PMC10043206_01
Magnetic resonance imaging (MRI) images of the patient. Muscle MRI shows the geographic distribution of muscular fatty infiltration. The predominantly affected muscles included the quadriceps femoris and posterior compartment in the thighs (A) and medial gastrocnemius with sparing of anterior tibialis in the lower legs (B). No muscle edema was seen in the proband (C, D).
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F1
MRI of the thighs indicated significant fatty replacement on the right side, which was more prominent in the posterior compartment; only the medial heads of the gastrocnemius and right soleus revealed mild degenerative changes in the lower legs (Figure 1).
Muscle biopsy of the biceps brachii muscle demonstrated a rimmed vacuolar myopathy. Light microscopy of muscle biopsy shows size variability, excess internal nuclei, necrotizing fiber, degenerating fibers, basophilic deposition (*), and rimmed vacuoles (arrowhead) under hematoxylin and eosin staining (A). Gomori trichrome staining revealed rimmed vacuoles (arrowhead) and darkly stained materials (*) (B). Nicotinamide adenine dinucleotide dehydrogenase-tetrazolium reductase staining revealed decreased oxidative enzyme reactivity (arrow) and darkly stained materials (arrowhead) in some myofibers (C). Combined cytochrome c oxidase (COX)/succinate dehydrogenase (SDH) stain demonstrates several COX-negative fibers (arrows) (D). The fibers show the checkerboard-like distribution in ATPase 4.5 (E) and ATPase 9.4 stain (F).
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F2
Left biceps' biopsy showed an extensive presence of basophilic deposition and rimmed vacuoles (Figure 2A). Rimmed vacuoles were further confirmed using Gomori trichrome staining, and darkly stained materials were remarkable (Figure 2B); fibers focally devoid of enzyme activity were observed in NADH-stained sections (Figure 2C). Several fibers showed absent COX activity (Figure 2D). No fiber-type grouping was observed via ATPase 4.5 and 9.4 staining (Figures 2E, F).
Immunofluorescence staining reveals SQSTM1-positive inclusions in rimmed vacuole marked with arrows.
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F3
Immunohistochemistry revealed that the cytoplasmic inclusions were p62/SQSTM1-positive (Figure 3).
Chromatograms showing sequencing analysis of heterozygous frameshift mutation c.542_549delACAGCCGC (p. H181Lfs*66) in SQSTM1 in the proband and her parents' sequences. The red frame was the deletion part of the proband (A). Schematic view of p62/SQSTM1 functional domains. PB1, Phox and Bem1; ZZ, zinc finger motif; TRAF6, TNF receptor-associated factor 6; PEST, proline, glutamic acid, serine, and threonine domain; LIR, LC3 interaction region; KIR, KEAP1 interaction region; UBA, ubiquitin association domain. The amino acid number indicates the proband's mutation (B).
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F4
Neither of the healthy parents was a carrier, suggesting that it was a de novo mutation (Figure 4A).