Alzheimer

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© 1997 Springer-Verlag Neurogenetics, Vol. 1, No. 1
73–80
ORIGINAL ARTICLE
Rediscovery of the case described by Alois Alzheimer
in 1911: historical, histological and molecular genetic
analysis
M. B. Graeber*, S. K ¨ sel
1
, R. Egensperger
2
, R. B. Banati
3
,U.M
¨
ller
4
, K. Bise
1
, P. Hoff
5
,
H. J. M ¨ ller
5
, K. Fujisawa
6
and P. Mehraein
1
Max Planck Institute of Psychiatry, Department of Neuromorphology, Am Klopferspitz 18a, 82152 Martinsried,
Germany,
1
Institute of Neuropathology, Molecular Neuropathology Laboratory, Ludwig Maximilians University,
Thalkirchner Str. 36, 80337 Munich, Germany,
2
Institute of Neuropathology, Molecular Neuropathology Laboratory,
Hannover Medical School, Carl-Neuberg-Str. 1, 30625 Hannover, Germany,
3
Hammersmith Hospital, Cyclotron Unit,
MRC Clinical Sciences Centre, Royal Postgraduate Medical School, London, W12 ONN, UK,
4
Institute of Human
Genetics, Justus Liebig University, Schlangenzahl 14, 35392 Giessen, Germany,
5
Psychiatric Clinic, Ludwig
Maximilians University, Nußbaumstr. 7, 80336 Munich, Germany and
6
Tokyo Metropolitan Institute for
Neuroscience, 2-6 Musashidai, Fuchu Tokyo, 183 Japan
Received January 16, 1997; Revised and Accepted February 13, 1997
ABSTRACT
INTRODUCTION
In 1911, Alois Alzheimer published a detailed report
In 1907, Alzheimer published his now famous report on a 51-
(Zbl. ges. Neurol. Psych. 4: 356–385) on a peculiar
year-old woman who had come under his care in 1901 while
he worked as an attending physician in the Frankfurt Asylum
case of the disease that had been named after him
by Emil Kraepelin in 1910. Alzheimer describes a 56-
(1). The original case file of this patient, Auguste D., was
year-old male patient (Johann F.) who suffered from
discovered recently, and it has been speculated that the patient’s
dementia was not caused by the typical neurodegeneration of
presenile dementia and who was hospitalized in Krae-
Alzheimer disease but by arteriosclerosis of the brain (2).
pelin’s clinic for more than 3 years. Post-mortem
Alzheimer’s report on Auguste D. is not a full-size research
examination of the patient’s brain revealed numerous
paper but an abstract summarizing the presentation he gave at
amyloid plaques but no neurofibrillary tangles in the
the
37. Versammlung s¨ dwestdeutscher Irren¨ rzte
(37
th
Meet-
cerebral cortex, corresponding to a less common
ing of the Southwest German Psychiatrists) in T¨bingen on
form of Alzheimer disease which may be referred to
November 3, 1906 (1). Therefore, the first report by Alzheimer
as ‘plaque only’. We have identified well-preserved
on the morphology of the disease that was named after him
histological sections of this case and performed
by Emil Kraepelin in 1910 (3) does not contain any illustrations.
mutational screening of exon 17 of the amyloid pre-
Yet numerous figures, mainly drawings, which include several
cursor protein
gene and genotyping
for apolipo-
examples of the histopathology of his first case were published
protein
E
alleles.
The
patient
was
shown
to
be
by Alzheimer in 1911 together with a second case report (4).
homozygous for apolipoprotein allele
e
3 and lacked
Both publications are available in English (5,6).
APP mutations at codons 692, 693, 713 and 717.
In the report on his second published case, Alzheimer gives
This case is of historical importance as it may have
a detailed description of the clinical history of a 56-year-old
convinced Kraepelin to name the disease after his
demented man (Johann F.). According to Alzheimer’s notes
co-worker, Alois Alzheimer.
(4), the patient was admitted to the psychiatric clinic on
November
12,
1907.
There
was
no
history
of
excessive
Keywords
: Alzheimer disease, amyloid plaques, amyloid
drinking. In the previous 6 months he had become forgetful,
precursor protein gene, apolipoprotein E gene,
could not find his way, could not perform simple tasks or
carried these out with difficulty. He stood around, did not
neurofibrillary tangles
*To whom correspondence should be addressed. Tel: 149 89 8578 3666; Fax: 149 89 8995 0077; Email: u792201@sunmail.lrz-muenchen.de
 74
Neurogenetics, 1997, Vol. 1, No. 1
bother about food, but ate greedily whatever was put in front
Primer sequences and PCR reaction conditions for amplification
of exon 17 of the APP gene and for genotyping of APOE
of him. He was not capable of buying anything for himself
and did not wash. He was admitted by the service for the poor
alleles were essentially as described (7,9).
Genomic amplification of exon 17 of the APP gene was
(4,6). After 3 years of hospitalization and repeated clinical
examinations, Johann F. died on October 3, 1910, in the Royal
performed with primers
Aint3
59-TAAGAAATGAAATTCTT-
CTAATTGC-3
Psychiatric Clinic in Munich showing features of pneumonia.
Neuropathological examination of his brain revealed the wide-
9
and
Aint4
5
9
-GCAGTCAAGTTTACCTACC-
TCCACC-3
after denaturation for 3 min at 94°C, and 32
cycles at 91°C for 30 s, 55°C for 30 s and 72°C for 1 min,
9
spread presence of amyloid plaques but not a single neuron
showing neurofibrillary change (4). Alzheimer provides ample
followed by a 10 min final extension step at 72°C on a TC
480 thermal cycler (Perkin-Elmer). GeneAmp nucleotides were
clinical, biographic and neuropathological data of this patient
which have allowed us to identify histological sections found
used in the buffer supplied with
Ta q
polymerase (Perkin-
Elmer) or a buffer containing 300 mM Tris–HCl, pH 9.0, 75
among archival material at the Institute of Neuropathology of
the University of Munich. Using recently established methods
mM (NH
4
)
2
SO
4
, and 10 mM MgCl
2
(Invitrogen).
For the determination of APOE alleles, a new polymerase
for the molecular genetic analysis of neuropathological tissue
(7–9), we have been able to perform mutational screening of
chain reaction assay was employed which allows genotyping
of archival neuropathological tissue (9). Primer pairs were
exon 17 of the amyloid precursor protein (APP) gene and
genotyping for apolipoprotein E (APOE) alleles. The history
Rup1 and Rup2 (5
9
-CTGGGCGCGGACATGGAG-3
9
,5
9
-GC-
of this discovery will be detailed elsewhere.
AGGTGGGAGGCGAGGC-3
9
)
resulting
in
a
115
bp
amplification product, and Rup3 and Rup4 (5
9
-GGCCAGAGC-
ACCGAGGAG-3
) amp-
lifying a 119 bp DNA fragment. Reaction conditions were
9
,5
9
-GCCCCGGCCTGGTACACT-3
9
MATERIALS AND METHODS
denaturation at 94°C for 5 min, followed by 43 cycles at 94°C
Selection of tissue sections
for 30 s, 68°C for 30 s and 72°C for 1 min, and final extension
at 72°C for 10 min. The reaction buffer contained 300 mM
Histological sections belonging to this case were identified as
described below. All tissue sections to be used for molecular
Tris-HCl, pH 9.5, 75 mM (NH
4
)
2
SO
4
, and 10 mM MgCl
2
genetic analysis were first studied under the light microscope.
(Invitrogen).
The following tissue sections were selected for DNA extraction:
Hha
I digestion of the Rup 1/2 and Rup 3/4 amplicons
no. 28 cerebral cortex (Mann’s stain?); no. 35 spinal cord
distinguishes between 3/4 and 2/3 alleles, respectively (9). For
control, the primers described by Wenham
et al.
(10) were
(Mann’s stain?); no. 45 cerebral cortex (Mann’s stain?); no.
used.
Following
digestion
with
Hha
I
(Boehringer),
DNA
67 cerebral cortex (Mann’s stain?); no. 43 cerebellum (Mann’s
fragments were separated on a 5% intermediate melting temper-
stain?); no. 8 cerebellum (Nissl’s stain); no. 18 cerebral cortex
ature agarose gel (Metaphor, Biozym) containing 0.15
m
g/ml
(Nissl’s stain).
ethidium bromide. Fragment sizes generated by
Hha
I digestion
are given in the legend to Figure 3b. Negative (‘no DNA’)
DNA extraction
PCR
samples
were
always
run
in
parallel
to
control
for
All sections were processed separately. Glass coverslips were
contamination.
removed by overnight incubation in 100% xylene at room
temperature. Tissue sections were then transferred to fresh
Non-radioactive direct sequencing of PCR products
Eppendorf tubes under sterile conditions and washed with 95
and 70% ethanol for 30 min each, followed by brief centrifuga-
The genomic sequence of exon 17 of the APP gene was
tion in an Eppendorf centrifuge. Pellets were dried at 68°C in
determined between nucleotides 2065 and 2211 (11), encom-
a TRIO-Thermoblock (Biometra) and resuspended in 1
3
TE.
passing codons 692, 693, 713, and 717, where point mutations
Digestions were performed in 200
m
l aliquots containing 0.2
have
been
described
in
some
individuals
(12–16).
Cycle
M Tris, pH 8.0, 10 mM EDTA, 10% SDS, and 4 mg/ml
sequencing of PCR products was performed using 5
9
-digoxig-
proteinase K (Cat. no. 161519, Boehringer Mannheim) at 50°C
enin end-labeled oligonucleotide primers 5
9
-TAAGAAATG-
for 16–24 h (8).
AAATTCTTCTAATTGC-3
9
and
5
9
-GCAGTCAAGTTTA-
For purification of DNA, digested tissue was extracted twice
CCTACCTCCACC-3
9
, followed by separation of the sequen-
with phenol-chloroform-isoamylalcohol (25:24:1) and once
cing reactions on an 8% denaturating polyacrylamide gel
with chloroform. Volumes obtained after extraction were con-
(7,17). PCR reaction conditions were 3 min at 94°C, followed
centrated using either ethanol precipitation at –20°C overnight
by 32 cycles at 91°C for 30 s, 55°C for 30 s and 72°C for 1
or using a Microcon-30 concentrator (Cat. no. 42409, Amicon)
min, with 10 min final extension at 72°C. Sequencing bands
as described (8). ‘Mock’ DNA extractions were included
were visualized using immunological detection with alkaline
for control.
phosphatase conjugated Fab fragments and nitroblue tetrazol-
ium/X-phospate as an enzymatic substrate (17).
Polymerase Chain Reaction (PCR)
PCR reactions were performed in 50
l volumes using thin-
walled reaction tubes and GeneAmp reagents (Perkin-Elmer)
m
RESULTS
following the manufacturer’s recommendation. A quantity of
Identification of the case
200
M of each deoxynucleotide, 200 ng of each primer, and
1.25 U of
Ta q
m
DNA polymerase were used in each PCR
Entry no. 784 in the autopsy book of Kraepelin’s clinic
identifies a male patient bearing the last name ‘Feigl’ who
reaction. Samples were covered with 20
m
l of mineral oil, and
1
m
l of the concentrated DNA solution was finally added.
died on October 3, 1910, and who came to autopsy from the
Neurogenetics, 1997, Vol. 1, No. 1
75
Figure 1.
(
a
) Autopsy book (‘Sektionsbuch’) of the psychiatric clinic in Munich. (
b
) Entry no. 784 lists a male individual (‘Mann’) bearing the name ‘Feigl’
who died on October 3, 1910, in the psychiatric clinic (‘Klinik M¨ nchen’). The diagnosis reads ‘Alzheimer’sche Krankheit’ (Alzheimer’s disease). (
c
)
Alzheimer’s signature (upper part of figure) taken from his curriculum vitae which was written after he had joined Kraepelin’s clinic (31). For comparison, the
diagnosis written in the autopsy book has been enlarged (lower part of figure).
Munich psychiatric hospital (Fig. 1a, b). Based on the case
but we have not yet completed our morphological analysis of
all tissue sections since some of the stains require verification.
number in the autopsy book and the family name of the patient,
we
were
able
to
identify
histological
sections,
found
on
Therefore, a detailed report on the extensive histology of this
case will be published elsewhere.
December 30, 1992, among archival material at the Institute
of Neuropathology of the University of Munich. Many of
these sections are labeled by both the case number and the
Molecular genetic analysis
last name of the patient (Fig. 2a). The admission report of
At the time when this case was rediscovered, the frequency of
Johann F. (Fig. 4) shows his first and his last name together,
APP mutations in Alzheimer disease was just being established.
several important dates as well as clinical information which
Using direct non-radioactive sequencing of polymerase chain
appear in Alzheimer’s paper (4). The autopsy book states
reaction (PCR) products (7,17), we did not detect mutations
‘Alzheimer’sche
Krankheit’
(Alzheimer
disease)
as
the
at codons 692, 693, 713, and 717 or at other nucleotides within
patient’s diagnosis with the handwriting closely resembling
exon 17 of the APP gene (Fig. 3a). Consequently, a pathogenic
that of Alois Alzheimer (Fig. 1c).
role of exon 17
APP
mutations in this case of severe amyloid
deposition can be excluded. The apolipoprotein E genotype of
Histological examination
Alzheimer’s patient was found to be
e
3/
e
3 (Fig. 3b). We have
Examination of the histological sections in the light microscope
also started screening for mutations in the presenilin genes.
yielded morphological results which are in complete agreement
Yet, given the limited amount of tissue available, this study was
with Alzheimer’s paper (4). Although many amyloid plaques
postponed in order to save material for additional histological
were present in the cerebral cortex of this patient (Fig. 2b–d),
analyses
and
until
our
knowledge
on
genetic
defects
in
we did not find neurofibrillary tangles. It should be noted that
Alzheimer disease is more complete.
sections of the hippocampus and the entorhinal region were
not available. Silver impregnations performed over 2 days in
Alzheimer’s laboratory using Bielschowsky’s method (Fig. 2a)
DISCUSSION
were found together with a number of Nissl stained specimens.
In addition, numerous sections apparently prepared according
Alzheimer gives a very detailed description of his patient,
Johann F. (4). Starting with this information, we were able to
to the methods of Mann, Herxheimer and Weigert were present
76
Neurogenetics, 1997, Vol. 1, No. 1
a
c
b
d
Neurogenetics, 1997, Vol. 1, No. 1
77
Figure 3. (a
) Genomic sequence of part of exon 17 of the APP gene. Known mutation sites, codon numbers and the corresponding peptide sequence are set in
bold type. No mutations were found. (
b
) APOE genotyping. Gel electrophoresis of PCR amplicons obtained with the primers Rup 1/2, Rup 3/4, and Wbp 1/2
before (lanes 2, 4, and 6) and after (lanes 3, 5, and 7)
Hha
I digestion. A 10 bp DNA ladder was loaded on lanes 1 and 8. The presence of an e4 allele would
create a 72 bp restriction fragment in lanes 3 and 7, whereas the presence of an e2 allele would result in a 72 bp and 81 bp restriction fragment in lanes 5 and
7, respectively. As can be seen in lanes 3, 5 and 7, the patient shows homozygosity for apolipoprotein allele
e3.
identify tissue sections which belong to this case based on the
difference in pronunciation. In addition, the clinical history
given in the admission sheet also appears in Alzheimer’s paper,
matching dates and on the first and the last name of the patient
in Alzheimer’s paper as well as in the autopsy book, on the
confirming the identity of the case. Finally, the ink used by
Alzheimer’s laboratory to label the tissue sections was com-
tissue sections and in the patient’s admission report. The latter
document (Fig. 4) contains additional interesting information.
pared to that of three other cases listed in the autopsy book,
and they were shown to be of identical origin (opinion from
Although most of its data match exactly, there are two
exceptions. The first difference concerns the date of the hospital
the Bavarian State Bureau of Criminal Investigation, data not
shown). Thus, the tissue sections were confirmed to be older
admission of the patient which was September 12, 1907 (Fig.
4), and not November 12, 1907, as reported by Alzheimer. On
than 80 years.
It is unknown whether Kraepelin ever saw Auguste D.,
reading the clinical history of the patient, this typographical
error in Alzheimer’s publication (4) is readily apparent. The
Alzheimer’s first patient, who died in the Frankfurt asylum
(1,2). However, Kraepelin was most likely familiar with Johann
second discrepancy concerns the misspelling of the name
‘Feigl’ in the admission report as there is an additional ‘e’
F. as Kraepelin and Alzheimer used to work together very
closely which is gratefully acknowledged by Kraepelin in
(Feigel). However, ‘Feigl’ is a common Bavarian name of
which several different spellings exist without an associated
the introduction to the second volume of his textbook (3).
Figure 2.
(
a
) Bielschowsky-stained tissue sections processed for two days (‘2 Tage Silber’). (
b–d
) Staining of classical amyloid plaques (with core) in the
cerebral cortex of Alzheimer’s patient. Pyramidal neurons and neurites within the plaque do not show neurofibrillary change. Magnification: 3700 (b), 31000
(c), 3500 (d).
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