PLCγ1 knockdown in BMSCs enhances bone repair via IL-4-JAK2-STAT3-mediated reparative macrophage polarization.

Severe bone defects pose significant clinical challenges. While mesenchymal stem cell (MSC)-based therapies are promising, directing MSC differentiation and modulating the immune microenvironment remain major hurdles. This study identifies phospholipase Cγ1 (PLCγ1) inhibition in bone marrow MSCs (BMSCs) as a novel strategy to enhance bone repair through synergistic crosstalk with macrophages. In a rat femoral bone defect model, implantation of alginate hydrogels containing PLCγ1-knockdown BMSCs significantly improved bone regeneration and induced an reparative macrophage (M2-like macrophage) phenotype (increased CD206+, decreased CD80+) within the marrow cavity. In vitro, PLCγ1 knockdown in BMSCs robustly promoted osteogenic differentiation without affecting proliferation. Conditioned medium (CM) from siPLCγ1 BMSCs polarized bone marrow-derived macrophages (BMDMs) toward an reparative macrophage phenotype (reduced CD80, CD86, iNOS; increased CD206 and Arg1). Co-culture experiments revealed a reciprocal interaction: siPLCγ1 BMSCs polarized BMDMs, which in turn further enhanced BMSC osteogenesis and migration-exceeding the effects of classically induced reparative macrophage. PLCγ1 knockdown increased IL-4 expression and secretion, activating the JAK-STAT signaling pathway. Corresponding JAK2-STAT3 activation and reparative polarization (M2 polarization) were observed in BMDMs treated with siPLCγ1 BMSC CM. We propose that PLCγ1 knockdown in BMSCs promotes reparative polarization via IL-4-JAK2-STAT3 signaling and facilitates bone repair through reciprocal BMSC-macrophage interaction. This study identifies PLCγ1 as a key regulator of BMSC-macrophage crosstalk and a potential therapeutic target for bone regeneration.