S100A8/A9 drives monocytes towards M2-like macrophage differentiation and associates with M2-like macrophages in osteoarthritic synovium

Objectives Macrophages are key orchestrators of the osteoarthritis (OA)-associated inflammatory response. Macrophage phenotype is dependent on environmental cues like the inflammatory factor S100A8/A9. Here, we investigated how S100A9 exposure during monocyte-to-macrophage differentiation affects macrophage phenotype and function.Methods OA synovium cellular composition was determined using flow cytometry and multiplex immunohistochemistry. Healthy donor monocytes were differentiated towards M1- and M2-like macrophages in the presence of S100A9. Macrophage markers were measured using flow cytometry, and phagocytic activity was determined using pHrodo Red Zymosan A BioParticles. Gene expression was determined using qPCR. Protein secretion was measured using Luminex multianalyte analysis and ELISA.Results Macrophages were the dominant leucocyte subpopulation in OA synovium. They mainly presented with an M2-like phenotype, although the majority also expressed M1-like macrophage markers. Long-term exposure to S100A9 during monocyte-to-macrophage differentiation increased M2-like macrophage markers CD163 and CD206 in M1-like and M2-like differentiated cells. In addition, M1-like macrophage markers were increased in M1-like, but decreased in M2-like differentiated macrophages. In agreement with this mixed phenotype, S100A9 stimulation modestly increased expression and secretion of pro-inflammatory markers and catabolic enzymes, but also increased expression and secretion of anti-inflammatory/anabolic markers. In accordance with the upregulation of M2-like macrophage markers, S100A9 increased phagocytic activity. Finally, we indeed observed a strong association between S100A8 and S100A9 expression and the M2-like/M1-like macrophage ratio in end-stage OA synovium.Conclusion Chronic S100A8/A9 exposure during monocyte-to-macrophage differentiation favours differentiation towards an M2-like macrophage phenotype. The properties of these cells could help explain the catabolic/anabolic dualism in established OA joints with low-grade inflammation.