Fabrication of an electrospun nanofibrous scaffold for use in the field of tissue engineering

1. Fabrication of an electrospunnanofibrous scaffold for use in the field of tissue engineering By: Shannon Daily & Tyler Crawford

2. Progress since last meeting • Met with Singaporeans! • Presented final proposal of collaboration to AOS staff • Finished Experimental Design • Turned in final and revised editions • Finished ISEF forms • Started updates to Background Research paper

3. Polycaprolactone (PCL) • Biocompatible polymer • Biodegradable at a slow enough rate to allow increased cell growth and stability • Proven to have potential for scaffolds in relation to tissue regeneration • Has created scaffolds w/ ideal conditions • High porosities • Large amounts of surface areas

4. Additional Polymer for AOS: • Chitosan • Advantages • natural polymer, biocompatible and biodegradable • Cellular binding capabilities • Accelerates wound healing • Anti-bacterial properties

5. Chitosan continued: • Chitosan: • Disadvantages • high viscosity limits spinnability • Fibers can swell in aqueous solution- need to be cross linked to maintain structural qualities

6. Additional Polymer for HCI: • Alginate • Advantages • Good for health reasons (low toxicity, immunogenic) • Low cost • Disadvantages • Poor spinnability (possibly be fixed with addition of a synthetic polymer)

7. Solvent • Universal solvent- can be used for PCL, CHT, & Alg • Acetic Acid • Previous research for PCL & CHT • Alginate? • Another possibility: formic acid/acetone • If acetic acid doesn’t work

8. Experimental Design Procedure: • Create control meshes of pure PCL • Mix solution • PCL & acetic acid • Electrospin • Starting parameters: 15 wt.% concentration, 20 cm from tip of syringe to collector plate, & 20 kV

9. Procedure continued: • Vary voltage to create 9 meshes • 3 Voltages- 3 trials for each • 20 kV • 15 kV • 25 kV • Examine pieces of meshes under scanning electron microscope (SEM) • Culture fibroblast cells onto other pieces of meshes

10. Procedure continued: • Observing cells • Examine under TC inverted light microscope • Analyze cell growth • Conduct cell counts in cells per unit area (mm2) • Means and standard deviations • ANOVA (Analysis of Variants) tests

11. Procedure continued: • Create solutions of PCL and chitosan • Electrospin • Vary concentration of chitosan to PCL • 5% CHT • 15% CHT • 25% CHT • Total of 9 meshes (3 trials of each concentration)

12. Procedure continued: • Analyze with Scanning Electron Microscope (SEM) • Culture fibroblast cells and seed into meshes created • Determine cell density • Analyze with means, standard deviations, and ANOVA tests

13. Data and Analysis: • Data obtained: • Fiber diameter and pore diameter of mesh • Cell density amounts • Analysis includes: • Means* • Standard Deviations* • ANOVA tests • 3 comparisons *5-7 measurements/areas for these methods

14. Comparisons • Pure PCL mesh vs. chitosan/PCL mesh • AOS & HCI • With pure PCL mesh: • AFM (atomic force microscope) vs. SEM • Fibroblast cells vs. Drosophila cells • pure PCL mesh vs. alginate/PCL mesh • Chitosan/PCL mesh vs. alginate/PCL mesh

15. Wikipage • Basic Timeline • Includes any work either side does during each month • Background Research Paper • Experimental Design drafts • Any PowerPoint presentations • References • Links to all relevant journal articles either have found

16. Timeline

17. To be worked on: • Finish last revision to ISEF forms • Finish the update on background research • Research more into alginate for potential universal solvent • Acquisition forms

18. Bibliography Akhyari, P., Kamiya, H., Haverich, A., Karck, M., & Lichtenberg, A. (2008). Myocardial tissue engineering: The extracellular matrix. European Journal of Cardio-Thoracic Surgery, 34, 229-241. doi: 10.1016/j.ejcts.2008.03.062 Bhardwaj, N. & Kundu, S. C. (2010). Electrospinning: A fascinating fiber fabrication technique. Biotechnology Advances, 28, 325-347. doi: 10.1016/j.biotechadv.2010.01.004 Chong, E.J., Phan, T.T., Lim, I.J., Zhang, Y.Z., Bay, B.H., Ramakrishna, S., & Lim, C.T. (2007). Evaluation of electrospun PCL/gelatin nanofibrous scaffold for wound healing and layered dermal reconstitution. ActaBiomaterialia, 3, 321-330. doi: 10.1016/j.actbio.2007.01.002 Geng, X., Kwon, O-H., & Jang, J. (2005). Electrospinning of chitosan dissolved in concentrated acetic acid solution. Biomaterials, 26, 5427-5432. Han, J., Branford-White, C.J., & Zhu, L.M. (2010). Preparation of poly(є-caprolactone)/poly(trimethylene carbonate) blend nanofibers by electrospinning. Carbohydrate Polymers, 79, 214-218. doi: 10.1016/j.carbpol.2009.07.052 Homayoni, H., Ravandi, S.A.H., & Valizadeh, M. (2009). Electrospinning of chitosannanofibers: Processing optimization. Carbohydrate Polymers, 77, 656-661. Lowery, J.L., Datta, N., & Rutledge, G.C. (2010). Effect of fiber diameter, pore size and seeding method on growth of human dermal fibroblasts in electrospun poly(є-caprolactone) fibrous mats. Biomaterials, 31, 491-504. doi: 10.1016/j.biomaterials.2009.09.072 Nisbet, D.R., Forsythe, J.S., Shen, W., Finkelstein, D.I., & Horne, M.K. (2009). A review of the cellular response on electrospun nanofibers for tissue engineering. Journal of Biomaterials Application, 24, 7-29. Pham, Q.P., Sharama, V., & Mikos, A.G. (2006). Electrospinning of polymeric nanofibers for tissue engineering applications: A review. Tissue Engineering, 12,1197-1211. Shevchenko, R.V., James, S.L., & James, S.E. (2010). A review of tissue-engineered skin bioconstructs available for skin reconstruction. Journal of the Royal Society Interface, 7, 229-258. doi: 10.1098/rsif.2009.0403 Sill, T.J., & von Recum, H.A. (2008). Electrospinning: Applications in drug delivery and tissue engineering. Biomaterials, 29, 1989-2006. doi: 10.1016/j.biomaterials.2008.01.011 Woodruff, M.A., & Hutmacher, D.W. (in press). The return of a forgotten polymer- Polycaprolactone in the 21st century. Progress in Polymer Science. doi: 10.1016/j.progpolymsci.2010.04.002