Abstract
Extracellular vesicles (EVs) have garnered growing attention as promising acellular tools for bone repair. Although EVs’ potential for bone regeneration has been shown, issues associated with their therapeutic potency and short half-life in vivo hinders their clinical utility. Epigenetic reprogramming with the histone deacetylase inhibitor Trichostatin A (TSA) has been reported to promote the osteoinductive potency of osteoblast-derived EVs. Gelatin methacryloyl (GelMA) hydrogels functionalised with the synthetic nanoclay laponite (LAP) have been shown to effectively bind, stabilise, and improve the retention of bioactive factors. This study investigated the potential of utilising a GelMA-LAP hydrogel to improve local retention and control delivery of epigenetically enhanced osteoblast-derived EVs as a novel bone repair strategy. LAP was found to elicit a dose-dependent increase in GelMA compressive modulus and shear-thinning properties. Incorporation of the nanoclay was also found to enhance shape fidelity when 3D printed compared to LAP-free gels. Interestingly, GelMA hydrogels containing LAP displayed increased mineralisation capacity (1.41-fold) (p ≤ 0.01) over 14 days. EV release kinetics from these nanocomposite systems were also strongly influenced by LAP concentration with significantly more vesicles being released from GelMA constructs as detected by a CD63 ELISA (p ≤ 0.001). EVs derived from TSA-treated osteoblasts (TSA-EVs) enhanced proliferation (1.09-fold), migration (1.83-fold), histone acetylation (1.32-fold) and mineralisation (1.87-fold) of human bone marrow stromal cells (hBMSCs) when released from the GelMA-LAP hydrogel compared to the untreated EV gels (p ≤ 0.01). Importantly, the TSA-EV functionalised GelMA-LAP hydrogel significantly promoted encapsulated hBMSCs extracellular matrix collagen production (≥1.3-fold) and mineralisation (≥1.78-fold) in a dose-dependent manner compared to untreated EV constructs (p ≤ 0.001). Taken together, these findings demonstrate the potential of combining epigenetically enhanced osteoblast-derived EVs with a nanocomposite photocurable hydrogel to promote the therapeutic efficacy of acellular vesicle approaches for bone regeneration.
Highlights
Introduction conditions of the Creative CommonsThe treatment of bone fractures represents a tremendous socioeconomic burden worldwide, with approximately 10 million people in the UK afflicted with musculoskeletal disorders [1]
A growing body of evidence has demonstrated the potential of harnessing Extracellular vesicles (EVs) as novel acellular tools to promote bone regeneration [24,26,32]
Having confirmed a similar quantity of MO-EVs and Trichostatin A (TSA)-EVs released from these Gelatin methacryloyl (GelMA)-LAP hydrogels, the enhanced stimulation observed at recipient human bone marrow stromal cells (hBMSCs) is likely due to TSA-EVs increased biological potency rather than differences in the concentration of nanoparticles released. These findings demonstrate the GelMA-LAP system controlled the EV release kinetics and preserved the biological potency of these epigenetically modified EVs when released from the hydrogel, indicating the GelMA-LAP system provides a suitable vehicle to deliver TSA-EVs without sacrificing their functionality
Summary
Introduction conditions of the Creative CommonsThe treatment of bone fractures represents a tremendous socioeconomic burden worldwide, with approximately 10 million people in the UK afflicted with musculoskeletal disorders [1]. The combination of bone graft substitutes with osteoinductive growth factors such as bone morphogenic protein 2 (BMP2) have shown positive clinical results [3,4]. There is a significant demand for novel approaches to regenerate damaged bone, overcoming the limitations of current strategies [8]. Cell-based tissue engineering approaches have shown great promise in recent years, with methods combining osteoinductive biomaterials with mesenchymal stromal cells (MSCs) seen as an attractive bone augmentation strategy [9]. Encouraging results have been observed, the direct transplantation of MSC-based therapies are associated with numerous complications including their uncontrolled differentiation, immunological rejection, inherent heterogeneity, functional tissue engraftment and neoplasm formation [10,11]. There is a growing interest in utilising cell-free approaches as an alternative to stimulate bone repair
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