Browsing by Author "Hazenbiller, Olesja"
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- ItemOpen AccessMicro-environmental factors directing differentiation of murine embryonic stem cells down osteogenic and chondrogenic lineages(2013-02-15) Hazenbiller, Olesja; Duncan, Neil; Krawetz, RomanThe field of bone tissue engineering (BTE) aims to develop graft substitutes for diseased or difficult to heal fractures. We used a BTE construct made of collagen type I and murine embryonic stem cells (mESCs) which has been shown to trigger mESCs differentiation into osteoblasts, and successfully contribute to fracture repair in vivo within a mouse model system. Bone healing is a complex process involving the interplay of biochemical and biomechanical cues. Therefore, this project aimed to systematically emulate the roles of chemical and mechanical cues present during fracture repair on the differentiation of mESCs in vitro in order to optimize treatment strategies for BTE. Further characterization of this cell/gel construct revealed that mESCs differentiate into a heterogeneous cell population of chondrocytes and osteoblasts, replicating the process of endochondral ossification that normally occurs during fracture repair. To study the effect of biomechanical cues, a loading system was specifically designed and characterized to apply confined compressive load to a soft, viscoelastic cell/gel construct. Mechanical stimuli enhanced chondrogenic differentiation but had no effect on osteogenic differentiation. Moreover, the role of integrins in directing mESCs differentiation and transducing mechanical signals was evaluated. Finally, the synergistic effect between extracellular matrix mediated differentiation, mechanical stimulation and BMP-2 delivery to the system using nano-particles was studied.
- ItemOpen AccessReduction of pluripotent gene expression in murine embryonic stem cells exposed to mechanical loading or Cyclo RGD peptide(2017-11-14) Hazenbiller, Olesja; Duncan, Neil A; Krawetz, Roman JAbstract Background Self-renewal and differentiation of embryonic stem cells (ESCs) is directed by biological and/or physical cues that regulate multiple signaling cascades. We have previously shown that mESCs seeded in a type I collagen matrix demonstrate a loss of pluripotent marker expression and differentiate towards an osteogenic lineage. In this study, we examined if this effect was mediated in part through Arginylglycylaspartic acid (RGD) dependent integrin activity and/or mechano-transduction. Results The results from this study suggest that mESC interaction with the local microenvironment through RGD dependent integrins play a role in the regulation of mESC core transcription factors (TF), Oct-4, Sox 2 and Nanog. Disruption of this interaction with a cyclic RGD peptide (cRGDfC) was sufficient to mimic the effect of a mechanical stimulus in terms of pluripotent gene expression, specifically, we observed that supplementation with cRGDfC, or mechanical stimulus, significantly influenced mESC pluripotency by down-regulating core transcription factors. Moreover, our results indicated that the presence of the cRGDfC peptide inhibited integrin expression and up-regulated early lineage markers (mesoderm and ectoderm) in a Leukemia inhibitory factor (LIF) dependent manner. When cRGDfC treated mESCs were injected in Severe combined immunodeficiency (SCID) mice, no tissue growth and/or teratoma formation was observed, suggesting that the process of mESC tumor formation in vivo is potentially dependent on integrin interaction. Conclusions Overall, the disruption of cell-integrin interaction via cRGDfC peptide can mimic the effect of mechanical stimulation on mESC pluripotency gene expression and also inhibit the tumorigenic potential of mESCs in vivo.