Basic Essentials of CNS Histopathology (In Less Than 1 Megabyte) Marie Beckner, MD

1. Basic Essentials of CNS Histopathology (In Less Than 1 Megabyte) Marie Beckner, MD UPMC Neuropathology March 1, 2002 Please e-mail suggestions to: becknerme@msx.upmc.edu

2. Cell Lineage in the CNS Peripheral blood Ependymal cells Microglia Astrocytes Oligodendrocytes O2 A cells Neurons Mesenchymal cells form blood vessels & meninges Neuroepithelial cell in germinal zone

3. Overview Neurons: Morphology Stains Cellular responses Inclusions Glia: Astrocytes Oligodendrocytes Ependymal cells Microglia and macrophages Meninges

4. Morphology of neurons, H&E Projection neurons Granular cell neurons Motor neurons large cell body nucleus with single large nucleolus prominent basophilic Nissl substance (RER & polyribosomes) axons & dendrites embedded in surrounding neuropil Example of “local circuit” neurons Naked nuclei seen Lack Nissl substance

5. Neurons: Conventional Stains Hematoxylin and Eosin (H&E) for general cytoarchitecture Cresyl violet for staining Nissl substance Silver stains (Bielschowsky & others) for staining axons and some inclusions Many others but technically difficult and used less now that immunohistochemical stains are widely available

6. Neurons: Immunohistochemical Stains Immunoperoxidase (ImP) or “brown” stains (can also be red, blue, black) Neurofilament proteins: perikaryal (cell body) and axonal cytoplasm Synaptophysin: vesicles at synapses so that punctate granular staining is seen diffusely in the neuropil and at the edges of neuronal bodies. Most useful and widely used. Neuron specific enolase (NSE): non-specific Others: Chromogranin, PGP9.5, -synuclein, NeuN, others (growing list)

7. Neurons: Cellular Responses Cell body Chromatolysis Acute necrosis Atrophy Ballooning change Neuron loss

8. Neurons: Chromatolytic Changes Central chromatolysis (usual response to axonal damage that disrupts basic cell functions) - loss of basophilic Nissl substance from central part of cell body, only at edges - enlargement of cell body with rounding - nucleus displaced to periphery - can also see in pellagra and Wernicke’s encephalopathy Peripheral chromatolysis (loss of Nissl substance from the periphery (less common)

9. Neurons: Mimics of Central Chromatolysis Large, normal neurons (Betz cells, mesen- cephalic nucleus of cranial nerve V) Lipofuscin displacement of nucleus and Nissl substance Neurons with eccentric nuclei (Clarke’s nucleus, paraventricular & supraoptic nuclei) Diseases where neurons have displaced nuclei and Nissl substance (storage diseases and ganglion cell tumors)

10. Neurons: Acute Necrosis Ischemic/hypoxic damage (6-8 hrs) leads to “red neurons” (not just bright pink) - Pronounced, eosinophilic cytoplasm - Shrunken cell body - Shrunken, darkly pyknotic nucleus that no longer contains a prominent, large nucleolus - Seen more easily in large neurons (vs. small) Nonspecific finding that does not rule out malignancy in a biopsy. Confused with “handling” artifact (cells have dark purple cytoplasm)

11. Neurons: Selective Vulnerabilities Mature > immature neurons Neurons > endothelial cells > glial cells Regions of neurons: Hippocampal sclerosis (neuronsglia) - pyramidal neurons of hippocampal CA1 sector (susceptible) > CA2 sector Laminar necrosis - middle & deep cortical > superficial layer Sulcal depths Watershed areas

12. Neurons: Atrophy Decrease in size of cell body and then loss (minimal & variable in aging) Degenerative diseases (many) Degeneration in response to injury 1. Retrograde (distal  proximal) ex. Infarct in occipital visual cortex  ipsilateral LGN neurons die 2. Transynaptic (neuron  neuron) ex. Loss of retinal ganglion cells  loss of neurons in LGNs

13. Neurons: Ballooning Change Metabolic derangement  undigestable product  engorgement of cell body  cell death Lysosomal storage disorders Ex. Tay-Sachs disease Sandhoff Disease Nieman - Pick Disease

14. Neuron Loss Final common pathway for many conditions Time course - acute or slow Apoptosis - rapid nuclear fragmentation Mineralization (Fe, Ca, etc.) may occur - very dark purple encrustation of dead neuron and/or axon - “ferrugination” - damaged axons with mineralization can resemble fungal hyphae - may see mineralized neurons adjacent to cystic cavities of remote infarcts

15. Neurons: Axonal Changes to Injury (1) Wallerian degeneration (proximal to distal) with or without degeneration of cell body - beaded appearance of axon - injury of axon or cell body ex. Nerve transection, metabolic injury, and ischemia CNS - no reinnervation (2) Retrograde degeneration (distal to proximal) -“Dying back” process - Metabolic derangement of entire neuron - Distal axon most vulnerable - Usually slow progressive clinical course ex. “Stocking glove” neuropathy in diabetes mellitus

16. Neurons: Axonal Changes to Injury (3) Focal axonal swelling (axonal spheroids, axonal retraction balls, axon torpedos for Purkinje cells) - distended axons filled with neurofilaments & organelles - nonspecific response (disturbed metabolism) - etiologies include severe axonal injury (ischemia, trama, degenerative, normal aging) - rarely inherited (neuroaxonal dystrophy) - “Herring bodies” - storage of hormones in infundibulum and neurohypophysis - normally seen in fasciculus gracilis in medulla with aging, may be mineralized

17. Neuronal Inclusions - Normal (1) Neuromelanin (starts around 5 yrs of age) - By-product of tyrosine needed in cate- cholamine (dopamine & norepinephrine) synthesis - Substantia nigra, locus ceruleus, & dorsal motor nucleus of vagus - Abundant brown, cytoplasmic pigment (much darker than lipofuscin) on H&E - Lost in substantia nigra in Parkinson’s dz

18. Neuronal Inclusions - Normal (2) Lipofuscin, “aging pigment”, common - golden brown on H&E in neurons & glia - displaces cytoplasmic organelles and may mimic central chromatolysis - lipids, proteins, carbohydrates (PAS+) - grossly a mahogany hue (LGBs) - lateral geniculate bodies (LGBs), inferior olives, dentate nuclei of cerebellum, and anterior horn cells of spinal cord - rare inherited, fatal disease - Ceroid lipofuscinosis

19. Neuronal Inclusions - Normal (3) Marinesco bodies - intranuclear, increase with age - small, multiple, no halos, no effacement of the nucleus, size of nucleolus - Cowdry type B (not viral !!) - immunoreactive for ubiquitin - substantia nigra and locus ceruleus

20. Neuronal Inclusions - Normal (4) Hyaline colloid inclusions - cytoplasmic, homogenous - hypoglossal nucleus (common) - anterior horn cells (rare) - dilated endoplasmic reticulum with amorphous material (5) Eosinophilic inclusions of inferior olives - immunoreactive for ubiquitin (6) Others - describe with diseases where they are increased

21. Neurons- Inclusions in Infection Nuclear (1) Cowdry type A - Herpes simplex (HSV), herpes simiae Varicella zoster (VZV), cyto- megalovirus (CMV), measles virus (subacute sclerosing panencephalitis) “owl’s eye” large, solitary, clear halo peripheral margination of nuclear chromatin (2) Cowdry type B - anterior horn cells in poliomyelitis (3) Ground glass - Herpes

22. Neurons - Inclusions in Infection cont. Cytoplasmic Rabies - Negri bodies (Round, eosinophilic, hyaline, well-defined) - Lyssa bodies (Irregular, eosinophilic) - Most easily seen in hippocampal pyramidal cells or Purkinje cells Viral antigen demonstrated also with immunostaining

23. Neurons - Pathologic inclusions, non- infectious diseases cont. (1) Neurofibrillary tangles (NFTs) - many in Alzheimer’s disease but also seen in other degenerative conditions, a few in some older patients, and rarely in a few other conditions NFTs assume shape of cell classically “flame” shape but may be round, “globoid” H&E - faint basophilic wisps Bielschowsky - dark brown, more prominent Immunoreactive for P-tau

24. Neurons - Pathologic inclusions cont. (2) Granulovacuolar degeneration of Simchowicz - cytoplasmic, basophilic, dot-like granules in small clear vacuoles (resemble marbles) - single or multiple (few to many) - hippocampal pyramidal neurons - few in older non-demented patients but may be extensive in Alzheimer’s disease If more than occasional rule out Alzheimer’s disease

25. Neurons - Pathologic inclusions cont. (3) Hirano bodies - Oval to elongated rod-shaped, eosinophilic - Cytoplasmic but may appear to be in neuropil - Hippocampal pyramidal neurons, CA1 sector - Cytoskeletal elements including -actinin - Few in normal elderly patients, increased in Alzheimer’s disease

26. Neurons - Pathologic Inclusions (4) Lewy bodies: - Large, homogenous, eosinophilic, - Halos +/-, cytoplasmic - Immunoreactive for -synuclein, ubiquitin, neurofilament, & crystallin - Parkinson’s disease - substantia nigra, locus ceruleus locations - Lewy body dementia - Cortical (more difficult to see without special stains

27. Neurons - Pathologic Inclusions cont. (5) Pick bodies: - Cytoplasmic, round to oval, slightly basophilic and difficult to see on H&E - In swollen neurons in Pick’s disease - Easily demonstrated with silver stains - Immunoreactive for P-tau, ubiquitin, neurofilament Bielschowsky H & E

28. Neuronal Differentiation in Tumors Nissl substance (RER) - Cresyl violet stain ImP stains (synaptophysin, neurofilament, etc.) EM (dense - core vesicles, etc.) Neuronal tumor cell rosettes: “True” rosette with central lumen “Pseudorosette - no central lumen Flexner-Wintersteiner rosette (retinoblastomas and PNETs) Homer-Wright rosette (Neuroblastoma, Medulloblastoma, PNETs)

29. Glia GFAP S100 Astrocytes (“star” cells) Oligodendrocytes (“few branch glia”) Ependymal cells Tanycytes Choroid plexus epithelium Microglia + + - + - + + + - + - -

30. Astrocytes H & E: Oval nuclei floating in a fibrillar matrix GFAP or S100: Radiating cytoplasmic processes Two main types: Fibrillary (white matter) - majority with numerous and extensive branches Protoplasmic (gray matter) - fewer branches Subtypes - Bergman glia (cerebellar cortex) - Pilocytic (periventricular, cerebellar, & spinal cord white matter) GFAP + reactive astrocyte

31. Astrocytes - Cellular Responses Gliosis - rapid primary reaction to CNS injury - can be chronic leading to dense fibrillary gliosis (CNS version of scar) - typical and special types - diffuse swelling of cortical astrocytes can contribute to the risk of edema (normally 1/3 volume of cerebral cortex) Astrocytic reaction in progressive multifocal leukoencephalopathy (PML) - bizarre nuclei with atypia that suggests a malignant astrocytoma Inclusions

32. Gliosis continued Typical - well-defined cytoplasm on H&E that is slightly to markedly increased. The distribution of reactive astrocytes is best seen with GFAP (brown, stellate) and is uniform. Gemistocytes are “stuffed cells” as shown. Special types: - Bergman gliosis (seen around cerebellar infarcts) - Chaslin gliosis - subpial - Alzheimer type II (“empty” irregularly shaped nuclei, liver disease)

33. Astrocytes - Cellular Responses Inclusions - Age related Lipofuscin pigment accumulation Corpora amylacea (polyglucosan bodies) - concentrically laminated basophilic spheres - glucose polymers in astrocytes (PAS +) - prominent accumulations subpial & perivascular - may resemble fungi (+ stains) but no budding - no harm except in rare Polyglucosan Body Disease Subpial corpora amylacea

34. Astrocytes - Cellular Responses Inclusions continued... Rosenthal Fibers H&E: Brightly eosinophilic (magenta), hyalinized, elongated, carrot-like, corkscrew, or sausage- like, lumpy-bumpy profiles in astrocytes. Not as orange-red as stacks of erythrocytes. Masson trichrome: Bright red - crystallin 3 etiologies: Reactive (chronic conditions) Neoplastic (low-grade, slow growth) Alexander’s disease (mutated GFAP)

35. Astrocytes - Cellular Responses Inclusions cont... Rosenthal fibers cont... Reactive - typically around cystic lesions (ex. Pineal cyst, craniopharyngioma, vascular malformation) Neoplastic - characteristic of pilocytic astrocytomas and helpful diagnostically, especially when nuclear pleomorphism is suggestive of a higher grade tumor. Helps greatly in correctly identifying tumors with slow growth. Worthy of big searches.

36. Astrocytes - Cellular Responses Inclusions cont... Eosinophilic Granular Bodies, EGBs Round, finely granular, pink Also indicative of slow-growing, well- differentiated neoplasms (pilocytic astrocytomas, pleomorphic xanthoastro- cytomas, and gangliogliomas PAS + Variant: protein droplets clustered intracellularly, eosinophilic hyaline globules Herring bodies in neuro- hypophysis look similar but represent stored hormones

37. Oligodendrocytes (“few branch glia”) Produce and maintain CNS myelin “Satellite” around neurons in gray matter Columns between bundles of myelinated axons in white matter H&E: naked, small, dark, uniformly rounded, nuclei - frozen section - no halos - formalin fixed - perinuclear halos (“fried egg” appearance) ImmunoP: GFAP - S100 +

38. Oligodendrocytes - Cellular Responses Loss: leads to demyelination - Multiple sclerosis (MS) plaques Proliferation: - Edges of active MS plaque - “Myelination gliosis”, normal prolifer- ation of oligodendrocytes during develop- ment in preparation for myelination Inclusions: - JC virus in PML (enlarged, glassy nuclei due to viral inclusions) - intracytoplasmic inclusion bodies in some degenerative diseases seen with special stains

39. Astrocytes and Oligodendrocytes Secondary structures of Scherer - dependent upon interaction of infiltrating tumor cells with normal host tissue elements Ex. Perineural (perineuronal) satellitosis Surface growth (subpial accumulation) Perivascular satellitosis Others

40. Ependymal Cells Lining of ventricular system Cuboidal, columnar, ciliated epithelium No basement membrane, sits on neuropil Tanycytes - long processes contact blood vessels Ependymal granulations are nodular proliferations of astrocytes with focal loss of ependymal lining. Non-specific reaction to injury. Atrophy - flattened, decreased cilia, can be seen in hydrocephaly Inclusions - herpes, CMV Ependymal cell clusters (developmental rests)

41. Ependymal Cells Neoplasms - can see 2 types of rosettes Perivascular, pseudorosette - hallmark - neoplastic ependymal cells surround a blood vessel but leave a nuclear free zone around the vessel filled with cytoplasmic processes (shown below) “True” ependymal rosette, less common - central well-defined lumen rather than a blood vessel Blood Vessel

42. Choroid Plexus Specialized ependymal cells Layer of plump, cuboidal/columnar epithelial cells surrounding fibrovascular cores Frond-like projections into ventricles Secretes CSF (500 ml per day) S100 +, GFAP -, transthyretin (prealbumin) + Meningothelial whorls and nests - can see calcifications, psammoma bodies Xanthomatous change - foamy cells can form xanthogranulomas Neoplasms: Choroid plexus papillomas Choroid plexus carcinomas

43. Microglia Primary immune cells of the CNS Derived from monocytes (CD68+) Antigen presentation, phagocytosis, cytokine secretion, etc. Resting- oval to elongated nuclei, inconspicuous Activated- very elongated nuclei, “rod cells” Response to CNS injury - diffuse microgliosis or microglial nodules (ex. viral infection) Activated Resting

44. Tissue Macrophages of the CNS Closely related to microglia, CD68 + Gitter cells (“lattice cells”) Derived from circulating monocytes and indigenous microglia Spherical cells with well-defined cell borders Large clusters (easily identified) in destructive processes or scattered cells (can mimic the hypercellularity of gliomas) Increased mitotic activity & proliferation

45. Meninges Dura (pachymeninx) - dense connective tissue Leptomeninges (pia mater and arachnoid) Arachnoid: - spindled cells under dura - loosely arranged meningothelial cells (EMA+), collagen, fibroblasts, and blood vessels Pia: thin, membrane overlying the brain “Subdural space” - does not really exist, a potential space or path of least resistance for pathologic processes disrupting meninges Arachnoid villi - whorled groups of meningo- thelial cells that absorb CSF May see pigmented melanocytes Fibrosis with age

46. Meninges - Neoplasms Meningiomas most common Cellular whorls Psammoma bodies Difficult to distinguish cell borders EMA + Desmosomes on electron microscopy Whorl Psamomma body

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