Articular cartilage (AC) possesses a limited healing potential, meaning that untreated focal joint defects typically progress, leading to the development of degenerative diseases such as osteoarthritis. Several clinical strategies exist that aim to regenerate AC; however, recapitulation of a fully functional, load-bearing tissue remains a significant challenge. This can be attributed, at least in part, to a paucity of biomaterials that truly mimic the native tissue and provide appropriate cues to direct its regeneration. The main structural component of articular cartilage, type II collagen, does not readily gelate at body temperature, challenging the development of cartilage extracellular matrix (cECM)-derived injectable hydrogels and bioinks for AC tissue engineering and bioprinting applications. Here, we describe the development and rheological characterisation of a methacrylated cartilage ECM-based hydrogel/bioink (cECM-MA), which could be photocrosslinked when exposed to ultraviolet (UV) light. Functionalisation of the collagen backbone with methacryloyl groups had a negligible effect on triple helix stability, as demonstrated by circular dichroism spectroscopy. These cECM-MA bioinks demonstrated shear-thinning properties and could be loaded with bone marrow mesenchymal stem cells (BM-MSCs), micro-extruded to generate self-supporting 3D constructs of predefined size and shape, and then photocrosslinked using UV light. Analysis of the cell-laden constructs showed that the BM-MSCs were viable post-printing and underwent chondrogenesis in vitro, generating a tissue rich in sulphated glycosaminoglycans and collagens. These results support the use of methacrylated, tissue-specific ECM-derived hydrogels as bioinks for 3D bioprinting and/or as injectables for cartilage tissue engineering applications.

Photodynamic therapy (PDT) is an anti-cancer therapy with proven efficacy; however, its application is often limited by prolonged skin photosensitivity and solubility issues associated with the phototherapeutic agents. Injectable hydrogels that can effectively provide intra-tumoral delivery of photosensitizers with sustained release is attracting increased interest for photodynamic cancer therapies. However, most of the hydrogels for PDT applications are based on systems with high complexity and often, pre-clinical validation is not provided. Herein, we provide a simple and reliable pH-sensitive hydrogel formulation that presents proper rheological properties for intra-tumoral injection. For this, Temoporfin (m-THPC), which is one of the most potent clinical photosensitizers, was chemically modified to introduce functional groups that act as cross-linkers in the formation of chitosan-based hydrogels. The introduction of -COOH groups gave rise to a water-soluble derivative, named as PS 2, that was the most promising candidate. Although PS 2 was not internalized by the target cells, its extracellular activation cause effective damage of the cancer cells which was likely mediated by lipid peroxidation. The injection of the hydrogel containing PS 2 in the tumors was monitored by high frequency ultrasounds and in vivo fluorescence imaging which confirmed the sustained release of PS 2 for at least 72 h. Following local administration, light exposure was conducted one (single irradiation protocol) or three (multiple irradiation protocol) times. The latter deliver the best therapeutic outcomes which included complete tumor regression and systemic anti-cancer immune responses. Immunological memory was developed as ~ 75% of the mice cured with our strategy rejected a second rechallenge with live cancer cells. Additionally, failure of PDT to treat immunocompromised mice bearing tumors reinforces the relevance of the host immune system. Finally, our strategy promotes anti-cancer immune responses that lead to the abscopal protection against distant metastases.