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Richelle C. Thomas Department of Chemical Engineering University of Texas at Austin April 20, 2011. Data Blitz. Motivation: Peripheral Neuropathy. Soft tissue scaffold Focus on injury Supports wound healing Reduces inflammation Promotes tissue reorganization. Motivation.
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Richelle C. Thomas Department of Chemical Engineering University of Texas at Austin April 20, 2011 Data Blitz
Motivation: Peripheral Neuropathy • Soft tissue scaffold • Focus on injury • Supports wound healing • Reduces inflammation • Promotes tissue reorganization
Motivation • Statement: 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
Goal Develop natural polymer hydrogel with 3D internal architecture Three-Dimensional Hydrogel with Internal Architecture Colloidal Crystal Amorphous Hydrogel
Specific Aim 1 Specific Aim 2 Specific Aim 3 Synthesize and characterize photocrosslinkable three-dimensionally patterned natural polymer Hyaluronic Acid hydrogels Optimize colloidal crystal templating protocol for three-dimensional construct. Evaluate degradation profile, degree of swelling and void area of templated gels relative to virgin scaffolds. Assess viscoelastic differences between amorphous and templated gels. Acquire confocal images of poragen structure within bulk hydrogel.
HA-Urea Film 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
HA-Urea Hydrogel Porous Structure Pore size 50-100 μ m FITC labeled, Scale bar: 150 μm Scale Bar: 250 μm
HA-Urea Hydrogel Porous Structure FITC labeled, scale bar = 200 μm
Specific Aim 1 Specific Aim 2 Specific Aim 3 • Cell studies • - Peripheral nerve regeneration (urea) • Vascularized systems (less dense, highly branched) • KH2PO4 • Guanidine • Glycine • CHAPS
Guanidine TRITC labeled, 4X Magnification
KH2PO4 FITC labeled, 4X Magnification, 3D Projection
KH2PO4 * Scale bar 200 um, (* 150 um)
Cytotoxicity Astrocytes 50 mg/ml GMHA 4X Control 10 X 4X Experiment 10 X Redo with sterile HA, DAPI stain
Thiol films • Form covalent bonds chemically • Form thiol-acrylate (ester) bonds • Hydrolyzable at physiological pH • Acrylate:thiol mixed in 7:3 ratio by mass • Rydholm, A.E., Anseth, K.S. and Bowman, C.N. “Effects of neighboring sulfides and pH on ester hydrolysis in thiol–acrylate photopolymers” ActaBiomaterialia, Volume 3, Issue 4, (2007) Pages 449-455
Thiol-Capsaicin Films 30 mg/ml GMHA, 10 ug/ml Thiol 7.2 mg/ml Capsaicin 80% MeOH 4.5mg/ml Capsaicin 80% MeOH 4.5 mg/ml Capsaicin 50% MeOH 2.78mg/ml Capsaicin 50% EtOH 4X 10X 20X
Thiol-Capsaicin Data How to identify/separate different phases within film? 30mg/ml GMHA | 2.78mg/ml Capsacin| 50% EtOH | 10ug/ml thiol Both images at 10X Magnification
Thiol-capsacindata Empty domains within film 30mg/ml GMHA | 2.78mg/ml Capsaicin | 50% EtOH | 10ug/ml thiol 4X 10X
Thiol-Capsaicin Data 30 mg/ml GMHA, 7.2 mg/ml Capsaicin, 80% MeOH, 10mg/ml thiol Ordered domains within film… 4X
Future Work • Peripheral Nerve Regeneration • Cytoxicity study • Evaluate cell neurite extension within hydrogel • Screen remaining vascular poragen candidates • Thiol films • Characterize • Evaluate ability of thiol-capsaicin films to provide sustained drug release over time