Development of hyaluronic acid – poly(ethylene glycol) hydrogels towards hematopoietic differentiation of mouse embryonic stem cells
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The fields of tissue engineering, regenerative medicine, and stem cell engineering are rapidly growing. However, these fields must overcome several obstacles before they can make a significant impact on treating cellular disorders. Two major hurdles that must be addressed are: determining how to control the pluripotency of stem cells and developing systems for high-throughput culture of stem cells. The prospect of using a cell source capable of differentiating into cells of any tissue in the body (embryonic stem cells) has received enormous interest in recent years. The pluripotent attribute of embryonic stem cells seems ideal but developing methods to drive embryonic stem cells to specific lineages, including the hematopoietic lineage, is a complex process dependent on multiple intrinsic and extrinsic factors including chemical, cellular, and environmental signaling. With regards to environmental signaling, the use of three-dimensional culture systems such as scaffolds and hydrogels, have been utilized in an attempt to drive lineage-specific differentiation in a synthetic, biomimetic microenvironment. To determine specific environmental factors responsible for hematopoietic differentiation a systematic biological and engineering process must be implemented. A biodegradable hydrogel composed of the hyaluronic acid, a polysaccharide abundant in the bone marrow microenvironment, and the synthetic polymer, poly(ethylene glycol) was formulated to culture mouse embryonic stem cells (mESCs). Photoencapsulation of mESCs did not significantly decrease cellular viability or proliferation. The FACS data was inconclusive however, from gene expression studies, it was determined that the hydrogel culture system promoted differentiation of mESCs as evidenced by a down-regulation of the gene encoding for stem cell maintenance transcription factor, Oct-3/4. Furthermore, embryoid bodies, necessary for in vitro differentiation were observed in the hydrogel systems. Although an increase in the gene encoding for the cell surface marker, c-kit was up-regulated, the surface marker, sca-1 was not up-regulated. Up-regulation of both c-kit and sca-1 is necessary for the development of hematopoietic progenitor cells. Results indicate that the differentiation of mESCs into the hematopoietic lineage was unsuccessful but differentiation in these hydrogel systems did occur. Future cell marker and gene expression studies are necessary to determine which cell lineage the encapsulated mESCs are differentiating into before the effects of incorporating other environmental, cellular, and chemical factors can be investigated.