BACKGROUND: Melatonin plays a role in repairing damaged cartilage and regulating immune cells. The anti-inflammatory effect of Melatonin involves multiple pathways and molecular activation, which directly or indirectly inhibits inflammatory reaction. M2 macrophages have the ability to anti-inflammatory response and repair damaged tissues, secrete IL10 and IL-4, and participate in tissue repair and remodeling. Erk5 is a recently discovered member of the MAPK family and one of the least studied members. It plays an important role in cell differentiation, proliferation, secretion and other functions. This experiment aims to study how Melatonin affects M2 Macrophage polarization and secretion through ERK5 signaling pathway. METHODS: The RAW 264.7 macrophages were used for cell culture. The cells were cultured according to the pre-experimental results. The effects of Melatonin on M2 macrophages were comprehensively evaluated by CCK8 activity detection, RT-PCR, ELISA, cellular immunofluorescence, and WB.SD mice were selected to evaluate the effect of Melatonin on cartilage damage in rats with knee Osteoarthritis through HE staining, immunohistochemistry and immunofluorescence. RESULTS: Melatonin cultivates RAW 264.7 macrophages. Without affecting the polarization ratio of M2 Macrophage polarization, Melatonin may reduce Erk5 gene expression, reduce Erk5 and p-Erk5 protein synthesis, and cooperate with BIX 02189 to enhance the secretion function of existing M2 macrophages and increase the secretion of cytokines IL10. Immunohistochemistry of rat knee Osteoarthritis model confirmed that the expression of IL10 was up-regulated and the synthesis of type II collagen was enhanced, but immunofluorescence found that the polarization of M2 Macrophage polarization in subchondral bone was not obvious. CONCLUSION: Melatonin enhances the ability of M2 macrophages to secrete IL10 by inhibiting Erk5 signaling pathway, but has no effect on M2 Macrophage polarization.