Purpose: The osteogenic effects of BMPs on mesenchymal stem cells (MSCs) are less profound in human as compared to rodent. The mechanism for this phenomenon is unclear. This study evaluated the effects of macrophages on proliferation and BMP-2 induced osteogenic differentiation of human MSCs.

Method: MSCs were isolated from human bone marrow. Human monocytes THP-1 (human acute monocytic leukemia cell line) were induced into macrophages by phorbol myristate acetate. The conditioned media (CM) from monocytes and macrophages were collected separately. After treated with CM from monocytes or macrophages for 5 and 7 days, the proliferation rate of human MSCs was determined by WST-8 assay. A group without CM served as control. Pretreated human MSCs were then induced towards osteogenic differentiation by osteoinductive medium supplemented with 0.1ug/ml BMP-2. Expression levels of osteogenic markers were determined by real-time quantitative PCR. Alkaline phosphatase (ALP) activity and mineral deposition were assessed by p-NPP colorimetric kinetic assay and calcium assay, respectively.

Results: The number of MSCs was significantly decreased in the group with macrophage CM at both 5 and 7 days (both p< 0.001) as compared with control group, but not in the group with monocytes CM. Expression levels of ALP and bone sialoprotein 2 in the macrophage CM group were significantly lower than those in the control group (p=0.003 and p< 0.001, respectively). ALP activity was also significantly lower in the group with macrophage CM than control group (p< 0.001). Although the expression levels of osteocalcin and RUNX2 as well as calcium deposition in the macrophage CM group were reduced, they did not reach statistical significance.

Conclusion: Macrophages suppressed the proliferation of MSCs and inhibited BMP-2 induced osteogenic differentiation of human MSCs. In addition to known BMP antagonists, macrophages might be another important factor in suppressing the osteogenic effect of BMP-2 on human MSCs.