Distinct Immunomodulatory Strategies Guide Mesenchymal Stromal/Stem Cell-Mediated Bone Regeneration.

The bone regenerative potential of bone marrow-derived mesenchymal stromal/stem cells (MSC) is critically shaped by their interaction with the host immune system. Here, we compare two MSC priming strategies: coculture with regulatory T cell (Treg) or exposure to the proresolving mediator Resolvin E1 (RvE1) to enhance ectopic osteogenesis in mice. Both approaches enhance MSC-mediated bone formation through distinct local and systemic immunomodulatory mechanisms. Treg priming promotes a regulated anti-inflammatory environment characterized by reduced IL-6 protein levels (3.6-fold), sustained IL-13 elevation up to 10 weeks (1.5-fold compared with RvE1), early suppression of cytotoxic CD8⁺ T cells and IFN-γ production by CD4⁺ T cells in lymph nodes, a >25% reduction in fibrous capsule formation, and enhanced B cell activity, collectively supporting ossification. In contrast, RvE1 priming promotes early TNF-α and late granulocyte-macrophage colony-stimulating factor (GM-CSF) production (1.2-fold compared with Treg) within the scaffold, facilitating osteoblast maturation. Transcriptomic profiling revealed distinct osteogenic signatures, with amphiregulin uniquely upregulated in the Treg group (>7-fold). Using a Treg depletion model, we further demonstrate that RvE1 retains partial immunomodulatory capacity and supports matrix organization. Together, these findings underscore the versatility of immune modulation in directing MSC fate and highlight the potential of tailoring immunoregulatory strategies to optimize bone regeneration.