Fig. 3.

Collagen fibers in the soft callus of DIO mice have normal orientation index but aberrant fiber diameter. (A) Representative BSHG images of collagen fibers within the soft callus of either lean or DIO mice. Images were captured at the indicated post-fracture time points. Scale bar = 70μm. (B) Quantitation of collagen volume fraction in BSHG images. Collagen volume fraction within each region of interest (ROI) was determined by normalizing the sum of BSHG voxels to the total ROI voxels. Collagen fraction volume was determined in 6 lean and 6 DIO callus tissues; 3 sections were analyzed in each sample, and enough ROIs were selected within each section to cover the entire studied area. (C) Representative examples of Fourier Transform (FT) power spectra generated by analyzing BSHG images using MATLAB. (D) Quantitation of orientation index of the soft-callus fibrillar collagen computed from FT power spectra shown in (C). Orientation index was calculated in 6 lean and 6 DIO callus tissues; 3 sections were analyzed in each sample, and 5 ROIs were selected within each section. (E) Quantitation of collagen-fiber diameter in the same ROIs utilized to compute orientation index in (D).

For all analyses, IF images shown in Fig. 2A were utilized as a guide to select ROIs where Col II, but not Col I, was expressed (exemplified by white arrows in Fig. 2A). All bar graphs represent average ± SEM. (*) P < 0.05; (**) P < 0.01 using student t-test and time-matched lean callus as a control.