells that have lost their contractile phenotype. The phenotypic change of SMCs to non- functional dedifferentiated cells has been discovered in vascular injuries ~50 years ago, but the phenotypic and genetic identity of the transformed cells are still unknown. We have found that the transformed cells in the hypertrophic small intestine are myofibroblast like cells that weakly express platelet-derived growth factor receptor alpha and beta (PDGFRa/ß) (called "PDGFRalowßhigh cells"). We have isolated differentiated PDGFRahigh cells and SMCs from colon and jejunum, and obtained genome scale gene expression profiles. By analyzing the transcriptomes, we identified SMC-specific transcription factors including SRF and MYOCD. Moreover, we found that the phenotype of SMCs is regulated by SRF and an epigenetic modulator DNA methyltransferase 1 and 3a (DNMT1/3a). The present project seeks to uncover a cellar and genetic mechanism for understanding how SMCs are dedifferentiated during the development of intestinal SM hypertrophy and how dedifferentiated cells are redifferentiated into SMCs during the recovery process. Furthermore, this project aims to isolate dedifferentiated cells, establish a cell line in culture, and genetically engineer SMCs from the cell line using SMC-specific transcription factors and the epigenetic regulator DNMT1/3a. To achieve these aims, we have formed a collaborative, multi-institutional research team with Drs. Seungil Ro (PhD, PI), Moon young Lee (MD/PhD, studying the plasticity of SMCs in the hypertrophic condition), a to-be-hired postdoc (PhD, studying the engineering of SMCs from PDGRFalow cells), Douglas Redelman (PhD, cell isolation), Gabsang Lee (PhD/DVM, cell line establishment), Terence Smith (PhD, calcium imaging), Laren Becker (MD/PhD, tissue supply and consulting), Kent Sasse (MD, tissue supply), and Joseph Miano and Kenton Sanders (PhDs, research tool supply and project consulting). These studies will significantly advance our understanding on the phenotypic transition of SMCs at the transcriptome level and could offer a human SMC line for clinical use to repair damaged or non-functional SMCs found in GI SM diseases.