Figure 6. Collagen architecture determined using second harmonic generation (SHG) microscopy.

A, example SHG images of a single slice of wildtype and mdx EDL muscles at 870 nm. B, a max projection of the stack with a colour map of the collagen fibre angles from OrientationJ from the same wildtype and mdx images as in (A). C, the number of collagen fibres visible with SHG was not significantly different in either muscle for mdx. The EDL had significantly more collagen fibres than soleus. D, collagen deviation was similar between EDL and soleus muscles. The mdx muscles had significantly lower deviation angle than the wildtype mice, with post hoc tests significant in soleus. E, sarcomere length determined from Fourier transform of SHG images. There is no significant difference in sarcomere length between wildtype and mdx mice for either muscle, but soleus had significantly longer sarcomere lengths than EDL. F, the angle between mean collagen angle and the myofibre angle was generally small (<10°) and did not significantly vary across muscle groups. G, all groups combined had a significant negative correlation of sarcomere length and collagen deviation angle. The soleus muscle had an independent significant negative correlation. H, there was a significant positive correlation between collagen fibre area and stiffness across all groups. I, there is also a relationship between collagen deviation and elastic stiffness. All groups combined had a significant negative correlation and the EDL muscle also had a significant negative correlation independently. *P < 0.05. Scale bar = 100 μm.