Engineering, science and medicine: transforming healthcare

The treatment of disease is being revolutionised by the increasing use and capabilities of intelligent medical devices. Currently, there is a translational drive from basic research into clinical realisation, with multidisciplinary teams responsible for this synergistic effort. This work is forming the era of biomedical engineering, which is manifested in many aspects of medicine at both the bench and the bedside. Here, we focus on two different categories: imaging and medical robotics; and, tissue engineering. We introduce the underlying tenets on which these fields are built before discussing current research and the possibilities for tomorrow’s practice.</p

Therapeutics on the clock: circadian medicine in the treatment of chronic inflammatory diseases

The circadian clock is a collection of endogenous oscillators with a periodicity of ~ 24 h. Recently, our understanding of circadian rhythms and their regulation at genomic and physiologic scales has grown significantly. Knowledge of the circadian influence on biological processes has provided new possibilities for novel pharmacological strategies. Directly targeting the biological clock or its downstream targets, and/or using timing as a variable in drug therapy are now important pharmacological considerations. The circadian machinery mediates many aspects of the inflammatory response and, reciprocally, an inflammatory environment can disrupt circadian rhythms. Therefore, intense interest exists in leveraging circadian biology as a means to treat chronic inflammatory diseases such as sepsis, asthma, rheumatoid arthritis, osteoarthritis, and cardiovascular disease, which all display some type of circadian signature. The purpose of this review is to evaluate the crosstalk between circadian rhythms, inflammatory diseases, and their pharmacological treatment. Evidence suggests that carefully rationalized application of chronotherapy strategies – alone or in combination with small molecule modulators of circadian clock components – can improve efficacy and reduce toxicity, thus warranting further investigation and use

The development of tissue engineering scaffolds using matrix from iPS-reprogrammed fibroblasts.

Tissue engineering solutions have been widely explored for enhanced healing of skin wounds. Diabetic foot ulcers (DFU) are particularly challenging wounds to heal for a variety of reasons, including aberrant ECM, dysregulation of vascularization, and persistent inflammation. Tissue engineering approaches, such as porous collagen-based scaffolds, have shown promise in replacing the current treatments of surgical debridement and topical treatments. Collagen-glycosaminoglycan scaffolds, which are FDA approved for diabetic foot ulcers, can benefit from further functionalization by incorporation of additional signaling factors or extracellular matrix molecules. One option for this is to incorporate matrix from a rejuvenated cell source, as wounds in younger patients heal more quickly. Induced pluripotent stem cells (iPS) are generated from somatic cells and share many functional similarities with embryonic stem cells (ES), while avoiding the ethical concerns. Fibroblasts differentiated from iPS cells have been shown to enrich their ECM with glycosaminoglycan (GAGs), collagen Type III and fibronectin, to have an increased ECM production, and to be pro-angiogenic. Here we describe a technique to grow matrix from post-iPS fibroblasts, and to develop a scaffold from this matrix, in combination with collagen, with the goal of enhancing wound healing. By activating scaffolds with extracellular matrix (ECM) from fibroblasts derived from an iPS source (post-iPSF), the scaffolds are enriched with beneficial elements like GAGs, collagen type III, fibronectin, and VEGF. We believe these scaffolds can enhance skin regeneration and that the techniques can be modified for other tissue engineering applications

Platelet-derived growth factor stabilises vascularisation in collagen-glycosaminoglycan scaffolds in vitro.

Collagen-glycosaminoglycan (CG) scaffolds have been widely developed for a range of regenerative medicine applications. To enhance their efficacy, CG scaffolds have previously been prevascularised in vitro using human umbilical vein endothelial cells and human mesenchymal stromal cells (hMSCs); however, at later timepoints, a regression of vascularisation is observed. This is undesirable for longer preculture periods (e.g., for partial/full organ regeneration) and for in vitro vascularised tissue model systems (e.g., for drug testing/modelling). We hypothesised that delayed platelet-derived growth factor (PDGF)-BB addition could stabilise vessels, preventing their regression. In 2D, we identified 25 ng/ml as a suitable dose that enhanced hMSC metabolic activity and proliferation, without affecting endothelial cells, or migration in either cell type. In our 3D model of CG scaffold vascularisation, early addition of PDGF (Day 3) behaved similarly to no PDGF controls. However, PDGF addition at later timepoints (i.e., Days 4 and 5), with a second addition on Day 10, prevented vascular regression. In quantifying our observations, we identified a need for a tool to measure in vitro vascularisation in porous scaffolds. This was a second key objective of this work. A novel ImageJ macro was developed, which allowed us to analyse vessel-like structures, evaluating their number and morphology, and confirmed our qualitative observations. Finally, upregulation of angiogenic genes (ANG1, KDR, and TEK2) involved in vessel maturation illustrated how PDGF addition contributed to vascular stability. Taken together, the results suggest that addition of PDGF at specific timepoints can be used to stabilise vasculature in CG scaffolds.</p

Development of magnetically active scaffolds as intrinsically-deformable bioreactors

Mesenchymal stem cell behavior can be regulated through mechanical signaling, either by dynamic loading or through biomaterial properties. We developed intrinsically responsive tissue engineering scaffolds that can dynamically load cells. Porous collagen- and alginate-based scaffolds were functionalized with iron oxide to produce magnetically active scaffolds. Reversible deformations in response to magnetic stimulation of up to 50% were recorded by tuning the material properties. Cells could attach to these scaffolds and magnetically induced compressive deformation did not adversely affect viability or cause cell release. This platform should have broad application in the mechanical stimulation of cells for tissue engineering applications

Synthesis of bilayer films from regenerated cellulose nanofibers and poly(globalide) for skin tissue engineering applications

Commercial cell-based skin regenerative products are highly expensive, carry the risk of rejection and require a long cell culture period to manufacture. This work describes the synthesis of bilayer films from poly(globalide) (PGl) and regenerated cellulose nanofibers (rCNFs) and their use as a cell-free scaffold to support keratinocyte attachment and proliferation. The method is simple, eco-friendly (as the cellulose precursor is obtained from agricultural waste) and of low cost. The rCNFs were produced by acid hydrolysis and PGl was obtained via enzymatic ring-opening polymerization. The bilayer films were synthesized by layer-by-layer casting at ambient temperature. All the films showed a well-defined interface between PGl and cellulose. The produced rCNF/PGl bilayer films showed cell metabolic activity far superior in comparison with pristine PGl regarding the keratinocyte growth, which illustrates the potential use of these materials in skin tissue engineering

Versatility of unsaturated polyesters from electrospun macrolactones: RGD immobilization to increase cell attachment

Poly(globalide) (PGl), an aliphatic polyester derived from unsaturated macrocylic lactone, can be cross-linked during electrospinning and drug-loaded for regenerative medicine applications. However, it lacks intrinsic recognition sites for cell adhesion and proliferation. In order to improve their cell adhesiveness, and therefore their therapeutic potential, we aimed to functionalize electrospun PGl fibers with RGD sequence generating a biomimetic scaffold. First, an amine compound was attached to the surface double bonds of the PGl fibers. Subsequently, the amino groups were coupled with RGD sequences. X-ray photoelectron spectroscopy (XPS) analysis confirmed the functionalization. The obtained fibers were more hydrophilic, as observed by contact angle analysis, and presented smaller Young's modulus, although similar tensile strength compared with non-functionalized cross-linked fibers. In addition, the functionalization process did not significantly alter fibers morphology, as observed by scanning electron microscopy (SEM). Finally, in vitro analysis evidenced the increase in human mesenchymal stromal cells (hMSC) adhesion (9.88 times higher DNA content after 1 day of culture) and proliferation (3.57 times higher DNA content after 8 days of culture) compared with non-functionalized non-cross-linked fibers. This is the first report demonstrating the functionalization of PGl fibers with RGD sequence, improving PGl therapeutic potential and further corroborating the use of this highly versatile material toward regenerative medicine applications

Personalized scaffolds for diabetic foot ulcer healing using extracellular matrix from induced pluripotent stem-reprogrammed patient cells

Diabetic foot ulcers (DFUs) are chronic wounds sustained by pathological fibroblasts and aberrant extracellular matrix (ECM). Porous collagen-based scaffolds (CS) have shown clinical promise for treating DFUs but may benefit from functional enhancements. Our previous work showed fibroblasts differentiated from induced pluripotent stem cells are an effective source of new ECM mimicking fetal matrix, which notably promotes scar-free healing. Likewise, functionalizing CS with this rejuvenated ECM shows potential for DFU healing. Herein, an approach to DFU healing is demonstrated for the first time using biopsied cells from DFU patients, reprogramming those cells, and functionalizing CS with patient-specific ECM as a personalized acellular tissue-engineered scaffold. A two-pronged approach is taken: 1) direct ECM blending into scaffold fabrication, and 2) seeding scaffolds with reprogrammed fibroblasts for ECM deposition followed by decellularization. The decellularization approach reduces cell number requirements and maintains naturally deposited ECM proteins. Both approaches show enhanced ECM deposition from DFU fibroblasts. Decellularized scaffolds additionally enhance glycosaminoglycan deposition and subsequent vascularization. Finally, reprogrammed ECM scaffolds from patient-matched DFU fibroblasts outperform those from healthy fibroblasts in several metrics, suggesting ECM is in fact able to redirect resident pathological fibroblasts in DFUs toward healing, and a patient-specific ECM signature may be beneficial.</p

Collagen scaffolds functionalised with copper-eluting bioactive glass reduce infection and enhance osteogenesis and angiogenesis both in vitro and in vivo.

The bone infection osteomyelitis (typically by Staphylococcus aureus) usually requires a multistep procedure of surgical debridement, long-term systemic high-dose antibiotics, and - for larger defects - bone grafting. This, combined with the alarming rise in antibiotic resistance, necessitates development of alternative approaches. Herein, we describe a one-step treatment for osteomyelitis that combines local, controlled release of non-antibiotic antibacterials with a regenerative collagen-based scaffold. To maximise efficacy, we utilised bioactive glass, an established osteoconductive material with immense capacity for bone repair, as a delivery platform for copper ions (proven antibacterial, angiogenic, and osteogenic properties). Multifunctional collagen-copper-doped bioactive glass scaffolds (CuBG-CS) were fabricated with favourable microarchitectural and mechanical properties (up to 1.9-fold increase in compressive modulus over CS) within the ideal range for bone tissue engineering. Scaffolds demonstrated antibacterial activity against Staphylococcus aureus (up to 66% inhibition) whilst also enhancing osteogenesis (up to 3.6-fold increase in calcium deposition) and angiogenesis in vitro. Most significantly, when assessed in a chick embryo in vivo model, CuBG-CS not only demonstrated biocompatibility, but also a significant angiogenic and osteogenic response, consistent with in vitro studies. Collectively, these results indicate that the CuBG-CS developed here show potential as a one-step osteomyelitis treatment: reducing infection, whilst enhancing bone healing.</p