Quantitative Analysis of Muscle Histologic Method in Rodent Facial Nerve Injury

Figure 1. After Masson trichrome staining (MTS) of tissue sections containing dilator naris muscle and levator labii superioris muscles, a sequential color deconvolution algorithm was applied, using Image J software, to measure muscle content of the specimens, fiber surface area, and fiber counts. A through C, This example of a control specimen at ×20 magnification after MTS demonstrated distinct 3-colored staining patterns. B, The entire specimen was then outlined with the Image J software.¹² C, The muscle area (red stain) was selectively outlined through color-based image thresholding. D, The same specimen is shown at ×400 magnification. E, Individual muscle fiber staining is selectively outlined using the same thresholding approach.

Figure 2. Mean maximal horizontal whisking amplitude at postrepair day 28. Recovery was significantly greater for the control side (58.5°) compared with the experimental side (17.1°). Limit lines indicate 95% confidence intervals.

Figure 3. Stereotypical features of muscle atrophy seen in dilator naris muscle and levator labii superioris on the experimental side. A and B, Control side reveals healthy features of muscle fibers, including polygonal shape with overall uniform fiber size. C and D, Experimental side reveals decreased volume of muscle replaced by collagen (blue stain) and adipose tissue (white space with septation). Individual fibers display irregular round appearance (original magnification ×20 [A and C] and ×400 [B and D]).

Figure 4. Quantitative comparison of muscle architectures. A and B, Statistically significant differences in muscle content and total fiber count were noted between the control and experimental sides. C, The mean cross-sectional area of individual muscle fibers was larger on the control side; however, this difference was not statistically significant. Limit lines indicate 95% confidence intervals.