Post mortem brain studies indicate that alterations in serotonergic and catecholaminergic systems might be associated with Alzheimer's disease (AD). The aim of the study was to determine serotonin (5-HT) levels and monoamine oxidase type B (MAO-B) activity in platelets of psychotic and non-psychotic patients with AD, established according to the NINCDS-ADRDA and DSM-IV-TR criteria. Cognitive impairment and psychotic features were evaluated using Mini Mental Status Examination and Neuropsychiatric Inventory. Platelet 5-HT concentration and MAO-B activity were determined spectrofluorimetrically in 116 (51 male, 65 female) healthy subjects and 70 psychotic (10 male, 60 female) and 151 non-psychotic (32 male, 119 female) patients. Psychotic and non-psychotic female and psychotic male patients had significantly lower platelet 5-HT concentration than corresponding sex matched control subjects. Platelet MAO-B activity was significantly increased in both male and female non-psychotic patients compared to the sex matched controls. Non-psychotic female patients had significantly higher platelet MAO-B activity than psychotic female patients. Our data suggest that platelet MAO-B activity, but not platelet 5-HT concentration, could differentiate between psychotic and non-psychotic subtypes of AD

Deželjin, Martina

Folnegović-Šmalc, Vera

Mimica, Ninoslav

Mustapić, Maja

Mück-Šeler, Dorotea

Pivac, Nela

Presečki, Paola

Radonić, Elizabeta

Stipčević, Tamara

English

Post mortem brain studies indicate that alterations in serotonergic and catecholaminergic systems might be associated
with Alzheimer’s disease (AD). The aim of the study was to determine serotonin (5-HT) levels and monoamine
oxidase type B (MAO-B) activity in platelets of psychotic and non-psychotic patients with AD, established according to the
NINCDS-ADRDA and DSM-IV-TR criteria. Cognitive impairment and psychotic features were evaluated using Mini
Mental Status Examination and Neuropsychiatric Inventory. Platelet 5-HT concentration and MAO-B activity were determined
spectrofluorimetrically in 116 (51 male, 65 female) healthy subjects and 70 psychotic (10 male, 60 female) and
151 non-psychotic (32 male, 119 female) patients. Psychotic and non-psychotic female and psychotic male patients had
significantly lower platelet 5-HT concentration than corresponding sex matched control subjects. Platelet MAO-B activity
was significantly increased in both male and female non-psychotic patients compared to the sex matched controls.
Non-psychotic female patients had significantly higher platelet MAO-B activity than psychotic female patients. Our data
suggest that platelet MAO-B activity, but not platelet 5-HT concentration, could differentiate between psychotic and non-
-psychotic subtypes of AD

Ninoslav Mimica

Dorotea Mück-Šeler

Nela Pivac

Maja Mustapić

Martina Deželjin

Tamara Stipčević

Paola Presečki

Elizabeta Radonić

Vera Folnegović-Šmalc

Hrčak - Portal of scientific journals of Croatia

Coll. Antropol. 32 (2008) Suppl. 1: 119–122Original scientific papersPlatelet Serotonin and Monoamine Oxidase inAlzheimer’s Disease with Psychotic FeaturesNinoslav Mimica1,2, Dorotea Mück-[eler3, Nela Pivac3, Maja Mustapi}3, Martina De`eljin3,Tamara Stip~evi}3, Paola Prese~ki4, Elizabeta Radoni}5 and Vera Folnegovi}-[malc1,21 Department of Psychiatry, Psychiatric Hospital »Vrap~e«, Zagreb, Croatia2 School of Medicine, University of Zagreb, Zagreb, Croatia3 Division of Molecular Medicine, »Ru|er Bo{kovi}« Institute, Zagreb, Croatia4 Department of Psychiatry, General Hospital »Pula«, Pula, Croatia5 Croatian Institute for Brain Research, School of Medicine, University of Zagreb, Zagreb, CroatiaA B S T R A C TPost mortem brain studies indicate that alterations in serotonergic and catecholaminergic systems might be associ-ated with Alzheimer’s disease (AD). The aim of the study was to determine serotonin (5-HT) levels and monoamineoxidase type B (MAO-B) activity in platelets of psychotic and non-psychotic patients with AD, established according to theNINCDS-ADRDA and DSM-IV-TR criteria. Cognitive impairment and psychotic features were evaluated using MiniMental Status Examination and Neuropsychiatric Inventory. Platelet 5-HT concentration and MAO-B activity were de-termined spectrofluorimetrically in 116 (51 male, 65 female) healthy subjects and 70 psychotic (10 male, 60 female) and151 non-psychotic (32 male, 119 female) patients. Psychotic and non-psychotic female and psychotic male patients hadsignificantly lower platelet 5-HT concentration than corresponding sex matched control subjects. Platelet MAO-B activ-ity was significantly increased in both male and female non-psychotic patients compared to the sex matched controls.Non-psychotic female patients had significantly higher platelet MAO-B activity than psychotic female patients. Our datasuggest that platelet MAO-B activity, but not platelet 5-HT concentration, could differentiate between psychotic and non--psychotic subtypes of AD.Key words: platelets, serotonin, monoamine oxidase, Alzheimer’s disease, psychotic featuresIntroductionAlzheimer’s disease (AD) is a complex neurodege-nerative disorder with different cognitive and behav-ioural abnormalities1. Various studies have found thatmajority of AD patients suffer from psychotic symptoms,like auditory and/or visual hallucinations and delusions2.Psychotic features upset the patients, complicate thetreatment response, and are frequently associated withthe rapid progress of the disease2.The aetiology of AD is still not clear. Post mortembrain studies in patients with AD3 showed neuropatho-logical and neurochemical alterations in catecholaminer-gic4 and serotonergic5 systems. The decrease in braindopamine6, serotonin (5-hydroxytryptamine, 5-HT)6, andits main metabolite 5-hydroxyindoleacetic acid (5-HIAA)concentrations, and the loss of 5-HT2 receptors7, wasfound in patients with AD. Blood platelets have been pro-posed as an easy obtainable peripheral model for someprocesses in the central serotonergic neurons8–10 and forthe expression of the brain amyloid precursor protein11.The studies on platelet 5-HT uptake12,13, and plateletmonoamine oxidase type B (MAO-B) in AD yielded in-consistent results14. MAO is a flavin-containing oxygenoxidoreductase. Two isoenzymes (MAO-A and MAO-B)differ in localization, substrates and inhibitors15. MAO-Bexists in platelets, astrocytes and 5-HT neurons. Its sub-strates are b-phenylethylamine, benzylamine, dopamine,tyramine, and tryptamine, and its inhibitor is deprenyl15.It is assumed that altered levels of platelet MAO-B areassociated with different psychopathologies and vulnera-bility to psychiatric disorders15,16. An increase in platelet119Received for publication June 1, 2007.MAO-B activity was observed in AD17. Recently we havefound18 that platelet MAO-B activity might be used as aperipheral biomarker for the early and late onset AD.We have previously found that platelet 5-HT mightdifferentiate between psychotic and non-psychotic de-pression19, posttraumatic stress disorder (PTSD)20, bipo-lar affective disorder in a manic phase21. In addition, ahigher platelet MAO-B activity was found in psychoticcompared to non-psychotic subtype of PTSD22. The hy-pothesis of the present study was that platelet biochemi-cal markers would differ in AD patients with or withoutpsychotic features. The aim of the present study was todetermine peripheral biochemical markers (platelet 5-HTconcentration and platelet MAO-B activity) in male andfemale patients with AD, subdivided according to thepresence of psychotic features, and in sex matched heal-thy controls.Subjects and MethodsThe study included 221 (42 male, 179 female) medica-tion-free patients (mean age 60.6±9.9 years) with AD,hospitalized in Psychiatric Hospital Vrap~e, Zagreb. Thediagnosis of the probable AD (NINCDS-ADRDA crite-ria23) was established by two psychiatrists according tothe DSM-IV-TR criteria24. Cognitive impairment was eval-uated using Mini Mental Status Examination (MMSE)25.Mean MMSE scores in AD patients was 18.9±3.2. Accord-ing to the Neuropsychiatric Inventory26, patients weresubdivided into two groups: 70 patients (10 male and 60female) with psychotic features and 151 patients (32male, 119 female) without psychotic features. Controlgroup consisted of sex and age-matched, medication-freehealthy subjects (65 female, 51 male) (mean age 60.3±11.2 years), with no history of psychiatric illness. All sub-jects were nonsmokers. The study was approved by theEthic committee and all participants gave informed con-sent.Blood samples (8 mL) were obtained from a jugularvein, after an overnight fasting, in a plastic syringe with2 ml of acid citrate dextrose anticoagulant. Platelet richplasma (PRP) was obtained after centrifugation of wholeblood, and platelets were sedimented by further centri-fugation of PRP. Platelet 5-HT concentration was deter-mined by the spectrofluorimetric method, as previouslydescribed27, using Varian Cary Eclipse spectrofluorime-ter Platelet MAO-B activity was determined spectro-fluorimetrically using kynuramine as a substrate27,28.Platelet protein levels were measured by the method ofLowry et al.29.The results were expressed as mean±SD. The differ-ences between groups were analyzed using Kruskal-–Wallis one-way analysis of variance (ANOVA) by ranksfollowed by Mann-Whitney rank sum test for pairwisecomparisons. The statistical package used was Statistica 6and SPSS 10.0 analysis.ResultsPlatelet 5-HT concentration was significantly (H=36.5, df=5, p<0.001; Kruskal Wallis ANOVA) differentamong healthy controls and patients with AD, subdi-vided according to the presence of psychotic features(Figure 1). A significant decrease in platelet 5-HT con-centration was observed in psychotic (p=0.002, MannWhitney test) and non-psychotic (p<0.001) female pa-tients and non-psychotic (p=0.002) male patients com-pared to platelet 5-HT concentration in correspondingsex-matched healthy controls. There was no significantdifference in platelet 5-HT concentration between male(p=0.712, Mann Whitney test) or female (p=0.422, MannWhitney test) psychotic or non-psychotic AD patients.N. Mimica et al.: Biochemical Markers in Alzheimer’s Disease, Coll. Antropol. 32 (2008) Suppl. 1: 119–122120Fig. 1. Platelet serotonin (5-HT) in male and female healthy con-trols and patients with Alzheimer’s disease (AD) with or withoutpsychotic features. *p=0.002 vs. healthy male; **p=0.002 vs. heal-thy female; #p<0.001 vs. healthy female (Kruskal Wallis ANOVAon ranks followed by Mann Whitney test)Fig. 2. Platelet monoamine oxidase (MAO) in healthy controlsand patients Alzheimer’s disease (AD) with or without psychoticfeatures. *p=0.017 vs. female psychotic patients; **p<0.006 vs.healthy male; #p<0.001 vs healthy female (Kruskal Wallis ANOVAon ranks followed by Mann Whitney test)Platelet MAO-B activity differed significantly (H=28.8, df=5, p<0.001, Kruskal Wallis ANOVA) in patientswith AD, subdivided into groups with or without psy-chotic features, and in healthy control subjects (Figure2). Platelet MAO-B activity was significantly higher inboth male (p<0.006) and female (p<0.001) non-psycho-tic patients with AD than in sex matched healthy con-trols. Non-psychotic female AD patients had significantly(p<0.017) higher platelet MAO-B activity than psychoticfemale AD patients. There was no significant difference(p=0.73, Mann Whitney test) in platelet MAO activitybetween psychotic and non-psychotic male AD patients.DiscussionThe results of the present study show that patientswith AD have different platelet 5-HT concentration andplatelet MAO-B activity when compared to sex- andage-matched control subjects. To our knowledge this isthe first report showing a decreased platelet 5-HT con-centration in patients with AD. Although platelets repre-sent a limited peripheral model for the central serotoner-gic neurons, it is noteworthy that similar reductions in5-HT and 5-HIAA concentrations have also been foundin the areas of the frontal and temporal cortices of ADpatients5. In addition, our results showed that the alter-ations in platelet 5-HT values in AD patients were not re-lated to the presence of psychotic features. This result isin contrast with the increased platelet 5-HT concentra-tions in psychotic unipolar depressed patients19, bipolarpatients in manic phase21, war veterans PTSD with psy-chotic symptoms20, or schizophrenic patients with pre-dominantly positive symptoms30, when compared to cor-responding control subjects.The discrepancies in platelet 5-HT concentration inpatients with AD might be explained by the various fac-tors, including a decrease in 5-HT synthesis, an increasein 5-HT metabolism and a change in 5-HT transporter. Ithas been suggested31 that aging may be associated withlower activity of the tryptophan hydroxylase, a rate lim-iting enzyme in 5-HT synthesis. Since our study includedage matched patients and control subjects, the differencein platelet 5-HT values between groups is presumablynot related to the difference in age of the subjects.Platelet 5-HT transporter is the most important mem-brane protein, responsible for the active transport of5-HT from plasma into platelets, and consequently forplatelet 5-HT concentrations. However, recent molecularstudy32 did not find an association between a deletion/in-sertion polymorphism within promoter region of the5-HT transporter gene and AD.Although blood platelets contain only MAO type B,which is not entirely specific for the metabolism of the5-HT, we can not exclude the possibility that the decreasein platelet 5-HT concentrations in patients with AD is inpart a consequence of the higher platelet MAO -B activity.In the present study we have found an increase inplatelet MAO-B activity in both male and female patientswith AD compared to control subjects. This is in line withour previous finding of the high platelet MAO-B activityin patients with early onset AD, and with other studiesshowing increased MAO activity in brain14,33 and pla-telets14,17,18,34 of patients with AD. In addition, we havefound that the increase in platelet MAO-B activity wasrestricted only to patients without psychotic features.Several factors such as sex, smoking, age, race, someneurodegenerative disorders, pernicious anaemia, andpsychotropic drugs15 affect platelet MAO-B activity. Tocontrol for these variables, we subdivided patients ac-cording to the sex, they were matched for race, age, andmedication, and none of the patients were smokers.Higher platelet MAO-B activity reported in our non-psy-chotic AD patients might be connected with some hiddenpsychopatologies15, personality traits35,36, such as neuro-ticism37 or high anxiety36. The increased platelet MAO-Bactivity in non-psychotic AD patients does not agree withthe recent finding of an elevated activity of plateletMAO-B in war veterans PTSD with psychotic symp-toms22 when compared to corresponding control subjects.This discrepancy might be due to the differences in diag-noses (AD vs. PTSD), age15 or smoking20. In addition, al-tered platelet MAO-B activity may be related to differ-ence in yet unknown transcriptional factor(s)38, ratherthan genotypic variation22, that act on transcriptionalregulation38 of the amount of enzyme and/or in the ki-netic regulation of the molecular activity of MAO-B inplatelets39.Our results of the increased MAO-B activity in AD pa-tients support the presumption that neurotoxic and reac-tive metabolites of catecholamine neurotransmitterscould be involved in the aetiology and progress of AD. Re-cently, neurotoxicity of MAO-B metabolites of noradre-nalin, adrenaline or dopamine has been shown in vitroand in vivo40. The serious consequence of the alteredMAO activity is the increase in catecholamine metabo-lites like 3, 4-dihydroxyphenylglycholaldehyde (DOPE-GAL) and 3,4-dihydroxyphenylacetaldehyde (DOPAL),which are highly reactive and toxic to neuronal cells invitro and in vivo40.In conclusion, we have found altered biochemical pa-rameters in platelets of patients with AD. Our results ofdecreased platelet 5-HT concentrations and increasedplatelet MAO-B activity suggest that both serotonergicand catecholaminergic system could be involved in thepathophysiology and progress of AD. In addition, plateletMAO-B activity, but not 5-HT concentration, might beused as a peripheral biological marker that can distin-guish between psychotic and non-psychotic subtypes ofAD.N. Mimica et al.: Biochemical Markers in Alzheimer’s Disease, Coll. Antropol. 32 (2008) Suppl. 1: 119–122121R E F E R E N C E S1. THE RONALD AND NANCY REAGAN RESEARCH INSTITUTEOF THE ALZHEIMER’S ASSOCIATION AND THE NATIONAL INSTI-TUTE ON AGING WORKING GROUP, Neurobiol Aging, 19 (1998) 109.— 2. SCHNEIDER LS, DAGERMAN KS, J Psychiatr Res, 38 (2004) 105.— 3. LANARI A, AMENTA F, SILVESTRELLI G, TOMASSONI D, PAR-NETTI L, Mech Ageing Dev, 127 (2006) 158. — 4. HERRMANN N, LAN-CTOT KL, KHAN LR, J Neuropsychiatry Clin Neurosci, 16 (2004) 261.— 5. GARZIA-ALLOZA M, GIL-BEA FJ, DIEZ-ARIZA M, CHEN CPLH,FRANCIS PT, LASHERAS B, RAMIREZ MJ, Neuropsychologia, 43(2005) 442. — 6. STORGA D, VRECKO K, BIRKMAYER JGD, REIBNE-GGER G, Neurosci Lett, 203 (1996) 29. — 7. BLIN J, BARON JC, DU-BOIS B, CROUZEL C, FIORELLI M, ATTAR-LEVY D, PILLON B,FOURNIER D, VIDAILHET M, AGID Y, Brain, 116 (1993) 497. — 8. AN-DRES AH, RAO MA, OSTROWITZKI S, ENZIAN W, Life Sci, 52 (1993)313. — 9. FRANKE L, SCHEWE HJ, MÜLLER B, CAMPMAN V, KITZ--ROW W, UEBELHACK R, BERGHÖFER A, MÜLLER-OERLINGHAU-SEN B, Life Sci, 47 (2000) 301. — 10. PLAIN H, BERK M, J Affect Dis-ord, 16 (2001) 229. — 11. CATTABENI F, COLCIAGHI F, DI LUCA M,Prog Neuropsychopharmacol Biol Psychiatry, 28 (2004) 763. — 12. ARO-RA RC, EMERY O, MELTZER HY, Neurology, 41 (1991) 1307. — 13. KO-REN P, DIVER-HARBER A, ADONSKY A, RABINOWITZ M, HERSH-KOWITZ M, J Gerontol, 48 (1993) B93. — 14. ADOLFSSON R, GOTT-FRIES CG, ORELAND L, WIBERG A, WINBLAD B, Life Sci, 27 (1980)1029. — 15. ORELAND L, Neurotoxicology, 25 (2004) 79. — 16. PAAVERM, EENSOO D, PULVER A, HARRO J, Psychopharmacology, 186 (2006)32. — 17. ORELAND L, GOTTFRIES CG, Prog NeuropsychopharmacolBiol Psychiatry, 10 (1986) 533. — 18. MIMICA N, MUCK- SELER D, PI-VAC N, MUSTAPIC M, FOLNEGOVIC-SMALC V, Period Biol, 106 (2004)126. — 19. MUCK-SELER D, JAKOVLJEVIC M, PIVAC N, J Affect Disord,39 (1986) 73. — 20. PIVAC N, KOZARIC-KOVACIC D, MUSTAPIC M,DEZELJIN M, BOROVECKI A, GRUBISIC-ILIC M, MUCK-SELER D, JAffect Disord, 93 (2006) 223. — 21 SAGUD M, MIHALJEVIC-PELES A,PIVAC N, JAKOVLJEVIC M, MUCK-SELER D, J Affect Disord, 97(2007), 247. — 22. PIVAC N, KNEZEVIC J, KOZARIC-KOVACIC D, DE-ZELJIN M, MUSTAPIC M, RAK D, MATIJEVIC T, PAVELIC J, MUCK-SELER D, J Affect Disord, (2007) doi: 10.1016/j.jad 2007.01.017. — 23.McKHANN G, DRACHMANN D, FOLSTEIN M, KATZMAN R, PRICED, STADLAN EM, Neurology, 34 (1984) 939. – 24. AMERICAN PSYCHI-ATRIC ASSOCIATION: Diagnostic and Statistical Manual of Mental Dis-orders, Fourth Edition, Text Revision (American Psychiatric Association,Washington, DC, 2000). — 25. FOLSTEIN MF, FOLSTEIN SE, Mc-HUGH PR, J Psychiatr Res, 12 (1975) 189. — 26. CUMMINGS JL, ME-GA M, GRAY K, ROSENBERG-THOMPSON S, CARUSI DA, GORN-BEIN J, Neurology, 44 (1994) 2308. — 27. MUCK-SELER D, PIVAC N,SAGUD M, JAKOVLJEVIC M, MIHALJEVIC-PELES A, Prog Neuropsy-chopharmacol Biol Psychiatry, 26 (2002) 1235. — 28. KRAJL M, BiochemPharmacol, 14 (1965) 1683. — 29. LOWRY OH, ROSENBROUGH NS,FARRA C, RANDALL RJ, J Biol Chem, 193 (1951) 265. — 30. PIVAC N,MUCK-SELER D, JAKOVLJEVI] M, Neuropsychobiology, 36 (1997) 19. —31. HUSSAIN AM, MITRA AK, Drug Metab Dispos, 28 (2000) 1038. —32. ZILL P, PADBERG F, DE JONGE S, HAMPEL H, BURGER K, STUB-NER S, BOETSCH B, JURGEN MOLLER H, ACKENHEIL M, BONDYB, Neurosci Lett, 284 (2000) 113. — 33. KENNEDY BP, ZIEGLER MG,HANSEN LA, THAL LJ, MASLIAH E, J Neural Transm, 110 (2003) 789.— 34. DANIELCZYK W, STREIFLER M, KONRADI C, RIEDERER P,MOLL G, Acta Psychiatr Scand, 78 (1988) 730. — 35. KIRK KM, WHIT-FIELD JB, PANG D, HEATH AC, MARTIN NG, Am J Med Genet, 105(2001) 700. — 36. IRVING JB, COURSWY RD, BUCHSBAUM MS, MUR-PHY DL, Psychol Med, 19 (1989) 79. — 37. SCHALLING D, ASBERG M,EDMAN G, ORELAND L, Acta Psychiatr Scand, 76 (1987) 172. — 38.EKBLOM J, GARPENSTRAND H, DAMBERG M, CHEN K, SHIH JC,ORELAND L, Neurosci Lett, 258 (1998) 101. — 39. BONGIOANNI P, F.GEMIGNANI, B. BOCCARDI, M. BORGNA B, ROSSI, Ital J Neurol Sci,18 (1997) 151. — 40. BURKE WJ, WEN LIS, CHUNG HD, RUGGIERODA, KRISTAL BS, JOHNSON EM, LAMPE P, KUMAR VB, FRANKO M,WILLIAMS EA, ZAHM DS, Neurotoxicol, 25 (2004) 101.D. Muck-[elerDivision of Molecular Medicine, Institute »Rudjer Bo{kovi}«, Bijeni~ka 54, HR-10000 Zagreb, Croatiae-mail: seler@irb.hrTROMBOCITNI SEROTONIN I MONOAMINOOKSIDAZA B U BOLESNIKAS AZHEIMEROVOM BOLESTI SA PSIHOTI^KIM SIMPTOMIMAS A @ E T A KPost mortem istra`ivanja su pokazala promjene serotoninskog i kateholaminskog sustava u mozgu oboljelih od Alz-heimerove bolesti (AB). Cilj istra`ivanja bio je odrediti koncentraciju trombocitnog serotonina (5-HT) i aktivnost trom-bocitne monoaminooksidaze tipa B (MAO-B) u bolesnika s AB sa ili bez psihoti~nih simptoma i u skupini zdravih osoba.Dijagnoza vjerojatne AB (NINCDS-ADRDA kriterij) postavljena je u skladu s DSM-IV kriterijima. Za procjenu kogni-tivnog o{te}enja kori{tena je Mini Mental Status Examination, a za odre|ivanje psihoti~nih simptoma Neuropsychiat-ric Inventory. U trombocitima 116 zdravih osoba (51mu{karaca, 65 `ena), 70 bolesnika (10 mu{karaca, 60 `ena) sapsihoti~kim simptomima i 151 bolesnika (32 mu{karca, 119 `ena) bez psihoti~kih simptoma odre|ena je koncentracija5-HT i aktivnost MAO-B pomo}u spektrofluorimetrijskih metoda. Koncentracija trombocitnog 5-HT bila je zna~ajnosni`ena u bolesnica sa psihoti~kim i nepsihoti~kim simptomima te u nepsihoti~kih bolesnika u odnosu na koncentracijutrombocitnog 5-HT u odgovaraju}oj skupini zdravih `ena i mu{karaca. Aktivnost trombocitne MAO-B bila je pove}anau mu{kih i `enskih nepsihoti~kih bolesnika s AB u usporedbi s aktivno{}u u zdravoj kontroli odgovaraju}eg spola. Utrombocitima nepsihoti~kih `enskih bolesnica prona|ena je zna~ajno pove}ana aktivnost MAO-B u odnosu na aktiv-nost enzima u trombocitima bolesnica sa psihoti~kim simptomima. Rezultati istra`ivanja upu}uju da bi aktivnost trom-bocitne MAO-B mogla biti biokemijski pokazatelj za razlikovanje psihoti~kog i nepsihoti~kog oblika AB.N. Mimica et al.: Biochemical Markers in Alzheimer’s Disease, Coll. Antropol. 32 (2008) Suppl. 1: 119–122122

Platelet Serotonin and Monoamine Oxidase in Alzheimer’s Disease with Psychotic Features

University of Zagreb Medical School Repository

Coll. Antropol. 32 (2008) Suppl. 1: 119–122Original scientific papersPlatelet Serotonin and Monoamine Oxidase inAlzheimer’s Disease with Psychotic FeaturesNinoslav Mimica1,2, Dorotea Mück-[eler3, Nela Pivac3, Maja Mustapi}3, Martina De`eljin3,Tamara Stip~evi}3, Paola Prese~ki4, Elizabeta Radoni}5 and Vera Folnegovi}-[malc1,21 Department of Psychiatry, Psychiatric Hospital »Vrap~e«, Zagreb, Croatia2 School of Medicine, University of Zagreb, Zagreb, Croatia3 Division of Molecular Medicine, »Ru|er Bo{kovi}« Institute, Zagreb, Croatia4 Department of Psychiatry, General Hospital »Pula«, Pula, Croatia5 Croatian Institute for Brain Research, School of Medicine, University of Zagreb, Zagreb, CroatiaA B S T R A C TPost mortem brain studies indicate that alterations in serotonergic and catecholaminergic systems might be associ-ated with Alzheimer’s disease (AD). The aim of the study was to determine serotonin (5-HT) levels and monoamineoxidase type B (MAO-B) activity in platelets of psychotic and non-psychotic patients with AD, established according to theNINCDS-ADRDA and DSM-IV-TR criteria. Cognitive impairment and psychotic features were evaluated using MiniMental Status Examination and Neuropsychiatric Inventory. Platelet 5-HT concentration and MAO-B activity were de-termined spectrofluorimetrically in 116 (51 male, 65 female) healthy subjects and 70 psychotic (10 male, 60 female) and151 non-psychotic (32 male, 119 female) patients. Psychotic and non-psychotic female and psychotic male patients hadsignificantly lower platelet 5-HT concentration than corresponding sex matched control subjects. Platelet MAO-B activ-ity was significantly increased in both male and female non-psychotic patients compared to the sex matched controls.Non-psychotic female patients had significantly higher platelet MAO-B activity than psychotic female patients. Our datasuggest that platelet MAO-B activity, but not platelet 5-HT concentration, could differentiate between psychotic and non--psychotic subtypes of AD.Key words: platelets, serotonin, monoamine oxidase, Alzheimer’s disease, psychotic featuresIntroductionAlzheimer’s disease (AD) is a complex neurodege-nerative disorder with different cognitive and behav-ioural abnormalities1. Various studies have found thatmajority of AD patients suffer from psychotic symptoms,like auditory and/or visual hallucinations and delusions2.Psychotic features upset the patients, complicate thetreatment response, and are frequently associated withthe rapid progress of the disease2.The aetiology of AD is still not clear. Post mortembrain studies in patients with AD3 showed neuropatho-logical and neurochemical alterations in catecholaminer-gic4 and serotonergic5 systems. The decrease in braindopamine6, serotonin (5-hydroxytryptamine, 5-HT)6, andits main metabolite 5-hydroxyindoleacetic acid (5-HIAA)concentrations, and the loss of 5-HT2 receptors7, wasfound in patients with AD. Blood platelets have been pro-posed as an easy obtainable peripheral model for someprocesses in the central serotonergic neurons8–10 and forthe expression of the brain amyloid precursor protein11.The studies on platelet 5-HT uptake12,13, and plateletmonoamine oxidase type B (MAO-B) in AD yielded in-consistent results14. MAO is a flavin-containing oxygenoxidoreductase. Two isoenzymes (MAO-A and MAO-B)differ in localization, substrates and inhibitors15. MAO-Bexists in platelets, astrocytes and 5-HT neurons. Its sub-strates are b-phenylethylamine, benzylamine, dopamine,tyramine, and tryptamine, and its inhibitor is deprenyl15.It is assumed that altered levels of platelet MAO-B areassociated with different psychopathologies and vulnera-bility to psychiatric disorders15,16. An increase in platelet119Received for publication June 1, 2007.MAO-B activity was observed in AD17. Recently we havefound18 that platelet MAO-B activity might be used as aperipheral biomarker for the early and late onset AD.We have previously found that platelet 5-HT mightdifferentiate between psychotic and non-psychotic de-pression19, posttraumatic stress disorder (PTSD)20, bipo-lar affective disorder in a manic phase21. In addition, ahigher platelet MAO-B activity was found in psychoticcompared to non-psychotic subtype of PTSD22. The hy-pothesis of the present study was that platelet biochemi-cal markers would differ in AD patients with or withoutpsychotic features. The aim of the present study was todetermine peripheral biochemical markers (platelet 5-HTconcentration and platelet MAO-B activity) in male andfemale patients with AD, subdivided according to thepresence of psychotic features, and in sex matched heal-thy controls.Subjects and MethodsThe study included 221 (42 male, 179 female) medica-tion-free patients (mean age 60.6±9.9 years) with AD,hospitalized in Psychiatric Hospital Vrap~e, Zagreb. Thediagnosis of the probable AD (NINCDS-ADRDA crite-ria23) was established by two psychiatrists according tothe DSM-IV-TR criteria24. Cognitive impairment was eval-uated using Mini Mental Status Examination (MMSE)25.Mean MMSE scores in AD patients was 18.9±3.2. Accord-ing to the Neuropsychiatric Inventory26, patients weresubdivided into two groups: 70 patients (10 male and 60female) with psychotic features and 151 patients (32male, 119 female) without psychotic features. Controlgroup consisted of sex and age-matched, medication-freehealthy subjects (65 female, 51 male) (mean age 60.3±11.2 years), with no history of psychiatric illness. All sub-jects were nonsmokers. The study was approved by theEthic committee and all participants gave informed con-sent.Blood samples (8 mL) were obtained from a jugularvein, after an overnight fasting, in a plastic syringe with2 ml of acid citrate dextrose anticoagulant. Platelet richplasma (PRP) was obtained after centrifugation of wholeblood, and platelets were sedimented by further centri-fugation of PRP. Platelet 5-HT concentration was deter-mined by the spectrofluorimetric method, as previouslydescribed27, using Varian Cary Eclipse spectrofluorime-ter Platelet MAO-B activity was determined spectro-fluorimetrically using kynuramine as a substrate27,28.Platelet protein levels were measured by the method ofLowry et al.29.The results were expressed as mean±SD. The differ-ences between groups were analyzed using Kruskal-–Wallis one-way analysis of variance (ANOVA) by ranksfollowed by Mann-Whitney rank sum test for pairwisecomparisons. The statistical package used was Statistica 6and SPSS 10.0 analysis.ResultsPlatelet 5-HT concentration was significantly (H=36.5, df=5, p<0.001; Kruskal Wallis ANOVA) differentamong healthy controls and patients with AD, subdi-vided according to the presence of psychotic features(Figure 1). A significant decrease in platelet 5-HT con-centration was observed in psychotic (p=0.002, MannWhitney test) and non-psychotic (p<0.001) female pa-tients and non-psychotic (p=0.002) male patients com-pared to platelet 5-HT concentration in correspondingsex-matched healthy controls. There was no significantdifference in platelet 5-HT concentration between male(p=0.712, Mann Whitney test) or female (p=0.422, MannWhitney test) psychotic or non-psychotic AD patients.N. Mimica et al.: Biochemical Markers in Alzheimer’s Disease, Coll. Antropol. 32 (2008) Suppl. 1: 119–122120Fig. 1. Platelet serotonin (5-HT) in male and female healthy con-trols and patients with Alzheimer’s disease (AD) with or withoutpsychotic features. *p=0.002 vs. healthy male; **p=0.002 vs. heal-thy female; #p<0.001 vs. healthy female (Kruskal Wallis ANOVAon ranks followed by Mann Whitney test)Fig. 2. Platelet monoamine oxidase (MAO) in healthy controlsand patients Alzheimer’s disease (AD) with or without psychoticfeatures. *p=0.017 vs. female psychotic patients; **p<0.006 vs.healthy male; #p<0.001 vs healthy female (Kruskal Wallis ANOVAon ranks followed by Mann Whitney test)Platelet MAO-B activity differed significantly (H=28.8, df=5, p<0.001, Kruskal Wallis ANOVA) in patientswith AD, subdivided into groups with or without psy-chotic features, and in healthy control subjects (Figure2). Platelet MAO-B activity was significantly higher inboth male (p<0.006) and female (p<0.001) non-psycho-tic patients with AD than in sex matched healthy con-trols. Non-psychotic female AD patients had significantly(p<0.017) higher platelet MAO-B activity than psychoticfemale AD patients. There was no significant difference(p=0.73, Mann Whitney test) in platelet MAO activitybetween psychotic and non-psychotic male AD patients.DiscussionThe results of the present study show that patientswith AD have different platelet 5-HT concentration andplatelet MAO-B activity when compared to sex- andage-matched control subjects. To our knowledge this isthe first report showing a decreased platelet 5-HT con-centration in patients with AD. Although platelets repre-sent a limited peripheral model for the central serotoner-gic neurons, it is noteworthy that similar reductions in5-HT and 5-HIAA concentrations have also been foundin the areas of the frontal and temporal cortices of ADpatients5. In addition, our results showed that the alter-ations in platelet 5-HT values in AD patients were not re-lated to the presence of psychotic features. This result isin contrast with the increased platelet 5-HT concentra-tions in psychotic unipolar depressed patients19, bipolarpatients in manic phase21, war veterans PTSD with psy-chotic symptoms20, or schizophrenic patients with pre-dominantly positive symptoms30, when compared to cor-responding control subjects.The discrepancies in platelet 5-HT concentration inpatients with AD might be explained by the various fac-tors, including a decrease in 5-HT synthesis, an increasein 5-HT metabolism and a change in 5-HT transporter. Ithas been suggested31 that aging may be associated withlower activity of the tryptophan hydroxylase, a rate lim-iting enzyme in 5-HT synthesis. Since our study includedage matched patients and control subjects, the differencein platelet 5-HT values between groups is presumablynot related to the difference in age of the subjects.Platelet 5-HT transporter is the most important mem-brane protein, responsible for the active transport of5-HT from plasma into platelets, and consequently forplatelet 5-HT concentrations. However, recent molecularstudy32 did not find an association between a deletion/in-sertion polymorphism within promoter region of the5-HT transporter gene and AD.Although blood platelets contain only MAO type B,which is not entirely specific for the metabolism of the5-HT, we can not exclude the possibility that the decreasein platelet 5-HT concentrations in patients with AD is inpart a consequence of the higher platelet MAO -B activity.In the present study we have found an increase inplatelet MAO-B activity in both male and female patientswith AD compared to control subjects. This is in line withour previous finding of the high platelet MAO-B activityin patients with early onset AD, and with other studiesshowing increased MAO activity in brain14,33 and pla-telets14,17,18,34 of patients with AD. In addition, we havefound that the increase in platelet MAO-B activity wasrestricted only to patients without psychotic features.Several factors such as sex, smoking, age, race, someneurodegenerative disorders, pernicious anaemia, andpsychotropic drugs15 affect platelet MAO-B activity. Tocontrol for these variables, we subdivided patients ac-cording to the sex, they were matched for race, age, andmedication, and none of the patients were smokers.Higher platelet MAO-B activity reported in our non-psy-chotic AD patients might be connected with some hiddenpsychopatologies15, personality traits35,36, such as neuro-ticism37 or high anxiety36. The increased platelet MAO-Bactivity in non-psychotic AD patients does not agree withthe recent finding of an elevated activity of plateletMAO-B in war veterans PTSD with psychotic symp-toms22when compared to corresponding control subjects.This discrepancy might be due to the differences in diag-noses (AD vs. PTSD), age15 or smoking20. In addition, al-tered platelet MAO-B activity may be related to differ-ence in yet unknown transcriptional factor(s)38, ratherthan genotypic variation22, that act on transcriptionalregulation38 of the amount of enzyme and/or in the ki-netic regulation of the molecular activity of MAO-B inplatelets39.Our results of the increased MAO-B activity in AD pa-tients support the presumption that neurotoxic and reac-tive metabolites of catecholamine neurotransmitterscould be involved in the aetiology and progress of AD. Re-cently, neurotoxicity of MAO-B metabolites of noradre-nalin, adrenaline or dopamine has been shown in vitroand in vivo40. The serious consequence of the alteredMAO activity is the increase in catecholamine metabo-lites like 3, 4-dihydroxyphenylglycholaldehyde (DOPE-GAL) and 3,4-dihydroxyphenylacetaldehyde (DOPAL),which are highly reactive and toxic to neuronal cells invitro and in vivo40.In conclusion, we have found altered biochemical pa-rameters in platelets of patients with AD. Our results ofdecreased platelet 5-HT concentrations and increasedplatelet MAO-B activity suggest that both serotonergicand catecholaminergic system could be involved in thepathophysiology and progress of AD. In addition, plateletMAO-B activity, but not 5-HT concentration, might beused as a peripheral biological marker that can distin-guish between psychotic and non-psychotic subtypes ofAD.N. Mimica et al.: Biochemical Markers in Alzheimer’s Disease, Coll. Antropol. 32 (2008) Suppl. 1: 119–122121R E F E R E N C E S1. 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SCHALLING D, ASBERG M,EDMAN G, ORELAND L, Acta Psychiatr Scand, 76 (1987) 172. — 38.EKBLOM J, GARPENSTRAND H, DAMBERG M, CHEN K, SHIH JC,ORELAND L, Neurosci Lett, 258 (1998) 101. — 39. BONGIOANNI P, F.GEMIGNANI, B. BOCCARDI, M. BORGNA B, ROSSI, Ital J Neurol Sci,18 (1997) 151. — 40. BURKE WJ, WEN LIS, CHUNG HD, RUGGIERODA, KRISTAL BS, JOHNSON EM, LAMPE P, KUMAR VB, FRANKO M,WILLIAMS EA, ZAHM DS, Neurotoxicol, 25 (2004) 101.D. Muck-[elerDivision of Molecular Medicine, Institute »Rudjer Bo{kovi}«, Bijeni~ka 54, HR-10000 Zagreb, Croatiae-mail: seler@irb.hrTROMBOCITNI SEROTONIN I MONOAMINOOKSIDAZA B U BOLESNIKAS AZHEIMEROVOM BOLESTI SA PSIHOTI^KIM SIMPTOMIMAS A @ E T A KPost mortem istra`ivanja su pokazala promjene serotoninskog i kateholaminskog sustava u mozgu oboljelih od Alz-heimerove bolesti (AB). Cilj istra`ivanja bio je odrediti koncentraciju trombocitnog serotonina (5-HT) i aktivnost trom-bocitne monoaminooksidaze tipa B (MAO-B) u bolesnika s AB sa ili bez psihoti~nih simptoma i u skupini zdravih osoba.Dijagnoza vjerojatne AB (NINCDS-ADRDA kriterij) postavljena je u skladu s DSM-IV kriterijima. Za procjenu kogni-tivnog o{te}enja kori{tena je Mini Mental Status Examination, a za odre|ivanje psihoti~nih simptoma Neuropsychiat-ric Inventory. U trombocitima 116 zdravih osoba (51mu{karaca, 65 `ena), 70 bolesnika (10 mu{karaca, 60 `ena) sapsihoti~kim simptomima i 151 bolesnika (32 mu{karca, 119 `ena) bez psihoti~kih simptoma odre|ena je koncentracija5-HT i aktivnost MAO-B pomo}u spektrofluorimetrijskih metoda. Koncentracija trombocitnog 5-HT bila je zna~ajnosni`ena u bolesnica sa psihoti~kim i nepsihoti~kim simptomima te u nepsihoti~kih bolesnika u odnosu na koncentracijutrombocitnog 5-HT u odgovaraju}oj skupini zdravih `ena i mu{karaca. Aktivnost trombocitne MAO-B bila je pove}anau mu{kih i `enskih nepsihoti~kih bolesnika s AB u usporedbi s aktivno{}u u zdravoj kontroli odgovaraju}eg spola. Utrombocitima nepsihoti~kih `enskih bolesnica prona|ena je zna~ajno pove}ana aktivnost MAO-B u odnosu na aktiv-nost enzima u trombocitima bolesnica sa psihoti~kim simptomima. Rezultati istra`ivanja upu}uju da bi aktivnost trom-bocitne MAO-B mogla biti biokemijski pokazatelj za razlikovanje psihoti~kog i nepsihoti~kog oblika AB.N. Mimica et al.: Biochemical Markers in Alzheimer’s Disease, Coll. Antropol. 32 (2008) Suppl. 1: 119–122122

Platelet serotonin and monoamine oxidase in Alzheimer's disease with psychotic features

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Platelet serotonin and monoamine oxidase in Alzheimer's disease with psychotic features

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