Harnessing 3D smart materials for targeted stem cell fate and function in skeletal repair

Abstract

The ability to regenerate damaged skeletal tissue is a significant unmet need for an increasingly ageing population. Stem cells offer an exciting approach to address the problem of tissue repair. Furthermore, it is critical to provide biocompatible scaffolds for regenerative medicine that can stimulate cell proliferation and differentiation to generate the tissue of interest. Currently, the field of materials research has provided a number of scaffolds, including scaffolds incorporating cells for repair, however, a significant challenge remains the generation and provision of efficacious scaffolds to support skeletal cell differentiation via delivery of osteogenic factors in a spatiotemporal manner. Furthermore, it is important to generate scaffolds that can aid vascular formation throughout the scaffold matrix to support nutrient delivery and to remove waste.The objectives of this study were to develop, characterise and optimise scaffolds able to support skeletal cell and osteogenic factor delivery through the use of collagen-derived gelatin methacryloyl (GelMA) hydrogel based bioinks, polymerised using visible light at a wavelength of 450 nm, to generate a highly compatible bioink for cell printing. The hydrogel maintained its shape post-printing and was able to support cell differentiation. Co-culture studies were performed using human adult bone marrow stromal cells (HBMSCs) with human umbilical vein endothelial cells (HUVECs) to investigate the potential to encapsulate the cell populations within hydrogel matrices to support vasculature formation in parallel with bone formation. Analysis showed 7.5% (w/v) GelMA was able to support the co-culture of HUVECs with HBMSCs, allowing expression of angiogenic markers including von Willebrand Factor (vWF) and CD31 (PECAM-1) in HUVECs. The current studies demonstrated that although co-culture constructs were able to maintain the expression of the early bone marker Alkaline Phosphatase (ALP), HUVECs failed to form any vasculature in the matrices in the absence of the angiogenic mitogen vascular endothelial growth factor (VEGF).In subsequent studies, a strategy was developed to deliver osteogenic factors, including ascorbic acid-2-phosphate (Vitamin C) and 1,25-OH2-Vitamin D3 (calcitriol), encapsulated in poly-lactic-co-glycolic (PLGA) nanoparticles that would efficiently release the incorporated factors and stimulate human bone marrow stromal cells (HBMSCs) differentiation into osteoblasts in vitro. The physical properties of the nanoparticles were optimised to prevent aggregation and to ensure uniform osteogenic factor delivery, although, the PLGA nanoparticles failed to encapsulate the osteogenic factors in the absence of stabilising compounds in the nanoparticle shells. In addition to investigation of natural polymer scaffolds for bone formation support, a study on synthetic polymers was undertaken using polyacrylate scaffolds, synthesised using a novel, high-throughput method to generate three-dimensional matrices. A total of six potential candidate materials were identified including a polymer comprising 40% (w/v) 2-(methylthio)ethyl methacrylate and 40% (w/v) isobornyl acrylate with a capacity to support extensive vasculature formation in the presence and absence of human foetal bone marrow stromal cells (FBMSCs). The polymer was able to support osteogenic differentiation in vitro evidenced by elevated levels of ALP and collagen expression. In vivo studies demonstrated the scaffolds were able to recruit host cells from the surrounding tissue and to support the formation of an extensive vasculature network. Harnessing the findings from both natural and synthetic polymer systems will ultimately allow for the fabrication of a support framework for bone formation tailored to both formation of vasculature and bone tissue. Such scaffolds offer an opportunity to address a number of orthopaedic challenges for an ageing population. Furthermore, these tailored scaffolds offer the potential to generate scaffolds at reduced cost and to establish simple scaffold platforms to aid bone reparation and recovery

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Identifiers

ORCID for Richard Oreffo: orcid.org/0000-0001-5995-6726

ORCID for Rahul Tare: orcid.org/0000-0001-8274-8837

ORCID for Shoufeng Yang: orcid.org/0000-0002-3888-3211

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