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Please use this identifier to cite or link to this item: http://hdl.handle.net/1993/11863

Title: MicroRNA-301a attenuates collagen gel contractility and promotes proliferation of human bone marrow derived mesenchymal stem cells
Authors: Klassen, Jessica
Supervisor: Dr. Darren Freed (Institute of Cardiac Sciences, St. Boniface Research Centre)
Examining Committee: Medicine
Graduation Date: October 2012
Keywords: Medicine
Issue Date: 23-Nov-2012
Abstract: Previous data in our lab suggests that human mesenchymal stem cells (MSCs) of bone marrow origin display a myofibroblast phenotype in vitro. Coincident with the onset of a myofibroblast phenotype, we observed a reduction in miR-301a expression. We hypothesized that miR-301a may govern the differentiation of bone marrow derived MSCs to myofibroblasts. MSCs were isolated from the sternum of patients undergoing open heart surgery and transfected with premiR-301a at early passage. We analyzed contractility through plating on flexible collagen gels and cell proliferation. Western blot and qRT-PCR were used to determine mRNA and protein expression of myosin isoforms and myofibroblast markers. Human MSCs were found to express non-muscle myosin (NMM)-IIA and -IIB, other myofibroblast markers such as α-smooth muscle actin (SMA), vimentin, FSP-1, ED-A Fibronectin, and procollagen I and contracted collagen gels in the basal state. Cells cultured in serum rich conditions displayed decreased contractility and increased proliferation whereas cells cultured in serum free conditions showed increased contractility and decreased proliferative capacity. The proliferative phenotype was associated with increased mRNA expression of miR-301a host gene, ska2. Overexpression of miR-301a caused a reduction in collagen gel contraction and an increase in proliferation when compared with a negative-scramble control. This was associated with decreased NMM-IIA and IIB mRNA. Western blot analysis confirmed decreased levels of NMM-IIA and IIB in addition to decreased protein expression of α-SMA, ED-A Fibronectin and procollagen I. In conclusion, MSCs display a dichotomous proliferative versus contractile phenotype in vitro. Our results suggest that miR-301a plays a mechanistic role in this dichotomy, such that it suppresses the myofibroblastic, contractile phenotype.
URI: http://hdl.handle.net/1993/11863
Appears in Collection(s):Faculty of Medicine, B.Sc. (Med) Projects

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