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The Journal of Neuroscience, May 1, 2001, 21(9):3017-3023
Mitochondrial Abnormalities in Alzheimer's Disease
Keisuke
Hirai1, 4,
Gjumrakch
Aliev2,
Akihiko
Nunomura1, 5,
Hisashi
Fujioka1,
Robert L.
Russell1,
Craig S.
Atwood1,
Anne B.
Johnson6,
Yvonne
Kress6,
Harry V.
Vinters7,
Massimo
Tabaton8,
Shun
Shimohama9,
Adam D.
Cash1,
Sandra L.
Siedlak1,
Peggy L. R.
Harris1,
Paul K.
Jones3,
Robert B.
Petersen1,
George
Perry1, and
Mark A.
Smith1
1 Institute of Pathology, 2 Department of
Neurology, and 3 Department of Epidemiology and
Biostatistics, Case Western Reserve University, Cleveland, Ohio 44106, 4 Pharmaceutical Research Laboratories I, Pharmaceutical
Research Division, Takeda Chemical Industries Ltd., Osaka, 532-8686 Japan, 5 Department of Psychiatry and Neurology, Asahikawa
Medical College, Asahikawa, 078-8510 Japan, 6 Department of
Pathology, Albert Einstein College of Medicine, Bronx, New York 10461, 7 Department of Pathology and Laboratory Medicine,
University of California, Los Angeles, California 90024, 8 Department of Neuroscience, University of Genova, 16132 Genova, Italy, and 9 Department of Neurology, Kyoto
University, Kyoto, 606-8507 Japan
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ABSTRACT |
The finding that oxidative damage, including that to nucleic acids,
in Alzheimer's disease is primarily limited to the cytoplasm of
susceptible neuronal populations suggests that mitochondrial abnormalities might be part of the spectrum of chronic oxidative stress
of Alzheimer's disease. In this study, we used in situ hybridization to mitochondrial DNA (mtDNA), immunocytochemistry of cytochrome oxidase, and morphometry of electron micrographs of
biopsy specimens to determine whether there are mitochondrial abnormalities in Alzheimer's disease and their relationship to oxidative damage marked by 8-hydroxyguanosine and nitrotyrosine. We
found that the same neurons showing increased oxidative damage in
Alzheimer's disease have a striking and significant increase in mtDNA
and cytochrome oxidase. Surprisingly, much of the mtDNA and cytochrome
oxidase is found in the neuronal cytoplasm and in the case of mtDNA,
the vacuoles associated with lipofuscin. Morphometric analysis showed
that mitochondria are significantly reduced in Alzheimer's disease.
The relationship shown here between the site and extent of
mitochondrial abnormalities and oxidative damage suggests an intimate
and early association between these features in Alzheimer's disease.
Key words:
Alzheimer's disease; free radicals; metabolism; mitochondria; neurodegeneration; oxidative stress
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INTRODUCTION |
The pathological presentation of
Alzheimer's disease (AD) involves selective pyramidal neuronal death
and an accumulation of intraneuronal and extracellular fibrils,
neurofibrillary tangles (NFT), and senile plaques, respectively
(Katzman, 1986 ; Smith, 1998). In a series of studies, we and others
have shown that oxidative stress is involved not only in damage to the
proteins of NFT and senile plaques but also involves extensive damage
to the cytoplasm of neuronal populations vulnerable to death during AD
(Montine et al., 1996; Smith et al., 1996, 1997; Sayre et al., 1997).
What is particularly striking regarding neuronal damage is that those neurons displaying oxidative damage show no overt signs of visible degeneration (Sayre et al., 1997; Smith et al., 1997), leading us to
consider whether more subtle cytological abnormalities might be
associated with oxidative damage.
We undertook this study to determine whether mitochondria could be
involved in this process because they can be both targets of oxidative
damage and sources of reactive oxygen. Dysfunction of mitochondrial
electron transport proteins has been associated with the
pathophysiology of AD (Blass and Gibson, 1991), as well as in
Parkinson's disease (Parker et al., 1989). Those studies that analyzed
mitochondrial function at the cellular level, through cytochrome
oxidase activity measurements, have consistently shown activity
deficits consistent with mitochondrial compromise (Wong-Riley et al.,
1997). Furthermore, cytoplasmic hybrid cells in which mitochondria from
sporadic cases of AD were fused with other cells also indicate a defect
in mitochondria function in AD (Ghosh et al., 1999; Khan et al., 2000;
Trimmer et al., 2000). However, these studies implicating mitochondrial
abnormalities in AD were not focused on specific neuronal cytological
abnormalities in mitochondria in vulnerable neurons and their
relationship to oxidative damage.
In this study, we use cytological in situ hybridization,
immunocytochemistry, and morphometry to determine whether mitochondrial abnormalities are associated with vulnerable neurons in AD. Our findings show major abnormalities in mitochondrial dynamics restricted to vulnerable neurons, suggesting an intimate relationship between mitochondria and oxidative damage in AD.
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MATERIALS AND METHODS |
Tissue
Brain tissue was obtained at autopsy from cases with a diagnosis
of AD (Khachaturian, 1985; Mirra et al., 1991) as well as control cases
with no clinical or pathological history of neurological disease.
Hippocampal tissue, including the adjacent temporal cortex, cerebellum,
and frontal cortex from 27 cases of AD (ages 57-93 years; postmortem
intervals, 2-20.5 hr; average, 6.4 hr), 12 old control cases (ages
54-85 years; postmortem intervals, 3-34 hr; average, 13.8 hr), and
eight young control cases (ages 3-49 years; postmortem intervals,
3-23 hr; average, 12.4 hr), was either fixed in methacarn
(methanol-chloroform-acetic acid, 60:30:10) or 2% paraformaldehyde-0.5% glutaraldehyde at 4°C for 16 hr for light or
electron microscopic examination, respectively. For light microscopy, after fixation, tissue was dehydrated through graded ethanol followed by xylene and embedded in paraffin. Sections, 6-µm-thick, were cut
and mounted on silane (Sigma, St. Louis, MO)-coated standard glass
microscope slides for in situ hybridization and
immunocytochemistry. For electron microscopy, tissue was sectioned at
60 µm using a Vibratome and immunodecorated with colloidal gold.
Biopsy
Tissue was taken for diagnostic procedures from the frontal or
temporal cortices of eight patients (age 53-84), most of which had
been included in other clinicopathological studies (Stewart et al.,
1992; Praprotnik et al., 1996a,b) and with a definite history (duration
3-11 years) and clinical presentation of dementia and fulfilling the
National Institute of Neurological and Communicative Disorders and
Stroke and the Alzheimer's Disease and Related Disorders Association working group criteria for probable AD (McKhann et al., 1984). Corresponding tissue from five patients (ages
62-80) suffering from various conditions such as hydrocephalus or
brain tumor were also examined and used as controls. Tissue was fixed in 1.5% glutaraldehyde in cacodylate buffer and post-fixed in 1%
osmium tetroxide for 1 hr immediately after removal from the brain.
After dehydration in graded ethanol and propylene oxide, the tissue was
embedded in Epon 812, sectioned at silver interference color,
electron-contrasted with uranyl acetate and lead citrate, and grids
were viewed at 80 kV using a JEOL 100CX electron microscope.
In situ hybridization
Light microscopy. In situ hybridization was performed
according to the method of Nakamura et al. (1996) with some
modifications. Probes used for in situ analysis of
mitochondrial DNA (mtDNA) were wild-type and with the
common 5 kb deletion (mtDNA 5kb). Four oligonucleotide probes, three
of 45 bp and one of 29 bp in length, were constructed for the present
study. The probes, designated "chimera", include the mtDNA region
from nucleotide coordinate 8454 to 8482 of ATPase subunit 8 and from
13460 to 13475 of NADH-coenzyme Q oxidoreductase subunit 5 and the
"chimera short" included the mtDNA region from 8454 to 8482. The
probes designated as "wild 1" and "wild 2" contain a fragment
that spans nucleotides from 10897 to 10941 and from 10981 to 11025 of
ND4L. The GC content of each 45 bp probe is 51.1%, and the sequences
of the probes lack long palindromic sequences. All probes to mtDNA were
synthesized, labeled by digoxigenin, and purified by Operon
Technologies (Alameda, CA): AluI and 2 repeat sequences were
found exclusively in nuclear DNA (Research Genetics, Huntsville, AL).
After deparaffinization, the sections were rinsed with 0.1 M PBS, pH 7.4, for 10 min. The sections were treated
with 10 µg/ml proteinase K (Boehringer Mannheim, Indianapolis, IN) in
0.1 M PBS for 20 min at 37°C, rinsed in 0.1 M
PBS, then incubated in 0.2 M HCl for 10 min at room
temperature (RT). After rinsing with 0.1 M PBS, the
sections were acetylated with 0.25% (v/v) acetic anhydride in 0.1 M triethanolamine for 10 min at RT. Next, the sections were
treated with RNase A (Sigma) (50 µg/ml) at 37°C for 30 min, rinsed
in 0.1 M PBS, and then dehydrated through a graded ethanol
series up to 100% and allowed to air dry. Hybridization solution
[50% (v/v) formamide, 2× SSC, 10% w/v dextran sulfate, 0.1 mg/ml
sonicated salmon testis DNA (Sigma), 0.2 mg/ml yeast tRNA (Sigma), and
0.4-0.6 µg/ml digoxigenin-labeled probe] was boiled for 10 min at
100°C. The sections were overlaid with 100 µl of the hybridization
solution, placed on a heating block for 5 min at 95°C, and then
hybridized overnight at 40°C. After hybridization, the specimens were
washed sequentially, once with 10 mM Tris-HCl, pH 7.4, 500 mM NaCl, 1 mM EDTA, twice with 2× SSC (1×
SSC: 0.15 M NaCl, 15 mM sodium citrate, pH
7.4), and once with 0.1× SSC. All washes were for 10 min at 37°C.
The sections were then washed twice with Tris-buffered saline (TBS; 50 mM Tris-HCl, pH 7.6, 150 mM NaCl) for 10 min at
RT. After incubation in 10% normal goat serum (NGS) for 1 hr at RT,
slides were incubated with a monoclonal antibody to digoxigenin (1:250;
Boehringer Mannheim) in 1% NGS overnight at 4°C and immunostained by
the peroxidase-anti-peroxidase method (Sternberger, 1986), with 3,3'
diaminobenzidine as cosubstrate. As controls, some specimens were
processed as above but without the oligonucleotide probes and, in other
cases, the hydrated sections were treated with a combination of 50 U/ml
S1 nuclease (Boehringer Mannheim) and 50 U/ml DNase I (Boehringer
Mannheim) overnight at 37°C. In both cases, no positive signal was
detected. Optical densities of in situ hybridization were
measured for manually outlined neuronal cell bodies with a Quantimet
570C Image Processing and Analysis Systems (Leica, Nussloch, Germany)
as previously described (Nunomura et al., 1999)
Electron microscopy. Vibratome sections were treated and
hybridized to probes as described above. Next, they were washed twice with TBS (50 mM Tris-HCl, pH 7.6, 150 mM NaCl) for 10 min at RT, incubated in 10% NGS
for 2 hr at RT, and then incubated with a monoclonal antibody to
digoxigenin (Boehringer Mannheim) diluted in 1% NGS overnight at
4°C. After rinses in 10% NGS, gold (17 nm)-conjugated antibody to
mouse IgG were applied for 4-24 hr and thoroughly rinsed in PBS.
Finally the sections are post-fixed in 2.5% glutaraldehyde for 1 hr
and again rinsed with PBS.
As controls, some specimens were processed as above but without the
oligonucleotide probes, and in other cases the hydrated sections were
treated with a combination of 50 U/ml S1 nuclease (Boehringer Mannheim)
and 50 U/ml DNase I (Boehringer Mannheim) or with omission of the
antibody to digoxigenin at 37°C for 1 hr. Finally, all sections were
exposed to osmium tetraoxide for 1 hr at RT, rinsed, dehydrated through
acetone, and flat-embedded in Spurr's embedding media. Ultrathin
sections were stained with uranylacetate and lead citrate and viewed in
a JEOL 100CX electron microscope at 80 kV.
Immunocytochemistry
To identify the pathology of AD, we used antisera to  (Perry
et al., 1991) to show NFT and a monoclonal antibody (4G8) to A (Kim
et al., 1988) for senile plaques. Nucleic acid and protein oxidative
damage was respectively identified with antibodies to 8-hydroxyguanosine (8-OHG; Trevigen) and nitrotyrosine (clone 7A2; gift
from J. S. Beckman, University of Alabama, Birmingham, AL). A
monoclonal antibody to cytochrome oxidase-1 (clone 1D6; Molecular
Probes, Eugene, OR) was also used in these studies. All immunostaining
for light microscopy was by the peroxidase-anti-peroxidase method
(Sternberger, 1986) and for electron microscopy by immunogold (Perry et
al., 1985). Additionally, Congo red was used to identify the
lesions of AD in some sections.
Morphometry
Micrographs were taken of neurons identified in the biopsy
tissue at the plane containing the nucleolus at magnification of 5000×
and additionally at 20,000× so that a montage including the entire
cytoplasm could be made. Between 2 and 10 neurons were examined for
each case. Photomicrographs were examined with a Zeiss stereomicroscope
at 10-20× and the following organelles were identified and counted
for each neuron: intact mitochondria, mitochondria with broken cristae,
vacuoles associated with lipofuscin, and lipofuscin. Total mitochondria
was the sum of intact mitochondria and mitochondria with broken
cristae. Each structure was outlined and the area was determined (NIH
Image J program) and compared to the total cytoplasmic area excluding
the nucleus.
Differences between groups were compared by the Student's t
test as well as two-way ANOVA (F test).
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RESULTS |
In situ analysis for mtDNA using oligonucleotide probes
revealed a consistent and significant increase in mtDNA levels in cases
of AD (Fig. 1A,C,E)
compared with age-matched (Fig. 1B,D,E) or young
controls (Fig. 1E) for all the probes used
(p = 0.0034; Student's t test) with
no significant difference between probes. Increases in both mtDNA
5kb (Fig. 2A,B) or
wild-type mtDNA (Fig. 2C,D) was restricted to neurons,
particularly those large vulnerable neurons of the hippocampus and
neocortex. Neuronal labeling was seen in granular structures in the
perinuclear cytoplasm and not noted in axons or dendrites. The sites
labeled in the cytoplasm were DNase- but not RNase-sensitive (data not
shown), whereas hybridization to nuclear sequences (AluI and
2) were restricted to the nucleus (data not shown). Importantly, in the
hippocampus, we found that although pyramidal neurons (py) show mtDNA
increase in AD (Fig. 2A,C) compared with controls
(Fig. 2B,D), other neuronal populations, e.g.,
granule cells of the dentate gyrus (gr) as well as glia, show no
increase. Importantly, a similar neuronal specificity was demonstrated
in the frontal and temporal cortex, whereas in the cerebellum, a region
only mildly affected by AD, there was no disease-related increase in
mtDNA, wild-type or deleted, in any cellular populations (data not
shown). The restriction of increased mtDNA to vulnerable neurons means
that although the increase is striking in vulnerable neurons (Fig. 1),
when examined in a regional context (Fig. 2), the change is small
explaining the inability of previous biochemical analysis of tissue
representing a variety of cell types to detect the substantial increase
in mtDNA shown here. In our own comparison of mtDNA in AD and control cases by PCR, we did not find a significant and consistent increase in
mtDNA (K. Hirai, M. A. Smith, and G. Perry, unpublished
observations). No statistically significant correlation was found
between our findings and postmortem interval or agonal status by
regression analysis.
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Figure 1.
Pyramidal neurons of the hippocampus, cells highly
vulnerable to death in AD, show increased mtDNA in all cases of AD
(A, mtDNA5kb, chimeric probe; C,
wild-type mtDNA) compared with controls (B, mtDNA5kb,
chimeric probe; D, wild-type mtDNA). E,
Quantitative densitometric analysis of the level of mtDNA shows the
increases are severalfold and significant (p = 0.0034; Student's t test) for AD compared with old or
young controls ± SEMs. Scale bar, 10 µm.
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Figure 2.
Both mtDNA5kb (chimeric probe) (A,
B) or wild-type mtDNA (wild type 1) (C, D) only
show an increase in vulnerable neurons in AD. In the hippocampus, we
found that, whereas pyramidal neurons (py) show
mtDNA increase in AD (A, C) compared with controls
(B, D), other neuronal populations, e.g., granule cells
of the dentate gyrus (gr), as well as glia, show
no detectable signals. Scale bar, 100 µm.
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The increase in mitochondrial components was not restricted to mtDNA
because the mitochondrial protein cytochrome oxidase was also
significantly elevated in the same neurons (Fig.
3) (p = 0.013;
Student's t test). To examine whether the increased mtDNA and mitochondrial protein noted in AD marked increased mitochondria or
more mtDNA and cytochrome oxidase 1 per mitochondria, we examined the
site of the increase by ultrastructure. We found only a weak in
situ hybridization or immunoreactivity signal from mitochondria in
AD (Fig. 4C) as well as
control samples, not unexpected based on the expectation of 2-10 mtDNA
for each individual mitochondria. However, we also found mtDNA and
cytochrome oxidase-1 (data not shown) in the cytosol and in the
case of mtDNA, also in vacuoles associated with lipofuscin (Fig.
4A,B). Confirming the light microscopic analysis, no
mitochondria in axons or dendrites showed marked mtDNA proliferation
(data not shown), and no signal was detected after the omission of
either oligonucleotide probe, antibody to digoxigenin, or with previous
treatment with DNases (data not shown). These findings indicate that
the increase in mitochondrial markers in AD is related to their
accumulation in the cytoplasm and the late autophagosomes of
lipofuscin.
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Figure 3.
Cytochrome oxidase 1 immunoreactivity is increased
severalfold in neurons in AD (A) compared with
controls (B) and as shown by quantitative
densitometric analysis (C)
(p = 0.013; Student's t
test) ± SEM. Scale bar, 50 µm.
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Figure 4.
Ultrastructural examination of a pyramidal neuron
in the hippocampus after in situ hybridization by using
probes to wild mtDNA (wild type 1) in AD (A, B) or
mtDNA5kb (chimeric probe) (insets, A, B). The high
density of gold particles was seen inside the vacuolar portions of
lipofuscin granules, which likely represents autophagocytosis of
damaged mitochondria in AD (A) and to a lesser
extent in controls (C, inset). In contrast, mitochondria
with cristae in both AD (B) and control cases
(C) showed a lower level of mtDNA labeling
(C). Scale bars: 1 µm; 0.25 µm
(insets).
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To evaluate the change in mitochondria in AD independent of DNA or
protein-related probes as well as whether these changes could be noted
in living patients before agonal state and soon after the onset of
dementia, we quantitatively examined mitochondria and lipofuscin in
specimens removed at biopsy (Fig. 5).
Morphometric analysis (Fig. 6) showed
that the area of intact mitochondria is significantly decreased in AD
(p = 0.012; F test) whereas there is
no difference between the area of damaged mitochondria in AD or control
cases. Because mitochondria are highly susceptible to morphological
artifacts such as broken cristae, through inadequate fixation, that
controls and AD cases showed similar frequency of mitochondria with
broken cristae suggests that fixation was similar for each group.
Whereas the percentage of area for vacuoles associated with lipofuscin
and lipofuscin were both increased in AD, the difference was not
significant. As reported earlier (Dowson et al., 1998), we noted that
the size of lipofuscin granules and vacuoles is larger in AD, but only
the latter reached significance (p = 0.029;
F test). Although only six of the neurons in the AD cases
contained neurofibrillary tangles, we found no significant differences
in mitochondria or lipofuscin in those neurons compared with
others.
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Figure 5.
Examination of the morphology of
mitochondria and lipofuscin in specimens removed at biopsy showed
intact mitochondria (A), mitochondria with broken
cristae (B), and vacuoles associates with
lipofuscin indicated by a V and lipofuscin indicated by
an L (C). Scale bars, 1 µm.
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Figure 6.
Morphometric analysis of the number
(A), size (B), and
percentage coverage (C) of cytoplasmic area by
intact mitochondria, mitochondria with broken cristae, total
mitochondria (intact plus mitochondria with broken cristae), vacuoles
associated with lipofuscin, and lipofuscin in cases of AD and controls.
Whereas percentage coverage of intact mitochondria decreases in AD
(p = 0.012), no significant changes were
noted in vacuoles associated with lipofuscin
(p = 0.056), mitochondria with broken
cristae (p > 0.10), or lipofuscin
(p > 0.08). Although the size of lipofuscin
did not change in AD (p = 0.095), that of
the vacuoles associated with lipofuscin significantly increased
(p = 0.029). Statistical comparison by the
F test. * indicates significant difference between AD
and control cases.
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Oxidative damage marked by 8-OHG and nitrotyrosine was increased in the
same neurons displaying mtDNA proliferation (Fig. 7). Also, as we previously noted for
8-OHG (Nunomura et al., 1999) and nitrotyrosine (Smith et al., 1997),
one of the most striking features of the mitochondrial abnormalities is
its relatively uniform effect on entire populations of vulnerable
neurons. Therefore, these data support an intimate topographic and
probably temporal relationship between neuronal oxidative damage and
mitochondrial abnormalities.
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Figure 7.
The distribution of neurons showing increased
mtDNA5kb (chimeric probe) (A), 8-OHG
(B), and nitrotyrosine (C)
immunoreactivity in AD completely overlaps. Number
indicates the same neurons in adjacent serial sections. Scale bar, 50 µm.
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DISCUSSION |
In this study, we addressed whether there are mitochondrial
abnormalities in vulnerable neurons by examining mtDNA, mitochondrial protein, and mitochondrial number in AD. We found both increased mtDNA
and protein in AD. Ultrastructural examination showed the increased
mtDNA and protein was found in the cytoplasm and in the vacuoles
associated with lipofuscin, a lysosome that, in previous studies of
cells, has been suggested as the site of mitochondrial degradation by
autophagy (Brunk et al., 1992). Morphometry of the organelles of
samples obtained at biopsy demonstrates there is, in fact, a
significant decrease in mitochondria of vulnerable neurons in AD. These
findings indicate vulnerable neurons in AD have increased mitochondrial
degradation products, suggesting either greater turnover of
mitochondria by autophagy or a reduction of proteolytic turnover
leading to accumulation of mtDNA and mitochondrial protein. These
mitochondrial components are likely damaged, because hydroxynonenal
adducts to lipoic acid (Humphries and Szweda, 1998), the prosthetic
group of two key Krebs cycle enzymes, can be found in the same vacuoles
substantiates the view that these components are nonfunctional (G. Perry, M. A. Smith, and L. Szweda, unpublished observations).
Also, because cytochrome oxidase must be membrane-bound to function,
our findings are consistent with the low functional activity of many
mitochondrial enzymes in AD (Wong-Riley et al., 1997).
The restriction of damaged mtDNA to neurons vulnerable to death in AD
means that although the increase is striking on a per cell basis, when
seen in the context of brain tissue, the change is very selective. The
selectivity likely explains the conflicting results of previous
biochemical analyses of mtDNA by PCR analysis. When we analyzed mtDNA
changes by PCR, we also found only small differences between samples
from AD and controls even in cases shown by in situ
hybridization to show a fourfold increase (data not presented). This
restriction to vulnerable neurons is the same found for oxidative
damage, suggesting that an intimate relationship exists between them.
The absence of mtDNA accumulation in non-neuronal cells does not mean
that they do not also show changes in mitochondria. In a previous
morphometric study, significant reduction in mitochondria density was
found in endothelial cells (Stewart et al., 1992). Blass et al. (1990)
and Blass and Gibson (1991) have also elegantly shown
mitochondrial abnormalities in fibroblasts and other cells obtained
from patients with AD. The striking changes noted here may reflect as
much how different categories of neurons deal with mitochondrial
abnormalities, and such an interpretation is consistent with the
findings of recent work with cytoplasmic hybrids that show mitochondria
derived from non-neuronal cells from cases of AD have substantial
energetic deficiencies (Ghosh et al., 1999; Khan et al., 2000; Trimmer
et al., 2000). It is tempting to consider that the range of oxidative
balance abnormalities noted in AD stem from a fundamental mitochondrial
deficiency, but clearly more work is necessary to establish such a
relationship. Although differences in mtDNA heredity may be important
to our observations, the restriction of changes to vulnerable neurons
only in cases of AD, rather than a more generalized change involving
all cell types, indicates mitochondrial inheritance alone is not the
only factor, but probably differences in mitochondrial turnover and metabolism as well as oxidant defense between different categories of
cells are involved and require further clarification in future studies
(Ito et al., 1999). Among the possible mechanisms underlying cellular
specificity, reactive oxygen in vulnerable neurons may damage
mitochondria while reducing their degradation (Friguet et al., 1994).
Alternatively, mitochondria are likely not being transported to the
axon properly consistent with the reduced number of microtubules seen
in neurons in AD (A. D. Cash, M. A. Smith and G. Perry, unpublished observations).
That abnormalities occur in neurons lacking neurofibrillary tangles
places mitochondria abnormalities as the earliest cytopathological change in AD. Other changes of AD could very well be linked to mitochondria because blockage of mitochondrial energy production shifts
amyloid -protein precursor metabolism to the production of more
amyloidgenic forms of amyloid- (Gabuzda et al., 1994), induces the
production of A68 antigen (Blass et al., 1990), and activates the
mitogen-activated protein kinase pathway (Luo et al., 1997;
Perry et al., 1999; Zhu et al., 2000, 2001), all features of AD.
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FOOTNOTES |
Received Oct. 13, 2000; revised Feb. 14, 2001; accepted Feb. 16, 2001.
This work was supported by National Institutes of Health Grants
AG09287, AG14249, P50 AG16570, and NS38648, the American Health Assistance Foundation and the United Mitochondrial Diseaese Foundation.
K.H. and G.A. contributed equally to this study.
Correspondence should be addressed to Dr. Mark A. Smith, Institute of
Pathology, Case Western Reserve University, 2085 Adelbert Road,
Cleveland, OH 44106. E-mail: mas21{at}po.cwru.edu.
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