Neuromuscular

Axons, Myelin & Schwann Cells: Molecular Features 1

Peripheral Nerve: Molecular Pathology of Schwann cells & Myelin

Axon associations with
  3 most abundant Myelin components: Myelin Basic Protein (MBP); P0 (MPZ); Periaxin (PRX)
  Non-myelinating Schwann cell marker: Nerve cell adhesion molecule (NCAM)

Myelin: Two types
  Large myelinated axons: Myelin contains both P0 & MBP
  Smaller myelinated axons: Myelin contains P0 but little MBP

Schwann cells
  Types: Several
  Development: Changes

Axon properties compared with Schwann cell/Myelin molecular features
Feature Axon Properties Schwann Cell/Myelin features
Control Nerves
Normal Adult Large-size; Myelinated P0 + MBP
Intermediate-size; Myelinated P0
Small size; Unmyelinated NCAM, Clustered distribution
Development,
  Perinatal		 Myelinated MBP + P0 + NCAM
Unmyelinated MBP + NCAM
Axon Disorders
Axon Degeneration,
  Ongoing		 Myelinated
Neurofilament stain: Lost		 MBP + P0 + NCAM
Phagocytic cells (Acid Phosphatase+)
Axon Loss,
  Chronic		 No Axon P0 + NCAM, or NCAM alone
Büngner bands
Small remaining Axons MBP (Pale)
Axon Regeneration Axon Size: Intermediate
Myelin sheath: Thin		 MBP + P0 ± NCAM
Schwann cells
  Reduced #s		 Small axons: ↓#s
Large axons: Demyelination		 NCAM cells ↓
Myelin Disorders
Demyelination, Ongoing Demyelination, Active Myelin: Acid phosphatase +
Large axons, Demyelinated No or ↓ P0
± MBP+ Schwann cells
Axon loss, Small NCAM: Normal or ↓#s
Demyelination, Chronic,
  Onion Bulb cells		 Remaining Larger axons
 P0 + NCAM
IgM vs MAG Large size MBP + P0 + NCAM
GBS-like
  Neurofascin-186 Ab		 Axon atrophy
Axon loss, Small		 NCAM: Reduced Schwann cells
Myelin structure
  Abnormal		   MBP > P0
MBP = Myelin Basic Protein; P0 = P0 protein; NCAM = Neural Cell Adhesion Molecule

Schwann cells (SC)/Myelin Plasticity
Molecular Markers of Stages
Non-Myelinating SC
  Normal Nerve
    MBP + NCAM: Perinatal nerve SC
      ↓
    NCAM: SC on Non-myelinated axons, Adult
  Damaged Nerve
    NCAM + P0: Denervated (Büngner bands) & Onion bulb SC
    MBP: SC on axons, Demyelinated or Remaining after Axon loss
Myelinating SC
  P0 + MBP: Myelin on Large axons
    ↓
  P0: Myelin on Intermediate-sized axons
    ↓
  P0 + MBP + NCAM: Myelin on Regenerated axons, or Damaged Myelin

Axon Types: Normal Adult Nerve


Toluidine blue stain
Normal Adult Nerve: 3 Axon Populations
  Myelinated axons
    Large size; Thick myelin
    Intermediate-size; Thinner myelin
  Unmyelinated axons: Small size; Grouped
Nfil = Neurofilaments
Normal Adult Nerve: 3 Axon Populations
Sizes
  Large
  Intermediate
  Small: Grouped; Not cut in cross section

Axons & Schwann cells/Myelin: Normal


Nfil = Neurofilaments; MBP = Myelin Basic Protein; P0 = P0 protein; NCAM = Neural Cell Adhesion Molecule
Normal Adult Nerve: 3 Myelin/Schwann cell Populations
  Myelinated axons
    Large size
      Myelin has both Myelin Basic Protein (MBP) & P0 protein (P0)
    Intermediate-size
      Myelin has mainly P0 with little MBP
  Small, Unmyelinated
    Schwann cells have NCAM but no P0 or MBP
  Note: MBP is never present alone in normal adult Myelin or Schwann cells: See below

Schwann cells & Myelin: Normal


P0 = P0 protein; NCAM = Neural Cell Adhesion Molecule
Normal Adult Nerve: Myelin/Schwann cell Populations
  Non-myelinating Schwann cells
    Stain for NCAM (Green), but not P0 or MBP
    Surround: Several, clustered small axons
  Myelin
    Most, or all, sheaths stain for P0 (Red)
  Few cells stain for both NCAM & P0 (No Yellow)
MBP = Myelin Basic Protein; P0 = P0 protein
Normal Adult Nerve: 2 Myelin Populations
  Large (Thick) Myelin sheaths: Myelin has abundant MBP & P0 (Yellow)
  Small (Thinner) Myelin sheaths: Myelin has mainly P0 (Green), but little MBP
  No sheaths have only MBP (Red)

Myelin & Periaxin: Normal Nerve

3 Most abundant myelin proteins in peripheral nerve
  MBP
  P0
  Periaxin


MBP = Myelin Basic Protein; Prx = Periaxin
Normal Adult Nerve: Myelin Populations
  Periaxin (Green) colocalizes (Yellow) mostly with MBP around large axons
P0 = P0 protein; Prx = Periaxin
Normal Adult Nerve: Myelin Populations
  P0 (Red) has some, patchy associated Periaxin

Infant Sural Nerve: Normal

  Axon Sizes: Smaller than adults
  Immature Schwann cells: Contain MBP
  Myelin: 1 population, Contains P0 & MBP

Axon types: Normal Infant


Toluidine blue stain
Infant Nerve: Axon Populations
  Large Myelinated: Smaller than adult large axons; Thin myelin sheath
  Intermediate-sized: Thinly Myelinated or Unmyelinated
Nfil = Neurofilaments
Infant Nerve: Axon Populations
  Large (Myelinated): Smaller than adult large axons
  Small Unmyelinated (Arrow): Present in clusters

Axons & Schwann cells/Myelin: Normal Infant


Nfil = Neurofilaments; MBP = Myelin Basic Protein; P0 = P0 protein; NCAM = Neural Cell Adhesion Molecule
Normal Infant Nerve
Largest Axons (F & G)
  Smaller than in adults

MBP (Left; E) (Red or Yellow): Present
  In most Schwann cells
  Alone, without P0, in some cells
 P0
  Present in sheaths around most myelinated axons (Center; F)
  Patchy presence in Schwann cells around unmyelinated axons

NCAM (Right; G)
  Present with most unmyelinated, but not myelinated axons

Schwann cells & Myelin: Normal Infant


P0 = P0 protein; NCAM = Neural Cell Adhesion Molecule
Schwann cells
  Stain for NCAM (Green; Yellow)
  Also contain patchy regions of P0 (Yellow), different from adults
Myelin
  Stains for P0 (Red)
MBP = Myelin Basic Protein; P0 = P0 protein
Schwann cells
  MBP (Red; Yellow)
    Present in all Schwann cells & Myelin
    Present in Schwann cells without P0 (Red)
Myelin
  Contents: MBP & P0

Adult Sural Nerve: Axon loss, Severe, Chronic

Remaining Small axons
  Structure
    Continuous or Beaded
    Single unmyelinated axon in a Schwann cell
  Associated Schwann cells
    Immature
    Contain MBP
    May have no NCAM or p0
Denervated Schwann cells (Büngner bands)
  May contain both NCAM & p0

Axon loss: Morphology


Toluidine blue stain
Severe axon loss
  Few, or No axons, are visible
Nfil = Neurofilaments
Neurofilament stain
  A few, scattered clusters of small axons are visualized

Myelinated Axon Loss
Schwann Cells surround Remaining Unmyelinated Singleton Axons

  Singleton Axon sizes: ~ 1μM
  No surrounding myelin
  Associated Schwann cells
    Express MBP but no P0
    One Axon/Schwann cell

Nfil = Neurofilaments; MBP = Myelin Basic Protein
Remaining Small axons without Myelin
  Structure: 2 patterns
    Beaded (Above)
    Continuous: Singletons
  Associated Schwann cells
    Stain for MBP (Yellow) on axons without myelin sheath
Myelin
  None visible around remaining singleton axons
Nfil = Neurofilaments; P0 = P0 protein; NCAM = Neural Cell Adhesion Molecule
Remaining Small Axons
  Size: Small
  May have no staining for p0 or NCAM (Arrows)
Schwann cells
  Abundant: Remain after axon loss
  Most have no associated axons
  Stain for: NCAM & P0
MBP on small axons with no myelin
  Some remaining axons may appear beaded with Neurofilament/MBP stain
  Normal small axons have no MBP staining

Neurofilament = Green; MBP = Red; Co-stain = Yellow

Singleton Unmyelinated Axons

"Singletons": Small Single Axons
  Surrounded by Schwann cell processes (Arrow)
  May contain small blebs (Sprouts; Right)
  Stain for MBP, even without myelin

From: R Schmidt
From: R Schmidt

From: R Schmidt
"Singletons": Small Single Axons
  Surrounded by Schwann cell processes but no myelin
  One axon per Schwann cell

From: R Schmidt

Denervated Schwann Cells (Büngner Band Cells)


P0 = P0 protein; NCAM = Neural Cell Adhesion Molecule
Schwann cells: Denervated
  No associated Axons
  Costain NCAM + P0 (Yellow)
  Also see: Onion bulb cells
MBP = Myelin Basic Protein;
NCAM = Neural Cell Adhesion Molecule
Schwann cells: Denervated
  No associated Axons
  Contain NCAM (Green) but not MBP
P0 = P0 protein;
NCAM = Neural Cell Adhesion Molecule
Schwann cells: Normal
  NCAM (Green)
    No co-staining with P0 (Red)
  P0 (Red)
    Stains myelin, not Schwann cells

Büngner bands: Clusters of Schwann cell processes with no axons


From: R Schmidt
Büngner bands (Arrow)
  Interdigitated Schwann cell processes    

Axon Regeneration

 Regenerated Myelinated Axons
  Size: Intermediate
  Myelin Thickness: Thin; Similar thickness in each axon cluster
  Myelin molecular components
    MBP + P0 ± NCAM
    Smaller myelinated axons have MBP in myelin

Toluidine blue stain
Regenerated Axons
  Many clusters of intermediate-sized thinly myelinated axons
    Axons in individual clusters have similar myelin thickness
Nfil = Neurofilaments
Regenerated Axons (Neurofilament stain)
  Clusters of regenerated larger axons are present (Arrow)

Axons & Schwann cells/Myelin: Axon Regeneration

Nfil = Neurofilaments; MBP = Myelin Basic Protein; P0 = P0 protein; NCAM = Neural Cell Adhesion Molecule
MBP: In myelin around many very small axons (Right; E)
P0: Stains myelin around many axons (Center; F)
NCAM (Right; G)
  Stains around some, but not all, larger axons
  Some NCAM profiles have no associated axon: Suggests incomplete axon regeneration with remaining denervated Schwann cells

Axon Regeneration: Myelin Features


P0 = P0 protein; NCAM = Neural Cell Adhesion Molecule
NCAM
  Patchy co-stain (Yellow) with P0 (Red) in myelin sheaths
  Stains many non-myelinating Schwann cells (Green)
MBP = Myelin Basic Protein; P0 = P0 protein
Myelin
  Most myelin sheaths, even small, costain for both P0 & MBP

Adult Sural Nerve: Wallerian Degeneration

Wallerian Degeneration: Active

Toluidine blue stain
Phagocytic Cells
  Types: Phagocytic Schwann cells; Macrophages
  Contents: Myelin debris; Lipid droplets
Myelin sheaths: Collapsed
Nfil = Neurofilaments
Degenerated axons: No neurofilament stain (Arrow)
    Neurofilament fragments, or
    Empty regions with no axon

Axon Degeneration: Acute


Nfil = Neurofilaments; MBP = Myelin Basic Protein; P0 = P0 protein; NCAM = Neural Cell Adhesion Molecule
Axon Degeneration, Early
  No axon staining within MBP stained myelin (Left; G) (Arrow)
  No axon staining within P0 stained myelin (Center; H) (Arrow)
  Minor loss of small axons: Scattered empty NCAM (Red) sheaths (Right; I)

Wallerian Degeneration: Active, Phagocytic cells


  Phagocytic Cells
    Types
      Phagocytic Schwann cells (Left)
      Macrophages (Center)
    Contents: Myelin debris (Left; Arrow); Lipid droplets (Center; Arrow)
AcP = Acid Phosphatase
Phagocyte Stain (Acid phosphatase): Endoneurial cells, scattered (Red)
  Compare to: Demyelination

Adult Sural Nerve: Demyelination

Demyelination: Macrophage-Mediated


Nfil = Neurofilaments
Demyelinated Large Axons
  Large axons: No surrounding myelin space
From: R Schmidt
Macrophage-mediated Demyelination
  Macrophage process (Arrow) extends underneath Schwann cell basal lamina

Demyelinated Axons


Toluidine blue stain
Large Demyelinated Axons (Arrow)
  Other axons often have thin myelin sheath for size
From: R Schmidt
Demyelinated Axon: Surrounded by Schwann cell processes

Demyelinated Large Axons: No surrounding Myelin sheath

Schwann cells surrounding Demyelinated axons: MBP+; P0 & NCAM-

Nfil = Neurofilaments; P0 = P0 protein
Demyelinated Large Axons (P0 stain)
  No surrounding myelin (Arrow; Below)
Nfil = Neurofilaments; MBP = Myelin Basic Protein
Demyelinated Large Axons
  Co-stain (Yellow) for MBP: Immature non-myelinating Schwann cells

Nfil = Neurofilaments; MBP = Myelin Basic Protein; P0 = P0 protein

Myelin Disorders: MAG Neuropathy


Toluidine blue stain
Myelin sheaths: Irregular structure
From: R Schmidt
Myelin Wide-spacing
  Cleft, contatining a few layers, within compact myelin
MAG Neuropathy

Nfil = Neurofilaments; P0 = P0 protein
No demyelinated axons
Nfil = Neurofilaments; MBP = Myelin Basic Protein
Small axons: Many have associated MBP co-staining (Immature Schwann cells)

NCAM = Neural Cell Adhesion Molecule; MBP = Myelin Basic Protein
NCAM (Yellow): Abnormally expressed within MBP-containing (Red) myelin sheaths
MBP = Myelin Basic Protein; P0 = P0 protein
MBP & P0 expressed in Large & Smaller myelin sheaths
  Smaller myelinated axons normally have myelin that stains only for P0

Chronic Demyelinating Neuropathy: Onion Bulbs


Toluidine blue stain
Onion bulbs
  Structure: Concentric layers of Schwann cells & Collagen (Arrow)
  Center: Some have myelinated axon
Neurofilament stain
Onion bulbs (OB): Relation to axons
  Some have large central axon
  Others have no central axon
  Small, unmyelinated axons
    May be present within onion bulb layers or around OB periphery

Onion Bulbs: Ultrastructure

Onion Bulb with Thinly Myelinated Central Axon

From: R Schmidt
Onion Bulb structure
  Alternating layers of Schwann Cell processes & Collagen
 "Obsolete" Onion bulb: No remaining Central axon

From: R Schmidt
Onion Bulb structure
  Alternating layers of Schwann Cell processes & Collagen

Onion Bulb Schwann cells: Molecular features


Nfil = Neurofilaments; NCAM = Neural Cell Adhesion Molecule
Onion Bulb cells
  NCAM
    Abundant in onion bulb Schwann cell processes
  Some onion bulbs contain central axons (Yellow)
Nfil = Neurofilaments; P0 = P0 protein
Onion Bulb cells
  P0
    Present in most onion bulb Schwann cell processes
  Some onion bulbs contain central axons (Yellow)

NCAM = Neural Cell Adhesion Molecule; P0 = P0 protein
Onion bulb Cells
  Contain both NCAM & P0 (Yellow)
  P0 (Red) also stains myelin around central axon in some onion bulbs
Nfil = Neurofilaments; MBP = Myelin Basic Protein
Onion bulb Cells
  MBP (Red)
    Present in myelin around a few large axons
    Not in onion bulb cells
    Costains unmyelinated small axons (Immature SC)

Neurofascin-186 antibodies: Axon Atrophy & Small Axon Loss

Axon Atrophy

Larger myelinated axons: Axon atrophy
  Morphology: Small & Slit-like shape
  Myelin sheath: Thick for axon size
See: Normal

Axon atrophy


Nfil = Neurofilaments; MBP = Myelin Basic Protein
Axons within MBP myelin sheaths
  Many are smaller than Normal

Myelin: Irregular Structure


MBP = Myelin Basic Protein; P0 = P0 protein
Larger myelin sheaths
  Irregular co-stain for P0 & MBP

Neurofascin-186 antibody: Reduced numbers of Non-myelinating Schwann cells & Small axons


Nfil = Neurofilaments; P0 = P0 protein
Small Axons (Green): Reduced numbers
Nfil = Neurofilaments; NCAM = Neural Cell Adhesion Molecule
Non-myelinating Schwann cells (Red): Reduced numbers
Small Axons: Reduced numbers (Many empty non-myelinating Schwann cells (Red))
Myelinated axons (Green): Generally small size

Myelin Disorders: Other Pathology Patterns


Nfil = Neurofilaments; NCAM = Neural Cell Adhesion Molecule
Schwann Cells (Red): Reduced numbers
Abnormal Myelin Composition
  P0 lost from myelin on large axons; MBP (Red) remains (Arrow)
  Also see: PN with IgM vs MAG

MBP = Myelin Basic Protein; P0 = P0 protein

Active Demyelination: Histiocytic Schwann cells & Myelin


Acid phosphatase stain
Most Myelin sheaths: Acid phosphatase+
  Compare to: Wallerian degeneration
MBP = Myelin Basic Protein; P0 = P0 protein
Myelin Structure: Irregular P0 & P0+MBP Sheaths
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Pathology Index

References
1. Neuropathol Appl Neurobiol 2023;49:e12898

4/28/2024