Nano-layered tendon patch to orchestrate timely interactions between tendon stem/progenitor cells and macrophages.

Tendon regeneration remains limited by the unmet challenge of precisely controlling the intensity and duration of local inflammation, often resulting in scarred healing. Here, we present a novel, micro-thin, nano-layered patch that employs Layer-by-Layer (LbL) self-assembly for the precisely controlled release of small molecules, Oxo-M and 4-PPBP, to address this technical challenge. This novel combination of small molecules exhibits distinct functions in modulating the polarization of macrophages and the tenogenic differentiation of tendon stem/progenitor cells (TSCs). The customized, temporal release of these small molecules effectively modulated the critical crosstalk between TSCs and macrophages, thereby promoting regenerative tendon healing. The nano-LbL tendon patch successfully mitigated early inflammation, promoted matrix synthesis, and regulated later tissue remodeling in vivo. Robust scRNA-seq and CellChat analysis delineated the temporal-orchestrated communication between cell types across healing phases, confirming the critical interplays between TSCs, tenocytes, and macrophages. We then validated the key signaling pathways regulating cell-cell communication modulating inflammation. Our nano-layered LbL nanopatch represents a highly translational approach for achieving scarless tendon regeneration. In addition, this study elucidated the essential signaling pathways regulating TSC-macrophage crosstalk, advancing our understanding of tendon biology and pathology.