Scaffold Degradation Part II Effect of Ionic Strength on Scaffold Biodegradability

1. Scaffold Degradation Part IIEffect of Ionic Strength on Scaffold Biodegradability Jay Sehgal North Allegheny Intermediate High School

2. Scaffold DegradationBackground Information • Scaffold - temporary framework used to support a structure • In tissue engineering, used to temporarily support the growth of tissue

3. What is Tissue Engineering/ Regenerative Medicine? Replacing diseased or injured tissues with tissue constructs designed and fabricated for the specific needs of each individual patient.

4. An Ultimate Vision for Regenerative Medicine: Complete Tissue Regeneration Spinal Cord Upper and Lower Jaw Tail Heart Limb

5. Tissue structure and function may be compromised by: • Inherent design flaws • Hereditary defects or conditions • Disease • Trauma • Environmental influences • Aging

6. Potential Solutions • Surgical or physical manipulation • Drug therapy • Diet/lifestyle changes • Transplants • Artificial tissues/organs • Gene Therapy • Tissue Engineering/Regenerative Medicine

7. Matrices Cells Healing Signaling Molecules Normal Wound Repair

8. The Basic Three R’s of Tissue Engineering Right Hormones Right ECM Right Cells

9. Guided Tissue Repair If needed, harvest cells from patient. Hormones Cells Implant Matrix Culture

10. Scaffold DegradationIntroduction • Scaffold degradation rate is important to tissue engineering. • If the rate is not studied and regulated, scaffold may interfere with the successful replacement of biological functions.

11. Scaffold DegradationIntroduction • The rate of degradation of scaffolding material can be affected by factors such as: • Temperature • pH • Size • Weight • Degree of porosity • Physical stresses

12. Scaffold DegradationSynopsis of Part I • The purpose of Scaffold Degradation Part I was to determine whether or not the scaffold’s biodegradation would be affected if it’s environment was acidic or basic. • Hypothesis: If the pH of the scaffold’s environment is acidic or basic, then the biodegradation process will be affected. • Polycaprolactone (PCL) was incubated for 14 days in acidic, neutral, and basic environments. The degradation was monitored. • The data concluded that the more acidic the environment was, the faster the PCL biodegraded. The data supported the hypothesis.

13. Scaffold DegradationScaffolding Materials • Polylactic acid (PLA): • Biodegradable polyester • Sutures • Drug delivery device • Stents • Polyglycolic acid (PGA): • Biodegradable polyester • Sutures • Drug delivery device

14. Scaffold DegradationPCL • Polycaprolactone (PCL) – biodegradable polyester • Created by polymerization of caprolactone using a catalyst and heat • Sutures • Drug delivery device • Used in this experiment as the scaffolding material

15. Scaffold DegradationIonic Strength (NaCl) • Ionic strength is a function of all ions present in a solution • NaCl’s ionic strength is equal to it’s concentration • Molar Mass (NaCl)≈58.5g/mol • 58.5g (NaCl) + 1.0L H20 = 1M solution of NaCl • 1M solution can be diluted to obtained levels of high or low ionic strength

16. Scaffold DegradationPurpose • Ionic strength of NaCl in the body’s natural cellular matrix doesn’t vary much from zero • Knowing the affects of various ionic strength’s on a scaffold’s degradation is very helpful in designing the scaffold in vitro • Using this knowledge, the rate of degradation can be regulated.

17. Scaffold DegradationHypothesis • Null Hypothesis • If biodegradable scaffolds will be submerged in water with varying ionic strength, then the degradation will not be affected.

18. Scaffold DegradationMaterials • Polycaprolactone (PCL): 3.2g • NaCl • Tetrahydrofuran (THF): 32mL • Teflon-coated muffin tins • Stirrer plates and stirrer bars • 15mL test tubes • Large flasks and beakers • Distilled water • Incubator • Fume hood • Drying oven • Electronic metric scale • Volumetric flasks

19. Scaffold DegradationCreate Scaffolds Procedure • Day 1 • 32mL THF added to 3.2g PCL in large flask • Flask stopped and placed under fume hood on stirrer overnight • Day 2 • 8mL solution into Teflon-coated muffin tin mold • 7.2g NaCl stirred in until slurry formed. • Repeated for 3 more molds • Molds placed under fume hood to allow THF to evaporate overnight

20. PCL being dissolved by THF on stirrer

21. Scaffold DegradationCreate Scaffolds Procedure • Day 3 • Scaffolds removed from mold and flipped • Allowed THF to evaporate overnight • Day 4 • Scaffolds placed in large beaker of H20 to leech out NaCl • Leeched for 2 days • Water changed 2x daily

22. Scaffold before degradation

23. Scaffold DegradationCreate Scaffolds Procedure • Day 6 • Scaffolds removed from water and towel dried • Each scaffold cut into 8 pieces • Pieces were placed in drying oven overnight

24. Scaffolds in drying oven

25. Scaffold DegradationDegradation Procedure • 3 sets of tubes: • Set A—100mM (10 tubes) • Set B—10mM (10 tubes) • Set C—DW Control (10 tubes) • 1 Molar stock solution • 100mL DW • 5.85g NaCl • 100mM solution • 90mL DW • 10mL 1M stock • 10mM solution • 90mL DW • 10mL 100mM solution

26. Scaffold DegradationDegradation Procedure • Tubes filled with appropriate solution • Dried scaffolds weighed and data recorded • Scaffolds placed in all 30 tubes • Tubes capped and incubated at 37° C (body temp) for 3 days • Scaffolds removed, washed, dried, and weighed

27. Test tubes ready for incubation

28. Scaffold DegradationData Analysis

29. Scaffold DegradationData Analysis

30. Scaffold DegradationConclusion • Data does not support hypothesis • Results from 10mM PCL and 100mM PCL show significantly more biodegradation than distilled water control • PCL biodegrades faster in environments with ionic strength’s of 10mM and 100mM

31. Scaffold DegradationSources of Error • Degradation time too short to determine full effects of ionic strength • NaCl not fully leeched out, remained part of mass • Concentration of molar stock solutions not perfectly accurate

32. Scaffold DegradationFuture Research • Test degradation rate at different ionic strength levels • Test degradation rate and bio-compatibility of scaffolds with varying factors using live cells • Test strength of scaffolds with varying factors • Investigate degradation with varying degrees of porosity