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According to established nomenclature,19 osteoclast surface per bone surface (OcS/BS) and osteoclast number (OcN/BS) were determined from TRACP-stained http://www.selleckchem.com/products/PD-0332991.html sections at the mid-diaphysis. As an index of bone formation, mineralizing surface (MS/BS) was determined from calcein-labeled sections as described previously.5 Because of the scarcity of double calcein labels in the mid-diaphysis, mineral apposition rate or bone-formation rate could not be reliably estimated.20 Right and left femurs were tested in three-point bending to failure using methods described previously.21, 22 Specimens were stabilized over two supports placed 7?mm apart in an Instron 8841 apparatus (Instron, Norwood, MA, USA). A loading force was applied in the anteroposterior direction midway between the two supports by a displacement ramp at a rate of 0.03?mm/s. Force and displacement data were collected at 100?Hz (Labview 5.0, National Instruments, Austin, TX, USA), and http://www.selleck.cn/products/bmn-673.html test curves were analyzed to determine ultimate force to failure and stiffness. Two-way analysis of variance (ANOVA) was used to assess differences for each genotype using treatment and time as the independent variables after testing for normality and equality of variance. When the F value was significant, the Holm-Sidak post hoc test was used for group comparisons at specific time points. Where appropriate, an unpaired Student's t test was used for group comparisons. Data were managed in Microsoft Excel and analyzed using the SigmaPlot 11.0 statistical package (Systat http://www.selleckchem.com/products/Everolimus(RAD001).html Software, Inc., Chicago, IL, USA). Group values are expressed as the mean?��?SEM unless otherwise noted. Consistent with previous reports,5, 6 cKO mice weighed significantly less than WT mice at baseline (27.57?��?1.01?g versus 30.33?��?1.79?g, respectively, p?