Osteoinductivity of demineralized bone matrix in immunocompromised mice and rats is decreased by ovariectomy and restored by estrogen replacement

Abstract

The osteoinduction potential of human demineralized bone matrix (DBM) in females with low estrogen (E 2) is unknown. Moreover, the osteoinductivity of commercial human DBM is tested in male athymic rats and mice, but DBM performance in these animals may not reflect performance in female animals or provide information on E 2's role in the process. To gain insight, human DBM was implanted bilaterally in the gastrocnemius of twenty-four athymic female mice (10 mg/implant) and twenty-four athymic female rats (15 mg/implant). Eight animals in each group were sham-operated (SHAM), ovariectomized (OVX), or ovariectomized with E 2-replacement (OVX + E 2) via subcutaneous slow release capsules of 17β-estradiol. OVX and OVX + E 2 animals were pair-fed to SHAM animals. Four animals from each group were euthanized at 35 days and four at 56 days. Animal weight, uterine weight, and blood estrogen levels confirmed that pair feeding, ovariectomy, and E 2 replacement were successful. Histological sections of implanted tissues were evaluated qualitatively for absence or presence of DBM, ossicle formation, and new bone or cartilage using a previously developed qualitative scoring system (QS) and by histomorphometry to obtain a quantitative assessment of osteoinduction. OVX mice had a small but significant QS decrease at 35 days compared to SHAM mice, confirmed by quantitative measurement of ossicle, marrow space, and new bone areas. The QS in rats was not affected by OVX but histomorphometry showed decreased new bone in OVX rats, which was restored by E 2. The QS indicated that the number of new bone sites was not reduced by OVX in rats or mice at 56 days, but the relative amount of new bone v. marrow space was affected and differed with animal species. Residual DBM was less in OVX animals, indicating that DBM resorption was affected. Cartilage was present in rats but not in mice, suggesting that endochondral ossification was slower and indicating that bone graft studies in these species are not necessarily comparable. These results show the importance of E 2 in human DBM-induced bone formation and suggest that E 2 may be needed for clinical effectiveness in post-menopausal women.