Cytoskeletal expression and remodeling during differentiation & redifferentiation

• Main Author: Guidry, Julia B
• Format: Dissertation
• Language: English
• Published: Ann Arbor ProQuest Dissertations & Theses 2015
• Subjects: Biomechanics; Biomedical engineering
• ISBN: 9781321751901; 1321751907

Because pluripotent stem cells (PSCs) can differentiate to any somatic cell type, they are highly studied as a source for cell-based therapies. Two types of PSCs are embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs). The use of ESCs in clinical applications are limited due to their inherently allogeneic nature and restriction in generating patient-specific or disease-specific cell lines. iPSCs hold potential as an autologous cell source for personalized stem cell-based therapies. Mechanical cues have been shown to direct differentiation of PSCs. In vitro studies expand the understanding of how iPSCs and ESCs respond to mechanical cues via cytoskeletal proteins. These studies will bring to light the potential role of the cytoskeleton in differentiation. Overall, the objective was to determine the relative cytoskeletal gene expression during differentiation and re-differentiation of ESCs and iPSCs, respectively. Spontaneous differentiation and force-mediated differentiation models were used to assess cytoskeletal expression and mesodermal differentiation in PSCs. First, we developed a model of differentiation for iPSCs comparable to an established ESC model of differentiation. When spontaneously differentiated as embryoid bodies, iPSCs were found to have higher mesodermal and cytoskeletal gene expression than that of ESCs. Under force-mediated differentiation, however, fluid shear stress increased mesodermal gene expression for both ESCs and iPSCs but resulted in different cytoskeletal responses. Differences in mesodermal and cytoskeletal expression in iPSCs and ESCs prompted further investigation into the cytoskeleton's role during differentiation. More specifically, we analyzed vimentin knockout ESCs (VIM KO-EBs) and wild-type ESCs (ESC-EBs) differentiated in an AggreWell EB model. VIM KO-EBs had lower mesodermal gene expression levels than that of ESC-EBs, indicating that vimentin may play a role in differentiation Overall, these studies indicate that cytoskeletal remodeling of iPSCs differed from that of ESCs during differentiation, which may be due to residual properties from the fibroblastic parental cell source of iPSCs. In addition, the cytoskeleton may play a role in a cell's differentiation capabilities. Consequently, therapies based on iPSCs may need to take into account residual properties from the parental cell source, such as cytoskeletal state and the mechanoresponse.