Discipline: Biological Sciences
Subcategory: Biomedical Engineering
Room: Exhibit Hall
Stephany R. Maldonado - University of Arizona
Bones have the ability to self-repair and will respond to increased mechanical loads with enhanced osteoblast activity to make new bone. Attached strain gauges are the best way to directly measure bone loading by measuring the amount of bone deformation, but standard adhesives degrade rapidly when implanted, limiting the in vivo utility. Our lab has previously used different types of Calcium Phosphate Ceramic (CPC) particles to facilitate attachment of strain gauges to bone in vivo to collect long-term measurements. Although the CPC particles we have originally used for this purpose are no longer commercially available, our lab has been exploring the effect of different CPC particles on adipose derived mesenchymal stem cells (MSCs). The purpose of this experiment is to test 6 different CPCs to compare cell proliferation and osteoblastic differentiation of MSCs. Results will be compared with CPC particles that are no longer available. Our goal is to determine the CPC type that provides the highest differentiation rate in order to induce rapid bone attachment to CPC coated stain gauges in vivo.The CPC particles used are labeled as CPC 2, 4, 5, 6, 7 and 9 from Matexcel and CPC 6 from an older batch of particles, which will be called Old CPC in this experiment. Eight 48-well plates were coated with 17 uL of Master Bond EP42HT-2 Epoxy and different types of CPC particles, which were compressed and set to dry. After 24 hours the well plates were inverted to remove excess CPC. Seven well plates with each CPC coating were prepared. In addition, wells with only an epoxy coating and uncoated wells were used as controls, having a total of 9 well plates. At weeks 2, 4 and 6, eight wells were used to measure cell proliferation with an XTT assay and the medium from the wells was used to measure Alkaline Phosphatase (ALP) activity. Additional well plates with the CPCs and controls were made for imaging. Three wells were imaged with Scanning Electron Microscopy (SEM), and 3 wells were stained for collagen with Direct Red 80. Well plates were sterilized using ethylene oxide and seeded with 2,400 MSCs in 500 uL of media, which was changed 3 times a week. The epoxy, CPC coated, and uncoated well plates demonstrated successful cell attachment. Direct Red 80 staining for week 2, CPC 7 demonstrated higher quantities of collagen compared to the other samples. ALP assay data shows an overall increase in ALP activity in all new CPC wells, starting with an average of 1.5±0.1 U/mL in week 2 compared to 1.7±0.8 U/mL from the uncoated wells. The CPC ALP average activity was 2.5±0.1 U/mL in week 6 and 2.7±0.8 U/mL in the uncoated wells. This indicates that CPC particles do not interfere with MSC differentiation and are nontoxic.
Funder Acknowledgement(s): This study was supported, in part, by a grant from LSAMP awarded to David S. Margolis, MD PhD.
Faculty Advisor: David S. Margolis, email@example.com
Role: I coated the well plates, seeded the cells into the plates and fed them, performed the assays and the staining. I also analyzed my data.