Comparison of Histopathologic Criteria and Skeletal Muscle Fixation Techniques for the Diagnosis of Polysaccharide Storage Myopathy in Horses

Abstract

The purpose of the study reported here was to determine the effect of three methods of fixation of skeletal muscle biopsy specimens on the histopathologic appearance of muscle sections and to determine criteria that were most consistently associated with a diagnosis of polysaccharide storage myopathy (PSSM) in horses. Surgically excised semimembranosus muscle biopsy specimens were obtained from nine horses previously diagnosed with PSSM and from 15 control horses. Portions of each specimen were fixed in formalin, frozen immediately, and chilled for 24 hours prior to freezing. Sections stained with hematoxylin and eosin (HE), periodic acid-Schiff (PAS), and amylase-PAS were scored for histopathologic criteria by three investigators blinded to the sample origin. The presence of amylase-resistant, abnormal polysaccharide was found to be the most sensitive and specific diagnostic indicator for PSSM, and was readily detected regardless of the fixation technique or investigator. Other less-specific features associated with PSSM included atrophy and cytoplasmic and subsarcolemmal vacuoles; however, their histologic scores varied among fixation technique and investigators. Scores for subsarcolemmal and cytoplasmic amylase-sensitive glycogen in horses with PSSM were similar to those for control horses and varied among fixation techniques. In conclusion, PSSM is most accurately diagnosed in muscle biopsy specimens on the basis of appearance of amylase-resistant, abnormal polysaccharide, not amylase-sensitive glycogen, regardless of fixation technique. In general, frozen sections appeared to be better suited for studying myopathies because many histopathologic features of skeletal muscle were obscured by formalin fixation.