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Regular Articles |
From the Departments of Pathology,*
Pharmacology,
Neurology,
Nuclear
Medicine,§
and Radiology,¶
State University of New York Health Science Center, Syracuse, New York;
the Departments of Neurology, Public Health, and Obstetrics and
Gynecology,||
Oregon Health Sciences University, Portland,
Oregon; the Department of Pathology,**
University
of Washington School of Medicine, Seattle, Washington; the National
Human Genome Research Institute,
National Institutes of Health, Bethesda, Maryland; and the American Red
Cross Holland Laboratories,
Rockville, Maryland
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Introduction |
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Materials and Methods |
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Entire brains of two affected individuals from the New York family
were available for study; they were fixed in 10% neutral buffered
formalin for 2 weeks before gross examination and sectioning. Blocks
were obtained from representative cortical and subcortical areas,
embedded in paraffin, and stained with hematoxylin and eosin (H&E),
according to routine histological procedures. Prepared glass slides
from another case, stained with H&E/Luxol fast blue, as well as
flash-frozen sections of one hemisphere were provided by the Harvard
Brain Tissue Resource Center (PHSMH31862). Portions of fixed brain
tissue from an additional case, as well as fixed tissue from other
organs from two cases, were obtained from regional tissue archives.
Microscopic glass slides of formalin-fixed and paraffin-embedded
cerebral biopsy tissue from the Oregon family were supplied by the
University of Washington School of Medicine (Seattle, WA) and were the
only samples available for histological study of this family. Enzyme
digestion studies were conducted with porcine pancreas 
-amylase, 10
mg/ml, pH 6.8, at 37°C; barley ß-amylase, 0.1 mg/ml, pH 4.8, at
37°C; and Aspergillus nigra amyloglucosidase, 0.1 mg/ml,
pH 4.8, at 37°C. Dimedone (5,5-dimethyl-1,3 cyclo-hexanedione) was
obtained from Sigma Chemical Co., St. Louis, MO, and was used as a 5%
ethanolic solution at 60°C by the procedure of Bulmer.10
All other histochemical procedures were performed by routine
histological laboratory protocols. All light microscopy was done on an
Olympus microscope.
Immunohistochemistry
Immunostaining was performed on tissue fixed in 10% buffered
formalin for at least 2 weeks and embedded in paraffin before staining.
Commercially available antibodies were obtained and used as follows,
for standard epitope retrieval techniques, strepavidin-biotin
methodology, and 3–3' diaminobenzidine as
chromogen; ubiquitin (polyclonal, 1:400; Vector-NovoCastra, Burlingame,
CA); B-crystallin (polyclonal, 1:80; Vector-NovoCastra);
amyloid-ß (Aß) (monoclonal, 1:80; Vector-NovoCastra); tau
(monoclonal, 1:250; Labvision, Fremont, CA); neurofilament protein
(monoclonal, 1:200; DAKO, Carpenteria, CA); phosphorylated
neurofilament protein (SM1–31) (monoclonal, 1:200; Sternberger
Monoclonals, Baltimore, MD); nonphosphorylated neurofilament protein
(SM1–32) (monoclonal, 1:2000; Sternberger Monoclonals); synaptophysin
(polyclonal, 1:500; DAKO); neuron-specific enolase (polyclonal, 1:40;
Biogenex, San Ramon, CA); lysozyme (polyconal, 1:1000; Biogenex);
antichymotrypsin (polyclonal, prediluted; Signet, Dedham, MA);
antitrypsin (polyclonal, prediluted; Signet); actin (monoclonal, 1:400;
DAKO); GFAP (polyclonal, 1:2000; DAKO); PGP9.5 (polyclonal, 1:3000;
Vector); HSP27 (monoclonal, 1:100; Biogenesis, Portsmoth, NH); HSP70
(monoclonal, 1:200; Stressgen, Victoria, BC, Canada); superoxide
dismutase (monoclonal, 1:8000; Boehringer Mannheim, Indianapolis, IN);
amyloid-ß precursor protein (APP) (monclonal, 1:100; Boehringer
Mannheim); and -synuclein (1:500; Chemicon, Temecula, CA). An
affinity-purified, polyclonal rabbit antibody to recombinant human
neuroserpin was produced11-12
and was used at a 2000-fold
dilution.
Lectin Histochemistry
Lectin histochemistry studies were performed on unfixed, frozen sections of cerebral cortex from a case and an age-matched control. A total of 21 fluorescein-labeled lectins available in three kits (Vector) were applied and viewed by fluorescence microscopy on a Leitz microscope.
Electron Microscopy
Tissue from the brain of one affected individual, including cerebral cortex, cingulate gyrus, substantia nigra, and subcortical white matter, was processed for electron microscopy (EM). Tissue was fixed initially for at least 2 weeks in 10% buffered formalin and was subsequently fixed overnight in 2.5% glutaraldehyde, postfixed for 1 hour in 1% osmium tetroxide, and embedded in Araldite 502 from which 1-mm-thick sections were cut and stained with toluidine blue. Ultra-thin sections of selected areas were stained with lead citrate and uranyl acetate and examined with a Jeol 100 SX electron microscope.
Biochemical Methods
Homogenization and Isolation of Inclusions
Cortical tissue (2 g) was homogenized in 10 ml of 250 mmol/L sucrose and 10 mmol/L ethylenediaminetetraacetic acid, buffered to pH 7.4 with 10 mmol/L 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid containing 10 µl of Sigma protease inhibitor cocktail. The homogenate was filtered through gauze, and the retentate was washed with an additional 5 ml of homogenization medium. Verification that the inclusion bodies withstood the homogenization was accomplished by visualization using light microscopy after H&E staining by air drying a sample on a glass slide followed by fixing for 1 minute in absolute ethanol before H&E. Similarly, the assay used to monitor the fractionation of the inclusions throughout the isolation procedure was light-microscopic visualization after H&E staining. The inclusion bodies were isolated from the homogenate as follows: the homogenate was centrifuged at 1000 x g for 10 minutes, the supernatant decanted, and the soft pellet resuspended in 5 ml of homogenization medium; 5 ml of 1% sarcosyl in homogenization medium was added, the suspension was incubated at 37°C for 30 minutes with frequent mixing and centrifuged at 45,000 x g for 20 minutes; the pellet was resuspended in 10 ml of the 1% sarcosyl solution and incubated at 37°C for 30 minutes with frequent mixing and again subjected to centrifugation at 45,000 x g for 20 minutes; the pellet was resuspended in 5 ml of homogenization medium, 5 ml of collagenase (2 mg/ml) was added, and the suspension was incubated at 37°C for 60 minutes followed by centrifugation at 45,000 x g for 20 minutes; and the resultant pellet was resuspended in 0.5 ml (1/20 original volume) of 4% sodium dodecyl sulfate (SDS), 125 mM Tris-HCl (pH 6.8), 20% glycerol, 10% mercaptoethanol and heated at 75°C for various periods of time. After heating, the samples were subjected to centrifugation at 14,000 x g for 10 minutes, and the supernatant was removed and analyzed by SDS-polyacrylamide gel electrophoresis (PAGE).
SDS-PAGE
The samples were diluted 2:1 in 4% SDS, 20% glycerol, 10% mercaptoethanol, 50 mmol/L Tris-HCl (pH 6.8) before being subjected to SDS-PAGE in a Bio-Rad 7.5% Redi-gel in Tris/glycine/SDS buffer at 150 V for 1.25 hours. Fifty microliters of the various samples were analyzed. After electrophoresis the gels were stained with Coomassie blue and destained by standard procedures.
Amino Acid Sequence
After SDS-PAGE the samples were electrophoretically transferred to a polyvinylidene difluoride membrane at 250 mA for 1.5 hours and stained with Coomassie blue. The membrane was sent to the Biotech Center at Cornell University, Ithaca, NY, where the 57-kd band was excised and either sequenced directly for N-terminal analysis or first subjected to endoproteinase C digestion (EC 3.4.99.30). The resulting peptides were separated by reverse-phase chromatography, and selected peaks were then sequenced.
Chemicals
All of the routine chemicals (Tris, SDS, glycine, N-lauroylsarcosine), as well as the enzymes, were obtained from Sigma.
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Neuropathological
Gross Examination
Two cases from the New York family were available for a complete neuropathological investigation of the brain. Both individuals, siblings aged 64 and 57 with clinical dementias of approximately 10 years’ duration, showed normal appearing sulci and gyri without atrophy and with transparent leptomeninges. Brain weights were 1530 and 1400 g, respectively. Examination of the bases of the brains disclosed no vascular malformations or significant atherosclerotic changes. Multiple coronal sections demonstrated slight ventricular dilatation together with normal appearing cortical gray, subcortical white, and deep gray matter structures free of encephalomalacia, hemorrhage, or discoloration. In both cases the substantia nigra, brainstem, and cerebellum were grossly unremarkable (data not shown).
Light Microscopy
Sections of brain obtained from these two individuals and from a
limited amount of autopsy material obtained from two additional family
members were studied using standard histopathological methods. The
neuropathological findings are presented in Figures 1 and 2
and
in Tables 1–3
. Sections of brain stained
with H&E were examined by light microscopy. At scanning magnification
the neocortex shows mild microvacuolar change and gliosis affecting
cortical layer II, principally in sections of the frontal, temporal,
and parietal cortices (Figure 1A)
. Numerous, eosinophilic, sharply
defined, round-to-oval inclusion bodies of variable size (approximately
5–50 µm) are scattered throughout the deeper layers of the cerebral
cortex primarily in layers III to V (Figure 1A)
. They occur both singly
and in clusters of three or more (Figure 1, B and C)
. Many of these
inclusions are homogeneous in appearance, although others have either a
dark core with a lighter halo or a variegated speckled appearance; a
thin rim of darker material surrounds a few of them (Figure 1, B and C)
. Some of the inclusions are found within neuronal cell bodies, but
many appear to lie free in the neuropil within vacuoles. The majority
of cortical neurons are free of this involvement. Inclusions are very
infrequently noted in the cerebral white matter. Deep gray matter is
variably affected with marked involvement of the substantia nigra where
many of the pigmented neurons show one or more inclusion bodies within
their perikarya. However, neither neuronal loss nor gliosis are
apparent. Many of the neurons affected with these bodies show no other
abnormalities; however, others are grossly distorted, having no
apparent cytoplasm, a displaced and compressed nucleus, and possibly an
overall reduction in cell size. Neuronal loss is evident only in
cortical layer II, and glial involvement is not seen except for the
reactive change in layer II, as well as a low-grade subcortical
gliosis. Histological examination of other organs disclosed only
extensive myocardial fibrosis, severe coronary atherosclerosis, and
pulmonary edema in one case, and hepatic steatosis in another case
(data not shown).
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and in Table 2
. The
inclusions are eosinophilic (Figure 1, A-D
; Table 2
) and are strongly
and uniformly stained by the PAS method (Figure 1E)
. Furthermore, they
retain their PAS reactivity after digestion by -amylase or other
glycosidases for up to 48 hours at 37°C (Table 2)
. Conversely,
preincubation in 5% ethanolic dimedone, an aldehyde blocker, inhibits
the PAS reaction (Table 2)
. The inclusions fail to stain with Best’s
carmine (Table 2)
. These findings suggest that the inclusion bodies
contain a carbohydrate component other than glycogen. Lack of staining
with either the Alcian blue or the high-iron diamine method shows that
the carbohydrate is not an acidic or a sulfated mucopolysaccharide
(Table 2)
. Stains for neutral lipid, including Sudan black B and oil
red O, are negative on unfixed frozen tissue (Table 2)
. Additional
histochemical studies show that the inclusions are negative for iron
and for calcium (Table 2)
. The inclusions were further analyzed using a
screening panel of fluorescein isothiocyanate-conjugated lectins
(Figure 1, G and H)
. By fluorescence microscopy, the presence of
neuronal lipofuscin was apparent by its bright autofluorescence.
Lectins showing moderate to strong affinity for the bodies included
those with relative specificity for mannose (Figure 1G)
and for
N-acetylglucosamine (Figure 1H)
. In summary, these findings
suggest that the inclusions are composed of a glycoprotein or
glycoproteins containing N-linked oligosaccharides.
Immunohistochemical methods were used to probe for known components of
other well-characterized neuronal inclusions. The results are presented
in Figure 1
and in Table 3
. As shown
(Table 3
; Figure 1F
) the inclusions are negative for both ubiquitin and
-synuclein, and thus they are distinguishable from Lewy
bodies13-15
; they contain neither Aß nor other
abnormal proteins or peptides characteristic of Alzheimer’s disease.
Furthermore, none of the following proteins are detected: intermediate
filaments or other cytoskeletal proteins, PGP9.5, B-crystallin,
heat shock proteins 27 or 70, superoxide dismutase, or tau (Table 3)
.
Neither Pick bodies nor ballooned neurons are seen. Tau-positive glial
inclusions are not identified. Rare tau-positive
neurofibrillary tangles are observed in the hippocampal formation. An
antibody to synaptophysin strongly labels the cortical neuropil but not
the inclusions. Thus, the inclusion bodies described here appear to be
distinctly different from any described previously; therefore we
propose the name Collins bodies to identify these unique inclusions.
Their neuroanatomical mapping is shown in Figure 2
.
Electron Microscopy
By transmission EM, Collins bodies appear as moderately
osmiophilic round globules with an amorphous to finely granular texture
(Figure 3)
. They have little internal
structure, aside from some speckling with darker material, and a few
demonstrate a darker inner core contrasting with a lighter outer shell.
Most are well delimited at their periphery, sometimes crowding aside
other cytoplasmic organelles; others appear to be enclosed within a
limiting membrane of rough endoplasmic reticulum (Figure 3)
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Isolation of Collins Bodies
The Collins bodies were isolated from unfixed, frozen cerebral
cortex tissue by homogenization in a buffered isotonic sucrose
solution, washed with detergent (N-lauroylsarcosine),
digested with collagenase, washed, and collected by centrifugation
(Figure 4
; see Materials and Methods for
complete details). The presence of Collins bodies was monitored
throughout the isolation procedure by light microscopy after H&E
staining; as noted, they are eosinophilic (Figure 5, A-C
). Clearly, a highly enriched
preparation is obtained (the H&E stain of the enriched fraction that is
labeled fraction P5 in Figure 4
is presented in
Figure 5 C
), although the bodies appear to be damaged somewhat (compare
Figures 5A and 5C
).
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In preliminary experiments the fraction enriched with Collins
bodies (fraction P5) was subjected to routine
SDS-PAGE analysis, ie, heating for 5 minutes in boiling water in a 2%
SDS-reducing buffer. However, this treatment released very little
protein detectable by Coomassie blue staining (Figure 6
, lane 2 represents SDS-PAGE of fraction
S6 obtained by heating for 5 minutes). Moreover,
microscopic examination revealed that Collins bodies remained intact
and could be collected by centrifugation (fraction
P6). Therefore, the Collins body-enriched
fraction (P5) was treated a second time and
subjected to harsher conditions, heating for 2 hours at 75°C in 4%
SDS. By this treatment the Collins bodies were disrupted, and SDS-PAGE
analysis showed one prominent protein band at 57 kd (Figure 6
, lane 3,
is fraction S6 obtained after prolonged heating).
A 57-kd protein was not found in an identically treated specimen
obtained from an age- and gender-matched control (Figure 6
, lane 4).
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The results show that Collins bodies are composed primarily of a
single protein. The identity of the 57-kd protein was determined by
transferring it to a polyvinylidene difluoride membrane and subjecting
it to amino acid sequence analysis before and after proteolytic
digestion (see Materials and Methods). The sequence information was
analyzed using the BLAST program and gave convincing evidence that the
57-kd protein is neuroserpin (Figure 7)
,
a serine protease inhibitor synthesized primarily in the CNS.
Differences between the amino acid composition of the isolate and the
wild-type neuroserpin were found: 1) asparagine 100 in the wild type is
identified as aspartic acid in the isolate (most likely, this
difference is an artifact because, when an asparagine residue is
followed by a glycine in the primary structure, the asparagine
frequently becomes deamidated during the sequencing procedure), and 2)
the N-terminal amino acid of the isolate corresponds to residue number
20, not number 17 as expected. (Reportedly the first 16 residues serve
as the single peptide.11,16
) Nonetheless there is
sufficient identity between the isolate and the amino acid sequence of
neuroserpin to confidently state that Collins bodies are composed of
neuroserpin.
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New York Family
The cellular distribution of neuroserpin is very different between
an age- and gender-matched control (Figure 5D)
and an affected subject
(Figure 5E)
. In the control, the neuroserpin-staining pattern is
generally diffuse, although small punctate foci of more intense
staining are occasionally seen. Conversely, in the affected individual
(Figure 5E)
neuroserpin is aggregated into inclusions that correspond
by size and distribution to Collins bodies. Furthermore, the inclusion
body-enriched fraction stains intensely with a neuroserpin antibody
(Figure 5F)
. These immunohistochemical findings support the conclusion
that Collins bodies contain neuroserpin.
Oregon Family
Pathological and immunohistochemical analyses of brain tissue
obtained by biopsy from one member of the Oregon family, who was
originally classified as suffering from an unclassified inclusion body
dementia,17
are shown (Figure 5, G and H)
. The inclusion
bodies are similar in size, H&E staining (Figure 5G)
, and distribution,
to those seen in the New York family. Furthermore, they stain
positively for neuroserpin (Figure 5H)
.
Pedigree
The familial occurrence of FENIB in the New York kindred is
apparent by examination of the pedigree (Figure 8)
. The disease appears in each of four
generations and in both genders, a pattern of inheritance consistent
with an autosomal dominant mode of transmission. Some affected
individuals were identified by either family history or clinical
evaluation. (The clinical evaluation of the family is continuing.)
Affected subjects present with deficits in attention and concentration,
perseveration, and restricted oral fluency. More severely affected
subjects demonstrate additional difficulties including restricted
visuospatial organization and visuoconstructional problem solving. They
experience progressive difficulties organizing their activities of
daily living. Although they demonstrate impaired learning and memory in
the earlier stages, it is to a lesser extent than typically seen in
early Alzheimer’s disease and is characterized by restricted learning
with more spared recall of information that has been learned. Single
photon emission computed tomography cerebral perfusion studies have
shown areas of cortical hypoperfusion, involving especially the
interior frontal and adjacent temporal and parietal cortices in
affected subjects. Magnetic resonance imaging studies are essentially
normal in younger, mildly affected subjects, but demonstrate diffuse
cortical atrophy in moderately to severely demented older individuals.
Electroencephalogram patterns show primarily generalized slowing,
consistent with diffuse cortical dysfunction, although one subject also
showed intermittent triphasic wave activity (data not shown). The
course of the disease can be quite long, exceeding 10 years in some
cases. Most patients will eventually require institutional care.
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; Figures 1 and 2
). We refer to these unique inclusions
as Collins bodies, in recognition of Dr. George H. Collins, who
initiated this study. We find that the Collins bodies are composed
primarily of neuroserpin, and consequently we name this disease
familial encephalopathy with neuroserpin inclusion bodies (FENIB).
The Collins bodies are distinguishable from other well-characterized
inclusions. Their PAS positivity, but negativity for ubiquitin and
-synuclein (as shown in Table 3
and Figure 1
) clearly differentiates
them from Lewy bodies.14,15,19
Collins bodies are also
easily distinguished from the more basophilic, argyrophilic, and
tau-positive bodies of classic Pick’s disease6,20
(Table 3)
. They do not appear to be related to protein dense microspheres or
spherons, which have recently been shown to contain the Aß precursor
protein and the Aß 1–40 peptide.21,22
Collins bodies
are nonreactive with a large panel of neuroimmunohistochemical reagents
(Table 3)
, and they defy precise classification by routine methods.
They are also different from other types of PAS-positive inclusions,
including corpora amylacea, Lafora bodies, Bielschowsky bodies, the
inclusions of type IV glycogenosis, and the polyglucosan bodies of
adult polyglucosan body disease.23-29
First, Collins
bodies retain PAS positivity even after prolonged incubation with
glycosidases in contrast to the aforementioned inclusions, which are
degraded enzymatically.24
Second, corpora amylacea, Lafora
bodies, and polyglucosan bodies are resistant to the aldehyde-blocking
reagent dimedone,28
in contrast to the Collins bodies,
which lose PAS staining after a brief exposure to this reagent (Table 2)
. Third, the lectin-binding specificities of the Collins bodies are
consistent with the conclusion that they contain significant amounts of
mannose and N-acetylglucosamine (Figure 1, G and H)
, as
might be found in an N-linked glycoprotein, in contrast to
other PAS-positive inclusions that contain a high proportion of a
glycogenlike substance and only a small percentage of
protein.28,29
Finally, Collins bodies are distinguished
ultrastructurally by a rather homogeneous, finely granular appearance,
sometimes bounded by a membrane, whereas polyglucosan bodies, such as
those of Lafora, show primarily fibrils and filaments, which may be
branched, with a variable granular component, and they never have a
limiting membrane.26,29
In addition to these classically described types of PAS-positive
inclusions, atypical myoclonus bodies (myoclonus body type II) have
been reported in a disorder that Berkovic calls atypical inclusion
body-progressive myoclonus epilepsy.30,31
Only a few case
reports have appeared describing this condition, which manifests with
myoclonus, seizures, and progressive neurological dysfunction,
including dementia, beginning from childhood to young
adulthood.32-35
The inclusions differ from Lafora bodies
and resemble Collins bodies in their staining properties (Alcian blue
negative), ultrastructure (nonfibrillar), and distribution (confined to
brain). Two of the present authors have previously described a case of
seizures and progressive neurological dysfunction in a young woman with
atypical neuronal inclusions.17
Since the initial case
report, one of the subject’s daughters has developed a seizure
disorder and progressive cognitive loss starting at age 18, suggesting
that this is a familial disease (Oregon family). Pathological and
immunohistochemical analyses of brain tissue obtained by biopsy from
one member of this family are shown (see Figure 5, G and H
). The
inclusion bodies are similar in size, histochemistry, and distribution
to Collins bodies (see Figure 5G
), and they stain positively with a
neuroserpin-specific antibody (see Figure 5H
). We therefore conclude
that the Oregon kindred is also affected by FENIB. The relationship of
FENIB to other cases demonstrating atypical inclusion bodies is
presently unknown.
The chemical composition of Collins bodies appears to be surprisingly
simple; they are composed primarily of neuroserpin (Figures 4–6)
.
Neuroserpin is a member of a large family of structurally related
proteins called serpins (serine protease inhibitors). The family is
divided into two subclasses: those with and those without
antiproteolytic activity.36,37
Those members that are
protease inhibitors share the property of unusual conformational
mobility—the capacity to undergo dramatic changes in
shape.38,39
This molecular mobility provides the serpins
with an evolutionary advantage of not only being capable of binding
their target protease, but also of entrapping it in a highly stable
complex that persists for the minutes or even hours needed to clear the
complex from the extracellular fluid compartment. The trapping
mechanism has been compared with that of a mousetrap—the serpin is
synthesized in a metastable state (ie, the trap is set during
biosynthesis) with an extended reactive site loop (ie, the cheese) that
acts as the substrate (ie, the bait) for the target protease (ie, the
prey). When the prey takes the bait, the serpin snaps closed (in
actuality, relaxes to a more stable conformation), pulling the target
protease into a stable complex and thus stopping the extracellular
proteolytic cascade. Conversely, an evolutionary disadvantage of the
serpin’s requirement of high molecular mobility is the propensity to
form dysfunctional molecules; a single amino acid change in certain key
domains of the molecule results in the loss of the crucial folding
information needed to form the metastable intermediate(s); the trap
closes prematurely. The end result is the polymerization of mutant
serpin molecules into intracellular aggregates.39,40
A
well-characterized example of a mutant serpin that results in a disease
is the 1-antitrypsin ( AT) deficiency syndrome commonly found in
Northern Europeans.40-42
Most individuals have the normal
M form of AT, but 4% are heterozygous for the Z variant. (The name
antitrypsin is a misnomer because the actual target protease is not
trypsin but elastase, an enzyme secreted by neutrophils in response to
inflammation. It is therefore now called 1-proteinase inhibitor.)
The Z mutation involves a single amino acid change at the base of the
extended reactive center such that the reactive loop of one molecule
binds or folds with adjacent molecules to form polymers. This
self-association prevents normal AT processing by the hepatocytes
and results in a loss of AT secretion. Although clinical disease is
manifested generally only in homozygous individuals, MZ heterozygotes
do have the hepatic inclusion bodies.43
It is more
interesting and most important that the histochemistry of the AT
inclusions produced in the liver of those individuals expressing the Z
variant is very similar to the neuroserpin aggregates or Collins bodies
observed in the brain in the neurodegenerative disease under study.
The findings presented are consistent with the hypothesis that FENIB is
caused by a mutation in the neuroserpin gene that results in the
polymerization and aggregation of the neuroserpin protein. The findings
supporting this conclusion are the following: first, FENIB is a
familial disease with apparent autosomal dominant transmission,
indicating that the expression of the mutant allele is sufficient to
produce the disease; second, there is a striking similarity in
appearance between Collins bodies and the AT inclusions found in
the liver of individuals affected with the Z variant; and third, the
chemical composition of Collins bodies is unique compared with other
neuronal inclusions, consisting almost entirely of neuroserpin. These
points are consistent with the interpretation that a single molecular
defect triggers the pathogenic cascade. The hypothesis was tested by
subjecting the two families experiencing FENIB to genetic analysis. We
have found that all of the clearly affected individuals in the New York
family express a missense mutation in the neuroserpin gene, a
serine-to-proline transition at amino acid position 49.44
Affected individuals in the Oregon family show a serine-to-arginine
change at position 52.44
Thus we conclude that FENIB is a
genetic disease caused by mutations in critical regions of the
neuroserpin protein resulting in polymerization and tissue
deposition—the identical sequence of events generated by mutations in
the gene encoding AT.
The causal relationship between neuroserpin deposition and the onset of the clinical syndrome is not clear. Neuroserpin is believed to play a vital role in controlling extracellular proteolysis in the nervous system, especially as an inhibitor of tissue-type plasminogen activator.45 Proteolytic cascades appear to be involved in the processes of synaptic remodeling, repair, and regeneration in the CNS.45-48 Consequently, a diminution in neuroserpin secretion could result in uncontrolled proteolysis with a subsequent decrease in synaptic contacts and the loss of neuronal function. It is equally conceivable that the inclusions themselves are neurotoxic. Future studies will have to determine how common neuroserpin mutations are, and why they cause neurological disease. We think that these are particularly pertinent questions because there is an emerging concept that many, if not all, neurodegenerative diseases are the result of intra- or extracellular deposits of misfolded or mutant proteins.49,50 FENIB may present a simple model to test this hypothesis.51 Ultimately it may be possible to propose common mechanisms of pathogenesis and to promote common therapies for this heterogeneous group of CNS disorders.49-51
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Acknowledgements |
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Footnotes |
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Supported by CAP Foundation Award (to R. L. D.), SUNY HSC Hendricks Fund #130283 (to A. E. S.), and SUNY-HSC Hendricks Fund #13230 and NIHAG16954 (to P. D. H.).
Accepted for publication August 24, 1999.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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B-crystallin immunohistochemistry define the principal causes of degenerative fronto temporal dementia. Arch Neurol 1995, 52:1011-1015-synucleinopathies. Phil Trans R Soc London Ser B 1999, 354:1101-1118This article has been cited by other articles:
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  M. J. Davies, E. Miranda, B. D. Roussel, R. J. Kaufman, S. J. Marciniak, and D. A. Lomas Neuroserpin Polymers Activate NF-{kappa}B by a Calcium Signaling Pathway That Is Independent of the Unfolded Protein Response J. Biol. Chem., July 3, 2009; 284(27): 18202 - 18209. [Abstract] [Full Text] [PDF]
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A. Takasawa, I. Kato, K. Takasawa, Y. Ishii, T. Yoshida, M. H. Shehata, H. Kawaguchi, O. M. M. Mohafez, M. Sasahara, and K. Hiraga Mutation-, Aging-, and Gene Dosage-dependent Accumulation of Neuroserpin (G392E) in Endoplasmic Reticula and Lysosomes of Neurons in Transgenic Mice J. Biol. Chem., December 19, 2008; 283(51): 35606 - 35613. [Abstract] [Full Text] [PDF]
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 E. Miranda, I. MacLeod, M. J. Davies, J. Perez, K. Romisch, D. C. Crowther, and D. A. Lomas The intracellular accumulation of polymeric neuroserpin explains the severity of the dementia FENIB Hum. Mol. Genet., June 1, 2008; 17(11): 1527 - 1539. [Abstract] [Full Text] [PDF]
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 H. M. Nielsen, L. Minthon, E. Londos, K. Blennow, E. Miranda, J. Perez, D. C. Crowther, D. A. Lomas, and S. M. Janciauskiene Plasma and CSF serpins in Alzheimer disease and dementia with Lewy bodies Neurology, October 16, 2007; 69(16): 1569 - 1579. [Abstract] [Full Text] [PDF]
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E. E. Benarroch Tissue plasminogen activator: Beyond thrombolysis Neurology, August 21, 2007; 69(8): 799 - 802. [Full Text] [PDF]
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I. Gourfinkel-An, C. Duyckaerts, A. Camuzat, C. Meyrignac, P. Sonderegger, M. Baulac, and A. Brice Clinical and neuropathologic study of a French family with a mutation in the neuroserpin gene Neurology, July 3, 2007; 69(1): 79 - 83. [Abstract] [Full Text] [PDF]
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 G. Galliciotti, M. Glatzel, J. Kinter, S. V. Kozlov, P. Cinelli, T. Rulicke, and P. Sonderegger Accumulation of Mutant Neuroserpin Precedes Development of Clinical Symptoms in Familial Encephalopathy with Neuroserpin Inclusion Bodies Am. J. Pathol., April 1, 2007; 170(4): 1305 - 1313. [Abstract] [Full Text] [PDF]
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 M. Yepes and D. A. Lawrence New Functions for an Old Enzyme: Nonhemostatic Roles for Tissue-Type Plasminogen Activator in the Central Nervous System Experimental Biology and Medicine, December 1, 2004; 229(11): 1097 - 1104. [Abstract] [Full Text] [PDF]
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E. Miranda, K. Romisch, and D. A. Lomas Mutants of Neuroserpin That Cause Dementia Accumulate as Polymers within the Endoplasmic Reticulum J. Biol. Chem., July 2, 2004; 279(27): 28283 - 28291. [Abstract] [Full Text] [PDF]
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A. Makarova, I. Mikhailenko, T. H. Bugge, K. List, D. A. Lawrence, and D. K. Strickland The Low Density Lipoprotein Receptor-related Protein Modulates Protease Activity in the Brain by Mediating the Cellular Internalization of Both Neuroserpin and Neuroserpin-Tissue-type Plasminogen Activator Complexes J. Biol. Chem., December 12, 2003; 278(50): 50250 - 50258. [Abstract] [Full Text] [PDF]
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 A. Williams Defining neurodegenerative diseases BMJ, June 22, 2002; 324(7352): 1465 - 1466. [Full Text] [PDF]
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D. Belorgey, D. C. Crowther, R. Mahadeva, and D. A. Lomas Mutant Neuroserpin (S49P) That Causes Familial Encephalopathy with Neuroserpin Inclusion Bodies Is a Poor Proteinase Inhibitor and Readily Forms Polymers in Vitro J. Biol. Chem., May 3, 2002; 277(19): 17367 - 17373. [Abstract] [Full Text] [PDF]
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 R. W. Carrell and D. A. Lomas Alpha1-Antitrypsin Deficiency -- A Model for Conformational Diseases N. Engl. J. Med., January 3, 2002; 346(1): 45 - 53. [Full Text] [PDF]
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 C. B. Bradshaw, R. L. Davis, A. E. Shrimpton, P. D. Holohan, C. B. Rea, D. Fieglin, P. Kent, and G. H. Collins Cognitive Deficits Associated With a Recently Reported Familial Neurodegenerative Disease: Familial Encephalopathy With Neuroserpin Inclusion Bodies Arch Neurol, September 1, 2001; 58(9): 1429 - 1434. [Abstract] [Full Text] [PDF]
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M. Yazaki, J. J. Liepnieks, J. R. Murrell, M. Takao, B. Guenther, P. Piccardo, M. R. Farlow, B. Ghetti, and M. D. Benson Biochemical Characterization of a Neuroserpin Variant Associated with Hereditary Dementia Am. J. Pathol., January 1, 2001; 158(1): 227 - 233. [Abstract] [Full Text] [PDF]
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);
(
);
or neuropathologically
(
).
Unaffected individuals are indicated by open symbols
(
).
The pedigree has been modified to protect the family’s privacy.
Deceased individuals are not indicated. Roman numerals refer to the
generation, Arabic numbers to individuals within that
generation.