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DYSTROPHINOPATHIES: Duchenne
DMD Muscle pathology: General features
- Myopathy: Ongoing & Chronic
- Fiber sizes: Varied
- Small fibers: Round
- Hypercontracted (opaque) muscle fbers
- Necrosis & Regeneration of muscle fibers
- More early in disease course
- Patterns
- Muscle fiber pathology
- "Delta lesions"
- Internal architecture
- Immaturity (2C fiber type): Many fibers
- Hypercontracted fibers
- Chronic changes: Other tissues
- Endomysial connective tissue: Increased
- Fat accumulation
- Locations: Endomysial & Perimysial
- Molecular
- Dystrophin staining
- Screening Antibodies: Target Epitopes
- N-terminus
- Rod domain
- 7G1: vs Exon 46 (Within deletion hotspot)
- 6A9: vs Exon 50 (Within deletion hotspot)
- Dys-1: vs Exons 26 to 30
- C-terminus
- Dys-2: vs Exon 79
- Less useful
- Few deletions in this region
- May be reduced in other myopathies
- Duchenne MD clinical phenotype: Staining patterns
- DMD Muscle
- Dystrophin: Absent staining for all epitopes
- Utrophin: Normal (Absent on muscle fibers) or Increased staining
- nNOS: Often absent with Exon 45-51 hotspot deletions
- Sarcoglycans: Often reduced in Duchenne MD
- Schwann cells: May stain for dystrophin with mutations proximal to Intron 55
- Becker MD clinical phenotype: Staining patterns
- Overall reduced staining for all antibodies
- Relative reduction in intensity of staining for some dystrophin epitopes
- Complete loss of some dystrophin epitopes
- Dystrophin female carriers
- Non-necrotic, dystrophin-negative muscle fibers
- Other sarcolemma-related proteins
- Sarcoglycans: Reduced
- Aquaporin 4: Reduced
- Laminin-α2: Normal or Increased
- Utrophin: Increased
- β-Spectrin: Normal
- Calcium
1
- Increased in myoplasm
- Pathomechanisms of Ca++ increase in DMD muscle fibers
- Enhanced Ca++ leak from sarcoplasmic reticulum (SR)
- Through oxidized Ryanodine receptor type 1 (RyR1) Ca++ release channels
- Hypernitrosylation of cysteine residues in RyR1
- FKBP1A
dissociation from RyR1
- Destabilization of RyR1 channel closed state
- Increased RyR1-dependent SR Ca++ leak
- Uncontrolled production of reactive species
- Oxygen (ROS)
- Nitrogen (RNS)
- Reduced activity of SR/ER Ca++ ATPase(SERCA) (Ca++pump)
- Excessive extracellular Ca++ trans-sarcolemmal influx
- Micro-tears: Delta lesions
- Abnormal Ca++-permeable ion channels
- Enhanced store-operated Ca2+ entry (SOCE)
- Related molecules
- SOCE trigger: Depletion of intracellular Ca++ stores
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From: AG Engel
Delta Lesions (Arrow)
Description: Focal wedge-shaped lesions
Locations: Subsarcolemmal regions of muscle fibers
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DMD
Muscle fibers
Size: Varied
Shape: Small fibers round or polygonal
Internal nuclei
Hypercontracted
Necrosis
Endomysial connective tissue: Increased between muscle fibers
Fat: Increased in endomysium & preimysium

H&E stain
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DMD Endomysial connective tissue: Increased between muscle fibers

VvG stain
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NADH stain
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DMD: Immature muscle fibers
Coarse internal architecture on NADH (Above)
2C fibers: Intermediate staining on ATPase pH 4.3 (Below)

ATPase pH 4.3 stain
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Duchenne Muscular Dystrophy: Early Pathology

H & E stain
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Muscle fibers
Sizes: Varied; Small fibers are rounded or polygonal; Occasional hypertrophic fiber
Necrosis (Left): Fibers scattered & in small groups
Myopathic groups (Below): Clusters of small necrotic (Black arrow) & regenerating (White arrow) muscle fibers
Internal nuclei: Occasional
Endomysial connective tissue: Normal to mildly increased
Perimysium: Early replacement by fat
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Acid phosphatase stain
Acid phosphatase + cells
Endomysium: Small Scattered cells
Necrotic muscle fibers: Clusters of phagocytic cells
Ring fibers (Arrow)
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Acid phosphatase stain
Acid phosphatase + cells
Necrotic muscle fibers: Clusters of phagocytic cells
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DMD: Muscle Fiber Necrosis
Necrotic Fibers: Scattered; Varied stages

H&E stain
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Gomori trichrome stain
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Necrotic Fibers: Scattered; Replaced by cells

VvG stain
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Myopathic Grouping
Myopathic grouping: Necrotic muscle fibers, clustered

H&E stain
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Necrosis, Early stage
Necrotic muscle fibers: Pale stained
Macrophages: Early invasion of muscle fibers

NADH stain
Necrotic muscle fibers are pale on NADH stain
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Myopathic grouping: Clusters of very small regenerating muscle fibers & histiocytic cells

VvG stain
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Myopathic groups: Clusters of small muscle fibers in similar, early, stage of regeneration or immaturity

VvG stain
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Myopathic groups: Clusters of small muscle fibers in similar, early, stage of regeneration or immaturity

H & E stain
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ATPase, pH 4.3
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NADH
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Clusters of small cells
Many stain for Acid phosphatase

Acid phosphatase
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Acid phosphatase
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H & E stain
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Myopathic Grouping: Late

H & E stain
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Myopathic grouping: Intermediate-sized, Immature muscle fibers in clusters

H & E stain
Immature fibers are small & basophilic
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ATPase, pH 4.3
Immature fibers are: Small & 2C (Intermediate staining)
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Alkaline phosphatase
Immature small fibers have: Cytoplasmic staining
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NADH
Immature fibers have coarse cytoplasmic staining
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NADH
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VvG
Immature small fibers have coarse cytoplasmic staining
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Immature muscle fibers: Many
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Staining properties of immature muscle fibers includes:
1. H & E (left): Basophilic fibers
2. Alkaline phosphatase positive (center)
3. 2C fibers: Intermediate staining on ATPase pH 4.3 (right)
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Duchenne Muscular Dystrophy: Severe at age 2 years

H&E stain
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DMD
Endomysial connective tissue: Increased
Muscle fibers
Necrosis & Regeneration
Size: Varied
DMD
Endomysial connective tissue: Increased
Muscle fibers
Necrosis & Regeneration
Size: Varied

Gomori trichrome stain
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Gomori trichrome stain
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VvG stain
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DMD
Many intermediate-stained muscle fibers

ATPase pH 4.3 stain
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Duchenne Muscular Dystrophy: Later Pathology (10 years)

H&E stain
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DMD: Later pathology
Fiber size: Varied
Endomysial connective tissue: Increased
Fat replacement of muscle: Prominent

H&E stain
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VvG stain
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Sudan stain
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Replacement of Muscle fibers & Perimysial connective tissue by Fat (Dark black)

Sudan stain
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H&E stain
Endomysial connective tissue: Increased
Fiber size: Variable
Small fibers: Rounded
Large or hypercontracted muscle fibers: Scattered
Necrotic fibers (Arrow): Scattered
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H&E stain

Endomysial connective tissue Increased between fibers
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Muscle fiber size: Varied
Small & Large fibers: May be are either type (I = Dark; II = Light)
Many type IIC fibers: Smaller size; Intermediate stain

ATPase pH 4.3 stain
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DMD late: Few necrotic muscle fibers

Acid phosphatase stain
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Muscle fiber internal architecture: Coarse

NADH stain
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DMD: Ring Fibers

H&E stain
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Gomori trichrome stain
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NADH stain
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DMD: Ring fibers

NADH stain
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Dystrophin staining
Normal
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Duchenne Muscular Dystrophy:
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Normal dystrophin staining
around the rim of muscle fibers.
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Absent dystrophin: Duchenne muscular dystrophy
Left: No staining around the rim of any muscle fibers
Right: No staining of most muscle fibers
One "revertant" fiber with dystrophin staining.
Revertant fibers reflect a somatic mutation allowing dystrophin expression
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Normal: Dystrophin present near surface of muscle fibers

Dys1 stain
Duchenne MD: Dystrophin absent from surface of muscle fibers

Dys1 stain
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Muscle from Duchenne MD male with large area of revertant muscle fibers
Muscle histology (Above): One fascicle (Arrow) has more normal muscle fibers & connective tissue amid otherwise myopathic muscle
Dystrophin staining (Below): Muscle fibers in preserved fascicle are revertants with dystrophin present around their rim
Some revertant muscle fibers are present in myopathic regions as well.
Western blot: Dystrophin from dystrophinopathies

from Novocastra
Lane 1: Becker dystrophy;
Dystrophin has reduced abundance but normal size.
Lane 2: Becker dystrophy; Dystrophin has reduced size and abundance.
Lane 3: Normal; Dystrophin has normal size and amount.
Lane 4: Duchenne dystrophy; Almost no protein is present.
Lane 5: Duchenne outlier; Dystrophin has severely reduced abundance.
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DMD muscle: After dystrophin mini-gene transfer

Dys3 antibody
Stains dystrophin N-terminal epitope present on transferred mini-protein
Present, irregularly, on surfaces of minority of muscle fibers
Most dystrophin positive muscle fibers are small & intermediate sized
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Dys2 antibody
Stains dystrophin C-terminal epitope not present on transferred mini-protein
Staining is generally not present on surfaces of muscle fibers
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Go to
Becker muscular dystrophy pathology
Return to
Dystrophinopathies.
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Neuromuscular syndromes
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References
1.
J Gen Physiol 2022;15:e202213081
2/25/2025