Abnormal myofiber morphology and limb dysfunction in claudication

Abstract

Background: Peripheral artery disease (PAD), which affects an estimated 27 million people in Europe and North America, is caused by atherosclerotic plaques that limit blood flow to the legs. Chronic, repeated ischemia in the lower leg muscles of PAD patients is associated with loss of normal myofiber morphology and myofiber degradation. In this study, we tested the hypothesis that myofiber morphometrics of PAD calf muscle are significantly different from normal calf muscle and correlate with reduced calf muscle strength and walking performance.

Methods: Gastrocnemius biopsies were collected from 154 PAD patients (Fontaine stage II) and 85 control subjects. Morphometric parameters of gastrocnemius fibers were determined and evaluated for associations with walking distances and calf muscle strength.

Results: Compared with control myofibers, PAD myofiber cross-sectional area, major and minor axes, equivalent diameter, perimeter, solidity, and density were significantly decreased (P < 0.005), whereas roundness was significantly increased (P < 0.005). Myofiber morphometric parameters correlated with walking distances and calf muscle strength. Multiple regression analyses demonstrated myofiber cross-sectional area, roundness, and solidity as the best predictors of calf muscle strength and 6-min walking distance, whereas cross-sectional area was the main predictor of maximum walking distance.

Conclusions: Myofiber morphometrics of PAD gastrocnemius differ significantly from those of control muscle and predict calf muscle strength and walking distances of the PAD patients. Morphometric parameters of gastrocnemius myofibers may serve as objective criteria for diagnosis, staging, and treatment of PAD.

Keywords: Claudication; Myofiber morphology; Walking distance.

Fig. 1

Normal histology of the gastrocnemius of a control patient. The muscle biopsy was fixed in methacarn, embedded in paraffin, and sectioned at 4 μm. Slide-mounted sections were stained with Masson trichrome and captured with a ×10 objective (panel A). Regions identified with black frames were captured with a ×40 objective and are presented as separate panels (B and C). The myofibers (red), exhibiting a regular polygonal shape, little size variation, and peripherally located nuclei (black), were tightly packed within a thin investment of connective tissue (blue). The white bars represent 100 μm (panel A) and 50 μm (panels B and C). (Color version of figure is available online.)

Fig. 2

Severe myopathy in the gastrocnemius of a patient with PAD. The muscle biopsy was fixed in methacarn, embedded in paraffin, and sectioned at 4 μm. Slide-mounted sections were stained with Masson trichrome and captured with a ×10 objective (panel A). The myofibers (red), exhibiting a nonpolygonal round shape, large variation in size and nuclei (black) often present in the interior, were embedded in an abundant collagenous matrix (blue). Regions identified with black frames were captured with a ×40 objective and are presented as separate panels (B–G; next page). Degradation and loss of myofibers, clearly apparent in panels (B–G), are associated with a remarkable increase of collagen deposition (arrow heads) in the extracellular matrix and the appearance of myofiber vacuoles (filled arrows), target lesions (chevrons), and more centrally located nuclei (unfilled arrows). The white bars represent 100 μm (panel A) and 50 μm (Panels B–G). (Color version of figure is available online.)

Fig. 3

Scatterplots showing correlations between calf muscle strength and myofiber cross-sectional area (μ²), roundness, and solidity in claudicating patients (n = 154). Calf muscle strength increased with increasing myofiber cross-sectional area and solidity and decreased with increasing myofiber roundness.