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Influence of 2D and 3D Environments on Osteogeneic Differentiation in hMSCs

Influence of 2D and 3D Environments on Osteogeneic Differentiation in hMSCs. Jacqueline Mimnaugh , RET Neuqua Valley High School Dr. Richard Gemeinhart Melanie Kollmer UIC Department of Biopharmaceutical Sciences Tracy Chuong , REU University of California Berkley.

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Influence of 2D and 3D Environments on Osteogeneic Differentiation in hMSCs

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  1. Influence of 2D and 3D Environments on Osteogeneic Differentiation in hMSCs Jacqueline Mimnaugh, RET Neuqua Valley High School Dr. Richard Gemeinhart Melanie Kollmer UIC Department of Biopharmaceutical Sciences Tracy Chuong, REU University of California Berkley

  2. What are hMSCs? Stem cells differentiate into other types of cells Two major categories: Embryonic (Pluripotent) Adult (Multipotent) Human Mesenchymal Stem Cells (hMSCs) Isolated from marrow Can differentiate into bone, cartilage, fat

  3. Why are hMSCs important? Induce hMSCs to develop into bone cells Osteogenesis Tissue Engineering • Bone diseases/defects, trauma, cancer, mal-union/non-union fractures How could we make new bone tissue for therapeutic uses?

  4. The Problem Cells in the lab are typically cultured on plates 2 D Cells in vivo (in an organism) 3 D It is possible that cells grown in a 3D scaffold would: • Be more like cells in vivo • Would not reach confluence as quickly • Could be implanted directly

  5. Research Project How will a 2D and 3D environment effect the osteogenic differentiation of hMSCs? Compare cells grown in a 2D and a 3D environment: • Viability? • Proteins? • Genes expressed? • Mineralization?

  6. Creating a 3d Scaffold Superporous Hydrogels • Poly (ethylene glycol) diacrylate or PEGDA • Polymer network, hydrophilic • Pores from 100 µm to 600 µm created by gel-foaming

  7. Project Overview 1. Seed hMSCs onto 2D plates and 3D hydrogels After 24 hours 2. Add osteogenic differentiation medium After 24 hours 3. Compare cells at day 2, 7, 14, 21 and 28

  8. Comparing the cells 1. MTS – Cell Viability 2. BCA – Protein Levels 3. Calcium 4. Alkaline Phosphatase – Early Marker 5. ELISA: Osteopontin – Early and Late Marker Osteocalcin– Late Marker 6. qPCR – Determine Gene Expression - ALPL, RUNX2, OC, OP, BMP2 7. Von Kossa Staining - Mineralization

  9. Accomplished Tasks Prep: Make gels, order supplies, review protocols 1. Seed hMSCs onto 2D plates and 3D hydrogels After 24 hours 2. Add osteogenic differentiation medium After 24 hours 3. Compare cells at day 2, 7, 14, 21 and 28

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