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Mobilizing endogenous progenitor cells to repair damaged tissue in situ has the potential to revolutionize the field of regenerative medicine, while the early establishment of a vascular network will ensure survival of newly generated tissue. In this study, a gene-activated scaffold containing a stromal derived factor 1α plasmid (pSDF1α), a pro-angiogenic gene that is also thought to be involved in the recruitment of mesenchymal stromal cells (MSCs) to sites of injury is described. It is shown that over-expression of SDF1α protein enhanced MSC recruitment and induced vessel-like structure formation by endothelial cells in vitro. When implanted subcutaneously, transcriptomic analysis reveals that endogenous MSCs are recruited and significant angiogenesis is stimulated. Just 1-week after implantation into a calvarial critical-sized bone defect, pSDF1α-activated scaffolds are recruited MSCs and rapidly activate angiogenic and osteogenic programs, upregulating Runx2, Dlx5, and Sp7. At the same time-point, pVEGF-activated scaffolds are recruited a variety of cell types, activating endochondral ossification. The early response induced by both scaffolds leads to complete bridging of the critical-sized bone defects within 4-weeks. The versatile cell-free gene-activated scaffold described in this study is capable of harnessing and enhancing the body's own regenerative capacity and has immense potential in a myriad of applications., (© 2024 The Author(s). Advanced Healthcare Materials published by Wiley‐VCH GmbH.)

After spinal cord injury (SCI), tissue engineering scaffolds offer a potential bridge for regeneration across the lesion and support repair through proregenerative signaling. Ideal biomaterial scaffolds that mimic the physicochemical properties of native tissue have the potential to provide innate trophic signaling while also minimizing damaging inflammation. To address this challenge, taking cues from the spinal cord's structure, the proregenerative signaling capabilities of native cord components are compared in vitro. A synergistic mix of collagen-IV and fibronectin (Coll-IV/Fn) is found to optimally enhance axonal extension from neuronal cell lines (SHSY-5Y and NSC-34) and induce morphological features typical of quiescent astrocytes. This optimal composition is incorporated into hyaluronic acid scaffolds with aligned pore architectures but varying stiffnesses (0.8-3 kPa). Scaffolds with biomimetic mechanical properties (<1 kPa), functionalized with Coll-IV/Fn, not only modulate primary astrocyte behavior but also stimulate the production of anti-inflammatory cytokine IL-10 in a stiffness-dependent manner. Seeded SHSY-5Y neurons generate distributed neuronal networks, while softer biomimetic scaffolds promote axonal outgrowth in an ex vivo model of axonal regrowth. These results indicate that the interaction of stiffness and biomaterial composition plays an essential role in vitro in generating repair-critical cellular responses and demonstrates the potential of biomimetic scaffold design., (© 2021 Wiley-VCH GmbH.)

Joint repair remains a major challenge in orthopaedics. Recent progress in biomaterial design has led to the fabrication of a plethora of promising devices. Pre-clinical testing of any joint repair strategy typically requires the use of large animal models (e.g., sheep, goat, pig or horse). Despite the key role of such models in clinical translation, there is still a lack of consensus regarding optimal experimental design, making it difficult to draw conclusions on their efficacy. In this context, the authors performed a systematic literature review and a risk of bias assessment on large animal models published between 2010 and 2020, to identify key experimental parameters that significantly affect the biomaterial therapeutic outcome and clinical translation potential (including defect localization, animal age/maturity, selection of controls, cell-free versus cell-laden). They determined that mechanically strong biomaterials perform better at the femoral condyles; while highlighted the importance of including native tissue controls to better evaluate the quality of the newly formed tissue. Finally, in cell-laded biomaterials, the pre-culture conditions played a more important role in defect repair than the cell type. In summary, here they present a systematic evaluation on how the experimental design of preclinical models influences biomaterial-based therapeutic outcomes in joint repair., (© 2021 The Authors. Advanced Healthcare Materials published by Wiley-VCH GmbH.)

Diabetic foot ulcers (DFUs) are chronic wounds, with 20% of cases resulting in amputation, despite intervention. A recently approved tissue engineering product-a cell-free collagen-glycosaminoglycan (GAG) scaffold-demonstrates 50% success, motivating its functionalization with extracellular matrix (ECM). Induced pluripotent stem cell (iPSC) technology reprograms somatic cells into an embryonic-like state. Recent findings describe how iPSCs-derived fibroblasts ("post-iPSF") are proangiogenic, produce more ECM than their somatic precursors ("pre-iPSF"), and their ECM has characteristics of foetal ECM (a wound regeneration advantage, as fetuses heal scar-free). ECM production is 45% higher from post-iPSF and has favorable components (e.g., Collagen I and III, and fibronectin). Herein, a freeze-dried scaffold using ECM grown by post-iPSF cells (Post-iPSF Coll) is developed and tested vs precursors ECM-activated scaffolds (Pre-iPSF Coll). When seeded with healthy or DFU fibroblasts, both ECM-derived scaffolds have more diverse ECM and more robust immune responses to cues. Post-iPSF-Coll had higher GAG, higher cell content, higher Vascular Endothelial Growth Factor (VEGF) in DFUs, and higher Interleukin-1-receptor antagonist (IL-1ra) vs. pre-iPSF Coll. This work constitutes the first step in exploiting ECM from iPSF for tissue engineering scaffolds., (© 2020 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Current treatments for articular cartilage defects relieve symptoms but often only delay cartilage degeneration. Mesenchymal stem cells (MSCs) have shown chondrogenic potential but tend to undergo endochondral ossification when implanted in vivo. Harnessing factors governing joint development to functionalize biomaterial scaffolds, termed developmental engineering, might allow to prime host MSCs to regenerate mature articular cartilage in situ without requiring cell isolation or ex vivo expansion. Therefore, the aim of this study is to develop a gene-activated scaffold capable of delivering developmental cues to host MSCs, thus priming MSCs for articular cartilage differentiation and inhibiting endochondral ossification. It is shown that delivery of the SOX-Trio induced MSCs to over-express COL2A1 and ACAN and deposit a sulfated and collagen type II rich extracellular matrix while hypertrophic gene expression and collagen type X deposition is inhibited. When cell-free SOX-Trio-activated scaffolds are implanted ectopically in vivo, they induced spontaneous chondrogenesis without evidence of hypertrophy. MSCs pre-cultured on SOX-Trio-activated scaffolds prior to implantation differentiate into phenotypically stable chondrocytes as evidenced by a lack of collagen X expression or vascular invasion. This SOX-trio-activated scaffold represents a potent, single treatment, developmentally inspired strategy to prime MSCs in situ for articular cartilage defect repair., (© 2020 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

microRNA-based therapies are an advantageous strategy with applications in both regenerative medicine (RM) and cancer treatments. microRNAs (miRNAs) are an evolutionary conserved class of small RNA molecules that modulate up to one third of the human nonprotein coding genome. Thus, synthetic miRNA activators and inhibitors hold immense potential to finely balance gene expression and reestablish tissue health. Ongoing industry-sponsored clinical trials inspire a new miRNA therapeutics era, but progress largely relies on the development of safe and efficient delivery systems. The emerging application of biomaterial scaffolds for this purpose offers spatiotemporal control and circumvents biological and mechanical barriers that impede successful miRNA delivery. The nascent research in scaffold-mediated miRNA therapies translates know-how learnt from studies in antitumoral and genetic disorders as well as work on plasmid (p)DNA/siRNA delivery to expand the miRNA therapies arena. In this progress report, the state of the art methods of regulating miRNAs are reviewed. Relevant miRNA delivery vectors and scaffold systems applied to-date for RM and cancer treatment applications are discussed, as well as the challenges involved in their design. Overall, this progress report demonstrates the opportunity that exists for the application of miRNA-activated scaffolds in the future of RM and cancer treatments., (© 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Clinically available hollow nerve guidance conduits (NGCs) have had limited success in treating large peripheral nerve injuries. This study aims to develop a biphasic NGC combining a physicochemically optimized collagen outer conduit to bridge the transected nerve, and a neuroconductive hyaluronic acid-based luminal filler to support regeneration. The outer conduit is mechanically optimized by manipulating crosslinking and collagen density, allowing the engineering of a high wall permeability to mitigate the risk of neuroma formation, while also maintaining physiologically relevant stiffness and enzymatic degradation tuned to coincide with regeneration rates. Freeze-drying is used to seamlessly integrate the luminal filler into the conduit, creating a longitudinally aligned pore microarchitecture. The luminal stiffness is modulated to support Schwann cells, with laminin incorporation further enhancing bioactivity by improving cell attachment and metabolic activity. Additionally, this biphasic NGC is shown to support neurogenesis and gliogenesis of neural progenitor cells and axonal outgrowth from dorsal root ganglia. These findings highlight the paradigm that a successful NGC requires the concerted optimization of both a mechanical support phase capable of bridging a nerve defect and a neuroconductive phase with an architecture capable of supporting both Schwann cells and neurons in order to achieve functional regenerative outcome., (© 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

The biofabrication of large natural biomaterial scaffolds into complex 3D shapes which have a controlled microarchitecture remains a major challenge. Freeze-drying (or lyophilization) is a technique used to generate scaffolds in planar 3D geometries. Here we report the development of a new biofabrication process to form a collagen-based scaffold into a large, complex geometry which has a large height to width ratio, and a controlled porous microarchitecture. This biofabrication process is validated through the successful development of a heart valve shaped scaffold, fabricated from a collagen-glycosaminoglycan co-polymer. Notably, despite the significant challenges in using freeze-drying to create such a structure, the resultant scaffold has a uniform, homogenous pore architecture throughout. This is achieved through optimization of the freeze-drying mold and the freezing parameters. We believe this to be the first demonstration of using freeze-drying to create a large, complex scaffold geometry with a controlled, porous architecture for natural biomaterials. This study validates the potential of using freeze-drying for development of organ-specific scaffold geometries for tissue engineering applications, which up until now might not have been considered feasible., (© 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Defects within bones caused by trauma and other pathological complications may often require the use of a range of therapeutics to facilitate tissue regeneration. A number of approaches have been widely utilized for the delivery of such therapeutics via physical encapsulation or chemical immobilization suggesting significant promise in the healing of bone defects. The study focuses on the chemical immobilization of osteostatin, a pentapeptide of the parathyroid hormone (PTHrP107-111), within a collagen-hydroxyapatite scaffold. The chemical attachment method via crosslinking supports as little as 4% release of the peptide from the scaffolds after 21 d whereas non-crosslinking leads to 100% of the peptide being released by as early as 4 d. In vitro characterization demonstrates that this cross-linking method of immobilization supports a pro-osteogenic effect on osteoblasts. Most importantly, when implanted in a critical-sized calvarial defect within a rat, these scaffolds promote significantly greater new bone volume and area compared to nonfunctionalized scaffolds (**p < 0.01) and an empty defect control (***p < 0.001). Collectively, this study suggests that such an approach of chemical immobilization offers greater spatiotemporal control over growth factors and can significantly modulate tissue regeneration. Such a system may be adopted for a range of different proteins and thus offers the potential for the treatment of various complex pathologies that require localized mediation of drug delivery., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Incorporation of therapeutics in the form of growth factors within biomaterials can enhance their biofunctionality. Two methods of incorporating transforming growth factor-beta 3 within collagen-hyaluronic acid scaffolds are described, markedly improving mesenchymal stem cell-mediated chondrogenic differentiation and matrix production. Such scaffolds offer control over the release of therapeutics, demonstrating their potential for repair of complex chondral defects requiring additional stimuli., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

An alternative strategy to the use of in vitro expanded cells in regenerative medicine is the use of freshly isolated stromal cells, where a bioactive scaffold is used to provide an environment conducive to proliferation and tissue-specific differentiation in vivo. The objective of this study is to develop a cartilage extracellular matrix (ECM)-derived scaffold that could facilitate the rapid proliferation and chondrogenic differentiation of freshly isolated stromal cells. By freeze-drying cryomilled cartilage ECM of differing concentrations, it is possible to produce scaffolds with a range of pore sizes. The migration, proliferation, and chondrogenic differentiation of infrapatellar fat pad-derived stem cells (FPSCs) depend on the concentration/porosity of these scaffolds, with greater sulphated glycosaminoglycan (sGAG) accumulation observed in scaffolds with larger-sized pores. It is then sought to determine if freshly isolated fat pad-derived stromal cells, seeded onto a transforming growth factor (TGF)-β3 eluting ECM-derived scaffold, could promote chondrogenesis in vivo. While a more cartilage-like tissue could be generated using culture expanded FPSCs compared to nonenriched freshly isolated cells, fresh CD44(+) stromal cells are capable of producing a tissue in vivo that stained strongly for sGAGs and type II collagen. These findings open up new possibilities for in-theatre cell-based therapies for joint regeneration., (© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Vascularization and bone repair are accelerated by a series of gene-activated scaffolds delivering both an angiogenic and an osteogenic gene. Stem cell-mediated osteogenesis in vitro, in addition to increased vascularization and bone repair by host cells in vivo, is enhanced using all systems while the use of the nanohydroxyapatite vector to deliver both genes markedly enhances bone healing., (© 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

A novel drug delivery system, enabling an in situ, thermally triggered drug release is described, consisting of an injectable thermoresponsive chitosan hydrogel containing doxorubicin-loaded thermosensitive liposomes. The design, fabrication, characterization, and an assessment of in vitro bioactivity of this formulation is detailed. Combining on-demand drug delivery with in situ gelation results in a promising candidate for local chemotherapy., (© 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)