A major challenge in cartilage tissue engineering (TE) is to develop scaffolds capable of providing an instructive biomimetic environment to effectively drive mesenchymal stromal cells (MSCs) differentiation. Hydrogels have emerged as promising biomaterials for this purpose, due to their biocompatibility and ability to mimic the tissue extracellular matrix. Recently, graphene oxide (GO) emerged as a promising nanomaterial for cartilage TE due to chondroinductive properties when embedded into polymeric formulations. It has been also shown that piezoelectric nanomaterials, like barium titanate (BaTiO3) nanoparticles, can be exploited as nanoscale transducers capable of inducing cell growth/differentiation. Ultrasound waves are an interesting tool to facilitate chondrogenesis. In particular, it has been demonstrated that low-intensity pulsed ultrasound (LIPUS) regulates the differentiation of ASCs.
The aim of this study was to investigate whether dose-controlled LIPUS is able to influence chondrogenic differentiation of ASCs embedded in a 3D piezoelectric hydrogel.
ASCs at 2*106 cells/mL were embedded in a 3D VitroGel RGD® hydrogel with or without nanoparticles (GO, 25 µg/ml, BaTiO3, 50 µg/ml) and subjected to LIPUS stimulation every 2 days, until day 10 of culture.
Hydrogels were cultured and chondrogenically differentiated for 2, 7, 10 and 28 days. At each time point cell viability, cytotoxicity, gene expression and immunohistochemistry (COL2, aggrecan, SOX9, and COL1) were evaluated.
In both 3D hydrogels we evidenced that LIPUS treatment did not affect negatively the viability of the embedded cells. LIPUS boosted the chondrogenic differentiation of ASCs laden in 3D piezoelectric hydrogel: the chondrogenic genes and proteins markers (COL2,aggrecan and SOX9) were increased while the fibrotic marker COL1 was decreased compared to control samples (non piezoelectric hydrogels and piezoelectric hydrogels not stimulated with LIPUS).
These results suggest that the combination of LIPUS and 3D piezoelectric hydrogels push the differentiation of ASCs and represent a promising tool in the field of cartilage TE.