Tailoring osteo-immunomodulatory micro-environments via a bioactive 3D PLA scaffold to potentiate regenerative...

The burden of long bone fractures impacts society profoundly. Polylactic acid (PLA) and PLA-based scaffolds have been widely used for bone tissue engineering due to its biodegradable and non-toxic properties. However, PLA scaffolds possess no biological and immunoregulatory activity. In addition, M2 phenotypic macrophages and M2 macrophage inducer have shown pro-healing effect. Therefore, in this study, we sought to harness the potent osteo-immunomodulatory properties of anti-inflammatory M2 macrophages and encapsulated bioactive interleukin-4 microdroplets (MDs) into the PLA scaffold matrix in order to enhance the biological activity and osteo-immunomodulatory properties of PLA scaffold. The MTT assay, live and dead staining, and phalloidin/DAPI dual staining were used to evaluate biocompatibility in human bone marrow mesenchymal stem cells (hBMSCs). Unstimulated macrophages and inflammatory macrophages were further used to test its immunoregulatory role. The immunoregulatory role of MDs-IL4/PLA hybrid scaffolds on hBMSCs differentiation was further investigated in vitro. The MDs-IL4/PLA hybrid scaffolds showed high biocompatibility in hBMSCs. In addition, MDs-IL4/PLA hybrid scaffolds demonstrated good immunoregulatory role in unstimulated and inflammatory macrophages, as evidenced by marked morphological changes, significantly increased M2 phenotypic markers, and reduction of pro-inflammatory marker expression, etc. Furthermore, our results indicate that the MDs-IL4/PLA hybrid scaffolds could significantly enhance the osteogenic differentiation of hBMSCs via its potential immunomodulatory properties. The subcutaneous implantation of MDs-IL4/PLA scaffold demonstrated significant increases in IL-4, IL-10, CCL2, CCL20, beta-NGF, and TIMP-1 expressions, indicating its notable in vivo immunomodulatory effects. By harnessing the synergistic effects of PLA and MDs-IL4, this novel scaffold holds promise for advancing regenerative medicine approaches, particularly in bone tissue engineering.