15 cognitively normal (CN), 15 amnestic mild cognitive impairment (MCI), 15 AD UPLC – TOF-MS – based non-targeted metabolomics approach Patients were enrolled and followed in the Mayo Clinic population-based epidemiologic study of normal aging, MCSA, and Mayo Clinic Alzheimer’s Disease Research Center (ADRC).
Acetylcholine, noradrenalin, dopamine and serotonin neurotransmitter systems are primarily affected in AD with subsequent loss of associated neurons ( 42 ). Consistent with that, we have found prominent, early changes in tryptophan biosynthesis in both CSF and plasma of MCI and AD patients (Fig. 3-5). Tryptophan is a precursor for serotonin, melatonin, and niacin synthesis ( 43-45 ). We also found alterations in the serotonin/melatonin pathway in CSF of both MCI and AD patients, and in plasma of MCI individuals (Fig. 3,4). These data are in agreement with recent studies indicating that loss of serotonergic neurons correlates with AD severity, memory impairment, and neuropsychiatric symptoms
Disorder in the hypothalamic-pituitary-adrenal (HPA) axis with increased cortisol levels in CSF and plasma is also well established for AD patients; and increased cortisol levels in CSF from AD patients have been recently demonstrated using metabolomic profiling . Our data confirmed that the pathway related to the cortisol biosynthesis from cholesterol was significantly affected in both CSF and plasma from AD patients (Fig. 5). However, we also found that cortisone biosynthesis and metabolism was among the pathways that, along with PGE2, most accurately separated the clinical groups in CSF (Fig. 6). Among pathways that were uniquely affected in plasma of AD patients were those related to obesity and type II diabetes mellitus (Fig. 5). This is an important observation taking in consideration the data demonstrating that type II diabetes mellitus is associated with an increased risk of cognitive dysfunction and dementia, and needs to be explored in future studies  Moreover, our study identified CSF PGE2 biosynthesis and metabolism as one of the key pathways that varied with AD severity (Fig. 7). Implication of PGE2 in neural injury in AD is well documented, and includes modulation of protein-lipid interactions, trans-membrane and trans-synaptic signaling . It was shown that levels of PGE2 measured in the CSF of control, MCI and AD patients enrolled in the longitudinal study inversely correlate with AD severity: PGE2 was higher in patients with mild memory impairment, but lower in those with more advanced AD .
3077463-MetabolomicsMetabolomics• The study of low molecular weightThe study of low molecular weightmolecules (molecules (<1500 Da) or metabolitesor metabolitesfound within cells and biological systemsfound within cells and biological systemson global level (metabolome)on global level (metabolome)• Measures changes downstream ofMeasures changes downstream ofgenomic, transcriptomic and proteomicgenomic, transcriptomic and proteomicalterations and, therefore, is consideredalterations and, therefore, is consideredmore representative of the functionalmore representative of the functionalstate of a cellstate of a cell• Can measure hundreds to thousandsCan measure hundreds to thousandsof unique chemical entities providingof unique chemical entities providingan overall understanding of metabolisman overall understanding of metabolism• Metabolites are conserved acrossMetabolites are conserved acrossvarious animal species, facilitating thevarious animal species, facilitating theextrapolation of research findings inextrapolation of research findings inlaboratory animals to humanslaboratory animals to humans• Is an integral part of system biologyIs an integral part of system biology
3077463-1. Sample collection2. Metabolite extraction3. Metabolite separation4. Metabolite identification5. Data analysis and canonical pathway enrichment analysis (MPP, SIMCA-P, MetacoreTM)
3077463-Identification of Altered Metabolic Pathways in PlasmaIdentification of Altered Metabolic Pathways in Plasmaand CSF in Mild Cognitive Impairment and Alzheimer’sand CSF in Mild Cognitive Impairment and Alzheimer’sDisease Using MetabolomicsDisease Using MetabolomicsE. Trushina, T. Dutta, X-M. T. Persson, M. M. Mielke, R. C. PetersenE. Trushina, T. Dutta, X-M. T. Persson, M. M. Mielke, R. C. PetersenPLoS ONE 8(5): e63644.PLoS ONE 8(5): e63644.Mayo Clinic Study of Aging and ADRCCSF andplasma
3077463-PLASMAPLASMA CSFCSFMCIvsCNMCIvsCNADvsCNADvsCNADvsMCIADvsMCIplasmaplasmaCSFCSFMCIAD CNMCIAD CNorthogonal two partial least squares-discriminant analysis (O2PLS-DA)Unsupervised Principal Component Analysis (PCA)
3077463-Plasma CSFLysineAndrostenedione and testosteroneTCA cycleSaturated fatty acidMitochondrial ketone bodiesEstroneProstaglandin 2Aminoacyl-tRNA BS in cytoplasmTryptophanLeucine, isoleucine and valineNeurophysiological process_Melatonin signalingPyruvateSerotonin-melatoninGABACholesterolPhospholipid p. 1ButanoatePlasmalogenPropionatePyruvate/rodent versionPhenylalaninePolyamine(L)-ArginineTCA cycleNicotine MB in liverAldosteroneSeratonin-melatoninCortisoneProstaglandin 2Methionine-cysteine-glutamateAspartate and asparagineVitamin B6Histidine-glutamate-glutamineArginine/rodent versionTryptophanUrea cycleAscorbateVitamin B7 (biotin)CholesterolSulfurAlanine, (L)-cysteine, (L)-methioninePyruvate/rodent version-log(p-value) -log(p-value)Role of Diethylhexyl Phthalate andTributyltin in fat differentiation6/853/413/513/692/272/353/943/973/1012/422/432/492/512/532/542/612/632/642/662/662/672/682/767/517/565/415/515/566/944/435/733/305/955/975/1014/702/183/474/883/523/562/293/66Metabolic changes in MCI vs. CNMetabolic changes in MCI vs. CN
3077463-Plasma CSF-log(p-value) -log(p-value)Cholesterol and sphingolipids transportVitamin D2PolyamineIntracellular cholesterol transport(L)-ArginineBeta-alanineAspartate and aspargineCortisoneGalactoseLipid metabolismFXR-regulated cholesterol and bile acid transportGlycolysis and gluconeogenesisCholesterolBile acidRegulation of CFTR gatingRole of VDR in regulation of genes involved in osteoporosisVitamin D3 metabolic C-23 and C-24 pathwaysTriacylglycerol BS in obesity and diabetes mellitus, type IIMuscle contraction_nNOS signaling in skeletal musclesNiacin-HDLAminoacyl-tRNA biosynthesis in mitochondriaTriacylglycerol metabolism p.2Urea cycleLysineTCA cycleProstaglandin 2Aminoacyl-tRNA BS in cytoplasmAndrostenedione and testosteroneAlanine, (L)-cysteine, (L)-methionineMechanism of action of DGAT1 in obesity and diabetes mellitus, type IIMethionine-cysteine-glutamateCortisol BS from cholesterolRegulation of lipid MB_FXR-dependent negative-feedback regulation ofbile acid concentrationRiboflavinAcetylcholineFructoseTryptophanDevelopment_Activation of astroglia cell proliferation by ACM3Fatty Acid Omega OxidationMethionineCholesterol and sphingolipids transportIntracellular cholesterol transport(L)-ArginineHistidine-glutamate-glutamineAspartate and aspargineCortisoneAscorbateMitochondrial ketone bodiesNicotine metabolism in liverGlycolysis and gluconeogenesisCholesterol and sphingolipids transport/transport from GolgiBile acidRegulation of CFTR gatingHETE and HPETEFXR-regulated cholesterol and bile acid cellulartransport PyruvatePropionateUrea cycleEstroneTCA cycleProstaglandin 2Saturated fatty acidsSerotonin-melatoninUMP(L)-Alanine, (L)-cysteine, (L)-methionineCortisol BS from cholesterolGABAGlycine, serine, cysteine and threonineBeta-AlanineTryptophan8/207/389/689/709/908/765/357/738/946/566/596/595/418/207/855/467/887/944/334/353/204/41 5/563/24 6/804/476/945/663/28 3/275/745/695/663/313/317/1232/143/333/343/315/425/565/613/244/433/286/976/1013/323/344/5113/948/519/738/709/908/764/185/336/516/565/414/327/944/426/956/1014/534/563/35
3077463-ConclusionsConclusions• Metabolomics offers novel approach to identify alterations in multiplebiochemical networks over the course of AD• It allows identification of both expected and non-expected changes inbiochemical pathways in animal models of AD and in human samples• Metabolic signatures in CSF and plasma correlate with AD severity• Metabolic signatures in plasma accurately reflect changes in CSF:MCI: 30% of the pathways altered in CSF and plasma were the sameAD: 60% of the pathways affected in CSF and plasma were the same• Application of metabolomics in conjunction with other currently availabletests could increase early AD diagnosis• Metabolomics could be conducted in readily available fluids such as bloodmaking it attractive for clinical application
3077463-1Future DirectionsFuture Directions• Studies in larger patient cohorts are needed• Test and sample validation studies need to be included in every project• Acquisition of the data using multiple analytical platforms should increasethe accuracy and reproducibility• Additional research is needed to reveal the role of metabolites linkedto AD pathology in the mechanisms of normal aging