Sclerostin Antibody Treatment Increases Bone Formation, Bone Mass, and Bone Strength of Intact Bones in Adult Male Rats

Abstract

We investigated the systemic effect of sclerostin monoclonal antibody (Scl-Ab) treatment on intact non-operated bones in an open osteotomy male Sprague Dawley (SD) rat model. Six-month-old male SD rats were subjected to transverse osteotomy at the right femur mid-shaft. Rats were injected subcutaneously with vehicle or Scl-Ab (25 mg/kg, 2 times per week) treatment for 9 weeks. Compared with vehicle control, Scl-Ab treatment significantly improved trabecular and cortical bone mass and microarchitecture at L5 vertebrae and left femora by micro-CT at week 6 and 9. Mechanical testing showed that Scl-Ab treatment resulted in significantly higher stiffness, energy to failure and ultimate load at the femora at week 9. Mineral apposition rate, mineralizing surface and bone formation rate on the trabecular bone in the distal femora was significantly increased in Scl-Ab group at week 6 and 9. The administered Scl-Ab was localized in the osteocytes and beta-catenin was strongly expressed in osteoblasts. Scl-Ab treatment significantly increased serum P1NP level and there was no between-group difference in serum level of CTX-1. In conclusion, Scl-Ab treatment could induce rapid and sustained increase in bone formation, bone mass and bone strength in non-operated bones. Sclerostin inhibition might be advantageous to prevent secondary fracture(s).

Figure 1. Representative micro-CT images of the L5 vertebrae of Scl-Ab treatment and the vehicle treatment.

Scl-Ab treatment significantly increased cortical thickness and trabecular thickness of the L5 vertebrae.

Figure 2. Micro-CT analysis of cortical and trabecular bone of the femora.

(A) Region of interest (ROI) chosen for analysis including cortical compartment of the femoral midshaft and trabecular compartment of distal femoral metaphyseal. (B) Representative 3D micro-CT images of Scl-Ab and vehicle treated samples. Scl-Ab treatment significantly increased cortical thickness at the femoral midshaft and trabecular bone volume and thickness at the distal femoral metaphyseal at week 6 and 9.

Figure 3. Mechanical test of the femora.

Three-point bending test showed that Scl-Ab treatment significantly increased ultimate load (A) and energy to failure (B) at week 6, and significantly increased stiffness (C) at week 9 compared with the vehicle group at the same time point. Bars represent Mean ± SD. *P < 0.05; **P < 0.01 compared with vehicle at the same time point.

Figure 4. Detection of the administered Scl-Ab and immunohistochemistry of beta-catenin.

(A) The administered Scl-Ab was concentrated in the osteocytes (arrows). Magnification: 400x. (B) Scl-Ab treatment induced the expression of beta-catenin in osteoblasts (arrowheads). Magnification: 400x.

Figure 5. Bone formation assessed by fluorescent labeling.

Scl-Ab treatment significantly enhanced new bone formation in the metaphyseal trabecular bone region of the distal femora (A) and increased the mineralizing surfaces (B). Magnification: 200x (A) and 25x (B).