The size and shape of skeletal muscle fibers are affected by various physiological and pathological conditions, such as muscle atrophy, hypertrophy, regeneration, and dystrophies. Hence, muscle fiber cross-sectional area (CSA) is an important determinant of muscle health and plasticity. We adapted the Imaris software to automatically segment muscle fibers based on fluorescent labeling of the plasma membrane, and measure muscle fiber CSA. Analysis of muscle cross sections by the Imaris semi-automated and manual approaches demonstrated a similar decrease in CSA of atrophying muscles from fasted mice compared with fed controls. In addition, we previously demonstrated that downregulation of the Ca²⁺-specific protease calpain-1 attenuates muscle atrophy. Accordingly, both the Imaris semi-automated and manual approaches showed a similar increase in CSA of fibers expressing calpain-1 shRNA compared with adjacent non-transfected fibers in the same muscle cross section. Although both approaches seem valid for measurements of muscle fiber size, the manual marking method is less preferable because it is highly time-consuming, subjective, and limits the number of cells that can be analyzed. The Imaris semi-automated approach is user-friendly, requires little training or optimization, and can be used to efficiently and accurately mark thousands of fibers in a short period of time. As a novel addition to the commonly used statistics, we also describe statistical tests that quantify the strength of an effect on fiber size, enabling detection of significant differences between skewed distributions that would otherwise not be detected using typical methods.

中文翻译：

---

使用 Imaris 软件半自动测量肌纤维大小

骨骼肌纤维的大小和形状受各种生理和病理条件的影响，如肌肉萎缩、肥大、再生和营养不良。因此，肌纤维横截面积 (CSA) 是肌肉健康和可塑性的重要决定因素。我们采用 Imaris 软件根据质膜的荧光标记自动分割肌纤维，并测量肌纤维 CSA。通过 Imaris 半自动和手动方法对肌肉横截面的分析表明，与进食对照相比，禁食小鼠的萎缩肌肉的 CSA 有类似的降低。此外，我们之前证明了 Ca ^{2+ 的}下调-特异性蛋白酶 calpain-1 减轻肌肉萎缩。因此，与相同肌肉横截面中的相邻非转染纤维相比，Imaris 半自动和手动方法都显示出表达 calpain-1 shRNA 的纤维的 CSA 类似的增加。虽然这两种方法似乎都适用于测量肌肉纤维大小，但手动标记方法不太可取，因为它非常耗时、主观，并且限制了可以分析的细胞数量。Imaris 半自动化方法用户友好，几乎不需要培训或优化，可用于在短时间内高效准确地标记数千根纤维。作为常用统计数据的新补充，我们还描述了量化对纤维尺寸影响强度的统计测试，