Fig. 2

Patient’s muscle biopsy. a–c Hematoxylin and eosin staining demonstrating a chronic myopathic process with wide-ranging muscle fiber size variability, fiber splitting, internalized nuclei, and increases in endomysial fatty connective tissue. b Rimmed vacuoles are seen in occasional muscle fibers (black arrows). c Atrophic fibers showing large subsarcolemmal nuclei (black arrow) or eosinophilic inclusions (blue arrows), which stain dark red (d) in trichrome and red (e) in acid phosphatase. f A higher magnification trichrome stained section shows that the large dark red central inclusions have a filamentous appearance and tiny rod-like structures. g Such filamentous tangles contain abundant alpha-actinin and h less myotilin, but not i p62, j desmin or k alphaB-crystallin, all of which are diffusely and often faintly increased (light brown) in some atrophic fibers. l MyHC-IIA immunohistochemical staining demonstrates paucity and atrophy of type 2A fibers, which harbor (m) filamentous tangles containing MyHC-IIA, as also demonstrated by (n) Z-stacked confocal microscopy in an immediately adjacent muscle section (bar = 50 microns). o NADH dehydrogenase reacted section showing numerous ring fibers (one is shown in the right upper corner at higher magnification, 40×) and overreactive (dark blue) atrophic fibers. Primary mouse anti-human MyHC-IIA monoclonal antibody [9, 35] (A4.74, 1:100 dilution, Developmental Studies Hybridoma Bank (DSHB) (University of Iowa, Iowa City, Iowa, USA) was visualized with Cy3-conjugated affinity purified donkey anti-mouse IgG (1:200, Millipore Chemicon, Temecula, California USA)