Chapter 8 Cellular mechanisms of neurological disease 91

Chapter 8
Cellular mechanisms of
neurological disease

CHAPTER CONTENTS Neuronal injury and death

Neuronal injury and death 91 Nerve cells have a limited capacity to withstand pathological
Cell death mechanisms 94 stimuli. Cell death occurs when the neuron reaches a ‘point
Inflammation and gliosis 95
of no return’ following irreversible damage to the plasma
Neurodegeneration 97
Prion diseases 100 membrane, nuclear DNA or mitochondria. Since neurons are
post-mitotic cells (meaning that they are unable to divide)
they cannot usually be replaced in most parts of the brain
and spinal cord. Exceptions include the hippocampus and
The nervous system is subject to the full range of pathological olfactory bulb (where neurons can be replenished from a
processes found in other organs, together with a number of pool of stem cells). The two main forms of cell death are
unique degenerative and demyelinating diseases. The basic illustrated in Figure 8.1 and discussed below.
pathological processes underlying these disorders will be
discussed in this chapter (including inflammation, gliosis Necrosis
and neuronal cell death) before moving on to specific Tissue death resulting from damage or disease is referred to
examples of neurological disorders in the chapters that as necrosis (Greek: nekros, corpse). This is the end-point of
follow. Demyelination is discussed separately in Chapter 14, numerous pathological processes and has features in common
in the context of multiple sclerosis. with the dissolution of the body that occurs after death

Golgi apparatus
Mitochondria
Nucleus

Endoplasmic reticulum

Programmed
Cell injury
cell death

Loss of
membrane
integrity

Dissolution of Apoptotic bodies

cellular
components Intact organelles

NECROSIS APOPTOSIS

Fig. 8.1 Types of cell death.

92 CLINICAL NEUROSCIENCE

(autolysis). Necrosis is associated with cellular swelling, loss

of membrane integrity and influx of sodium and calcium ions, Death ligand
with eventual rupture of the cell. This is followed by digestion
of the cellular constituents by lysosomal enzymes. As the
Death receptor trimer
necrotic cell breaks down, its internal components are
Death receptor
discharged into the extracellular space, eliciting an
inflammatory reaction. Several patterns of necrosis are
recognized, but the type usually seen in the brain is called
liquefactive necrosis. In this process, dead brain tissue is
gradually removed by macrophages and liquefied by
hydrolytic enzymes. Complete resorption may take several
years, ultimately leaving only a fluid-filled cyst.

Apoptosis
Deliberate deletion of unwanted cells is termed
Death domain
programmed cell death. This is essential for normal growth
and development, but is also responsible for cell loss in a Procaspase-8
number of CNS pathologies including neurodegeneration,
stroke and multiple sclerosis.
The most common and best characterized form of Caspase-8
programmed cell death is apoptosis (pronounced: apa-
TOSIS). The term derives from the Greek and alludes to the
deliberate shedding of autumn leaves (Greek: apo, away from; Effector caspases
ptosis, falling). Apoptosis has characteristic microscopic
features including cell shrinkage, condensation of the nuclear
chromatin, cytoplasmic blebbing and formation of membrane- APOPTOSIS
bound apoptotic bodies which contain viable organelles.
Most newly formed nerve cells are primed to commit Fig. 8.2 The extrinsic pathway for apoptosis is triggered by cell
death ligands binding to receptors at the cell surface. This leads to
programmed cell death (‘cellular suicide’) unless rescued by
activation of caspase-8 within the cell, which initiates apoptosis.
exposure to the appropriate trophic factors (Greek: trophē,
nourishment). One of the best known examples of this is
the dependence of dorsal root ganglion neurons on nerve
growth factor (NGF). During CNS development, up to 50%
of neurons are deliberately deleted because they fail to (i)
reach their intended targets or (ii) make appropriate Extrinsic pathway (Fig. 8.2)
connections with other nerve cells. Apoptosis can be triggered by cell death ligands in the
Apoptosis is also an important mechanism for the extracellular fluid. These act on cell-surface receptors
destruction of critically injured or abnormal cells and is belonging to the tumour necrosis factor (TNF) family.
triggered in diseases where pathological stress has After ligand binding, death receptors cluster within the
compromised key cellular elements beyond repair. Irreparable membrane to form trimers and a conformational change in
damage to the nuclear DNA is a potent trigger for apoptosis, the cytoplasmic part of the protein exposes an ominously
contributing to the prevention of tumours. It is also used by named death domain. The intracellular region of the cell
activated lymphocytes to sacrifice virus-infected cells and to death receptor binds apoptotic proteins in the cytoplasm via
delete self-reactive T-lymphocytes in the thymus gland adaptor proteins. The interaction is mediated by death
(preventing autoimmune disease). effector domains (DEDs) that are present in both the
adaptor proteins and the procaspases. These interactions lead
Caspases to formation of a death-inducing signalling complex
Apoptosis is orchestrated by a family of proteolytic enzymes (DISC) which activates caspase-8 and thereby initiates
called caspases (cysteine-dependent aspartate-specific programmed cell death.
proteases) which cleave target proteins at aspartate residues.
The caspases are synthesized as inactive procaspases and Intrinsic pathway (Fig. 8.3)
once activated are able to cleave parts of the neuronal Apoptosis can also be triggered from within the cell if it is
cytoskeleton and nuclear DNA. exposed to excessive pathological stress. Activation of the
intrinsic pathway depends upon the balance of pro-apoptotic
■ Initiator caspases are activated by intrinsic (cell stress)
and anti-apoptotic factors, many of which are members of the
or extrinsic (cell death) signals. Examples include
BCL (B-cell lymphoma) family of proteins. Members that
caspases 2, 8, 9 and 10.
tend to oppose programmed cell death include Bcl-2 and
■ Effector caspases mediate programmed cell death by
Bcl-XL; others, such as Bad and Bax are pro-apoptotic. These
digesting the nuclear DNA and other key cellular
molecules act as stress sensors in the cytoplasm, responding
elements. Examples include caspases 3, 6 and 7.
to pathological stimuli by translocating to mitochondria.
Many components of the caspase cascade converge on the A key event is formation of a permeability transition
effector (or executioner) proteins, such as caspase-3, which pore (PTP) in the mitochondrial membrane, which is a large
can be used as a marker of apoptosis. Programmed cell death transmembrane pore (or ‘megachannel’). Pore formation
can be triggered by extrinsic or intrinsic stimuli. allows cytochrome c oxidase and apoptosis inducing
 Chapter 8 Cellular mechanisms of neurological disease 93

Apoptotic which translocates from the inner to the outer leaflet of the
stimulus plasma membrane. Phagocytes recognize and bind these
molecules and internalize the apoptotic bodies for
degradation. The entire process is carefully orchestrated and,
in contrast to necrosis, there is no inflammatory reaction.

Key Points
■ The two main types of cell death are necrosis and apoptosis.
■ Necrotic cell death is characterized by dissolution of the cell with
release of lysosomal enzymes and an inflammatory reaction.
Pro-apoptotic Bax Bcl–2 Anti-apoptotic Liquefactive necrosis is the most common type in the CNS.
+ –
factors Bad Bcl–XL factors ■ Apoptosis is the best-understood form of programmed cell
death. It is triggered by extrinsic (cell death) and intrinsic (cell
stress) pathways and mediated by initiator and executioner
Cytochrome C AIF
caspases.
■ Programmed cell death is important in embryogenesis, tumour
prevention, deletion of self-reactive (autoaggressive)
Effector caspases lymphocytes and destruction of virus-infected cells.
■ It also contributes to nerve cell loss in a range of neurological
disorders including neurodegenerative diseases, stroke and
APOPTOSIS multiple sclerosis.

Fig. 8.3 The intrinsic pathway for apoptosis depends upon the
balance of pro-apoptotic and anti-apoptotic members of the Bcl-2 Axonal damage
family. A key event is formation of the permeability transition pore (shown It is possible for axons to undergo selective degeneration
here in orange) in the mitochondrial membrane (coloured purple). This without death of the cell body. For instance, following
allows apoptosis-inducing factors to be released into the cytoplasm. transection of a peripheral nerve, the distal portions of
the affected axons degenerate together with their myelin
sheaths. This is termed Wallerian degeneration, which is
factor (AIF) to escape from mitochondria and reach the accompanied by regenerative changes in the parent cell, called
cytoplasm. Cytochrome c then forms a complex with Apaf-1 the axon reaction. These include swelling of the cell body,
(apoptotic protease activating factor 1) which recruits dispersal of the Nissl substance (chromatolysis) and
caspase-9 to form a larger multi-protein complex called the displacement and enlargement of the nucleus, reflecting
apoptosome. Following formation of the apoptosome, increased gene transcription and protein synthesis. If the
caspase-9 is activated and apoptosis is triggered, culminating regenerative attempt fails (which is usually the case in the
in the upregulation of cell death genes. CNS) then the parent cell will eventually undergo apoptosis.

Disposal of the cell Transneuronal degeneration (Fig. 8.4)

Once a cell is committed to programmed cell death, its DNA Following axonal transection, neuronal degeneration
and cytoskeleton are dismantled in an orderly manner. This is may spread to involve other nerve cells, referred to as
an active process that expends energy. A key step is activation transneuronal degeneration. For instance, following
of the enzyme caspase-activated DNAse (CAD) by effector interruption of an anatomical pathway consisting of a linear
caspases, which breaks down the DNA into nucleosomal chain of neurons, there may be subsequent loss of nerve cells
units. Organelles are packaged into membrane-bound ‘downstream’ (anterograde) or ‘upstream’ (retrograde) of
apoptotic bodies (see Fig. 8.1) which contain viable the original injury, which may take months or years. This
mitochondria. These structures express cell-surface markers reflects the general principle that nerve cells need to be
that trigger their internalization by neighbouring cells. An integrated into a functional network and receive trophic
example is the membrane constituent phosphatidylserine, signals from other neurons in order to remain viable.

t ransneuronal degene
rade ratio
trog n Wallerian
Re
degeneration

Neuron Neuron
Neuron
2 3
1

An
te ro on
grad rati
Axonal e trans e ne
transection neuronal deg

Fig. 8.4 Degeneration following axonal injury. Following axonal transection (neuron 2) the distal part of the nerve fibre undergoes Wallerian
degeneration. Other neurons may subsequently be lost in an anatomical pathway by transneuronal degeneration (neurons 1 and 3).
94 CLINICAL NEUROSCIENCE

Microtubules
Cell death mechanisms
Axon Cells are continuously subjected to physiological and
Filopodium
pathological stimuli to which they must adapt in order to
survive. Pathological stimuli leading to neuronal cell death
include excitotoxicity and oxidative stress, with accumulation of
excessive intracellular free calcium as a final common event.
Actin microfilaments
Excitotoxicity
Excessive stimulation by excitatory neurotransmitters (such as
Lamellopodium glutamate) can cause neuronal cell death in a process known
as excitotoxicity. Intense glutamatergic stimulation leads to
prolonged neuronal depolarization, lifting the magnesium
blockade of NMDA (N-methyl D-aspartate) receptors. In
this situation, free calcium ions are able to flood the neuronal
cytoplasm via liberated NMDA receptors, as well as via
calcium-permeable AMPA (alpha-amino, 3-hydroxy-4-
Fig. 8.5 A growth cone. isoxasole-propionic acid) receptors and voltage-gated
calcium channels (see Ch. 7). This leads to further
depolarization, with additional glutamate release, generating a
vicious cycle. In addition to acute excitotoxic injury, there is
Axonal regrowth evidence that low-grade excitotoxicity may cause chronic
Axons are able to regenerate following peripheral nerve neuronal damage in some disorders (e.g. motor neuron
damage and may re-establish connections with muscle fibres disease; see Ch. 4, Clinical Box 4.9). Accumulation of
or glands. The distal tips of the severed axons form growth intracellular free calcium is an important final common event
cones (Fig. 8.5) which ‘crawl’ along residual Schwann in excitotoxicity neuronal cell death.
cell basement membranes to reinnervate target structures.
This process occurs after peripheral nerve injury (see The role of calcium
Clinical Box 8.1) but does not seem to be possible in the brain The free calcium ion concentration in the cytoplasm is
and spinal cord. normally kept very low by several mechanisms including
sequestration by calcium-binding proteins (e.g.
parvalbumin and calbindin) and export from the cell. The
high calcium influxes generated by excitotoxic stimulation
overwhelm buffering and extrusion mechanisms, leading to
Clinical Box 8.1: Bell’s palsy activation of harmful calcium-dependent enzymes, including:
Bell’s palsy is a unilateral facial paralysis resulting from ■ Calpains, which degrade the neuronal cytoskeleton.
inflammation of the facial nerve. The cause is uncertain, ■ Proteases, which digest structural proteins and enzymes.
but it may be related to a virus infection. Presentation is abrupt and ■ Phospholipases, which impair the integrity of cell
is sometimes preceded by otalgia (earache). Damage to the facial
membranes.
nerve may lead to changes in the sense of taste or sensitivity to loud ■ Endonucleases, leading to DNA fragmentation.
noises. This is because the facial nerve also supplies taste buds in
the anterior two thirds of the tongue and the stapedius muscle, Several pro-apoptotic genes are also upregulated by calcium-
which prevents excessive vibration of the ear drum. In most cases mediated cascades, which promotes degradation of the
symptoms improve spontaneously as the damaged axons slowly cytoskeleton and may initiate programmed cell death.
grow back. Recovery is often imperfect, with aberrant reinnervation Calcium-mediated activation of xanthine oxidase and nitric
of muscles and glands, leading to symptoms such as eye closure on oxide synthase may also lead to oxidative stress.
attempting to smile or crocodile tears in place of salivation.
Oxidative stress
Oxidative phosphorylation in normal mitochondria generates
potentially harmful reactive oxygen species (ROS) or
free radicals, including superoxide anion (O2−) and
Key Points hydroxyl radical (OH−). These are highly reactive species
with unpaired electrons that can damage cell membranes,
■ Axonal transection leads to degeneration of the distal portion of
proteins and DNA. The body has a number of scavenging
the axon, together with its myelin sheath (termed Wallerian
mechanisms and molecules to deal with free radicals,
degeneration).
including antioxidant vitamins (especially C and E) and three
■ The axon reaction (in the cell body) reflects an attempt by the
key enzymes:
cell to regrow the axon and establish new connections with the
denervated target structures ■ Superoxide dismutase.
■ Axonal regrowth is commonly seen following peripheral nerve ■ Catalase.
injury, but does not occur in the brain or spinal cord. ■ Glutathione peroxidase.
■ Interruption of an anatomical pathway may lead to subsequent
Mitochondria continuously produce superoxide anion which is
degeneration in nerve cells proximal or distal to the original
catabolized by superoxide dismutase to form hydrogen
lesion (termed transneuronal degeneration).
peroxide (H2O2). This is also a reactive oxygen species and is
 Chapter 8 Cellular mechanisms of neurological disease 95

Key Points
■ Glutamate-mediated excitotoxicity is a common cell death
mechanism in many neurological diseases, mediated by influx of
excessive free calcium via NMDA receptors.
Membrane damage ■ High intracellular calcium levels trigger enzymes such as
(lipid peroxidation) calpains, proteases, phospholipases and endonucleases that
digest key cellular elements including the cytoskeleton and DNA.
Reactive ■ Reactive oxygen species (ROS) are continuously produced by
oxygen mitochondria and removed by natural scavenging mechanisms
species (e.g. superoxide dismutase, catalase, glutathione peroxidase).
■ Free radicals (e.g. superoxide anion, O2−; and hydroxyl radical,
Oxidative stress OH−) are highly reactive and damage cells by forming abnormal
cross-linkages between lipids, carbohydrates and proteins.
■ Excessive production of free radicals including nitric oxide
contributes to cell death in neurodegenerative diseases, multiple
sclerosis and stroke.

Protein damage DNA damage

(cross-linking) (cross-linking/breakage)

Fig. 8.6 Oxidative stress. Reactive oxygen (and nitrogen) species damage
key cellular components such a proteins, cell membrane and DNA, leading
to cell death.

degraded by catalase. Excessive generation of free radicals or

reduced capacity of the normal scavenging mechanisms leads
to oxidative stress, with abnormal cross-linkages forming
between nucleic acids, lipids, carbohydrates and proteins as free
radicals react with them indiscriminately (Fig. 8.6). Oxidative
stress may be exacerbated by age-related mitochondrial
Fig. 8.7 Astrocytosis. Photomicrograph showing the spider-like
abnormalities due to mutations in mitochondrial DNA that appearance of reactive astrocytes using immunohistochemistry (antibody
accrue during the lifetime of an individual. labelling) for glial fibrillary acidic protein (GFAP).

Nitric oxide
Nitric oxide gas is synthesized from L-arginine by isoforms of Reactive gliosis
the enzyme nitric oxide synthase (NOS). It is a free radical Gliosis (also known as reactive gliosis) consists of activation
species with a number of important physiological roles (e.g. and proliferation of glial cells, stimulated by inflammatory
as a vasodilator, transmitter substance and regulator of cytokines including interleukin-1 (IL-1), tumour necrosis
inflammatory and immune responses). Its synthesis is factor alpha (TNF-α) and interleukin-6 (IL-6). It is a
induced by glutamate signalling via the calcium-permeable combination of astrocytosis and microgliosis.
NMDA receptor and excessive production of nitric oxide is a
feature of excitotoxicity. Astrocytosis (Fig. 8.7)
Nitric oxide reacts with superoxide anion to produce Following brain injury, nearby astrocytes enlarge, multiply
peroxynitrite (ONOO−). This is a reactive nitrogen species and increase their expression of glial fibrillary acid
that may damage proteins by interacting with cysteine and protein (GFAP). Proliferation of astrocytes may be sufficient
tyrosine residues. Although nitric oxide has an anti-apoptotic to fill in a small tissue defect, but larger areas of damage (e.g.
effect in many cell types, excessive production contributes to following a major stroke, see Ch. 10) are transformed into a
cell death in neurodegenerative diseases, multiple sclerosis fluid-filled cystic cavity lined by a glial scar. Astrocytes secrete
and stroke. (i) cytokines that recruit inflammatory cells from the blood
and (ii) various trophic factors, including:
■ Nerve growth factor (NGF).
Inflammation and gliosis ■ Brain-derived neurotrophic factor (BDNF).
■ Glial-cell-line-derived neurotrophic factor (GDNF).
The body responds to pathological insults with an
inflammatory reaction in which blood vessels become These are chemical mediators that promote neuronal survival
‘leaky’ so that protein-rich fluid and inflammatory cells can and axon sprouting. They are released into the extracellular
enter the tissues. Gliosis is a unique response to damage that fluid, but can also be delivered directly to the neuronal
only occurs in the brain and spinal cord. cytoplasm via intercellular gap junctions (see Ch. 7).
96 CLINICAL NEUROSCIENCE

Clinical Box 8.2: Acute

bacterial meningitis
The main clinical features of meningitis are fever,
headache and drowsiness, together with nausea, vomiting and
photophobia (sensitivity to light). Clinical examination may show
neck stiffness and evidence of raised intracranial pressure including
papilloedema (swelling of the optic discs; see Ch. 5, Fig. 5.16).
Unlike viral meningitis, a self-limiting illness from which most
patients make a complete recovery, the mortality in bacterial
meningitis (Fig. 8.9) is around 10% and a significant proportion of
survivors are left with neurological deficits, cranial nerve palsies,
hydrocephalus or epilepsy.

Fig. 8.8
Microgliosis. Photomicrograph of reactive microglial cells using
immunohistochemistry for the macrophage marker CD68.

Microgliosis (Fig. 8.8)

Microglia are the resident phagocytes (scavengers) of the
brain. They normally exist in a ramified, quiescent state,
but following tissue injury they become activated in response
to inflammatory cytokines and growth factors. Activated
microglia migrate towards injured tissues by following
chemotactic gradients. They differentiate into macrophages
and internalize cellular debris and microorganisms. Those
that have ingested myelin debris form lipid-laden foam cells.
Activated microglia are immunocompetent cells that
express MHC class II (major histocompatibility) proteins and
are antigen-presenting cells. They may therefore contribute
to T-cell-mediated immune responses and have been
implicated in the inflammatory demyelinating disease
multiple sclerosis (Ch. 14). Microglial activation is also a
component of most neurodegenerative disorders such as
Alzheimer’s disease and Parkinson’s disease (Chs 12
and 13).

Acute and chronic inflammation

A number of terms are used to describe the site and
distribution of acute and chronic inflammation in the
nervous system. Meningitis is inflammation of the
Fig. 8.9 Acute bacterial meningitis. Post-mortem photograph
showing thick green-yellow purulent exudate filling the subarachnoid
meninges (protective coverings of the brain). The term space over the surface of the cerebral hemispheres. From Kleinschmidt-
pachymeningitis is used if the dura is predominantly DeMasters, BK and Tyler, KL: Practical Surgical Neuropathology:
affected or leptomeningitis if inflammation is centred on A Diagnostic Approach (Churchill Livingstone 2010) with permission.
the arachnoid, pia and subarachnoid space (see Ch. 1). The
features of acute bacterial meningitis are discussed in
Clinical Box 8.2. Key Points
■ Reactive gliosis is the unique CNS response to damage or
Inflammation in the CNS
disease and consists of astrocytosis and microgliosis, coordinated
A small focus of inflammation in the brain is referred to as
by inflammatory cytokines such as IL-1, TNF-α and IL-6.
cerebritis. More extensive and diffuse brain inflammation is
■ Reactive astrocytes enlarge and proliferate, increasing synthesis
termed encephalitis. Encephalitic processes are further
of GFAP, to form a glial scar. Astrocytes also release
subdivided into three main types:
inflammatory mediators and trophic factors.
■ Polioencephalitis (Greek: polios, grey) is grey-matter ■ Activated microglia differentiate into macrophages and
predominant. internalize cellular debris and microorganisms. They are also
■ Leukoencephalitis (Greek: leukos, white) is white- immunocompetent antigen-presenting cells.
matter predominant. ■ Inflammation of the meninges is referred to as pachymeningitis
■ Panencephalitis (Greek: pan-, all) affects both grey and (if it affects the dura) or leptomeningitis (when it is centred on
white matter. the pia-arachnoid). Inflammation of the brain is termed cerebritis
(if focal) or encephalitis (if diffuse). Myelitis is inflammation of the
The term myelitis indicates inflammation of the spinal cord,
spinal cord.
such as the inflammatory spinal cord disease poliomyelitis
 Chapter 8 Cellular mechanisms of neurological disease 97

(caused by infection with a poliovirus); whereas combined Normal protein folding

inflammation of the brain and spinal cord is referred to as Nuclear DNA encodes the primary structure of proteins,
encephalomyelitis. It should be emphasized that these consisting of the basic amino acid sequence. Attainment of
terms are descriptive and do not indicate the underlying cause the correct three-dimensional conformation requires protein
of the inflammation. folding (Fig. 8.11). This transforms the linear amino acid
sequence into more complex spatial arrangements with
alpha-helices and beta-pleated sheets that make up the
secondary structure. Further folding gives rise to a three-
Neurodegeneration
dimensional globular protein with a particular tertiary
The neurodegenerative diseases are a heterogeneous group of structure. Association with other proteins may occur, to form
progressive, incurable neurological disorders that are more a multi-protein complex with its own quaternary structure.
common in later life. They often present with dementia (e.g. Protein folding relies on physical and chemical properties of
Alzheimer’s disease; Ch. 12) or as a movement disorder (e.g. the constituent amino acid residues (i.e. attraction and
Parkinson’s disease; Ch. 13). Most cases are classified as repulsion by hydrogen bonds, electrostatic forces and
sporadic or idiopathic, meaning that the cause is not known. hydrophobic interactions).
Inherited forms often exist, but they are less common and
tend to present at an earlier age. Unfolded protein response
The unfolded protein response (UPR) is triggered by the
General features presence of misfolded proteins in the endoplasmic reticulum
In most neurodegenerative diseases there is a selective loss of or as a result of errors in post-translational modification. It is
certain populations of nerve cells, associated with deposits of a type of cell stress response characterized by upregulation
an abnormal protein or peptide in neurons and/or glia. These of molecular chaperone proteins that attempt to refold
disorders are therefore referred to as proteinopathies (Fig. abnormally configured proteins. In the event of an
8.10). In cases where inherited (familial) forms of a disease overwhelming pathological stimulus or major dysfunction of
have been identified, the mutation often affects the protein the normal mechanisms for protein disposal, large numbers
itself or an enzyme involved in its processing. of misfolded proteins may aggregate in the cytoplasm. When
this happens they tend to precipitate out of solution as
Neuronal loss and gliosis insoluble clusters called micelles, since hydrophobic groups
The affected brain regions show neuronal loss and that would normally be buried are exposed to the aqueous
reactive gliosis in a disease-specific pattern, affecting environment of the cell.
particular groups of neurons, but sparing others. Importantly,
the clinical features in each case are determined by the Disposal of abnormal proteins
anatomical distribution of the pathological changes rather Abnormal proteins are earmarked for destruction by tagging
than the particular protein involved. In most cases it is not them with the 8.5 kDa protein ubiquitin (Fig. 8.12). This is
clear why specific populations of neurons are susceptible attached in a series of enzyme-catalysed steps involving
whilst others are resistant (referred to as selective activating (E1), conjugating (E2) and ligating (E3) enzymes.
vulnerability). These add a polyubiquitin chain that is composed of
multiple ubiquitin monomers. Once ubiquitinated, the
Protein folding and misfolding abnormal protein is targeted to the proteasome. This is a
An important component of many neurodegenerative diseases large multi-subunit protein complex that digests proteins into
is the accumulation of abnormally folded proteins – or failure peptide fragments and amino acids in an active (ATP-
of the normal cellular mechanisms for their disposal. dependent) process.

Main clinical features Protein

Parkinson’s disease Parkinsonism Alpha-synuclein

Dementia with Lewy bodies Dementia Alpha-synuclein Synucleinopathies
Multiple system atrophy Parkinsonism, cerebellar ataxia Alpha-synuclein

Alzheimer’s disease Dementia Tau

Progressive supranuclear palsy Parkinsonism Tau Tauopathies
Dementia pugilistica (boxers) Dementia, parkinsonism Tau

Motor neuron disease Progressive paralysis TDP-43

TDP-43 proteinopathies
Frontotemporal dementia* Dementia, personality change TDP-43

Huntington’s disease Dementia, chorea, personality change Huntingtin

Creutzfeldt-Jakob disease Dementia, ataxia Prion protein

Fig. 8.10 Proteinopathies. A number of common and/or important neurodegenerative disorders are shown, together with the main protein that
accumulates in neurons or glial cells. *[NB: approximately 50% of frontotemporal dementias are TDP43-proteinopathies; in other cases the pathological
inclusions are composed of tau (40%) or other proteins (10%)].
98 CLINICAL NEUROSCIENCE

Primary amino Beta-pleated

acid sequence sheet

Molecular
chaperones

Alpha helix

Unfolded protein Folded protein

Fig. 8.11 Protein folding. The amino acid sequence of an unfolded protein is encoded by the nuclear DNA (primary structure). This leads to spontaneous
formation of alpha helices and beta-pleated sheets (secondary structure) and further folding to produce a globular protein (tertiary structure). The process of
protein folding is promoted and accelerated by molecular chaperone proteins.

Ubiquitin-activating Fig. 8.12 The

Monomeric enzyme (E1) Activated ubiquitin-proteasome
ubiquitin ubiquitin system. Damaged or
incorrectly folded
proteins are tagged with
ubiquitin and earmarked
for degradation (to amino
acids) by the proteasome.
Ubiquitin-conjugating
Digestion within the
enzyme (E2)
proteasome is an
ATP-dependent process
Polyubiquitin that consumes cellular
chain energy. Conversion of
monomeric ubiquitin to
its active form (see upper
part of figure) is also an
ATP-dependent step.
Ubiquitin
ligase (E3) Damaged
protein
Proteasome
Ubiquitinated
protein

Amino acids

The proteasome consists of a cylindrical core region (the The autophagy-lysosomal pathway
20S core particle) composed of four rings stacked around A second disposal mechanism for proteins and other cellular
a central pore. This is capped at either end by regulatory components is autophagy (Greek: autos, self; phagein, eat).
particles (19S or 11S) which also have a central pore that The abnormal protein is first enveloped in an autophagic
allows access to a hollow degradation chamber. The entire vacuole. This then fuses with a primary lysosome which
assembly is the 26S proteasome. Proteins are fed into the contains powerful hydrolytic enzymes that digest the contents.
proteasome and digested, whilst the polyubiquitin chain is Any materials left over from the process, such as undigested
recycled. Accumulation of ubiquitin-tagged proteins is a cell membrane constituents, remain within membrane-bound
feature of many neurodegenerative disorders, sometimes due residual bodies. This mechanism is particularly important for
to disturbance or overload of the ubiquitin-proteasome protein aggregates that cannot be cleared by the proteasome.
system. Disruption of the autophagy pathway is important in some
 Chapter 8 Cellular mechanisms of neurological disease 99

Key Points
■ Deposition of abnormal proteins occurs in most
neurodegenerative diseases, which can therefore be classified as
proteinopathies.
■ This is accompanied by neuronal death and reactive gliosis and
the clinical features depend mainly on the extent and distribution
of neuronal loss rather than the particular protein involved.
■ In some degenerative diseases there is abnormal protein folding
or disturbance of the normal cellular mechanisms that deal with
(or dispose of) improperly folded proteins.
■ These include the unfolded protein response, molecular
chaperone proteins, the ubiquitin–proteasome system and the
autophagy–lysosomal pathway.

A
forms of familial Parkinson’s disease caused by mutation
in the ATP13A2 gene (encoding a lysosomal ATPase).

Protein aggregation
Aggregation of abnormal proteins in neurons (or glial cells)
gives rise to structures called inclusion bodies. These are
difficult to identify by routine light microscopy and are better
visualized by silver staining or immunohistochemistry
(antibody-labelling of specific proteins). Intracellular inclusions
are often cytoplasmic (e.g. in Alzheimer’s and Parkinson’s
diseases; Fig. 8.13) but in some cases are found within the
nucleus. Abnormal proteins may also accumulate in the
extracellular compartment (between nerve cells).
B
Amyloid
Fig. 8.13 Neuronal inclusions. (A) A neurofibrillary tangle (composed of
Many proteins and peptides that form pathological aggregates
abnormally phosphorylated tau protein) in the cytoplasm of a cortical
have a beta-pleated sheet structure. This enables monomers pyramidal neuron in a patient with Alzheimer’s disease [demonstrated using
to stack together to form elongating protofibrils and fibrils the modified Bielschowsky silver stain]. From Prayson, R: Neuropathology 1e
of around 10 nm in diameter, stabilized by hydrogen bonds. (Churchill Livingstone 2005) with permission; (B) Micrograph showing two
Deposits of these insoluble protein fibrils are referred to as neurons in the substantia nigra in a patient with Parkinson’s disease. The
amyloid. It is important to emphasize that the term ‘amyloid’ neuron on the left has a large nucleus and prominent nucleolus with
abundant brown (neuromelanin) pigment in the cytoplasm. The nucleus of
does not refer to one particular protein and that many different
the neuron on the right is not fully seen in this section, but the cytoplasm
peptides with a beta-sheet structure can form ‘amyloid deposits’. contains a bright pink Lewy body which is surrounded by a characteristic
The name derives from the Greek, meaning starch-like. pale halo [Routine haematoxylin and eosin (H&E)-stained section]. Courtesy
All forms of amyloid take up certain tissue stains such as of Professor Steve Gentleman.
Congo red and thioflavin S. Due to the regular, crystalline
arrangement of the amyloid fibrils, the deposits also have the Amyloid diseases
ability to rotate the plane of polarized light, termed Deposition of amyloid is responsible for diseases in many
birefringence. As a result, amyloid deposits stained with different organ systems, but the most common and best
Congo red have a characteristic apple green colour when understood amyloid disorder is Alzheimer’s disease. It is
viewed under polarized light (Fig. 8.14). characterized by the deposition of amyloid beta (Aβ)
peptide in the extracellular compartment of the brain
Amyloid fibril formation (between nerve cells) in the form of amyloid plaques.
The process of amyloid formation is illustrated in Figure 8.15. Secondary pathological changes occur in the neuronal
Fibrillogenesis is the process by which a peptide forms cytoplasm, with accumulation of an abnormally
insoluble aggregates of amyloid. It involves three sequential phosphorylated form of the microtubule-associated protein
steps. A peptide with a beta-pleated sheet structure must tau. Hyperphosphorylated tau is found within nerve cells as
first be produced in sufficient quantities. The second step is filamentous structures called neurofibrillary tangles. Since
nucleation, which starts the process of fibril formation. It neuronal injury and tau deposition appear to occur as a
requires a supportive microenvironment with a sufficiently consequence of amyloid accumulation, Alzheimer’s disease is
high protein concentration, together with various permissive referred to as a secondary tauopathy. This is in contrast to
factors including appropriate acidity (pH), temperature or the the primary tauopathies, in which abnormal tau-positive
presence of certain metallic ions. Finally, the phase of fibril inclusions are present in the absence of Aβ.
growth involves the sequential addition of monomeric
peptide units (each with a beta-sheet structure) to form an Amyloid toxicity
extending chain. This leads to the gradual assembly of It is not certain how abnormal protein aggregates cause
oligomeric species (or protofibrils) which associate to neuronal damage, but factors that have been implicated
form mature amyloid fibrils. include inflammation, gliosis, oxidative stress and production
100 CLINICAL NEUROSCIENCE

Amyloidogenic
monomers

+ +
Beta-
pleated
sheets

A
A Stacking monomers

B B Fibril formation C Amyloid fibrils

Fig. 8.15 Amyloid formation. Amyloidogenic monomers with a
Fig. 8.14 Amyloid. (A) Photomicrograph showing deposition of amyloid beta-pleated sheet structure are able to stack together (A) to form
in a blood vessel in a case of cerebral amyloid angiopathy [stained with elongating chains that are stabilized by hydrogen bonds. These gradually
Congo red]; (B) Microscopy using polarised light to show the characteristic grow to form protofibrils (B) and mature amyloid fibrils (C).
apple-green birefringence of amyloid [Congo red stain]. From Ellison and
Love: Neuropathology 2e (Mosby 2003) with permission.

Key Points
of toxic intermediates during fibrillogenesis. There is growing
■ Amyloid is a general term used to describe insoluble aggregates
evidence that oligomeric intermediates generated during
of various types of protein, all of which have a beta-pleated
amyloid fibril formation may be primarily responsible for
sheet structure that enables them to stack together and form
neurotoxicity in a number of neurodegenerative disorders
fibrils.
(including Alzheimer’s disease and Parkinson’s disease).
■ Many different proteins and peptides can form amyloid deposits,
In particular, it has been shown in cultured cells that
all with properties in common (e.g. staining with Congo red and
oligomeric prefibrillary species of amyloid beta are able to
showing apple-green birefringence under polarized light).
form a membrane attack complex that can perforate the
■ The most common amyloid disorder is Alzheimer’s disease, in
neuronal cell membrane. This leads to cellular swelling, loss
which Aβ (amyloid beta) peptide forms insoluble aggregates
of transmembrane gradients and influx of free calcium ions
(plaques) in the cerebral cortex.
– all of which promote neuronal cell death. A similar
■ Amyloid deposition is neurotoxic, but the mechanism of
phenomenon has been described with oligomers of alpha-
neuronal injury is not certain and may include oxidative stress,
synuclein protein (the main pathological species implicated
abnormal phosphorylation and calcium influx.
in Parkinson’s disease and related synucleinopathies).
■ Oligomeric intermediates created during the process of amyloid
formation have been particularly implicated and may form
‘pores’ in the plasma membrane that compromise the cell.
Prion diseases
The prion diseases are a group of extremely rare
neurodegenerative disorders characterized by vacuolar devastating neurological decline that quickly ends in death. In
degeneration of the cerebral cortex, which is termed humans, the classical form of prion disease is Creutzfeldt–
spongiosis. They are caused by a unique form of infectious Jakob disease or CJD, a sporadic disorder of later life
pathogenic agent composed only of protein: the prion characterized by an aggressive and rapidly fatal dementia
(‘proteinaceous infectious particle’). (Clinical Box 8.3). There are a few very rare genetic forms of
prion disease including fatal familial insomnia (FFI) and
General characteristics Gerstmann–Sträussler–Scheinker syndrome (GSS) that
Prion diseases have been described in a number of animals are all inherited in an autosomal dominant manner. Each of
including sheep (scrapie) and cattle (bovine spongiform these has distinguishing clinical and pathological features, but
encephalopathy or BSE) in which they cause a rapid and all are incurable and ultimately fatal.
 Chapter 8 Cellular mechanisms of neurological disease 101

Clinical Box 8.3: Sporadic CJD Clinical Box 8.4: New variant CJD
Creutzfeldt–Jakob disease is a sporadic The new variant of CJD was first identified by its
neurodegenerative disease which affects one in a million microscopic appearance. This differs from that of classical
people worldwide per year, most of whom are over the age of 65. It CJD and includes characteristic florid plaques, named for their
is characterized by a very aggressive dementia that leads to death resemblance to flowers (Fig. 8.17). The new variant affects younger
in a matter of weeks. This is often accompanied by abnormal people (usually below the age of 30) and has a longer duration,
movements including electric-shock-like myoclonic jerks. The EEG typically a year or more. Psychiatric features and cerebellar
shows characteristic periodic complexes in 60–80% of cases, ataxia are also common and myoclonic jerks are absent. MRI
consisting of biphasic or triphasic ‘sharp waves’ at a frequency of scanning characteristically shows T2-hyperintensity in the posterior
1–2 Hz. The cerebrospinal fluid may contain increased levels of thalamus (the pulvinar sign). As with classical CJD, the condition is
certain proteins (e.g. 14-3-3 and S100b). Examination of the brain incurable and fatal, but it is much rarer, with fewer than 180 cases
after death shows widespread spongiform degeneration of the reported in the UK (representing 80% of the worldwide total).
cerebral cortex (Fig. 8.16), accompanied by neuronal loss, gliosis and
deposition of abnormal prion protein.

Fig. 8.17 Florid plaques in new variant Creutzfeldt–Jakob

disease (vCJD). Several plaques are shown, consisting of a central
Fig. 8.16 Spongiform degeneration of the cerebral cortex amyloid core, composed of abnormal prion protein, surrounded by
(‘spongiosis’) in Creutzfeldt–Jakob disease (CJD). Routine coarse vacuoles, which creates a ‘florid’ or floral appearance [routine
haematoxylin and eosin (H&E) stain. haematoxylin and eosin (H&E) stain].

Infectivity
In addition to sporadic and inherited forms, prion diseases
Key Points
have the unique property (among degenerative diseases) ■ CJD is a very rare neurodegenerative process, affecting one in a
of infectivity. They are therefore also referred to as million people each year worldwide, characterized by aggressive
transmissible spongiform encephalopathies (TSEs). dementia, ending in death within a matter of weeks.
In the 1990s in the UK and Europe, bovine spongiform ■ The pathological features include spongiosis (vacuolar
encephalopathy entered the human food chain via degeneration of the cerebral cortex) and deposition of abnormal
contaminated beef and led to a new variant of CJD prion protein, together with neuronal loss and gliosis.
(Clinical Box 8.4). Prion diseases have also been transmitted ■ Most cases of prion disease are sporadic (and some are
iatrogenically (Greek: iatros, doctor) by blood transfusions inherited) but can also be transmitted as an infectious disease
and growth hormone supplements (obtained from human and are known as transmissible spongiform encephalopathies.
donors) and various neurosurgical procedures (via ■ In the UK in the 1990s, contaminated beef from BSE-infected
contaminated instruments or dural grafts). cattle entered the human food chain, leading to a new variant
of CJD which has distinct clinical and pathological features.
Prion protein
The infective agent in prion disease is unique since it is Codon 129
non-cellular, has no DNA or RNA and appears to be composed There is a common polymorphism in the general population
entirely of protein (the ‘protein only hypothesis’). It is an that affects codon 129 of the prion gene (PRNP, located on
abnormally folded form of cellular prion protein (or PrPc) chromosome 20) which codes either for methionine (M) or
which is present in many different tissue types. The pathogenic valine (V). Since each gene has two alleles, there are three
form has the same primary amino acid structure, but a possible genotypes with different population frequencies:
different secondary structure that is rich in beta-pleated
■ MV (50%)
sheets rather than alpha-helices and is able to form amyloid
■ MM (40%)
deposits. This infective form is designated PrP Sc (scrapie
■ VV (10%)
variant). The classical model of normal and abnormal prion
protein structure is illustrated in Figure 8.18, but the actual All pathologically confirmed cases of variant CJD have
three-dimensional structure is not known with certainty. occurred in people who are homozygous for methionine at
102 CLINICAL NEUROSCIENCE

Fig. 8.18 Structure of prion protein.

PrPc PrPsc (A) Normal prion protein, with a
secondary structure that is rich in alpha
helices; (B) Abnormal prion protein has
an identical amino acid sequence but is
abnormally folded; abnormal prion
protein is rich in beta-pleated sheets.

Alpha helix

Beta-pleated
sheet

A B

codon 129 (MM). This has therefore been referred to as the

susceptibility genotype, which implies that the other forms
confer resistance to infection. In keeping with this idea, mice
that are homozygous for valine (VV) are virtually immune to Amyloid
inoculation with abnormal prion protein whereas formation
heterozygotes (MV) show intermediate susceptibility.
However, since the incubation period for prion diseases is
sometimes measured in decades, the possibility remains that
new cases of variant CJD will eventually emerge in people
who do not have the susceptibility genotype. Normal prion protein
(PrPc)

Conversion of PrPc to PrPSc

In sporadic disease the abnormal prion protein is assumed
to arise by spontaneous transformation of native prion
protein to the abnormal (scrapie) form. This is Conversion
thermodynamically unfavourable and is an extremely
unlikely spontaneous event, which presumably explains why
Heterodimer Homodimer
sporadic disease is so rare.
Animal models have shown that propagation of abnormal
prion protein can only take place if the normal cellular
protein is present. For instance, prion-knockout mice are
completely resistant to infection, but infectivity can be
Abnormal prion protein
restored if native prion protein is reintroduced. (PrPsc)
Conversion models Fig. 8.19 A model of prion replication. One possible mechanism of
There are two main models that attempt to describe how prion formation [illustrated here] is called template-directed refolding. The
cellular prion protein may be converted to the abnormal abnormal prion protein recruits normal (cellular) prion protein and induces a
conformation change, converting it to abnormal prion protein. The
form. The first is the template-directed refolding
abnormal protein is rich in beta pleated sheets which can stack to form
(heterodimer) model (Fig. 8.19). This suggests that amyloid.
 Chapter 8 Cellular mechanisms of neurological disease 103

abnormal prion protein is able to recruit the cellular form

and form a heterodimer with it, acting as a template to
Key Points
catalyse its conversion to the scrapie form. The converted ■ The pathogenic agent in transmissible spongiform
prion can then recruit more cellular protein or polymerise to encephalopathies such as CJD and variant CJD is the prion
form amyloid fibrils. (proteinaceous infectious particle). This is a unique pathogen
Another possibility is described by the nucleation– that appears to be composed entirely of protein.
polymerization model which suggests that the critical event ■ The pathogenic (scrapie) form of prion has the same primary
is the formation of a nucleus (which consists of an oligomeric structure as the native protein, but a different secondary
aggregate of prion protein). This initial ‘seeding’ event is structure that is rich in beta-pleated sheets.
highly unlikely since it is thermodynamically unfavourable ■ Animal models show that infection and propagation can only
– but once the nucleus has formed, polymerization and fibril take place if the normal cellular protein is present, since
elongation proceed rapidly. prion-knockout mice are resistant.
■ The template-directed refolding and nucleation-polymerization
models are attempts to explain how abnormal prion protein
might recruit and convert native (cellular) prion protein.