The development of a 3D printed PEG-Chitosan-PCL dressing loaded with peptide-microRNA nanoparticles for wound healing

Abstract

Wound healing is a fundamental and dynamic process in the human body. Without prompt and adequate wound healing, the health of the patient is affected, leading to life-threatening consequences. Herein, microRNAs (miRs) were chosen as the genetic therapy by regulating multiple genes and associated pathways to promote cell proliferation and migration. The cell-penetrating peptide RALA was used for encapsulating plasmid encoding miR-31 and miR-132 (pmiR-31 and pmiR-132) to form self-assembled nanoparticles (NPs) with an average size ~80 nm and positive zeta-potential ~20 mV facilitating cellular entry. The miR expression level was increased in NCTC-929 cells transfected with heated and fresh RALA/pmiR NPs, indicating high temperature (<100°C) has no degradation effect on miR upregulation. The heating process is conducted to mimic the conditions of hot melt extrusion (HME) for fabricating polymer-based filaments, followed by Arburg Plastic Freeforming (APF) three-Dimensional Printing (3DP) process to produce 3D porous patches. Moreover, heated NPs effectively regulated target gene expression via RT-PCR and ELISA. PCL/PEG/chitosan-based 3D printed wound patches were fabricated with ideal physiochemical properties for wound healing via HME and 3DP, optimised through a Design of Experiments approach. Lyophilised peptide-DNA nanoparticles were successfully loaded into optimal PCL/PEG/chitosan filaments and the nanoparticles remained intact. The delivery of RALA/pmiR NPs from biocompatible 3D-printed patches to NCTC-929 and HacaT cells significantly improved cell migration, resulting in 50% wound reduction without hindering cell attachment, potentially preventing secondary injury upon patch removal. In the C57BL/6N murine full-thickness wound-healing model, NP-loaded 3D printed patches outperformed a commercial dressing, showing accelerated wound closure rate (80%), increased epidermal thickness (~80 µm) and blood vessel count (~2 fold) after 7 days. Taken together, these results demonstrated that peptide-DNA can be loaded in PCL/PEG/chitosan-based 3D printed patches without compromising the integrity of the biological cargo to result in improved wound closure and tissue repair.

Thesis is embargoed until 31 December 2029.

Date of Award	Dec 2024
Original language	English
Awarding Institution	Queen's University Belfast
Supervisor	Helen McCarthy (Supervisor), Peter Boyd (Supervisor) & Nicholas Dunne (Supervisor)