Melt Electrowriting with Nanodiamond Integration for Biomedical Advancements

Chapter 1 reviewed skin tissue engineering scaffolds and discussed the potential of the MEW technique for skin tissue engineering.
In Chapter 2, biocompatible fluorescent nanodiamonds (FNDs) were introduced into polycaprolactone (PCL) – the golden standard material in melt electrowriting (MEW), to fabricate scaffolds for biomedical applications with improved mechanical properties and the possibility of their real-time degradation tracking. In this study, compared to pure PCL scaffolds, the functionalized ones containing 0.001 wt.% of 70 nm-diameter nanodiamonds (PCL-FNDs) showed increased tensile moduli (1.25 fold) and improved cell proliferation during 7-day cell cultures (2.00 fold increase). Furthermore, the addition of FNDs slowed down the hydrolytic degradation process of the scaffolds, accelerated for the study by the addition of the enzyme lipase to deionized water. Additionally, due to the nitrogen vacancy (NV) centres present on the FNDs, their amount and location were able to be tracked in real-time in printed fibres using confocal microscopy. This research shows the possibility for high-resolution life-tracking of MEW PCL scaffolds’ degradation.
In Chapter 3, multifunctional wound dressings with biologically active agents to prevent infections and promote healing, while also having cell delivery capability were fabricated. The poly-lactic acid /nanodiamonds (PLA/ND) scaffolds were first printed using melt electrowriting (MEW) and then coated with quaternized β-chitin (QβC). The NDs were well-dispersed in the printed filaments and worked as fillers and bioactive additions to PLA material. Additionally, they improved coating effectiveness due to the interaction between the negative charges (from NDs) and positive charges (from QβC). The obtained results showed that NDs addition not only increased the thermal stability of PLA but also benefitted cellular behaviour and inhibited the growth of bacteria. Scaffolds coated with QβC increased the effect of bacteria growth inhibition and facilitated the proliferation of human dermal fibroblasts. Additionally, we have observed extracellular matrix (ECM) remodeling on QβC-coated PLA/NDs scaffolds. The scaffolds provided support for cell adhesion and could serve as a valuable tool for delivering cells to chronic wound sites. The proposed PLA/ND scaffold coated with QβC holds great potential for achieving fast healing in various types of wounds.
Engineered skin models are promising platforms to study skin disease or assess cosmetics and clinical skin care products, with high adequacy and without ethical concerns. Three-dimensional models allow cell-cell and cell-matrix interactions which mimic human tissues more closely compared to two-dimensional cell culture systems and animal models. Besides high reproducibility and biocompatibility, relevant tissue models require proper mechanical and structural characteristics. In Chapter 4, 3D skin models composed of cell-laden methacrylated gelatin hydrogel (GelMA) and porous polycaprolactone (PCL) melt electrowritten (MEW) scaffolds were proposed. GelMA was used to provide nutrients and a hydrated environment for embedded fibroblasts and keratinocytes. The MEW scaffolds were composed of two layers: random fibers for culturing the keratinocytes to fabricate the epidermis with waved architecture; and well-aligned shapes filled with cell-laden GelMA to mimic the dermis. Three dermal designs differed in porosities and pore sizes were compared to optimize the dermis reconstruction. Within one week, the design with bigger pore sizes achieved optimal cell distribution, penetration, and ECM deposition. The SEs can be utilized for toxicity testing, or exploration of fundamental biological processes of skin tissue.
The application of our skin model was further studied in Chapter 5, we combined the melt-electrowritten PCL scaffolds and cell-laden Matrigel to fabricate skin equivalents (FTSEs). Besides the confirmation of the close resemblance of the mechanical and biological properties, we used the developed SE to build a testing system, and the damage caused by UVA irradiation and antioxidant efficacy were evaluated. The effectiveness of Tea polyphenols (TPs) and L-ascorbic acid (Laa) is compared based on free radical generation. TPs were demonstrated to be more effective in downregulating free radical generation. Further, T1 relaxometry wass used to detect the generation of free radicals at a single-cell level, which allowed tracking of the same cell before and after UVA treatment.
To conclude, this work explored the applicability of FNDs and NDs as fillers to modify polymer properties without compromising printing resolution. Polymer scaffolds were combined with hydrogel matrix to fabricate artificial skin tissue. Studied modifications enhanced the biological properties of polymer scaffolds, which can benefit various applications. Additionally, the proposed skin models broadened the application of polymer scaffolds as platforms for in vitro testing, offering significant potential as alternatives to in vivo models.