- Email: [email protected]
Premortem diagnosis of cerebral amyloid angiopathy with deposition of cystatin C in Binswanger's type dementia
THIRD INTERNATIONAL
CONFERENCE
117
119
PREPARATIONOF NOMOCLONAL ANTIBODIES TO A BETA PROTEIN SUBPEPTIDE AND SCREENING FOR IMMUNOREACTIVITYIN THE SERA OF PATIENTS WITH ALZHEIMER'SDISEASE, T. Shinoda, Y. Kametani, K. Miyanaga and K. Iinuma. Dept.of Biochemistry,Tokyo MetropolitanUniversity; Dept. of Medicine, Gunma University,Maebashi; National Children' Hospital Research Center, Tokyo 192-03 , Japan. Monoclonalantibodies and polyclonalantisera were raised against synthetic subpeptidescorrespondingto residues (N: 1-28, SPOl) of a beta protein and APP 695 (N: 63-73, SPO3) reported to be a major constituentof the brain amyloid of patients with Alzheimer disease and also of adult Down syndrome brain amyloid. Specificities of these antibodies were characterizedby an ELISA and also by an immunoblottechnique.Titers of monoclonalantibodieswere between 10000-20000,and antisera being 4000-8000,respectively. Crossreactivitiesof antibodies to SPOl, and SP03 were examined at various antigen concentrations.Linear increases in the O&90 nm were attained in both cases over the range of antigen concentrations of 5-lOOng/ml in the test system employed. Using &PO1 and aSP03 in the sandwich ELISA technique,we determined crossreactivitiesin the aera for four different groups, three neurologically affected groups (Alzheimer'sdisease and seniledementia of the Alzheimer'stype(AD/SDAT),multi infarct dementia(MID), senile dementia unspecified(NOS)and a non-demented group. Number of serum specimens available for each group was as follows: ADJSDAT, 2f+;MID, 51; NOS, 25 and the non-demented,94. Mean value (year ks.d.) for each group was respectively72.83 + 1.92, 78. & 1.17, 80.15 f 1.50, and 76.74 f 0.75. Practicallyno aignificant difference in the mean value of the level of the crossreactivitywas seen between male and female groups (male: 16.04 k 3.51ng eq/ml, female: 14.99 k 2.73ng eq/ml). The mean concentration of the immunoreactivityin the sex-aof four different groups was a~ follows: AD/SDAT, 35.11 4 7.91; MID, 16.76 4 2.58; NOS, 13.28 4 2.20 and the non-demented,l/+.&l ?? 1.9l(mean I= s.d.) A highest value of the immunoreactivitywas attained with Alzheimer'sdisease cases. The differencewas significantbetween AD/SDAT and MID (p
SULPHATED CLYCOSAMINOCLYCANS IN THE
118 PREMORTEM DIAGNOSIS OF CEREBRAL AMYLOID ANGIOPATHY WITH DEPOSITION OF CYSTATIN C IN BINSWANGER'S TYPE DEMENTIA, K. Shimode, S. Fujihara. S. Kobayashi, K. Imaoka. A. Nagai. T. Tsunematsu. The 3rd Depts of Internal Medicine. Shimane Medical University, Izumo.693.Japan 1n 1972. Gudmundsson et al. reported the first cases of hereditary cerebral hemorrhage with amyloidosis in Iceland (HCHWA-I). Ghiso et al. demonstrated that this amyloid protein is a variant of cystatin C which is one of cysteine proteinase inhibitor in 1988. We reported the first Japanese case of cerebral amyloid angiopathy (CAA) with deposition of cystatin C like HCHWA-I in 1989, and we developed enzyme-linked immunosorbent assay (ELISA) of cystatin C in cerebrospinal fluid(CSF). Using this method. we confirmed that cases of cerebral hemorrhage due to CAA with cystatin C deposition showed abnormally low concentration of cvstatin C in CSF (Shimode. et al. 1991). 1n this paper. we investigated whether CAA with cystatin C deposition dose relate to multi-infarct dementia (MID) including Binswanger's rype dementia or not. Methods: We measured cystatin C concentration in CSF using ELISA in 40 patients with MID and 30 control patients with various other neurological diseases. MID was diagnosed by DSM-III-R and CT scan. We also examined the brain tissues immunohistologically in one case of MID. Results: Six cases with MID showed abnormally low concentration of cystatin C in CSF. and none of control patients showed low value. These 6 cases were supposed to be Binswanger's type dementia because they showed marked periventricular lucency on CT. We confirmed CAA with cystatin C and p-protein deposition in the cerebral small vessels immunohistologically in one of these patients. Histologically this patient was diagnosed as vascular dementia of Binswanger's type. Discussion: These results indicate that CAA with cystatin C deposition may related to white matter lesion in some case of Binswanger's type dementia, and measurement of cystatin C in CSF is useful to detect such patient in MID.
ON ALZHEIMER’S
DISEASE
CEBEBBOSPINAL FLUID OF INDIVIDUALS WITH DEMENTIA OF THE ALZHEIMEB TYPE, J.Willmer*, RKisilevsky”, D.Guzman*, N.Azad*, S.Al-Shamri*. Dept of Medicine (Neurology and Geriatrics), University of Ottawa*,
Ottawa, Ontario, CANADA and Dept of Pathology, Queen’s University**,
Kingston, Ontario,
CANADA. Solphated glycosaminoglycans(GAGS) have been shown to be aociated
with all forms of
amyloid including that found in the amyloidotic lesions of Alzheimer’s disease. GAG are also components of normal mammalian nervoos systems.The purpose of this study was to examine the levels of CACs in the cerebrospinal fluid ((SF) of individuals with dementia of the Alrheimer type (DAT) to determine whether levels would be higher than individuals who did not have DAT. Eighteen patients referred to a memory disorder clinic were used in this pilot study. These individosb were assessedusing a standardized clinical protocol and classified as probable DAT or non DAT using NINCDS
criteria for dementia. There were no significant differences
in age between the two groups (DAT mean age = 69.5 years, non DAT mean age=66.3 years). The non DAT group had a slight preponderance of females (DAT; 8 males: 1 female, non DAT; 5 males: 4 females). Lumbar punctures were carried out on all individuals and GAG concentrations were determined. Tritiated heparin was added to the (SF in a known quantity to accurately asses the percent GAG recovery. The GAG were precipitated out of the (SF and quantity determined by a calorimetric assayin conjunction with radioactive counts to determine yield. The mean concentration of GAG (expressed as micrograms uranic acid per millilitre) the probable DAT group was 9.97 f 1.73 (~9). (~9).
for
and for the non DAT group was 1286 f2.77
The yield for the probable DAT group was 68.2%* 9.2 and for the non DAT group was
55.8% f 7.2 These differences are statistically significant (p>984b). These results are somewhat different from those expected but show statistically significant differences between groups. The probable DAT group showed lower GAG concentrations and greater yields than the non DAT group. This perhaps suggeststhat the CACs are in the DAT sequestered
brains.
being
No explanation for the differences in yield are apparent at
present. This study is preliminary and needs to he confirmed with larger numbers. R.K. supported by Medical Research Council of Canada grant MA-10477
120 SEARCH FOR PROTEING;yANGES IN ALZHEIMER CSF BY WITH ELECTROPHORESIS TWO-DIMENSIONAL IMMOBILIZED pH GRADIENTS IN THE FIRST DIMENSION Kari Mattila, Tuula Pirttila, Anders Wallin* and Harry Fre University of Tampere, De artment of Clinical Sciences, I!O. BOX 607, SF-33101, Tampere, Finlan B University;, Department of Ps~~;~den and Gothenburg Neurochemistry, Jorgen s ospital, S-42203 Hisings Bat Alzheimer’s disease (AD) is the most common cause of dementia. No definitive antemortem marker has been established in the cerebrospinal fluid (CSF), serum or tissues of patients afflicted by this neurode enerative disorder the primary defect and pathogenesis of which sti9 1 remain obscure.
pH ranges,
among
So far, we have not been able to detect any changes in proteins D maps unique to AD.
them
in the 2-
CONFERENCE
117
119
PREPARATIONOF NOMOCLONAL ANTIBODIES TO A BETA PROTEIN SUBPEPTIDE AND SCREENING FOR IMMUNOREACTIVITYIN THE SERA OF PATIENTS WITH ALZHEIMER'SDISEASE, T. Shinoda, Y. Kametani, K. Miyanaga and K. Iinuma. Dept.of Biochemistry,Tokyo MetropolitanUniversity; Dept. of Medicine, Gunma University,Maebashi; National Children' Hospital Research Center, Tokyo 192-03 , Japan. Monoclonalantibodies and polyclonalantisera were raised against synthetic subpeptidescorrespondingto residues (N: 1-28, SPOl) of a beta protein and APP 695 (N: 63-73, SPO3) reported to be a major constituentof the brain amyloid of patients with Alzheimer disease and also of adult Down syndrome brain amyloid. Specificities of these antibodies were characterizedby an ELISA and also by an immunoblottechnique.Titers of monoclonalantibodieswere between 10000-20000,and antisera being 4000-8000,respectively. Crossreactivitiesof antibodies to SPOl, and SP03 were examined at various antigen concentrations.Linear increases in the O&90 nm were attained in both cases over the range of antigen concentrations of 5-lOOng/ml in the test system employed. Using &PO1 and aSP03 in the sandwich ELISA technique,we determined crossreactivitiesin the aera for four different groups, three neurologically affected groups (Alzheimer'sdisease and seniledementia of the Alzheimer'stype(AD/SDAT),multi infarct dementia(MID), senile dementia unspecified(NOS)and a non-demented group. Number of serum specimens available for each group was as follows: ADJSDAT, 2f+;MID, 51; NOS, 25 and the non-demented,94. Mean value (year ks.d.) for each group was respectively72.83 + 1.92, 78. & 1.17, 80.15 f 1.50, and 76.74 f 0.75. Practicallyno aignificant difference in the mean value of the level of the crossreactivitywas seen between male and female groups (male: 16.04 k 3.51ng eq/ml, female: 14.99 k 2.73ng eq/ml). The mean concentration of the immunoreactivityin the sex-aof four different groups was a~ follows: AD/SDAT, 35.11 4 7.91; MID, 16.76 4 2.58; NOS, 13.28 4 2.20 and the non-demented,l/+.&l ?? 1.9l(mean I= s.d.) A highest value of the immunoreactivitywas attained with Alzheimer'sdisease cases. The differencewas significantbetween AD/SDAT and MID (p
SULPHATED CLYCOSAMINOCLYCANS IN THE
118 PREMORTEM DIAGNOSIS OF CEREBRAL AMYLOID ANGIOPATHY WITH DEPOSITION OF CYSTATIN C IN BINSWANGER'S TYPE DEMENTIA, K. Shimode, S. Fujihara. S. Kobayashi, K. Imaoka. A. Nagai. T. Tsunematsu. The 3rd Depts of Internal Medicine. Shimane Medical University, Izumo.693.Japan 1n 1972. Gudmundsson et al. reported the first cases of hereditary cerebral hemorrhage with amyloidosis in Iceland (HCHWA-I). Ghiso et al. demonstrated that this amyloid protein is a variant of cystatin C which is one of cysteine proteinase inhibitor in 1988. We reported the first Japanese case of cerebral amyloid angiopathy (CAA) with deposition of cystatin C like HCHWA-I in 1989, and we developed enzyme-linked immunosorbent assay (ELISA) of cystatin C in cerebrospinal fluid(CSF). Using this method. we confirmed that cases of cerebral hemorrhage due to CAA with cystatin C deposition showed abnormally low concentration of cvstatin C in CSF (Shimode. et al. 1991). 1n this paper. we investigated whether CAA with cystatin C deposition dose relate to multi-infarct dementia (MID) including Binswanger's rype dementia or not. Methods: We measured cystatin C concentration in CSF using ELISA in 40 patients with MID and 30 control patients with various other neurological diseases. MID was diagnosed by DSM-III-R and CT scan. We also examined the brain tissues immunohistologically in one case of MID. Results: Six cases with MID showed abnormally low concentration of cystatin C in CSF. and none of control patients showed low value. These 6 cases were supposed to be Binswanger's type dementia because they showed marked periventricular lucency on CT. We confirmed CAA with cystatin C and p-protein deposition in the cerebral small vessels immunohistologically in one of these patients. Histologically this patient was diagnosed as vascular dementia of Binswanger's type. Discussion: These results indicate that CAA with cystatin C deposition may related to white matter lesion in some case of Binswanger's type dementia, and measurement of cystatin C in CSF is useful to detect such patient in MID.
ON ALZHEIMER’S
DISEASE
CEBEBBOSPINAL FLUID OF INDIVIDUALS WITH DEMENTIA OF THE ALZHEIMEB TYPE, J.Willmer*, RKisilevsky”, D.Guzman*, N.Azad*, S.Al-Shamri*. Dept of Medicine (Neurology and Geriatrics), University of Ottawa*,
Ottawa, Ontario, CANADA and Dept of Pathology, Queen’s University**,
Kingston, Ontario,
CANADA. Solphated glycosaminoglycans(GAGS) have been shown to be aociated
with all forms of
amyloid including that found in the amyloidotic lesions of Alzheimer’s disease. GAG are also components of normal mammalian nervoos systems.The purpose of this study was to examine the levels of CACs in the cerebrospinal fluid ((SF) of individuals with dementia of the Alrheimer type (DAT) to determine whether levels would be higher than individuals who did not have DAT. Eighteen patients referred to a memory disorder clinic were used in this pilot study. These individosb were assessedusing a standardized clinical protocol and classified as probable DAT or non DAT using NINCDS
criteria for dementia. There were no significant differences
in age between the two groups (DAT mean age = 69.5 years, non DAT mean age=66.3 years). The non DAT group had a slight preponderance of females (DAT; 8 males: 1 female, non DAT; 5 males: 4 females). Lumbar punctures were carried out on all individuals and GAG concentrations were determined. Tritiated heparin was added to the (SF in a known quantity to accurately asses the percent GAG recovery. The GAG were precipitated out of the (SF and quantity determined by a calorimetric assayin conjunction with radioactive counts to determine yield. The mean concentration of GAG (expressed as micrograms uranic acid per millilitre) the probable DAT group was 9.97 f 1.73 (~9). (~9).
for
and for the non DAT group was 1286 f2.77
The yield for the probable DAT group was 68.2%* 9.2 and for the non DAT group was
55.8% f 7.2 These differences are statistically significant (p>984b). These results are somewhat different from those expected but show statistically significant differences between groups. The probable DAT group showed lower GAG concentrations and greater yields than the non DAT group. This perhaps suggeststhat the CACs are in the DAT sequestered
brains.
being
No explanation for the differences in yield are apparent at
present. This study is preliminary and needs to he confirmed with larger numbers. R.K. supported by Medical Research Council of Canada grant MA-10477
120 SEARCH FOR PROTEING;yANGES IN ALZHEIMER CSF BY WITH ELECTROPHORESIS TWO-DIMENSIONAL IMMOBILIZED pH GRADIENTS IN THE FIRST DIMENSION Kari Mattila, Tuula Pirttila, Anders Wallin* and Harry Fre University of Tampere, De artment of Clinical Sciences, I!O. BOX 607, SF-33101, Tampere, Finlan B University;, Department of Ps~~;~den and Gothenburg Neurochemistry, Jorgen s ospital, S-42203 Hisings Bat Alzheimer’s disease (AD) is the most common cause of dementia. No definitive antemortem marker has been established in the cerebrospinal fluid (CSF), serum or tissues of patients afflicted by this neurode enerative disorder the primary defect and pathogenesis of which sti9 1 remain obscure.
pH ranges,
among
So far, we have not been able to detect any changes in proteins D maps unique to AD.
them
in the 2-