Figure 8: YKL-05-099 increases bone formation and bone mass in vivo.

(a) Ocy454 cells were treated with the indicated doses of YKL-05-093, YKL-05-099 or PTH for 20 min. Whole cell extracts were generated, followed by immunoblotting. (b) Control or shSIK2/3 Ocy454 cells were treated with YKL-05-093 or YKL-05-099 (1 μM) for 4 h. RNA was prepared, and gene expression analysed by RT-qPCR. Both YKL-05-093 and YKL-05-099 regulate SOST and RANKL expression in control, but not SIK2/3-deficient cells. (c) Male mice of 8 week age (n=5 per group) were treated with a single IP dose of YKL-05-099 (20 μmol kg⁻¹) or vehicle. After 2 h, animals were killed, RNA was prepared from femurs, and gene expression analysed by RT-qPCR. SOST down-regulation was observed in response to YKL-05-099, but the P-value for this difference was 0.105. * indicates P<0.01. (d) Male mice of 8 week age were treated with vehicle (n=8) or YKL-05-099 (n=7, 10 umol kg⁻¹, IP) once daily 5 days per week for 2 weeks. Animals were killed 2 h after the final dose, and RNA from femurs analysed for the indicated genes. BGLAP encodes osteocalcin. * indicated P<0.01 versus vehicle, ^# indicates P<0.05 versus vehicle. (e–k), static and dynamic histomorphometry were performed on the tibia from the same mice as in d. Each data point represents an individual mouse; P-values for each difference are shown on the graph. (l) Representative trichrome-stained photomicrograph showing increased osteoblasts on cancellous bone surfaces from YKL-05-099-treated mice. Scale bar, 20 μm. (m) Dual calcein/demeclocycline images demonstrating increased mineralizing surface in YKL-05-099-treated mice.