Modelling skeletal pain harnessing tissue engineering.

Bone pain typically occurs immediately following skeletal damage with mechanical distortion or rupture of nociceptive fibres. The pain mechanism is also associated with chronic pain conditions where the healing process is impaired. Any load impacting on the area of the fractured bone will stimulate the nociceptive response, necessitating rapid clinical intervention to relieve pain associated with the bone damage and appropriate mitigation of any processes involved with the loss of bone mass, muscle, and mobility and to prevent death. The following review has examined the mechanisms of pain associated with trauma or cancer-related skeletal damage focusing on new approaches for the development of innovative therapeutic interventions. In particular, the review highlights tissue engineering approaches that offer considerable promise in the application of functional biomimetic fabrication of bone and nerve tissues. The strategic combination of bone and nerve tissue engineered models provides significant potential to develop a new class of in vitro platforms, capable of replacing in vivo models and testing the safety and efficacy of novel drug treatments aimed at the resolution of bone-associated pain. To date, the field of bone pain research has centred on animal models, with a paucity of data correlating to the human physiological response. This review explores the evident gap in pain drug development research and suggests a step change in approach to harness tissue engineering technologies to recapitulate the complex pathophysiological environment of the damaged bone tissue enabling evaluation of the associated pain-mimicking mechanism with significant therapeutic potential therein for improved patient quality of life. Graphical abstract: Rationale underlying novel drug testing platform development. Pain detected by the central nervous system and following bone fracture cannot be treated or exclusively alleviated using standardised methods. The pain mechanism and specificity/efficacy of pain reduction drugs remain poorly understood. In vivo and ex vivo models are not yet able to recapitulate the various pain events associated with skeletal damage. In vitro models are currently limited by their inability to fully mimic the complex physiological mechanisms at play between nervous and skeletal tissue and any disruption in pathological states. Robust innovative tissue engineering models are needed to better understand pain events and to investigate therapeutic regimes.

De Novo Design of Functional Coassembling Organic-Inorganic Hydrogels for Hierarchical Mineralization and Neovascularization.

Synthetic nanostructured materials incorporating both organic and inorganic components offer a unique, powerful, and versatile class of materials for widespread applications due to the distinct, yet complementary, nature of the intrinsic properties of the different constituents. We report a supramolecular system based on synthetic nanoclay (Laponite, Lap) and peptide amphiphiles (PAs, PAH3) rationally designed to coassemble into nanostructured hydrogels with high structural integrity and a spectrum of bioactivities. Spectroscopic and scattering techniques and molecular dynamic simulation approaches were harnessed to confirm that PAH3 nanofibers electrostatically adsorbed and conformed to the surface of Lap nanodisks. Electron and atomic force microscopies also confirmed an increase in diameter and surface area of PAH3 nanofibers after coassembly with Lap. Dynamic oscillatory rheology revealed that the coassembled PAH3-Lap hydrogels displayed high stiffness and robust self-healing behavior while gas adsorption analysis confirmed a hierarchical and heterogeneous porosity. Furthermore, this distinctive structure within the three-dimensional (3D) matrix provided spatial confinement for the nucleation and hierarchical organization of high-aspect ratio hydroxyapatite nanorods into well-defined spherical clusters within the 3D matrix. Applicability of the organic-inorganic PAH3-Lap hydrogels was assessed in vitro using human bone marrow-derived stromal cells (hBMSCs) and ex vivo using a chick chorioallantoic membrane (CAM) assay. The results demonstrated that the organic-inorganic PAH3-Lap hydrogels promote human skeletal cell proliferation and, upon mineralization, integrate with the CAM, are infiltrated by blood vessels, stimulate extracellular matrix production, and facilitate extensive mineral deposition relative to the controls.

Osteogenic and angiogenic tissue formation in high fidelity nanocomposite Laponite-gelatin bioinks.

Bioprinting of living cells is rapidly developing as an advanced biofabrication approach to engineer tissues. Bioinks can be extruded in three-dimensions (3D) to fabricate complex and hierarchical constructs for implantation. However, a lack of functionality can often be attributed to poor bioink properties. Indeed, advanced bioinks encapsulating living cells should: (i) present optimal rheological properties and retain 3D structure post fabrication, (ii) promote cell viability and support cell differentiation, and (iii) localise proteins of interest (e.g. vascular endothelial growth factor (VEGF)) to stimulate encapsulated cell activity and tissue ingrowth upon implantation. In this study, we present the results of the inclusion of a synthetic nanoclay, Laponite® (LPN) together with a gelatin methacryloyl (GelMA) bioink and the development of a functional cell-instructive bioink. A nanocomposite bioink displaying enhanced shape fidelity retention and interconnected porosity within extrusion-bioprinted fibres was observed. Human bone marrow stromal cell (HBMSC) viability within the nanocomposite showed no significant changes over 21 days of culture in LPN-GelMA (85.60 ± 10.27%), compared to a significant decrease in GelMA from 7 (95.88 ± 2.90%) to 21 days (55.54 ± 14.72%) (p < 0.01). HBMSCs were observed to proliferate in LPN-GelMA with a significant increase in cell number over 21 days (p < 0.0001) compared to GelMA alone. HBMSC-laden LPN-GelMA scaffolds supported osteogenic differentiation evidenced by mineralised nodule formation, including in the absence of the osteogenic drug dexamethasone. Ex vivo implantation in a chick chorioallantoic membrane model, demonstrated excellent integration of the bioink constructs in the vascular chick embryo after 7 days of incubation. VEGF-loaded LPN-GelMA constructs demonstrated significantly higher vessel penetration than GelMA-VEGF (p < 0.0001) scaffolds. Integration and vascularisation was directly related to increased drug absorption and retention by LPN-GelMA compared to LPN-free GelMA. In summary, a novel light-curable nanocomposite bioink for 3D skeletal regeneration supportive of cell growth and growth factor retention and delivery, evidenced by ex vivo vasculogenesis, was developed with potential application in hard and soft tissue reparation.