Figure 5
Assessment of ECM protein levels in the cKO mice versus normal mice. (A–H1) immunohistochemistry for DMP1 (A–B1), OPN (C–D1), BSP (E–F1) and FGF23 (G–H1) in the femurs of 4-week-old mice; (A1,B1,C1,D1,E1,F1,G1 and H1) were the higher magnification views of the box areas in A,B,C,D,E,F,G and H, respectively. (A,B,C,D,E,F,G and H) were from consecutive sections. Bars in (A,B,C,D,E,F,G and H): 500 µm; bars in (A1,B1,C1,D1,E1,F1,G1 and H1) 50 µm. (I–K) Western immunoblotting analyses for DMP1, OPN and BSP in the protein extracts from the long bone of 12-week-old mice; equal amounts of total proteins from the normal and cKO groups were loaded for detecting each of the proteins. The cortical bone of the cKO mice (B,B1) contained many areas that lacked anti-DMP1 signals. Immunohistochemistry analyses showed that the levels of OPN, BSP and FGF23 were higher in the cortical bone of the cKO mice than in the normal mice (C,C1,D,D1,E,E1,F,F1,G,G1,H,H1). In the Western immunoblotting analyses, DMP1 band in the cKO group was weaker while that of OPN or BSP was stronger than in the normal mice (I,J,K). The samples in the control lanes were DMP1, OPN and BSP, respectively; these proteins were extracted and purified from the rat long bone35.
