interpretation of nerve biopsy

1. INTERPRETATION OF
NERVE BIOPSY
PRESENTER: DR. HAJRA K. MEHDI
MODERATOR: DR. KALYANI R
Sri Devaraj Urs Medical College, Kolar

2. CONTENT
• Introduction
• Indication of nerve biopsy
• Selection and preparation of nerve
• Vasculitic neuropathy
• Sarcoid neuropathy
• Neuropathy associated with infection
• Inflammatory demyelinating polyneuropathy
• Polyneuropathy associated with Monoclonal
gammopathy
• Lymphomatous neuropathy
• Hereditary neuropathies
• Metabolic neuropathy
• Toxic polyneuropathy
• Summary
• Reference

3. INTRODUCTION
• Connective tissue: endoneurium, perineurium, epineurium
Nerve components:
• Myelinated fibers: myelinated axons range in diameter from 2 – 18 micrometer
• Small ~ 4micrometer
• Large ~ 12micrometer
• Unmyelinated fibers
• Internode
• Node of Ranvier

4. Normal Anatomy of a Single Axon:
• The myelinated segments are called internodes.
• Internode length increases as does the axon diameter.
• For large axons, the internodal length ranges between 0.35 and 1.83 mm and is
approximately 1/3 of that size for smaller myelinated axons.
• Especially in the elderly, there is significant variability in internodal length.
• The unmyelinated fibers actually outnumber myelinated fibers approximately 4 to 1.
INTRODUCTION

5. Sural Nerve Cross-Section
(H&E)

6. Sural Nerve Cross-Section
(Trichrome)

8. Semithin section (Toluidine Blue)
5 year: axon diameter reaches adult volume
10-15 years: normal myelin thickness

9. • There is normally a bimodal distribution of myelinated fiber diameters.
• The thickness of the myelin sheath is generally proportional to the axon diameter.
• Measured by “g ratio” on morphometry.
• g ratio = axon diameter/fiber diameter (0.6-0.7 is optimal for nerve conduction)
• In some forms of polyneuropathy, such as hereditary motor sensory neuropathy type I, the
large myelinated fibers may be preferentially affected, while in amyloidosis, for example,
the small myelinated fibers may be preferentially affected.

10. INTRODUCTION
• Depending on which component of the peripheral nerve is predominantly involved in the
pathological process, peripheral neuropathy can be classified into two main categories.
AXONAL DEGENERATION DEMYELINATION
• Alcohol • GBS
• Nutritional • CIDP
• Vasculitic • Hypertrophic
• Giant axonal • Metachromatic
• Diabetic • Tomaculous
• Leprosy

12. WALLERIAN DEGENERATION

14. BANDS OF BUNGNER
• Linear bands of interdigitating Schwann cells
• Surrounded by basal lamina (Schwann tube)
• Size: > 3 μm
• Basement membrane: Irregular
• Due to: Loss of Myelinated axons OR Proliferation of associated Schwann cells

15. BANDS OF BUNGNER

16. • Hypertrophic neuropathy
• Refers to the concentric laminated layers surrounding the nerve fiber.
• Best detected in the semithin section
• Paraffin section is to look for an increased number of Schwann cell nuclei.
• Pathogenetically, onionbulb formation is indicative of repeated demyelination and
remyelination.
ONION-BULB FORMATION

17. ONION-BULB FORMATION

18. RENAUT BODIES
• Renaut bodies are fibrillary structures present in the subperineurial area.
• They are composed predominantly of fibroblasts.
• They are lightly staining with H&E, Toluidine blue and Alcian blue,

19. REGENERATION CLUSTERS

20. Vasculitic Neuropathy
Sarcoid Neuropathy
Neuropathies Associated With Infection
Inflammatory Demyelinating Polyneuropathy
Polyneuropathy Associated With Monoclonal
Gammopathy
Ischemic Neuropathy
Amyloid Neuropathy
Metachromatic Leukodystrophy
Lymphoma
Tomaculous Neuropathy
Hereditary Neuropathy
INDICATION FOR NERVE BIOPSY

21. • Distal lower limbs - Sural nerve or superficial peroneal nerve
• Upper limbs - Superficial radial nerve or a branch of the ulnar nerve
• Progressive optic neuropathy – Optic nerve biopsy
TYPES OF NERVE BIOPSY
• Fascicular biopsy
• Whole biopsy
SELECTING THE NERVE FOR BIOPSY

22.  Easily identifiable and relatively protected
from compression injury.
 Purely sensory thus producing no motor deficit
following biopsy.
 Liable to be affected by neuropathy because it
is a distal branch of a long nerve.
SURAL NERVE BIOPSY

24. PROCESSING OF NERVE BIOPSY

25. Paraffin sections:
• Nerve is fixed in a neutral buffered formalin solution
• Cut into two pieces, one-third for the transverse section and two-thirds for the
longitudinal section.
• Sections are cut at 5 μm, except for Congo-red stain, which should be cut at 8 to 10 μm.
• Stained with H & E, modified trichrome, and Congo-red stains.

26. H&E Morphology, Vasculitis, Inflammation,
Myelin ovoids, axonal degeneration.
Masson’s Trichrome Fibrosis, Hyalinisation, Vessels
Kpal Myelinated fibers
Cresyl violet Sulfatide storage material
Osmium tetroxide Myelin
Toulidine Myelin
Congo Red Amyloid
IHC Axons, Lymphocytes, Macrophages,
Immunoglobulin
STAINING

27. • Cut the nerve at 1 μm with the EM microtome for transverse sections.
• Toluidine blue stain:
Collagen — pale blue
Cytoplasm — pale blue
Nuclei — dark blue
Myelin — very dark blue to black
SEMITHIN SECTIONS

28. TEASED NERVE FIBERS

31. ARTIFACTS
Myelin like bodies

32. ARTIFACTS
Myelin sheaths mechanically damaged

33. ARTIFACTS

34. 1. Status of the epineurium including the blood vessels
2. Alterations in the perineurium
3. Endoneurium edema
4. Density of the large and small myelinated nerve fibers
5. Extent of axonal degeneration and atrophy
6. Frequency of bands of Bungner and Myelin degeneration
chambers
7. Number of macrophages clusters
8. Onion bulb formation
9. Inflammatory infiltrates
10. Presence/absence of amyloid
WHAT TO
LOOK
FOR?

35. • Most common inflammatory cause of a neuropathy.
• Clinically, these can be of acute, subacute and chronic onset and often are painful and
multifocal.
• It affects the peripheral nerve vasculature, resulting in ischaemic damage to the nerve.
• Vasculitic neuropathy is defined by an inflammation within the vessel wall and signs of
active or chronic vascular damage, whilst perivascular or mural inflammation alone is
considered non-specific.
VASCULITIC NEUROPATHY

36. Probable vasculitic neuropathy can be diagnosed
• (a) Predominantly axonal damage
• (b) Perivascular inflammation with active or chronic vessel damage or
• (b) (Peri-) vascular inflammation in combination with one of the following five
histological predictors
(i) Vascular IgM, complement or fibrinogen deposits by immunofluorescence
(ii) Hemosiderin deposits
(iii) Patchy/multifocal nerve fibre loss
(iv) Prominent axon degeneration.
VASCULITIC NEUROPATHY

37. VASCULITIC NEUROPATHY

38. ACTIVE VASCULITIC NEUROPATHY

39. INACTIVE VASCULITIC NEUROPATHY

40. AXONAL DEGENERATION

41. SUBPERINEURIAL EDEMA

42. Larger (100–250 μm) epineurial arterioles:
• Polyarteritis Nodosa
• Churg–Strauss syndrome
• Wegener’s granulomatosis
• Rheumatoid vasculitis.
Smaller (<100 μm) epineurial arterioles:
• Sjögren’s syndrome
• SLE
VASCULITIC NEUROPATHY

43. • Noncaseating granuloma in then epineurium or in muscle biopsy.
• Axonal degeneration.
• Numerous myelin ovoids.
• Nerve fiber degeneration due to compression by granuloma.
SARCOID NEUROPATHY

44. • M. leprae is the only bacterium that invades peripheral nerves in man and animals.
Common nerves:
• Ulnar nerve at the elbow
• Deep peroneal branch at the ankle
• Superficial branches of the facial nerve
• Median nerve at the wrist.
• Nerve biopsy is imperative for the diagnosis of primary neuritic leprosy in which neuropathy is
the sole clinical manifestation without typical skin lesions or a positive skin smear.
LEPROSY

45. • Pathological hallmark is an intense inflammatory noncaseating or caseating
granulomatous lesion that severely damages the neural architecture.
• Granulomas in the epineural and perineural spaces & endoneural space.
• Caseation may occur and produce large abscesses within the nerve.
• Bacilli are scanty and, when found, are almost always in the nerve.
• Healing - Fibrosis and hyalization in the endoneurium and thick perineurial and
epineural sheaths
TUBERCULOID LEPROSY

46. TUBERCULOID LEPROSY

47. Pathological hallmark:
• Perineural and endoneural infiltration of enlarged macrophages and Schwann cells with
M. leprae bacilli (foamy or leprae cells) and inflammatory cells are the cardinal features.
• In severe cases, the epineurium may be infiltrated by huge numbers of foamy cells,
especially around blood vessels.
• Granulomatous inflammatory response - minimal
LEPROMATOUS LEPROSY

48. LEPROMATOUS LEPROSY

49. LEPROMATOUS LEPROSY

50. • Acute – Gillian Barre Syndrome (GBS)
• Chronic – Chronic Inflammatory Demyelinating Polyradiculoneuropathy (CIDP)
• Hallmark of inflammatory neuropathy- presence of inflammatory cells in the
endoneural space of the nerve.
• Inflammatory cells are primarily responsible for the macrophage induced demyelination
in these neuropathies.
INFLAMMATORY DEMYELINATING
POLYNEUROPATHY

52. CIDP

53. CIDP

55. • Neuropathies are usually distally accentuated and symmetrical, but multiple mono-
neuropathies may also occur.
• Predominantly of the axonal type.
• Definitive diagnosis: demonstration of amyloid in the nerve.
• Two forms: Primary Amyloidosis and Familial Amyloid Polyneuropathies (FAP)
• Stains: Congo Red, Thioflavin S or T
AMYLOID NEUROPATHY

56. Three patterns of deposition:
1. Extraneurial connective tissue deposition of amyloid – Carpal Tunnel Syndrome
2. Widespread endoneurial deposition of amyloid – Familial forms
3. Deposition within the walls of the vasa nervosum of both the epineurium and
endoneurium.
AMYLOID NEUROPATHY

57. AMYLOID NEUROPATHY

58. AMYLOID NEUROPATHY

59. AMYLOID NEUROPATHY

60. • At early stages, unmyelinated and small myelinated fibres are selectively lost.
• Later stages large myelinated fibres are also affected.
• Neuropathological differential diagnosis: CIDP and other inflammatory neuropathies.
• Structural differential diagnosis: Oxytalan fibres which can be found in Renaut bodies
AMYLOID NEUROPATHY

61. • Subacute progressive sensorimotor polyneuropathy, cranial neuropathy, mononeuropathy
multiplex, or an isolated median or sciatic nerve palsy.
• CSF -mildly increased protein and cells , positive for tumor cells.
• Cardinal histological feature: Diffuse, massive infiltration of lymphomatous cells in all
three compartments of the nerve.
• Perivascular cuffing of lymphomatous cells and striking angiocentricity of the tumor
cells.
LYMPHOMATOUS NEUROPATHY

62. LYMPHOMATOUS NEUROPATHY

63. • Accumulation of galactosyl-3-sulfate (sulfatide) in the brain, kidney, gallbladder, and
peripheral nerves.
• Sulfatide deposits: Metachromasia with cresyl violet and toluidine blue
• 4 forms - Late infantile, Juvenile, Adult, Multiple sulfatase deficiency.
• CNS involvement predominates in all forms.
• Diagnosis: Nerve biopsy
Biochemical assay - Arylsulfatase A assay
METACHROMATIC LEUKODYSTROPHY

64. • Biopsied nerve should be stained on frozen sections.
• Accumulation of metachromatic granules of 0.5 to 1 μm in diameter in the perinuclear
cytoplasm of Schwann cells.
• Reduction in the myelinated fiber population.
• Occasional small onion-bulb formations.
• Presence of metachromatic granules in the sural nerve biopsy is diagnostic.
METACHROMATIC LEUKODYSTROPHY

66. METACHROMATIC LEUKODYSTROPHY

69. HMSN I - Charcot-Marie-Tooth (CMT):
• Demyelinating neuropathy
• Hypertrophic neuropathy
• Multiple onion bulbs (distal nerves > proximal nerves)
HEREDITARY MOTOR & SENSORY
NEUROPATHY

70. CMT-1 Normal

71. Onion Bulbs in CMT-1

72. HMSN II - Neuronal form:
• Autosomal dominant
• Loss of myelinated axons as the predominant finding.
HMSN III - Dejerine-Sottas Disease:
• Autosomal ressesive
• Enlarged peripheral nerve
• Multiple onion bulb formation
HEREDITARY MOTOR & SENSORY
NEUROPATHY

73. • Early onset
• Slowly progressive distal polyneuropathy
• Strikingly curly hair,
• Nerve biopsy is recommended in any patient in whom this neuropathy is suspected since
it shows a definite diagnostic finding of giant axons
GIANT AXONAL NEUROPATHY

74. • Dramatic axonal swelling.
• Giant axons are scattered in the nerve fascicle and are easily recognizable.
• On longitudinal sections these giant axons are shown as cigar-shaped ballooning, often
near a node of Ranvier.
• Teased nerve fibers characteristically show single or multiple spindle-shaped or fusiform
swellings along axons measuring from 40 to 350 μm in length and 10 to 30 μm in
diameter.
GIANT AXONAL NEUROPATHY

75. GIANT AXONAL NEUROPATHY

76. • Axonal neuropathy
• Hallmarks: Fiber atrophy and loss of myelinated & unmyelinated fibers associated with
axonal degeneration & segmental demyelination.
• Focal loss of myelinated fibers (central fascicular degeneration or selective nerve
fascicular degeneration).
• Vascular abnormality in the endoneural or epineural space (thickening, occlusion, medial
sclerosis, and even fragmentation of the internal elastica).
DIABETIC NEUROPATHY

77. • Nerve biopsy is indicated in any diabetic
neuropathy if other treatable diseases such
as vasculitic neuropathy or CIDP are
suspected.
DIABETIC NEUROPATHY

79. • 5th to 7th decade, peripheral neuropathy, dementia, neurogenic bladder, and upper motor
neuron disease.
• Large number of polyglucosan bodies in the central and peripheral nervous system
• Intra-axonal, round, range from 5 to 70 μm in diameter, and usually occur in myelinated
fibers.
• Many polyglucosan bodies are required for diagnosing PGBD.
POLYGLUCOSAN BODY NEUROPATHY

80. POLYGLUCOSAN BODY NEUROPATHY

81. • Tomacula: Focal sausage-shaped swellings of myelin sheaths.
• No accompanying axonal swelling.
• Segmental demyelination – uniform finding.
• Represents demyelinating neuropathy
• Most commonly and typically seen in Hereditary neuropathy with liability for
Pressure Palsy (HNPP) and familial recurrent brachial plexopathy
TOMACULOUS NEUROPATHY

82. TOMACULOUS NEUROPATHY

84. TOXIC NEUROPATHY

85. • A 42-year-old male presented to the emergency department following acute onset of
breathlessness.
• He had a history of unexplained lower limb paraplegia at 29 years of age.
• Over 4 months prior to admission, the patient had multiple presentations with variable
non-specific relapsing complaints including progressive upper limbs weakness, transient
left eye diplopia, vague complaints of lethargy, musculoskeletal pain, and alternating
urinary retention and incontinence.
CASE 1

87. CHRONIC INFLAMMATORY DEMYELINATING
POLYNEUROPATHY

88. • 11-year-old, otherwise healthy male presented with progressive right-hand weakness and
numbness with no cutaneous abnormalities.
• Physical examination and electrodiagnostic testing revealed findings consistent with a
severe ulnar neuropathy at the elbow.
• MRI revealed diffuse thickening and enhancement of the ulnar nerve and narrowing at
the cubital tunnel.
CASE 2

90. LEPROMATOUS NEUROPATHY

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92. ??????

93. ??????

94. • Nerve biopsy is still very useful in a wide range of conditions, although molecular
genetic and neurophysiological findings are diagnostic in an increasing number of
genetic and immune-mediated neuropathies.
• Nerve biopsy must be considered on a case-by-case basis because the results may help
confirm a suspected diagnosis and/or guide the therapeutic strategy.
SUMMARY

95. • Chinnamma Thomas. Color Atlas of Nerve Biopsy Pathology. Archives of Pathology &
Laboratory Medicine 2002;126:1000.
• Sommer C, Brandner S, Dyck PJ, et al. Guideline on processing and evaluation of sural
nerve biopsies. Neuromuscul Disord 2008;18:90-96.
• Vallat JM, Vallat-Decouvelaere AV. Nerve biopsy: Current indications and results. In:
Cros D, ed. Peripheral Neuropathy. A Practical Approach to Diagnosis and Management.
Philadelphia, PA: Lippincott, Williams & Wilkins, 2001;20-42.
• Vallat JM. An Update on Nerve Biopsy. J Neuropathol Exp Neurol 2009;68:833-44.
• Internet sources
REFERENCE