Rotator cuff repair is a common surgery in sports medicine. During the surgery, torn tendon was re-fixed onto the bony surface. The majority of patients gain good results. However, re-tear occurs in some patients. The reason under this phenomenon is that the normal tendon-bone enthesis cannot be reconstructed. In order to strengthen the tendon-bone healing and promote enthesis regeneration, numerous manners are tested, among which stem cell related therapies are preferred. Stem cells, due to the ability of multi-lineage differentiation, are widely used in regenerative medicine. However, safety and ethics concerns limit its clinical use. Recent studies found that it is the secretome of stem cells that is biologically effective. On ground of this, we, in the current study, collected the conditioned medium of human bone marrow-derived stem cells (hBMSC-CM) and tested whether this acellular method could promote tendon-bone healing in a rat model of rotator cuff repair. By using histological, radiological, and biomechanical methods, we found that hBMSC-CM promoted tendon-bone healing of the rat rotator cuff. Then, we noticed that hBMSC-CM exerted an impact on macrophage polarization both in vivo and in vitro by inhibiting M1 phenotype and promoting M2 phenotype. Further, we proved that the benefit of hBMSC-CM on tendon-bone healing was related to its regulation on macrophage. Finally, we proved that, hBMSC-CM influenced macrophage polarization, which was, at least partially, related to Smad2/3 signaling pathway. Based on the experiments above, we confirmed the benefit of hBMSC-CM on tendon-bone healing, which relied on its immune-regulative property. Considering the accessibility and safety of acellular hBMSC-CM, we believe it is a promising candidate clinically for tendon-bone healing.