Discipline: Biological Sciences
Subcategory: Biomedical Engineering
Room: Exhibit Hall A
Victoria Reyes - Madison Area Technical College
Co-Author(s): Tom Tubon, Madison Area Technical College; Bill Murphy, University, Wisconsin, Madison; Nhi Le, University, Wisconsin, Madison; Leona Liu; University, Wisconsin, Madison.
Human mesenchymal stromal cells (hMSCs) can be isolated preferably from bone marrow. HMSCs are incredibly useful in translational research due to their properties of self-renewal and their ability to differentiate into multiple cell lineages, including; bone, skin, cartilage, and muscle cells. The regenerative properties of hMSCs allow for the development of cell-based therapies that are specific to the patient’s body and immune system. The advancement of cell-based technologies is incredibly important and may one day solve the issue of low supply and high demand of organs needed for transplantation. A recent survey by the Health Resources and Services Administration indicates that 113,000 men, women and children on the national transplant waiting list as of January 2019.
Our goal in this research study is to identify the optimal conditions for hMSC growth under defined conditions to help inform fidelity and reproducibility as the process is scaled up for manufacturing. This study investigates the use of bioprinted hydrogel arrays to assay the effect of varying concentrations of vascular endothelial growth factor binding peptide (VBP), and varying substrate stiffness (1 kPa and 8 kPa) on the growth of cells. In addition, cells were cultured for 24 hours in serum-free and serum-containing media under normoxic (20% oxygen) and hypoxic conditions (1%) within a specialized CO2 chamber. Exposing cells to hypoxic conditions and varying levels of sequestering factors are critical parameters to consider because the levels of oxygen and nutrients that cells and tissues encounter when transplanted are extremely low. Previous studies have shown that pre-exposing hMSCs to vascular endothelial growth factor (VEGF) and hypoxia leads to higher proliferation and increased retention as compared to cells that are not pre-exposed. While our observations show no significant increase in cell number percentage with increasing VBP concentrations, we did see a large increase in cell expansion with increasing stiffness, regardless of whether the cells were cultured under hypoxic or normoxic conditions. Our data indicates that this is likely due to mechanical stimulation of specific growth factor signaling, involving pathways such as ERK, Akt and ROCK. These pathways may also trigger essential factors that are able to counteract apoptosis. To help us identify what specific pathways are triggered with increasing stiffness, we plan on inhibiting cell signaling pathways involved in mechanosening and cytoskeleton activity, such as the ROCK/Rho signaling pathway. Additionally, we plan on culturing the cells for longer than 24 hours to determine how long-term effects can impact the proliferation and health of hMSCs. Finally, in future experiments, we plan on using a more accurate way to measure cell count, instead of measuring by cell percentage.
References: Organdonor.gov. (2018). Organ Donation Statistics | Organ Donor. [online] Available at: https://www.organdonor.gov/statistics-stories/statistics.html.
Funder Acknowledgement(s): This material is based upon work supported by the National Science Foundation under Grant No. EEC-1648035. Special thanks to Tom Tubon, Leona Liu, Nhi Le, Bill Murphy, and the Murphy lab team at UW Madison.
Faculty Advisor: Tom Tubon, email@example.com
Role: I assisted in the making of the hydrogels, and data analysis.