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Updated: Dec 28, 2025
Mesenchymal Stem Cell Regulation of Macrophage Phagocytosis; Quantitation and Imaging
Published on: July 16, 2021
Macrophages enhance mesenchymal stem cell osteogenesis via down-regulation of reactive oxygen species
Meng-Lin Luo1, Yang Jiao2, Wen-Ping Gong3
1State Key Laboratory of Military Stomatology & National Clinical Research Center for Oral Diseases & Shaanxi Key Laboratory of Oral Diseases, Department of Prosthodontics, School of Stomatology, the Fourth Military Medical University, Xi'an, 710032, PR China.
Objectives:
The role played by macrophages in regulating the differentiation of mesenchymal stem cells (MSCs) during wound healing and bone regeneration is increasingly being recognized. The present study compared the pro-osteogenic effects of three co-culture methods, conditioned medium generated by macrophages (CM), indirect culture (IC) or direct culture (DC) with macrophages, on bone marrow MSCs (BMMSCs).
Methods:
Primary BMMSCs were isolated, characterized and co-cultured with RAW264.7 mouse macrophages. Cell morphology and intracellular reactive oxygen species (ROS) levels were determined by scanning electron microscopy (SEM) and flow cytometry, respectively. Alkaline phosphatase (ALP) staining and assay, Alizarin red staining (ARS) and quantitative real-time polymerase chain reaction (qRT-PCR) were performed to evaluate osteogenic differentiation.
Results:
Inclusion of macrophages in any of the three co-culture methods resulted in improvement in osteogenic differentiation and mineralization of BMMSCs (DC > IC > CM), as measured by ALP staining and activity, ARS and osteoblastic gene expression (Runx2, Alp, Ocn and Bmp2). The enhanced osteogenesis was reversed with hydrogen peroxide. Macrophages reduced the increased levels of intracellular ROS generated by BMMSCs during osteogenic differentiation in a manner similar to the use of an antioxidant, N-acetyl cysteine.
Conclusions:
Macrophages exert an osteogenesis-enhancing effect to accelerate BMMSC osteogenesis via ROS downregulation.
Clinical Significance:
The present findings suggest that targeting MSC-macrophage interaction is an effective strategy for regulating stem cell fate and facilitating bone regeneration.
