Tohoku J. Exp. Med., 2010, 221(1)

BMP-7 in Combination with Estrogen Enhances Bone Formation in a Fracture Callus Explant Culture

AIQUN WEI,¹ ANTHONY LEONG,¹ LISA WILLIAMS,¹ SYLVIA CHUNG,¹ BOJIANG SHEN,¹ DIVYA BHARGAV¹ and ASHISH D. DIWAN¹

¹Orthopaedic Research Institute, St George Hospital Campus, University of New South Wales, Sydney, NSW, Australia

In postmenopausal women, estrogen withdrawal results in decrease in bone density or osteoporosis. Osteoporosis leads to fracture and retards bone-healing response. Bone morphogenetic protein-7 (BMP-7), a member of the transforming-growth factor-β superfamily, has been shown as a promising candidate that stimulates bone growth in its application to fracture healing. The purpose of this study was to determine whether BMP-7 could enhance bone formation in the absence of estrogen. Female rats underwent a controlled closed fracture at the midshaft of the right femur. The callus tissues were harvested from the fracture site eight days following the fracture, and were cultured in serum-free media. The explanted callus tissues were then treated with BMP-7, estrogen (E2) or both. We assessed bone formation by measuring alkaline phosphatase (AP) activity, expression of an osteogenic transcription factor, Runt-related transcription factor-2 (Runx2), production of nitric oxide (NO), and calcium mineralization. Supplementation of serum-free cultures with BMP-7 alone increased cell proliferation by twofold, caused a 6.5-fold increase in AP activity, and enhanced calcium mineralization after 48 h. Moreover, BMP-7 in combination with E2 caused a 8.2-fold increase in the AP activity. Runx2 protein expression was increased following stimulation with BMP-7 and E2. Interestingly, E2 induced the amount of NO production by twofold, whereas BMP-7 did not, either alone or with E2. Thus, BMP-7 could enhance early and late markers of bone fracture healing in callus explant cultures, except for NO. BMP-7 could be a promising growth factor in the treatment of fractures as a consequence of osteoporosis.

keywords —— osteoporosis; callus explant culture; BMP-7; estrogen; alkaline phosphatase; fracture healing

© 2010 Tohoku University Medical Press

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Tohoku J. Exp. Med., 2010, 221, 61-68

Correspondence: Ashish D. Diwan, Orthopaedic Research Institute and Department of Orthopaedic Surgery, St George Hospital Sydney, Kogarah, NSW 2217, Australia.

e-mail: a.diwan@spine-service.org