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Influence of 2D and 3D Environments on Osteogeneic Differentiation in hMSCs - PowerPoint PPT Presentation


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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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slide1

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

slide2

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

slide3

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?

slide4

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
slide5

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?
slide6

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
slide7

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

slide8

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

slide9

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