Fig. 3. Short-range anchoring tendons and ligaments are not affected in Scx^-/- mice. (A-D) Frontal sections through the rib cage of wild-type (WT) and Scx^-/- embryos at E18.5 stained with a myosin heavy chain (MHC; red) antibody to detect intercostal muscle attachments to the ribs. (A,B) ScxGFP signal (green) at the intercostal muscle attachments. (C,D) Section in situ hybridization (ISH) detecting collagen I expression in rib osteoblasts and in the tendinous attachment layer (yellow arrowheads) of wild-type and mutant embryos. (E,F) Sagittal sections through the knees of wild-type and Scx^-/- embryos at E18.5. Collagen I was detected by section ISH followed by antibodies to MHC (red) and collagen II (green). Fluorescent antibody signals were merged with the ISH image in Photoshop. Yellow arrowheads show the cruciate ligaments of the knee; black arrowheads show the patellar ligament; purple arrowheads show the rectus femoris tendon and vastus medialis tendon. (G-I) Superficial sagittal sections through the tail of wild-type and Scx^-/- embryos at E18.5 that are carrying the ScxGFP reporter. (G) The cartilage of tail bones visualized by DIC optics and overlaid with ScxGFP signal in the annulus fibrosis. (H,I) ScxGFP signal in the annulus fibrosis at high magnification. (J-L) Transmission electron microscope images of a medial sagittal section through the tail of an E18.5 Scx^-/- embryo. (J) Low magnification reveals the nucleus pulposus (NP; the intervertebral disc), which is flanked laterally by cartilage cells and by dorsal and ventral tightly stacked layers of ligament cells of the annulus fibrosus (AF, red arrowheads). (K,L) The annulus fibrosus at a higher magnification. The collagen fibers (purple arrowheads) are aligned along the section plane (K) and perpendicular to the section plane (L). AF, annulus fibrosus; NP, nucleus pulposus; P, patella; Rf, rectus femoris muscle; Vm, vastus medialis muscle.