The human neurodegenerative
diseases, including amyotrophic lateral sclerosis (ALS)
and Alzheimer’s disease (AD), are adult-onset, chronic,
progressive disorders whose clinical features reflect
the vulnerability of specific populations of neurons in
each disease. In ALS, weakness and atrophy reflect dysfunction/death
of motor neurons; in AD, memory loss and dementia are
the result of neurofibrillary tangles, Ab 42 amyloid
deposits, and death of neurons in basal forebrain, hippocampus,
and cortex. Subsets of cases of familial ALS (FALS) and
AD (FAD), often show dominant inheritance; some cases
of FALS are linked to mutations in the superoxide dismutase
1 (SOD1) gene; and some pedigrees with FAD exhibit mutations
in genes encoding either the amyloid precursor protein
(APP) or presenilins (PS1 and PS2). Much has been learned
about the biology of these mutant transgene products by
recent in vitro and in vivo studies. This
lecture describes some of this work, with particular emphasis
on exciting advances from studies of transgenic (Tg) mice
that show many features of these human disorders.
Amyotrophic
Lateral Sclerosis and Tg Models
ALS is characterized
by paralysis, muscular atrophy, spasticity, and a variety
of other motor signs; electrodiagnostic studies disclose
evidence of denervation of muscle. Weakness and atrophy
are related to abnormalities of large a-motor neurons
of the brainstem and spinal cord, and spasticity reflects
alterations in upper motor neurons. Lower motor neurons
show a variety of abnormalities including ubiquitin and
phosphorylated neurofilament immunoreactivities in cell
bodies and swollen axons (spheroids) with maloriented
arrays of neurofilament. In some cases of SOD1-linked
FALS, motor neurons may also contain SOD1-immunoreactive
intracytoplasmic inclusions. It has been estimated that
10% of adult-onset cases of ALS are familial with autosomal
dominant inheritance and age-dependent penetrance. Approximately
20% of cases of FALS are linked to mutations in SOD1,
a member of a family of metalloenzymes that acts as a
free radical scavengers.
Recently, several
groups of investigators produced Tg mice with FALS-linked
SOD1 mutations. Lines of Tg mice that express the G37R
HuSOD1 mutation at 3-12x levels of endogenous SOD1 in
the spinal cord invariably develop progressive motor neuron
disease. At four months of age, these Tg mice begin to
show reduced spontaneous movements, difficulty moving
their hindlimbs, and muscle wasting; eventually, forelimbs
become weak, and hindlimbs are completely paralyzed. Electromyographic
patterns and muscle biopsies show features identical to
those documented in patients with ALS. G37R HuSOD1 mice
have significantly elevated levels of SOD1, and activity
gels show increases in SOD1 activity, confirming that
the G37R mutation synthesized in vivo retains full
specific activity. In mice that express mutant SOD1, motor
axons accumulate SOD1 immuno-reactivity. In motor neurons,
SOD1 is transported anterograde in axons as part of the
slow component. Recent studies in Tg mice expressing mutant
SOD1 have shown the toxic protein is transported into
axons. In the early preclinical period, motor axons as
well as some dendrites exhibit very small vacuoles, usually
associated with enlarged degenerating mitochondria and
swollen endoplasmic reticulum. Subsequently, cell bodies
show abnormal patterns of ubiquitin and phosphorylated
neuro-filament immunoreactivities. Motor axons, some of
which develop abnormalities of the cyloskeleton, undergo
Wallerian degeneration; muscle fibers are denervated.
The observation
that Tg mice expressing mutant SOD1 show no loss of enzymatic
activity yet develop disease strongly suggests that, at
least for these mutations, SOD1-linked FALS is caused
by the gain of a toxic property by the mutant enzyme.
This concept is supported by several other lines of evidence:
some mutant SOD1 possess near-normal levels of enzyme
activity/stability in vitro and/or restore SOD1
null yeast to the wild-type phenotype but accelerate the
death of nerve cells in vitro; SOD1 mutations do
not have a dominant negative effect on wild-type SOD1;
and wild-type HuSOD1 Tg mice do not develop overt motor
neuron disease. SOD1 null mice develop normally and do
not develop a FALS-like disease. A major question in the
field is the nature of the toxic properties acquired by
mutant SOD1 and the mechanisms of cell injury: one hypothesis
is that a peroxidase activity by mutant SOD1 catalyzes
conversion to H2 O2 to OH, which is capable of
oxidizing a variety of targets; and a second hypothesis
is that mutant SOD1 has the enhanced ability to utilize
peroxynitrite to form nitronium ions that can nitrate
tyrosine residues. Both of these hypotheses are consistent
with the premise that FALS mutations alter protein structure
in such a way that mutant SOD1 has a toxic effect on substrates
critical for the survival of motor neurons. Because SOD1
is abundant in spinal motor neurons and transported anterograde
in axons, we have suggested that the toxic mutant protein
damages a variety of molecular targets with significant
consequences for motor neurons.
Alzheimer’s
Disease and Tg Models
AD, the most
common cause of senile dementia, is the result of selective
vulnerability of subsets of neurons and the presence of
neurofibrillary tangles, Ab amyloid deposits, and
death of nerve cells in the basal forebrain, hippocampus,
and cortex. Risk factors for AD incllude: age; mutations
in APP and PS1 and PS2 genes, which cause autosomal dominant
disease; and the presence of apolipoprotein E4 allele,
which is a susceptibility factor. Animal models are critical
for investigating a variety of processes that involve
normal brain function, for analyzing the mechanisms of
disease-related abnormalities in vivo, and for
testing novel therapies. The character, evolution, and
mechanisms of some of the cellular abnormalities in AD
have been clarified in studies of aged monkeys and Tg
mice that overexpress mutant transgenes. Both aged nonhuman
primates and APP and APP-PS1 mutant Tg mice show cellular
abnormalities, including neuritic plaques consisting of
dystrophic neurites and deposits of Ab 42 (a putative
toxic peptide) similar to those that occur in individuals
with AD. In vitro and in vivo studies have
shown that different APP mutations increase the amount,
length, and fibrillogenic properties of Ab . PS1 mutations
influence APP processing and increase levels of Ab
42 and accelerate amyloidogenesis in vivo. Tg mice
overexpressing mutant APP transgenes show increased ratios
of Ab 42:40 and develop Ab deposits in the cortex
and hippocampus. Recently, we have examined the extent
and frequency of Ab deposits in: 12-month-old Tg mice
that coexpress HuPSI-A246F and APPswe: mice that coexpress
wild-type HuPS1-and APPswe: mice that express APPswe alone;
and mice that express mutant PS1 alone. In double Tg mice,
coexpression of HuPS1-A246E with APPswe reduces the interval
of the formation of initial Ab deposits from 12 months
in mice expressing APPswe alone to <9 months in mice
expressing APPswe with HuPS1-A246E. These data provide
evidence to support the hypothesis that a principal pathway
by which mutations in PS1 predispose individuals to FAD
is to accelerate Ab depostion.
Conclusion
Enormous progress
has been made in understanding these neurodegenerative
diseases. In particular, Tg model systems provide extraordinary
opportunities to clarify some of the mechanisms leading
to cellular abnormalities that occur in ALS and AD and
to define some of the pathogenic pathways that represent
targets for therapy. Finally, these models are critical
for testing novel treatment strategies that, if effective
in animals, can be rapidly introduced into human clinical
trials.
