Automated Methods for the Analysis of Skeletal Muscle Fiber Size and Metabolic Type - International Review of Cell and Molecular Biology - page 298

Table 7.2 Validation of CSA calculations by CyteSeer ® —cont'd

CSA
(CyteSeer ® )

CSA
(Image J)

% Difference
(from Image J)

Muscle sample

Monkey VL ( N ¼ 207) 8385 8414 0.34

Monkey VL ( N ¼ 234) 8263 8573 3.62

Monkey VL ( N ¼ 72) 11,943 12,065 1.02

Average 10399.4 10512.7 5.26

SD 2290.2 2758.2 3.98

a GTN, SOL, PLT, and VL denote gastrocnemius, soleus, plantaris, and vastus lateralis, respectively.
Images from skeletal muscle tissue sections were analyzed with CyteSeer ® (utilizing the Skeletal Muscle
Algorithm) and in a semiautomated fashion utilizing Image J. The number of fibers analyzed per sample is
shown ( N ).

upregulated in denervated skeletal muscle fibers. Progressive denervation
and atrophy of skeletal muscle fibers commonly occur during aging
( Doppler et al., 2008; Fidzianska and Kaminska, 1995; Grumbles et al.,
2008; Kostrominova et al., 2005; Lapalombella et al., 2008 ). To analyze
the sample, laminin was visualized in the green fluorescent channel and
NCAMwas visualized in the red channel. Notably, fibers that displayed pos-
itive labeling with NCAM averaged just 22% of the CSA as the NCAM
negative fibers ( Fig. 7.9 ) consistent with previous observations that
denervated atrophic muscle fibers have upregulated expression of NCAM.
This example illustrates the generality of the overall approach, as the fiber
outlines identified from the laminin channel could be used to quantify
one or more biomarkers on a fiber-by-fiber basis, utilizing the separate opti-
cal channels afforded by fluorescence microscopy.

6.1.4 Analysis of muscle fiber damage
During the etiology of DMD, muscle fibers become damaged, undergo
degeneration, and subsequently are replaced by regenerating fibers. In these
cases, the muscle fiber outlines are often complex and irregular. Similar mor-
phological changes can be obtained in mouse models by the injection of cer-
tain venoms or anesthetics, which initiate fiber degradation and subsequent
regeneration ( D'Avila et al., 2006; Ryall et al., 2008 ). The irregularities in
the fibers of damaged muscle represent a challenge to automated image-
analysis algorithms. To illustrate this difficulty, a tissue section from a
mouse injected with snake venom was analyzed with CyteSeer ® 's Skeletal