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714 Alzheimer's Disease: Variables in clinico-pathological correlations
FIFTH INTERNATIONAL CONFERENCE ON ALZHEIMER'S DISEASE
S177
White matter microglia produce Gelatinase A and its activator, membrane-type matrix metalloproteinase-I in human brain tissues Y. Yoshiyama*, T. Yamada and T Hattori Department of Neurology, School of Medicine, Chiba University, l-l-8 Inohana, Chuo-ku, Chiba 260, Japan Gelatinase A (GelA) is, one of the metalloproteinases, an enzyme capable of cleaving beta-amyloid protein, and may function as an alpha-secretase to produce secretary form of amyloid precursor protein. Membrane-type matrix metalloproteinase-I (MT-MMP-t) which has tans-menbrane domain at C terminal activates an inactive precursor of GelA (pro-gelatinase A) in vitro. We examined non-neurological and Alzheimer disease brain tissues for the localization of gelatinase A and MT-MMP-1, and their expression of mRNA, using immunohistochemistry and in situ hybridization. The specificity of the antibodies were evaluated by immunoblot analysis. Double immunostaining was used for determination of the responsible cells. The anti-GelA antibody gave positive stainning of microglial cells in all the brain tissue. Positively stained microglia were found only in the white matter. The mRNA of GelA was also detected in white matter microglia. The two anti-MT-MMP-1 antibodies also gave positive staining in white matter microlial cells in all the brain tissues, and its mRNA was detected only in the same type of cells. Theses results suggest interaction of GelA and MT-MMP-1 in microglial cells in human central nervous system, and may indicate that GelA is produced in white matter microglia through the activation of pro-GelA by MT-MMP-t on the cell membrane.
Department of Neurology, Faculty of Medicine, Kyoto University, 54 Shogoin-kawaharacho, Sakyo-ku, Kyoto 606, and tDepartement of Neurobiology, Kyoto Pharmaceutical University,Kyoto 607, Japan Protein kinase C (PKC) is a phospholipid-dependent protein kinase that plays an important rolein various cellularfunctions. P K C is abundant in the neuronal tissue,and are involved in neuronal survival and the functions of neuronal trophicfactors.P K C is divided into Ca2+-dependent and CaS+-independent PKC. Recent studies revealed that Ca2+-independent P K C have distinctenzymatic properties as for substrate specificityor phospholipid dependency to those of CaZ+-dependent PKC, suggesting that Ca2+-independent P K C plays distinctroles in cellularfunction to Ca2+-dependent PKC. W e have previously shown that PKC-13 levelswere significantly lowered in the temporal cortex ofAlzheimer's disease (AD) patients. In the present study, we analyzed the Ca2+-dependent P K C activityand the Ca2+-independent P K C activityof the brain from A D patients and age-matched controls.W e also examined the protein levels of Ca2+-independent PKC-5, -£ and -4 in A D and controlbrains. In the cytosolicand membranous fractions,Ca2+-dependent and -independent P K C activitieswere significantlylower in the A D brain than the controls.Particularly,the reduction of Ca 2+independent P K C activityin the membranous fractionof the A D brain was most enhanced when cardiolipin,the optimal stimulator of PKC-£, was used in the assay. PKC-£ levelin both the cytosolicand membranous fractions was significantlyreduced in A D brains, although PKC-5 and -~ levelswere not changed compared to controls. In conclusion,Ca2+-dependent and -independent P K C are changed in AD, and among the Ca2+-independent P K C isozymes, the alteration of PKC-£ is a specificevent in A D brain, suggesting its crucial role in A D pathophysiology.
712
714
T h e Role of I m m u n e Disturbances in the Pathogenesis of ALzheimer's Disease L. Groppa*, S. Groppa and V. Lisnic Department o f Neurology, Medical State University N. Testemitseanu, 43-36 Cuza Voda str., Chisinau 2072, Republic o f Moldova Seventy two patients with Alzheimer's Disease (AD) were studied. The diagnosis was established according to N I N C D S A D R D A criteria. A reliable increase o f ceils with supressor-killer phenotype (CD 8+) and corresponding reliable lowering o f the helpersuppressor (CD4+/CD8+) index associated with a reliable increase in the a m o u n t o f serum immunoglobulins A and G were found out. The proliferative ability o f mononuclears was significantly reduced (p<0,01). This fact was manifested not only by a low index o f the blast-transformation reaction (BTR) in response to phytoagglutinin, but also with a low level o f spontaneous cell proliferation (p<0,01). The analysis o f results o f the specific BTR, u s i n g myelin, the c o m m o n protein, in the capacity o f antigen revealed the response to more quickly increase. The study o f B-lymphocytes abilities for policlonal production o f immunoglobulins showed the reliable lowering of the a m o u n t o f IgM and IgG spontaneous production. Also significantly increased index o f IgA and IgG production after antigen stimulation was established. The obtained results indicates the impairment of the proliferative and effector B-lymphocytes abilities. Diminish possibility o f proliferation and immunoglobulins production, sensitivity to the antigens o f the nervous tissue and increasing of the immunoglobulins production point out the disbalance o f regulatory, possible supressor influence upon patients i m m u n e system.
Alzheimer's Disease: Variables in Clinico-Pathological Correlations P. Mohanakrishnan l*, A.H. Fowler~, M.M. Husain2, R.A. Komoroski 2, P. Liem2, PR. lolles 3, and J.P. Vonsattel 4 XLSUMedical Center, New Orleans, LA ; 2University of Arkansas for Medical Sciences, Little Rock, AR; 3Medical College of Virginia, Richmond, VA; and 4Brain Tissue Resource Center, McLean Hospital, Belmont, MA, U.S.A. Metabolic changes in the brains of patients with Alzheimer's disease (AD) can be observed using in vivo M R spectroscopy. Yet, the changes seen cannot be correlated to pathology of the disease, since the histopathologie information is generally not available from living people. The objectives of our studies were to: 1) identify the changes in the brain metabolite levels in AD using high resolution proton (*H) NMR spectroscopy of autopsy brain extracts, and 2) correlate the metabolite levels with semiquantitative assessment of senile plaques (SPs), neurofibrillary tangles (NFTs) and neuronal loss (NL). The posterior temporoparietal cortex (PTPC), the cerebellum and the hippocampus/medial temporal cortex (HMTC) of 13 AD and four nondemented (ND) brains of individuals between the ages of 63 and 95 were stuclied. *H NMR studies of 90% methanol extracts of these brain regions were performed at 300 MHz. AD in the autopsy cases was confirmed by conventional histopathologic examination of the contralateral brain halves. The postmortem intervals (PMIs) varied between 1.4 and 14 hours for AD cases (between 9 and 16.8 hours for ND cases). Age and PMI were not matched between the two groups. The PMIs of a subset (mean age = 77 years) of seven AD brains (PMI > 6 hrs) were matched to those of the ND group (mean age = 68 yrs). For this subset, the concentrations ofNAA, creatine, and GABA of PTPC, and NAA and GABA of both HMTC and cerebellum were significantly lower than (p < 0.05) those for the control group. Significant negative linear correlations were observed between the NFTs and NL, and the NAA-estimates for HMTC." The estimates of creatine, glutamate, GABA, and NAA (a putative neuronal marker) of PTPC also had significant negative correlations with the qualitative assessment of NFTs. Our observation of the NAA and GABA decreases in the cerebella of the AD brains compared to the controls may be due to contralateral cerebral lesions (via crossed cerebellar diaschisis). On the basis of our results we conclude that there is an association between the regional decreases in brain metabolite levels and histopathologic changes and that ~HNMR studies of autopsy brains furnish useful information on pathophysiology of AD.
711
713 C h a n g e s in Ca2÷-dependent a n d - i n d e p e n d e n t P r o t e i n K i n a s e C in A l z h e i m e r ' s D i s e a s e H. Matsushima*, S. Shimohama, M. Chachin?, T. Taniguchit, J. Kimura
715 lmmunocytochemical Localization of Cathepsin D, a Bovine Protease, in Alzheimer Disease, Down Syndrome, and Aging Brains. G. Y. Wen*, M. N. Malik, K. S. Kim and H. M. Wisniewski.
S177
White matter microglia produce Gelatinase A and its activator, membrane-type matrix metalloproteinase-I in human brain tissues Y. Yoshiyama*, T. Yamada and T Hattori Department of Neurology, School of Medicine, Chiba University, l-l-8 Inohana, Chuo-ku, Chiba 260, Japan Gelatinase A (GelA) is, one of the metalloproteinases, an enzyme capable of cleaving beta-amyloid protein, and may function as an alpha-secretase to produce secretary form of amyloid precursor protein. Membrane-type matrix metalloproteinase-I (MT-MMP-t) which has tans-menbrane domain at C terminal activates an inactive precursor of GelA (pro-gelatinase A) in vitro. We examined non-neurological and Alzheimer disease brain tissues for the localization of gelatinase A and MT-MMP-1, and their expression of mRNA, using immunohistochemistry and in situ hybridization. The specificity of the antibodies were evaluated by immunoblot analysis. Double immunostaining was used for determination of the responsible cells. The anti-GelA antibody gave positive stainning of microglial cells in all the brain tissue. Positively stained microglia were found only in the white matter. The mRNA of GelA was also detected in white matter microglia. The two anti-MT-MMP-1 antibodies also gave positive staining in white matter microlial cells in all the brain tissues, and its mRNA was detected only in the same type of cells. Theses results suggest interaction of GelA and MT-MMP-1 in microglial cells in human central nervous system, and may indicate that GelA is produced in white matter microglia through the activation of pro-GelA by MT-MMP-t on the cell membrane.
Department of Neurology, Faculty of Medicine, Kyoto University, 54 Shogoin-kawaharacho, Sakyo-ku, Kyoto 606, and tDepartement of Neurobiology, Kyoto Pharmaceutical University,Kyoto 607, Japan Protein kinase C (PKC) is a phospholipid-dependent protein kinase that plays an important rolein various cellularfunctions. P K C is abundant in the neuronal tissue,and are involved in neuronal survival and the functions of neuronal trophicfactors.P K C is divided into Ca2+-dependent and CaS+-independent PKC. Recent studies revealed that Ca2+-independent P K C have distinctenzymatic properties as for substrate specificityor phospholipid dependency to those of CaZ+-dependent PKC, suggesting that Ca2+-independent P K C plays distinctroles in cellularfunction to Ca2+-dependent PKC. W e have previously shown that PKC-13 levelswere significantly lowered in the temporal cortex ofAlzheimer's disease (AD) patients. In the present study, we analyzed the Ca2+-dependent P K C activityand the Ca2+-independent P K C activityof the brain from A D patients and age-matched controls.W e also examined the protein levels of Ca2+-independent PKC-5, -£ and -4 in A D and controlbrains. In the cytosolicand membranous fractions,Ca2+-dependent and -independent P K C activitieswere significantlylower in the A D brain than the controls.Particularly,the reduction of Ca 2+independent P K C activityin the membranous fractionof the A D brain was most enhanced when cardiolipin,the optimal stimulator of PKC-£, was used in the assay. PKC-£ levelin both the cytosolicand membranous fractions was significantlyreduced in A D brains, although PKC-5 and -~ levelswere not changed compared to controls. In conclusion,Ca2+-dependent and -independent P K C are changed in AD, and among the Ca2+-independent P K C isozymes, the alteration of PKC-£ is a specificevent in A D brain, suggesting its crucial role in A D pathophysiology.
712
714
T h e Role of I m m u n e Disturbances in the Pathogenesis of ALzheimer's Disease L. Groppa*, S. Groppa and V. Lisnic Department o f Neurology, Medical State University N. Testemitseanu, 43-36 Cuza Voda str., Chisinau 2072, Republic o f Moldova Seventy two patients with Alzheimer's Disease (AD) were studied. The diagnosis was established according to N I N C D S A D R D A criteria. A reliable increase o f ceils with supressor-killer phenotype (CD 8+) and corresponding reliable lowering o f the helpersuppressor (CD4+/CD8+) index associated with a reliable increase in the a m o u n t o f serum immunoglobulins A and G were found out. The proliferative ability o f mononuclears was significantly reduced (p<0,01). This fact was manifested not only by a low index o f the blast-transformation reaction (BTR) in response to phytoagglutinin, but also with a low level o f spontaneous cell proliferation (p<0,01). The analysis o f results o f the specific BTR, u s i n g myelin, the c o m m o n protein, in the capacity o f antigen revealed the response to more quickly increase. The study o f B-lymphocytes abilities for policlonal production o f immunoglobulins showed the reliable lowering of the a m o u n t o f IgM and IgG spontaneous production. Also significantly increased index o f IgA and IgG production after antigen stimulation was established. The obtained results indicates the impairment of the proliferative and effector B-lymphocytes abilities. Diminish possibility o f proliferation and immunoglobulins production, sensitivity to the antigens o f the nervous tissue and increasing of the immunoglobulins production point out the disbalance o f regulatory, possible supressor influence upon patients i m m u n e system.
Alzheimer's Disease: Variables in Clinico-Pathological Correlations P. Mohanakrishnan l*, A.H. Fowler~, M.M. Husain2, R.A. Komoroski 2, P. Liem2, PR. lolles 3, and J.P. Vonsattel 4 XLSUMedical Center, New Orleans, LA ; 2University of Arkansas for Medical Sciences, Little Rock, AR; 3Medical College of Virginia, Richmond, VA; and 4Brain Tissue Resource Center, McLean Hospital, Belmont, MA, U.S.A. Metabolic changes in the brains of patients with Alzheimer's disease (AD) can be observed using in vivo M R spectroscopy. Yet, the changes seen cannot be correlated to pathology of the disease, since the histopathologie information is generally not available from living people. The objectives of our studies were to: 1) identify the changes in the brain metabolite levels in AD using high resolution proton (*H) NMR spectroscopy of autopsy brain extracts, and 2) correlate the metabolite levels with semiquantitative assessment of senile plaques (SPs), neurofibrillary tangles (NFTs) and neuronal loss (NL). The posterior temporoparietal cortex (PTPC), the cerebellum and the hippocampus/medial temporal cortex (HMTC) of 13 AD and four nondemented (ND) brains of individuals between the ages of 63 and 95 were stuclied. *H NMR studies of 90% methanol extracts of these brain regions were performed at 300 MHz. AD in the autopsy cases was confirmed by conventional histopathologic examination of the contralateral brain halves. The postmortem intervals (PMIs) varied between 1.4 and 14 hours for AD cases (between 9 and 16.8 hours for ND cases). Age and PMI were not matched between the two groups. The PMIs of a subset (mean age = 77 years) of seven AD brains (PMI > 6 hrs) were matched to those of the ND group (mean age = 68 yrs). For this subset, the concentrations ofNAA, creatine, and GABA of PTPC, and NAA and GABA of both HMTC and cerebellum were significantly lower than (p < 0.05) those for the control group. Significant negative linear correlations were observed between the NFTs and NL, and the NAA-estimates for HMTC." The estimates of creatine, glutamate, GABA, and NAA (a putative neuronal marker) of PTPC also had significant negative correlations with the qualitative assessment of NFTs. Our observation of the NAA and GABA decreases in the cerebella of the AD brains compared to the controls may be due to contralateral cerebral lesions (via crossed cerebellar diaschisis). On the basis of our results we conclude that there is an association between the regional decreases in brain metabolite levels and histopathologic changes and that ~HNMR studies of autopsy brains furnish useful information on pathophysiology of AD.
711
713 C h a n g e s in Ca2÷-dependent a n d - i n d e p e n d e n t P r o t e i n K i n a s e C in A l z h e i m e r ' s D i s e a s e H. Matsushima*, S. Shimohama, M. Chachin?, T. Taniguchit, J. Kimura
715 lmmunocytochemical Localization of Cathepsin D, a Bovine Protease, in Alzheimer Disease, Down Syndrome, and Aging Brains. G. Y. Wen*, M. N. Malik, K. S. Kim and H. M. Wisniewski.