Doctor of Philosophy
Intelligent Polymer Research Institute
Skin provides the protective surface for animals and humans and is therefore prone to physical, chemical, and biological injuries. In all but superficial wounds, the capacity to repair by regeneration is lost and the mechanisms involved in wound closure are unable to restore the skin’s original functions. In this context, skin repair is achieved using surgical techniques including skin grafts, and a range of synthetic or biological scaffolds. Wounds impact millions of patients every year and represent a serious cause of morbidity and mortality worldwide. The increase in need for better skin repair, in part due to issues such as the aging population coupled with chronic conditions has driven the development of products to enhance therapeutic outcomes, yet current treatment outcomes are far from ideal and complete replication of the cellular structure and tissue functional requirements of skin remains a challenge.
General aims: Address the major drawbacks of available skin substitutes and delivery system platforms. Herein two approaches are proposed, the first one (Chapters 2-3) involves the development and initial in vitro characterization of a 3D multifunctional bioprinted platform based on platelet lysate, which could be used to deliver cells and growth factors to the wound site while providing a supportive network that mimics the native ECM for skin cells to infiltrate and thrive. This system was designed with the aim of providing an advanced alternative to current skin grafts and skin substitutes available for clinical use. The second system proposed (Chapter 4) is based on an electrofluidic approach for control of bioactive molecule delivery into soft tissue model using threads and surgical sutures which was designed with the aim of being used in sutures for surgical wound closure.
Methods: (Chapter 2-3) 3D printed HDF-PLGMA bioink were fabricated using a pneumatic extrusion-based 3D Bioplotter. The epidermal-dermal model was fabricated by seeding HaCaT on top of 3D printed HDF-PLGMA constructs. The innervated dermal model was fabricated by seed hNSC H9 neurospheres to the bottom of 3D printed HDF-PLGMA constructs. (Chapter 4) Commonly employed surgical sutures were used to create an adequate fluid connection between the electrodes and a tissue-like 3D hydrogel support. The platform consisted of two reservoirs into which the ends of the thread/suture were immersed. The anode and cathode were placed separately into each reservoir. The thread/suture was taken from one reservoir to the other through the gel. When the current was applied, biomolecules loaded onto the thread/suture were directed into the gel, and the rate of movement of the biomolecules was dependent on the magnitude of the current.
Results: (Chapter 2) Briefly, the work described in this chapter relates to the development of a multifunctional bioink consisting of PL and GelMA (PLGMA) and the biofabrication of a 3D printed dermal-like structure. The data presented shows that the proposed PLGMA bioink meets essential requirements of printability in terms of rheological properties and shape fidelity. Moreover, its mechanical properties can be readily tuned to achieve stiffness that is equivalent to native skin tissue. Biologically relevant factors were successfully released in a sustainable manner and the bioavailability of those factors was demonstrated by high cell viability, good cell attachment, and improved proliferation of printed dermal fibroblasts, as well as by upregulation of ECM synthesis by dermal fibroblasts. (Chapter 3) Continuing the work described in chapter 2, chapter 3 relates the fabrication of a more complex skin equivalent based on the PLGMA platform previously established. Bilayer skin model: The expression of general keratinocyte differentiation markers was used to confirm the capacity of the platform to promote normal epithelial morphogenesis and differentiation of keratinocytes. Innervated skin model: The expression of development neuronal markers, as well as a general neuronal marker, was used to demonstrate that the proposed platform supported hNSC-H9 neurosphere neurite outgrowth and neuronal differentiation. Challenges faced in the co-incorporation of HaCaT and hNSC-H9 neurospheres to HDF-PLGMA construct that need to be considered to progress with this research will also be presented. (Chapter 4) A novel electrofludic system for the controlled release of bioactive molecules such as small molecules, drugs, polysaccharides, or proteins via an electric field is described for the first time. In a proof-of-concept study, the controlled delivery of dexamethasone 21-phosphate disodium salt (DSP), a clinically relevant anti-inflammatory prodrug, as well as other molecules were used to demonstrate the feasibility of the proposed system. Despite being peripherally related to the overall PhD theme; this system could be potentially integrated in the PLGMA matrix by printing the 3D skin cells-PLGMA on top of the suture for precise delivery of growth factors and cells to sutured wounds.
Taken together, although it is still at an early stage of development, the bioprinting platform, as well as the electrofludic systems demonstrated to hold potential as a foundation for the fabrication of complex and physiologically relevant delivery platforms for wound management and tissue regeneration.
Daikuara, Luciana Yumiko, Fabricating delivery platforms for wound management and tissue regeneration, Doctor of Philosophy thesis, Intelligent Polymer Research Institute, University of Wollongong, 2021. https://ro.uow.edu.au/theses1/1397
FoR codes (2008)
0903 BIOMEDICAL ENGINEERING, 100404 Regenerative Medicine (incl. Stem Cells and Tissue Engineering)
Unless otherwise indicated, the views expressed in this thesis are those of the author and do not necessarily represent the views of the University of Wollongong.