Matrigel and Its Applications

1. Matrigel and Its Applications Olga Filippova, Akash Patel

2. Proposal Temperature Control System for Solid Freeform Fabrication of Cell-Laden Matrigel Constructs • Temperature Control System • Cool Matrigel Reservoir • Heat substrate • Material Delivery System • Motion System

3. Liver Introduction • Lobule = Functional Unit • Hepatocytes • 70 – 80% of liver mass • Sinusoidal plates • Major liver functions • Drug metabolism • ECM • Collagens • I, III, IV, V, XVIII • Glycoproteins • Laminin, Fibronectin www.ece.ncsu.edu/imaging/MedImg/SIMS/GE1_3.html

4. Matrigel Gelain et al • Natural hydrogel • Basement membrane from EHS mouse sarcoma • Advantages • Composition • Laminin • Collagen IV • Thermal Crosslinking • 4oC (liquid)  37oC (gel) • Outstanding biocompatibility • Promotes differentiation

5. Matrigel (cont) • Disadvantages • Uncontrolled composition • Poor gelation kinetics • Little cell proliferation • Lower mechanical integrity • Young’s Modulus 400 Pa – highly compliable • Shear Modulus 180 Pa – below natural liver tissue • Thermal Crosslinking • Reverse characteristics – liquid at low temperatures, solid at physiological temperatures

6. Construct Characterization • Visualization • Proper sinusoidal shape • Proper width • Gelation • Time • Mass/Volume loss • Mechanical Testing • Compression

7. Cell Characterization • Cell Viability • Live/Dead Assay – membrane damage • Cell Proliferation • Alamar Blue Assay – growth environment analysis • Cell-Specific Function • Quanti Chrome Assay – urea production Huuskonen et al http://www.ab-direct.com/about/alamarblue-483.html

8. Great Promise for 3D Computer-Aided Deposition of Cell-Laden Hydrogel Structure • Especially useful in regenerative medicine • May help solve the problem of limited donor grafts • Scaffold optimization with tailored biological properties • Provide: • Supportive environment for cell attachment, proliferation, and differentiation • Biological cues to elicit desired cellular response • ECM composition, growth factors

9. 5 hrs Day 1 Day 2 viable cells (% of total) Day 3 Matrigel Lutrol E127 25% Alginate 2% Agarose 1% Bone Tissue Printing • 3D printed cell laden spatially organized hydrogel scaffolds • Demonstrate ability to print two distinct cell populations at predefined locations • Osteogenic and endothelial progenitors • Potentially use 3D printing to develop vascularized bone grafts for TE • * Air humidity during printing process may influence survival of extruded cells

10. Bone Tissue Printing

11. Drop-on-Demand Printing of Cell and Materials for Designer Tissue Constructs • Simultaneous printing of cells and biomaterials allows precise placement of cells and proteins within 3D hydrogel structures • Created contractile cardiac hybrids have properties that can be tailored in 3D to achieve desired porosities, mechanical and chemical properties • Results suggest that the printing method could be used for hierarchical design of functional cardiac patches, balanced with porosity for mass transport and structural support

12. Tissue Engineering Replacement Skin • Use of synthetic degradable gels is emerging • Matrigel • Primarily as a way to deliver cells and/or molecules • In situ • Smart matrices • Highly specific 3D architecture for skin grafts

13. Cancer Research Models • 3D cell cultures have much greater potential than 2D cultures • Using 3D cultures, scientists discover patterns of gene expression and other biological activities that more closely mirror what happens in organisms • 3D ECM models are especially useful for monitoring mechanisms of tumor growth and metastasis

14. Other Demands for Microscale Accurate, Precise Matrigel Printing “3-D microwell culture of human embryonic stem cells” • Constructed a 3-D microwell system for long-term hESC culture • Physical and extracellular matrix patterning constraints limit colony growth • Matrigel used to culture cells

15. Other Uses for Matrigel Printing Device Growth of miniature pig parotid cells on biomaterials in vitro • Coating the surface of synthetic materials aids cell growth and maintenance of a morphology that more closely resembles normal epithelium Compatibility of human fetal neural stem cells with hydrogel biomaterials in vitro • Stroke and spinal cord or brain injury often result in cavity formation • Stem cell transplantation in combination with tissue engineering has the potential to fill cavity and replace lost neurons • Hydrogels (Matrigel) may enhance migration capability across the injured cavity in vivo due to their ability to provide a three-dimensional matrix scaffold suitable for cellular adhesion, migration and support

16. Other Uses for Matrigel Printing Device (cont) Craniofacial muscle engineering using a 3-dimensional phosphate glass fibre construct • A 3-dimensional mesh arrangement of the glass fibres was the best at encouraging cell attachment and proliferation • In addition, increasing the density of the seeded cells and using Matrigel and insulin-like growth factor I enhanced the formation of prototypic muscle fibers Beyond the Cells: Scaffold Matrix Character Affects the In Vivo Performance of Purified Adipocyte Fat Grafts • Focused on the effects of fat graft structure on in vivo performance • Adult adipocyte mixed with Matrigel were implanted as fat grafts • Purified Matrigel grafts showed statistically greater longevity and volume maintenance versus all other groups