вход по аккаунту


An immunochemical study of the pyruvate dehydrogenase deficit in Alzheimer's disease brain.

код для вставкиСкачать
An Immunochemical Study
of the Pyruvate Dehydrogenase Deficit in
Alzheimer's Disease Brain
Kwan-Fu Rex Sheu, PhD,"S Young-Tai G m , PhD,? John P. Blass, MD, PhD,"$ and Marc E. Weksler, MDt
The activity of the pyruvate dehydrogenase complex (PDHC; EC, EC, and EC was reduced to
about 30% of control values in histologically unaffected occipital cortex of the brains of patients with Alzheimer's
disease, as well as in histologically affected frontal cortex. In contrast, activity of another mitochondrial enzyme,
glutamate dehydrogenase, was normal. Neither age nor time until postmortem study correlated significantly with
PDHC activity in either Alzheimer or control samples, and PDHC was not inactivated significantly on incubation with
homogenates of either Alzheimer or control brain. Antibodies against the highly purified bovine PDHC inhibited
Alzheimer and control PDHC equally per unit of enzyme activity. Immunoblots also indicated that the PDHC antigens
were not different in normal and Alzheimer brains. This antibody, however, inhibited Alzheimer PDHC more effectively than it did control PDHC, based on milligrams of protein, suggesting a reduced amount of normal PDHC
protein. Other data suggest that the PDHC deficiency is related to mitochondrial damage and to impaired calcium
homeostasis in Alzheimer nerve cells, which may then mediate a variety of other cellular impairments.
Sheu K-FR, Kim Y-T, Blass JP, Weksler ME: An immunochemical study of the pyruvate dehydrogenase
deficit in Alzheimer's disease brain. Ann Neurol 17:444-449, 1985
Whereas the loss of cholinergic and other neurotransmitter markers is well documented in the brains of
patients with Alzheimer's disease [6, 271, the reason
for the death of the cells that make these neurotransmitters is not known. We [ S , 271 and others [I, 22)
have suggested that one of the mechanisms in a number of neurodegenerative disorders is histotoxic hypoxia resulting from damage to cellular components
required for oxidative metabolism, including key mitochondrial enzymes. One of these mitochondrial constituents, which may be a sensitive marker for mitochondrial damage, is the pyruvate dehydrogenase
complex (PDHC; EC, EC, and EC It has been reported to be reduced in histologically abnormal Alzheimer frontal [29f, parietal,
and temporal 1181 cortices. This enzyme complex is of
particular interest because it is linked not only to energy metabolism, but also to acetylcholine synthesis
14) and to cellular calcium homeostasis [S). Furthermore, quantitatively the major phosphoprotein whose
phosphorylation pitch changes with habituation 1, 81
or training 1163 is the 42,000 dalton a-peptide of
We now report that PDHC is as reduced in histologically normal occipital cortex from Alzheimer brain
as in histologically affected frontal cortex, and that this
PDHC appears to be antigenically and electrophoretically normal. The reduction in PDHC activity may be
related to loss of cellular calcium homeostasis, which
itself may be a pathophysiologically critical change in
Alzheimer's disease 117, 20).
From the Departments of *Neurology and tMedicine, Cornell University Medical College, New York, N Y 10021, and the $Altschul
Laboratory of Dementla Research, Burke Rehabilitation Center,
White Plains, NY 10605.
Received Aug 23, 1984, and in revised form Nov 21. Accepted for
publication Nov 25, 1984.
Materials and Methods
Preparation of Brain Specimens
Postmortem specimens of frontal cortex, occipital cortex,
and caudate nucleus from Alzheimer and matched control
brains were kindly supplied by Drs Peter Davies and Robert
Terry, Departments of Pathology and Neuroscience, Albert
Einstein College of Medicine, Bronx, NY, from the Alzheimer's Disease Research and Development Association research brain bank. Histological examination there reportedly
demonstrated no notable neuritic plaques or neurofibrillary
tangles in the samples of occipital cortex studied. Detailed
information on the agonal state was not available, although
more of the patients than of the controls had pneumonia
Tissue was routinely homogenized (50 mg wet weight per
milliliter) in 10 mM N-morpholinopropanesulfonic acid (pH
adjusted to 7.2 with Tris), 2 mM dithiothreitol, 100 FM
thiamin pyrophosphate, and 10 p~ leupeptin hemisulfate.
For enzyme activity measurements 1 g d L of Triton X-I00
Address reprint requests to: D~ Sheu, ~~~~~~i~ ~~~~~~h services,
Burke Rehabilitation Center, 785 Mamaroneck Ave, White Plans,
N Y 10605.
Enzyme Activities in Brain in Alzheimer'~Disease"
Age (yr)
Time until
Autopsy (hr)
Frontal Cortex
Occipital Cortex
80 t 3
9 5 2
12 -+ 3
10.6 +- 0.8
3.8 +- 1.2b
237 +- 9
212 +- 20
2.9 t 1.2'
160 2 38
186 t 13
"Brain regions from 7 controls and 7 patients with Alzheimer's disease were studied. Enzyme activities axe expressed in nanomoles per minute
per milligram of protein, measured at 30°C. All values are mean f SEM.
Significantly different from value in control group: 'p < 0.01; 'p < 0.001.
PDHC = pyruvate dehydrogenase complex; GDH = glutamate dehydrogenase.
and 1 gmlL of Lubrol P X (final concentrations) were further
added. For activity-antibody titrations PDHC was then solubilized (to be described). To test for proteolytic activity,
which might inactivate PDHC, brain homogenates (50 mg
wet weight per milliliter) were prepared in 10 mM N-morpholinopropanesulfonic acid (pH adjusted to 7.2 with Tris),
2 mM dichiothreitol, 1 gm/L of Triton X-100, and 1 gm/L of
Lubrol PX, in the absence of Ieupeptin and thiamin pyrophosphate. For enzyme activity measurements homogenates were frozen at - 80°C. Activity appeared stable when
homogenates were frozen for at least 40 hours. For immunoblotting analysis, the homogenates were solubilized in
20 g d of sodium dodecyl sulfate before they were frozen
at - 80°C. Immunoblots were obtained within a week.
Enzyme Activity Measurements
PDHC activity was assayed based on acetyl-coenzyme A
formation C14, 253. Homogenates were preincubated with 5
mM magnesium chloride and 0.1 mM calcium chloride for 10
minutes at room temperature to activate PDHC [26]. In a
few experiments PDHC was also activated by further addition of a phosphatase isolated from pigeon liver acetone
powder (12).
Glutamate dehydrogenase (EC was assayed according to the method of Plaitakis and colleagues 1211, except that NADH was used in a concentration of 0.15 mM.
Immunochemical Studies
Immunochemical studies were conducted with rabbit antibodies against the highly purified bovine kidney PDHC
[24]. As described elsewhere, these antibodies cross-react
with human PDHC [27] and do not discriminate between
PDHC and phospho-PDHC [24J
Protein in brain samples was
resolved by sodium dodecyl sulfate-polyacrylamide (95%)gel
electrophoresis according to the technique of Weber and
Osborn [3 11 and then transferred electrophoretically to a
nitrocellulose membrane [30]. The PDHC peptides on the
nitrocellulose blots were identified using the anti-PDHC and
horseradish peroxidase conjugated goat antirabbit immunoglobulin second antibodies (Bio-Rad Laboratory, Richmond,
CA) [ 111. These procedures have been described elsewhere
TITRATIONS. Brain homogenates
prepared as described and containing 20 gm/L of Triton
X-100 were thawed and centrifuged in an Eppendorf microfuge at top speed for 10 minutes. Aliquots of the clear
supernatants were incubated with anti-PUHC and control
immunoglobulins, as detailed in the legends to the figures.
Other Procedures
Protein was determined according to the method of Lowry
and co-workers [l5], using bovine serum albumin as standard. Statistical significance was evaluated by Student's twotailed t test.
It has been reported previously that the PDHC activities in frontal C291, parietal, and temporal 118) cortices
in Alzheimer brain are lower than those in control
brain. The present study demonstrates that PDHC activity in histologically unaffected occipital cortex was
also significantly ( p < 0.01) reduced in Alzheimer's
disease, as it was in histologically affected frontal cortex ( p < 0.001) (Table). Furthermore, the degree of
deficit in the former region (27% of control) was almost as pronounced as in the latter (35%). Treatment
with an exogenous PDHC-phosphatase did not increase PDHC activity in controls (from 12.5 k 1.7 to
12.6 4 2.3 nmollmidmg protein, mean t SEM) but
did increase it slightly in Alzheimer samples (from 3.4
+- 1.3 to 5.6 & 1.5 nmoYmidmg protein; p < 0.01).
(Samples were the same as described in Figure 4.)
Thus, phosphatase activation ameliorated the defect
only partially (from 27% of control to 44% of control). The twofold to threefold reduction in PDHC
activity in Alzheimer brain was unlikely to reflect a
similar degree of loss of mitochondria, because glutamate dehydrogenase, another enzyme localized in mitochondria, did not show significant differences between Alzheimer and control brains. Also, Sorbi and
colleagues 1291 have reported that the activity of another mitochondrial enzyme, fumarase, was unchanged
in Alzheimer frontal cortex. The deficit in PDHC
activity was also unlikely to be due to age or other
postmortem artifacts. Age and time until postmortem examination did not, on average, appear to be
significantly different in patients with Alzheimer's disease and controls (see the Table) ( p > 0.05). The cor-
Sheu et al: Pyruvate Dehydrogenase in Alzheimer's Disease 445
0 '
0 0 0
V, 60%
Fig 2. Incubation of Alzheimer and control brain homogenates
with exogenous pyruvate dehydrogenase complex (PDHC). Control (open circles) or Alzheimer (filled circles) brain homogenates (100 $; 5%, wlv) were incubated with 30 pg of purzj5ed
bovine kidney PDHC in a buffer containing 1 mM magnesium
chloride and 0.1 mM calcium chloride at room temperature. The
activity of PDHC was monitored at the times indicated. Results
are expressed as means & SEM ( n = 3). See the Materials and
Methods section for details.
AGE ( y r )
F i g 1 . Pyruvate dehydrogenase complex (PDHC) activities in
Alzheimer and normal brains as functions of postmortem delay
and patient age. PDHC activities in Alzheimer (filled symbols) and normal control (open symbols) brain are plotted as a
function of time until postmortem examination (A)and patient
age (B). In A the linear regression showed thefillowing values:
nomzal, slope (m) = 0.500, cowelation coefficient (r) = 0.357;
Alzheimer group, m = 0.190, r = 0.420. In B: normal,
m = 0.0449, r = 0.165; Alzheimer group, m = 0.192, r =
- 0.245. (Circles = frontal cortex; squares = occipital cortex.)
relations between age or postmortem delay and
PDHC activity were also poor (Fig l), accounting at
maximum for less than 20% of the variance.
A conceivable pathogenetic (or postmortem) cause
of this diminished PDHC in Alzheimer brain is that
affected brain contains more proteolytic enzymes that
degrade PDHC than does normal brain, either during
life or after death. To test this hypothesis, an assay
system was set up in which purified PDHC was added
exogenously to saturate this putative proteolytic enzyme in the absence of leupeptin and thiamin pyrophosphate (Fig 2). Over an extended period of incubation ( 3 hours), the exogenously added PDHC
remained relatively stable in both control and Alzheimer brains. This observation does not support the
existence of a proteolytic enzyme or other system that
446 Annals of Neurology VoI .I7 No 5
May 1985
selectively inactivates PDHC in Alzheimer brain,
either as part of the pathogenesis during life or post
In order to delineate further the molecular basis for
the low PDHC activity in Alzheimer brain, this enzyme was studied using rabbit antibodies raised against
the bovine kidney PDHC E24, 271. A typical immunoblot is shown in Figure 3. In all brains examined
(6 control and 6 Alzheimer brains), no apparent difference in the size or distribution of PDHC peptides was
observed between these two groups. These blots were
also very similar among frontal and occipital cortices
and the caudate nucleus. The three main immunogenic
peptides were, in decreasing size, the dihydrolipoyl
transacetylase (70,000 daltons), the dihydrolipoyl dehydrogenase (58,000 daltons), and the a-peptide of
pyruvate dehydrogenase (42,000 daltons). This antibody does not recognize the P-peptide of pyruvate
dehydrogenase on these blots [2 7 ) . Proteolysis was
also evident, as in one set of the control samples in
Figure 3 . The extra immunogenic peptide of 60,000
daltons was probably a proteolytic fragment of the
larger dihydrolipoyl transacetylase component. These
immunoblotting results suggest that the PDHC peptides in Alzheimer brain are not grossly different from
the normal PDHC peptides. The Alzheimer brain contained, if anything, fewer proteolytic fragments that
bound to the antibody than did the controls (see Fig 3),
supporting the conclusion that the lower activity does
not result from selective proteolysis. The lower enzyme activity may result from the presence of fewer
enzyme molecules.
g 80%.
p g ANTI-PDHClmg
4 0 0 6 0 0 70(
Fig 3. Immunoblots of pyruvate dehydrogenase complex (PDHC)
in Alzheimer brain. Brain homogenates of 40 pg o f protein each
were resolved by sodium dodecyl sulfate-polyacykzmide gel electrophoresis. The PDHC peptides were identified by immunoblotting, as detailed in Materials and Methods. Typical results are
shown, from two of the six Alzheimer brains and two of the six
control brains studied. (F = frontal cortex; 0 = occipital cortex;
C = caudate nucleus; CON = control brain; ALZ = Alzheimer brain. The PDHC peptides were: E2, dihydrolipoyl
transacetylase component, 70,000daltons; E 3, dihydrolipoyldehydrogenase component, 58,000 daltons; and a E l , a-peptide of
pyruvate dehydrogenase component, 42,000 daltons.)
To quantitate PDHC protein, enzyme activities in
control and Alzheimer brain extracts were titrated
with this polyclonal antibody. The binding of antibody
was estimated from the initial linear portions of the
activity inhibition curves (Fig 4). On a protein weight
basis (see Fig 4A), control brain samples required
significantly more antibody than did Alzheimer brains
( p < 0.01) to reach the same degree of inhibition.
When, however, the titration curves were compared
per unit of enzyme activity (see Fig 4B), no significant
differences were observed between these two groups
( p > 0.5). Both the immunoblots and the immunotitration findings are consistent with the existence of a
reduced quantity of normal PDHC antigens in Alzheimer brain, about 36% of that in the controls, as
determined by activity-antibody titration (see Fig 4A).
These results are in good agreement with the activity
The problem in interpreting alterations in chemical
constituents of Alzheimer brain is that so many of
them decrease-seventeen of thirty-five in the studies
of Bowen and colleagues 171. One must decide
whether the changes are postmortem artifacts, whether
they are primary or secondary, and whether they are
Fig 4. Titration of pyruvate dehydrogenase complex (PDHC) activity by anti-PDHC in Alzheimer brain. Alzheimer (filled
symbols) and control (open symbols) brain extracts (50 to 250
$; 0.2 to 3 nmollminlmg of PDHC) offrontaland occipital
cortices were incubated with various amounts of anti-PDHC
antibody. Control immunoglobulins were added t o equalize the
final volumes for each brain sample. The incubations were performed at room temperaturefor 25 minutes. The residual
PDHC activities were rekztive activities, compared with those before incubation. The amount of anti-PDHC required to neutralize the PDHC activity was estimated by extending the initial
linear portion of the titration curves t o the horizontal axes. (A)
Titration of PDHC activity per unit o f homogenate protein
The amounts of anti-PDHC required t o neutralize enzyme activity in control and Alzheimer brain samples were 239 33
and 86 t 6 pg anti-PDHC per milligram of brain protein, respectively (mean SEM; p < 0.01).(B) Titration of PDHC
activity per unit of enzyme activity. The amounts of antiPDHC wquived to neutralize enzyme activity in control and Alzheimw brain samples were 12.6 5 1.7 and 14.0 3.3 pg
anti-PDHC per milliunit of PDHC, respectively (mean
Sheu et al: Pyruvate Dehydrogenase in Alzheimer's Disease
important pathophysiologically. In fact, the chemical
changes reported to date that are not postmortem artifacts are all considered to be secondary, even if important {b, 29). For instance, the decrease in the cholinergic marker enzyme choline acetyltransferase is
generally considered to be secondary to loss of cholinergic nerve endings [b].
The decreases in PDHC activity in Alzheimer brain
do not appear to be due to postmortem artifact. Previous studies in this laboratory [14, 291 and elsewhere
[Is) indicate that PDHC is stable in brain post mortem. Incubation of purified PDHC with homogenates
of postmortem human brain caused the same slow rate
of inactivation in Alzheimer and control samples. The
activity level of PDHC did not correlate significantly
with either time until postmortem study or patient age.
The consistent decrease in PDHC activity found in
two previous independent studies 118, 291 as well as
the present one argues against a major contribution
of the agonal state. Animals chronically ill with pyrithiamine-induced thiamin deficiency do not lose
brain PDHC activity, when measured by a method
similar to that used in this study 19). The possibility
that the agonal state has important effects on PDHC
activity in human brain regions postmortem remains
open, however, until it is systematically investigated.
The data we have presented do not suggest a primary
structural abnormality in PDHC peptides in Alzheimer brain, although they do not rule out genetic
changes that alter the amount of protein or selectively
alter the P-peptide. Neither immunological nor electrophoretic studies provide evidence of abnormalities
in PDHC peptides in Alzheimer brain. Rather, the
reduced enzyme activity appears to be due to a reduced amount of normal antigen, as evidenced by increased enzyme inhibition per milligram of brain protein. Studies of PDHC in Alzheimer fibroblasts are
under way but as yet have shown no clear-cut abnormality (J. P. Blass et al, unpublished results, 1985).
Decreases in PDHC are not unique to Alzheimer’s
disease and have been reported in a number of other
system degenerations, including Huntington’s disease
1291 and hereditary ataxias C31 as well as several
neurological diseases of childhood (21. It is possible
that PDHC is a relatively labile constituent of mitochondria, so that secondary decreases in PDHC may
be a sensitive marker of mitochondrial dysfunction.
As has been pointed out elsewhere { 5 ] , disorders that
impair mitochondrial oxidative metabolism are likely
to present clinically as disease of the nervous system,
because the nervous system has the tightest, most immediate dependence on oxidative metabolism of any
organ system. The hypothesis has been advanced that
mitochondrial damage marked by PDHC deficiency is
a common pathophysiological mechanism in a variety
of degenerative disorders of the nervous system, even
448 Annals of Neurology
Vol 17 No 5
May 1785
though for unexplained reasons they have distinguishable patterns of damage neuropathologically.
Several lines of data suggest that mitochondrial damage is pathophysiologically important in Alzheimer’s
disease. The decreases in PDHC are as marked in histologically unaffected occipital cortex as in histologically affected frontal cortex, indicating that they are
not simply consequences of anatomical damage. They
occur without significant changes in the levels of the
other mitochondrial enzymes assayed to date, glutamate dehydrogenase (see the Table) and fumarase
1291, indicating that they are not simply nonspecific
markers of mitochondrial damage. They do not appear
to be due to differences in the activation state of
the degree of phosphorylation of its
42,000-dalton peptide-because
treatment with exogenous phosphatase ameliorated the deficit only
slightly. Cholinergic systems are exquisitely sensitive
to even small abnormalities in oxidative metabolism
13, 41, and cholinergic systems are characteristically
damaged in Alzheimer’s disease 16, 7, 29). The PDHC
also appears to be a critical portion of the mitochondrial calcium pump (I, 8). The sequestration of calcium in mitochondria is a major mechanism for maintaining cellular calcium homeostasis, and failure of this
mechanism can induce a cascade of effects deleterious
to neurons {13, 17, 19, 20, 23, 28). Indeed, Peterson
and Gibson { 19,201 have proposed that loss of cellular
calcium homeostasis is a critical lesion in brain aging,
affecting cholinergic as well as other systems.
It is tempting to propose that the mitochondrial
damage is related to alterations in neuronal calcium
homeostasis, which can lead to a variety of pathophysiologically important events. The loss of PDHC activity
appears large enough to impair mitochondrial calcium
sequestration and increase calcium levels in the rest of
the cell {I, 81. Per1 and Brody [I71 have reported that
the levels of calcium are elevated in Alzheimer
neurons that bear neurofibrillary tangles. Their
methods do not allow measurement of mitochondrial
calcium. Selkoe and co-workers t231 have proposed
that the neurofibrillary tangles are, in fact, formed by
the action of a calcium-stimulated transglutaminase on
normal neurofilament proteins. Peterson and Gibson
rI9, 20) have shown that the impairment of cholinergic function that accompanies even mild impairment
of Oxidative metabolism can be attributed largely to
changes in calcium-dependent release of acetylcholine.
They have shown that treatment designed to normalize
cellular calcium homeostasis ameliorates this effect of
hypoxia and aging on cholinergic function and behavior C19, 20). Siesjo C28) has emphasized that increases
in cytoplasmic calcium caused by impairment of the
mitochondrial calcium pump can activate a variety of
calcium-stimulated lipases and proteases, leading to
cell damage and, eventually, autolysis and cell death.
This is, in fact, one of the prominent mechanisms of
neuronal cell death now being examined in a number
of laboratories C281. Kandel and Schwartz 1131 and
Greengard El01 have emphasized the role of calcium
in mediating the molecular changes, including phosphorylation of membrane proteins, involved in processing information at the cellular level. Thus, reduction in the activity of PDHC, with impairment of the
mitochondrial calcium pump and increases in cytoplasmic calcium, might underlie a whole spectrum of
changes in Alzheimer’s disease: impairment of information processing, impairment of cholinergic function,
accumulation of tangles, and ultimate cell death. Only
future studies can tell whether this formulation will be
useful for understanding the pathophysiological mechanisms of Alzheimer’s disease.
Supported in part by Grants AG 00842 and NS 15125 from the
National Institutes of Health, and grants from the Will Rogers Institute, the Winifred Masterson Burke Relief Foundation, and the
George E. Link, Jr. Foundation.
The authors thank Drs Peter Davies and Robert Terry for providing
brain samples, and Mrs Lois Hinman for the gift of the liver phosphatase. They thank Mr Gary Dorante and Miss Jennifer B a g for
expert assistance. Dr Gary E. Gibson and other colleagues in the
Department of Neurology, Cornell University Medical College,
provided stimulating advice.
1. Baudry M, Gall C, Kessler M, et al: Denemation-induced decrease in mitochondrial calcium transport in rat hippocampus. J
Neurosci 3:252-259, 1983
2. Blass JP: Idborn error of pyruvate metabolism. In Stanbury JB,
Wyngaarden JB, Fredrickson DS, et al (eds): The Metabolic
Basis of Inherited Disease. Fifth edition. New York, McGrawHill, 1983, pp 193-203
3. Blass JP, Gibson GE: Studies of the pathophysiology of pyruvate dehydrogenase deficiency. Adv Neurol21:181-194, 1978
4. Blass JP, Gibson GE, Shimada M, et al: Brain carbohydrate
metabolism and dementia. In Roberts PJ (ed): Biochemistry of
Dementia. London, Wiley, 1980, pp 121-134
5. Blass JP, Zemcov A: Alzheimer disease: a metabolic systems
degeneration? Neurochem Pathol 2:103-114, 1984
6. Bowen DM, Allen SJ, Benton JS, et al: Biochemical assessment
of serotonergic and cholinergic dysfunction and cerebral atrophy in Alzheimer disease. J Neurochem 41:266-272, 1984
7. Bowen DM, White P, Spillane JA, et al: Accelerated aging or
selective neuronal loss as an important cause of dementia? Lancet 1:ll-14, 1979
8. Browning M, Baudry M, Bennett WF, et al: Phosphorylationmediated changes in pyruvate dehydrogenase activiry influence
pynwate-supported calcium accumulation by brain mitochondria. J Neurochem 361932-1940, 1981
9. Gibson GE, Ksiezak-Reding H, Sheu K-FR, et al: Correlation
of enzymatic, metabolic, and behavioral deficits in thiamin
deficiency and its reversal. Neurochem Res 9803-814, 1984
10. Greengard P lntracellular signals in the brain. Harvey Lect
75:277-331, 1981
11. Hawkes R, Niday E, Gordon J: A dot-immunobinding assay for
monoclonal and other antibodies. Anal Biochem 119:142-147,
12. Hinman LM,Baker AC, Blass JP: Protein phosphatase 1 from
pigeon liver: a convenient reagent for activation of pyruvate
dehydrogenase complex in uitro. Fed Proc 43:1869, 1984
13. Kandel ER, Schwartz JH: Molecular biology of learning: modulation of transmitter release. Science 218:433-443, 1982
14. Ksiezak-Reding H , Blass JP, Gibson GE: Studies on the pyruvate dehydrogenase complex in rat brain with the arylamine
aceryltransferase-coupled assay. J Neurochem 38:1627-1636,
15. Lowry OH, Rosebrough NJ, Farr AL, et al: Protein measurement with the Fohn phenol reagent. J Biol Chem 193:265-275,
16. Morgan DG, Routtenberg A: Brain pyruvate dehydrogenase:
phosphorylation and enzyme activity altered by a training experience. Science 214:470-471, 1981
17. Per1 DP, Brody AR: Alzheimer’s disease: x-ray spectrometric
evidence of aluminum accumulation in neurofibrillary tanglebearing neurons. Science 208:297, 1980
18. Perry EK, Perry RH, Tomlinson BE, et al: Coenzyme-A acerylating enzymes in Alzheimer disease: possible cholinergic “compartment” of pyruvate dehydrogenase. Neurosci Lett 18:105110, 1980
19. Peterson C, Gibson GE: Aging and 3,4-diaminopyridine alters
synaptosomal calcium uptake. J Biol Chem 258: 11,48211,486, 1983
20. Peterson C, Gibson GE: Amelioration of age-related neurochemical and behavioral deficit by 3,4-diaminopyridine.
Neurobiol Aging 4:25-30, 1983
21. Plaitakis A, Nicklas WJ, Desnick RJ: Glutamate dehydrogenase
deficiency in three patients with spinocerebellar syndrome. Ann
Neurol 7:297-303. 1980
22. Quastel JH: Biochemistry and mental disorder. Lancet 2: 14 171424, 1932
23. Selkoe DJ, Abraham C, Ihara Y:Brain transglutaminase: in
vitro cross-linking of human neurofilament proteins into insoluble polymers, Proc Natl Acad Sci USA 79:6070-6074, 1982
24. Sheu K-FR, Kim YT: Studies on the bovine brain pyruvate
dehydrogenase complex using the antibodies against bovine kidney enzyme complex. J Neurochem 43:1352-1358, 1984
25. Sheu K-FR, Lai JCK, Blass JP: Pyruvate dehydrogenase (PDHb)
phosphatase in brain: activity, properties and subcellular locahzation. J Neurochem 40:1366-1372, 1983
26. Sheu K-FR, Lai JCK, Blass J P Properties and regional distribution of pyruvate dehydrogenase kinase in rat brain. J
Neurochem 42:230-236, 1984
27. Sheu K-FR, Lai JCK, Kim YT, et al: Immunochemical characterization of pyruvate dehydrogenase complex in rat brain. J
Neurochem 44593-599, 1985
28. Siesjo BK: Cell damage in the brain: a speculative synthesis. J
Cerebral Blood Flow Metab 1:155-185, 1981
29. Sorbi S, Bird ED, Blass JP: Decreased pyruvate dehydrogenase
complex activity in Huntington and Alzheimer brain. Ann
Neurol 13:72-78, 1983
30. Towbin H , Staehelin T, Gordon J: Electrophoretic transfer of
protein from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci USA 7643504354, 1979
31. Weber K, Osborn M: Protein and sodium dodecyl sulfate:
molecular weight determination on polyacrylamide gels and reU (eds): The Proteins.
lated procedure. In Neurath H , Hill I
Third edition. New York, Academic, 1975, pp 179-223
Sheu e t al: Pyruvate Dehydrogenase in Alzheimer’s Disease
Без категории
Размер файла
666 Кб
stud, dehydrogenase, disease, pyruvate, brain, deficit, alzheimers, immunochemical
Пожаловаться на содержимое документа