Fig. 4. Defects in bone formation and cartilage vascularization in the DKO embryo. (A1-A4,B1-B4,C1-C4) Von kossa staining on longitudinal sections of the ribs (A1-A4), the humeri (B1-B4) and the femurs (C1-C4) from E18.5 embryos of wild type (A1-C1), Ihh^-/- (A2-C2) and DKO (A3-C3,A4-C4). (A1-A3,B1-B3,C1-C3) Images at a low magnification. The ribs (A1-A3) were oriented with the ventral side towards the left, and the humeri (B1-B3) and the femurs (C1-C3) the distal end towards the left. Arrows denote either the normal bone collar (A1-C1), or the corresponding perichondrium where the bone collar is missing (A2-C2,A3-C3). Red asterisks, mineralized cartilage; purple asterisk (A3), marrow cavity. (A4-C4) Images at a higher magnification of boxed areas in A3-C3, respectively. Arrows indicate ectopic bone. Double-headed arrows (B4,C4) indicate the perichondrium separating the ectopic bone from mineralized cartilage. (D1-D3) Hematoxylin and Eosin staining of sections through the equivalent ribs from E18.5 embryos of wild type (D1), Ihh^-/- (D2) and DKO (D3). Double-headed arrows indicate the length of the hypertrophy zone. Asterisks (D1,D3) indicate areas of vascularization.