Characterization of Salt Templated Hyaluronic Acid Hydrogels for Nerve Regeneration

1. Richelle C. Thomas, Christine E. Schmidt University of Texas at Austin Characterization of Salt Templated Hyaluronic Acid Hydrogels for Nerve Regeneration

2. Motivation: Peripheral Neuropathy • Over 20 million Americans suffer from some form of peripheral neuropathy The Neuropathy Association <http://www.neuropathy.org/site/PageServer?pagename=About_Facts>

3. Motivation: Peripheral Neuropathy

4. Motivation: Diabetes • Global Prevalence • 60-70% of diabetics have mild to severe nervous system damage • Diabetic foot ~80% of non-traumatic amputations • Largest diabetic population: • India, China, US • Largest cause of PN in west • US Prevalence • 10.7% US aged 20 & older • 23.1% US aged 60 &older • Cost • US $174 B in 2007 National Institute of Diabetes and Digestive and Kidney Diseases. National Diabetes Statistics, 2007 fact sheet. Bethesda, MD: U.S. Department of Health and Human Services, National Institutes of Health, 2008.

5. Motivation: Peripheral Neuropathy • Easy to injure • Trauma • 1.5-2.8% incidence rate • War • 35% troops extremities wounded in combat Journal of Craniofacial Surgery. 21(4):998-1001, July 2010.

6. Nerve Repair Options End-to-End Suturing Bridging Grafting Conduit/Implant Communication between nerve stumps Physical guidance to regenerating axons Ciardelli G, Chiono V. 2006. Materials for Peripheral Nerve Regeneration. Macromol. Biosci. 6:13-26

7. Biomaterials for PN requires proper integration of stimuli Chemical Electrical Contact Huang et al. (2008) Lee et al. (2002) Gomez et al. (2006) For more information on stimuli integration see Forcinitiet al. ABME. (2008).

8. Motivation: Peripheral Neuropathy • Soft tissue scaffold • Focus on injury • Supports wound healing • Reduces inflammation • Promotes tissue reorganization

9. Properties of an Ideal Scaffold Hudson TW, Evans GR, Schmidt CE. 1999. Engineering strategies for peripheral nerve repair. Clin. Plast. Surg. 26:617–28

10. Properties of an Ideal Scaffold Naturally Derived Polymer Hydrogels Biocompatible Inflammatory Response  Mechanically sound Not collapse   Biodegradability Room for tissue growth No need for 2nd surgery PBS Swollen Degradation rate Comparable to new tissue formation  Artif Organs V 30 No 7 2006

11. Hyaluronic Acid • Extracellular matrix component • Polyanionic • Hydrophillic • Involved in mediating wound repair • Non-cell adhesive

12. Photocrosslinkable Hydrogels High Swelling HA Hydrogels Advantages Biocompatible Non-cell adhesive Control local chemical properties Optically transparent 20 min 1 min Modulate Swelling Degradation Mechanical Properties 2 min 5 min 10 min UV Exposure

13. Motivation Hyaluronic Acid gels beneficial for wound healing applications Problem: Hydrogels are amorphous and do not provide any significant physical contact guidance to infiltrating cells beyond their inherent porosity

14. 25 mm 12 mm Role of Physical vs. Chemical Cues Microchannels vs. Laminin Gomez, Chen, Schmidt (2007). J. R. Soc Interface. 4(13): 223-233. Physical cues are preferred over chemical cues for axon initiation (polarization) 70 %  Physical Cues (microchannels) 30 %  Chemical Cues (NGF or Laminin) Gomez, N., Schmidt, C.E. (2007) Biomaterials. 28 (2): 271-284

15. Combined Chemical & Physical Cues Hippocampal cells on PDMS Microchannels w/immobilized NGF (0.11 ng/mm2) Scale bars=10 μm Gomez, N., Schmidt, C.E. (2007) Biomaterials. 28 (2): 271-284 Neurons extended longer and more oriented axons on surfaces with combinatorial cues.

16. Motivation Hyaluronic Acid gels beneficial for wound healing applications Problem: Hydrogels are amorphous and do not provide any significant physical contact guidance to infiltrating cells beyond their inherent porosity

17. Goal Develop natural polymer hydrogels that have 3D internal architecture Extend internally patterned films into 3D hydrogel constructs

18. Methods 32% Methacrylated Hyaluronic Acid 50 mg/ml GMHA 0.1-1% Photoinitiator

19. Photocrosslinkable Hydrogels High Swelling HA Hydrogels Advantages Biocompatible Non-cell adhesive Control local chemical properties Optically transparent 20 min 1 min Modulate Swelling Degradation Mechanical Properties 2 min 5 min 10 min UV Exposure

20. Modulate Enzymatic Degradation

21. UV Exposure & Photoinitiator Concentration Effect on Swelling

22. Storage/Loss Modului over Frequency Range

23. 2D Porous Structure Scale bar: 10 lm. TRITC- labeled Green fluorescent albumin–FITC • connectivity among the pores of the scaffold • protein diffusion was restricted to the pores • tightly crosslinked, low-permeability hydrogel Zawko, S.A. , et al. (2010. “Crystal templating dendritic pore networks and fibrillar microstructure into hydrogels.” ActaBiomaterialia 6(7): 2415-2421

24. Conclusions • Degradation rate, swelling dependent on UV exposure & PI concentration • Storage moduli consistent over frequency range • Tunability of mechanical properties makes system for variety of applications • Drug delivery • Vascularization

25. Future Work • Peripheral Nerve Regeneration • Determine swollen pore size of 3D hydrogels with confocal microscopy • Render porous network cell-adhesive and evaluate cell mobility within hydrogel. • Other applications • Incorporate anti-inflammatory agents into hydrogel matrix to evaluate ability to provide sustained drug release over time

26. Acknowledgements • Dr. Christine E. Schmidt • Labmates • Leandro Forciniti, Sarah Mayes, John Hardy • Undergraduate Researchers • Shan Modi, Paul Chung

27. Questions?