Lefu la Alzheimer's (AD) ha le na li-protein biomarkers tse bonts'ang pathophysiology ea lona e mengata, e sitisang tsoelopele ea tlhahlobo le kalafo. Mona, re sebelisa li-proteomics tse felletseng ho khetholla li-biomarker tsa cerebrospinal fluid (CSF) tse emelang mefuta e mengata ea AD pathophysiology. Multiplex mass spectrometry e supile liprotheine tse ka bang 3,500 le tse ka bang 12,000 ho AD CSF le boko, ka ho latellana. Tlhahlobo ea marang-rang ea proteome ea boko e ile ea rarolla li-module tse 44 tsa mefuta-futa ea lihloliloeng, tse 15 tsa tsona li ne li kopana le proteome ea cerebrospinal fluid. Matšoao a CSF AD ka har'a li-module tsena tse fetang a phuthoa ka lihlopha tse hlano tsa protheine, tse emelang mekhoa e fapaneng ea pathophysiological. Li-synapses le metabolites bokong ba AD lia fokotseha, empa CSF ea eketseha, ha li-myelination tse ruileng tsa glial le lihlopha tsa ho itšireletsa mafung bokong le CSF li eketseha. Ho tsitsa le ho khetheha ha mafu a liphetoho tsa phanele ho netefalitsoe ho lisampole tse ling tse fetang 500 tsa CSF. Lihlopha tsena li boetse li khethile lihlopha tse nyenyane tsa baeloji ho AD e sa tšoauoeng ke letho. Ka kakaretso, liphetho tsena ke mohato o ts'episang ho lithulusi tsa biomarker tse thehiloeng marang-rang bakeng sa lits'ebetso tsa bongaka ho AD.
Lefu la Alzheimer (AD) ke sesosa se atileng haholo sa 'dementia' ea methapo lefatšeng ka bophara 'me se tšoauoa ka mefuta e mengata e fapaneng ea ho se sebetse hantle ha tsamaiso ea baeloji, ho kenyeletsoa phetiso ea synaptic, immune-mediated immune system, le metabolism ea mitochondrial (1-3). Leha ho le joalo, li-biomarker tsa eona tse thehiloeng tsa protheine li ntse li tsepamisitse maikutlo ho boneng protheine ea amyloid le tau, ka hona ha e khone ho bonahatsa pathophysiology ena e fapaneng. Li-biomarker tsena tsa "core" tsa protheine tse lekantsoeng ka ho fetisisa ka cerebrospinal fluid (CSF) li kenyelletsa (i) amyloid beta peptide 1-42 (Aβ1-42), e bontšang ho thehoa ha cortical amyloid plaques; (ii) tau e felletseng, sesupo sa ho senyeha ha axon; (iii) phospho-tau (p-tau), moemeli oa pathological tau hyperphosphorylation (4-7). Le hoja li-biomarker tsena tsa cerebrospinal fluid li nolofalitse haholo ho lemoha ha rona mafu a "protein" ea AD (4-7), a emela feela karolo e nyenyane ea baeloji e rarahaneng ka mor'a lefu lena.
Ho haella ha mefuta e sa tšoaneng ea pathophysiological ea li-biomarker tsa AD ho lebisitse ho mathata a mangata, ho kenyelletsa (i) ho se khone ho tseba le ho lekanya liphatsa tsa lefutso tsa bakuli ba AD, (ii) tekanyo e sa lekaneng ea boima ba lefu le tsoelo-pele, haholo-holo sethaleng sa preclinical, Le ( iii) nts'etsopele ea meriana ea phekolo e ileng ea hlōleha ho rarolla ka ho feletseng likarolo tsohle tsa ho senyeha ha methapo ea kutlo. Ho itšetleha ha rona ho pathology ea bohlokoa ho hlalosa AD ho tsoa mafung a amanang le eona ho mpefatsa mathata ana. Bopaki bo ntseng bo eketseha bo bontša hore boholo ba batho ba tsofetseng ba nang le 'dementia' ba na le litšobotsi tse fetang tse le 'ngoe tsa ho fokotseha ha kelello (8). Hoo e ka bang 90% kapa ho feta ea batho ba nang le lefu la AD le bona ba na le lefu la vascular, TDP-43 inclusions, kapa mafu a mang a senyehang (9). Likarolo tsena tse phahameng tsa ho kopana ha mafu li sentse moralo oa rona oa hajoale oa tlhahlobo ea 'dementia', 'me ho hlokahala tlhaloso e felletseng ea lefu lena.
Ka lebaka la tlhoko e potlakileng ea mefuta e fapaneng ea li-biomarker tsa AD, tšimo e ntse e tsoela pele ho amohela mokhoa oa "omics" o ipapisitseng le sistimi e akaretsang ea ho sibolla li-biomarker. The Accelerated Pharmaceutical Partnership (AMP)-AD Alliance e thakhotsoe ka 2014 'me e ka pele-pele lenaneong. Boiteko bona ba mekhatlo e mengata e entsoeng ke Mekhatlo ea Sechaba ea Bophelo, thuto, le indasteri e ikemiselitse ho sebelisa mekhoa e thehiloeng tsamaisong ho hlalosa hantle pathophysiology ea AD le ho hlahisa tlhahlobo ea tlhahlobo ea mefuta-futa ea mefuta-futa le mekhoa ea phekolo (10). E le karolo ea morero ona, marang-rang a proteomics e se e le sesebelisoa se tšepisang bakeng sa tsoelo-pele ea li-biomarker tse thehiloeng tsamaisong ea AD. Mokhoa ona o se nang leeme oa data o hlophisa lihlopha tse rarahaneng tsa proteomics ka lihlopha kapa "module" ea liprotheine tse kopanetsoeng tse amanang le mefuta e itseng ea lisele, li-organelle le mesebetsi ea likokoana-hloko (11-13). Hoo e ka bang lithuto tsa 12 tse ruileng tsa marang-rang tsa proteomics li entsoe bokong ba AD (13-23). Ka kakaretso, litlhahlobo tsena li bonts'a hore proteome ea marang-rang ea boko ea AD e boloka mokhatlo o bolokehileng haholo oa lihlopha tse ikemetseng le libaka tse ngata tsa cortical. Ho phaella moo, tse ling tsa li-module tsena li bontša liphetoho tse ka ikatisang ka bongata bo amanang le AD ho pholletsa le lihlopha tsa data, tse bontšang pathophysiology ea maloetse a mangata. Ka kakaretso, liphuputso tsena li bonts'a ntlha e ts'episang ea ho sibolloa ha proteome ea marang-rang ea boko e le biomarker e thehiloeng tsamaisong ea AD.
E le ho fetola proteome ea marang-rang ea AD bokong hore e be li-biomarker tse thehiloeng ho tsamaiso ea bophelo bo botle, re ile ra kopanya marang-rang a tsoang bokong le tlhahlobo ea proteomic ea AD CSF. Mokhoa ona o kopantsoeng o lebisitse ho khethoeng ha lihlopha tse hlano tse tšepisang tsa li-biomarker tsa CSF tse amanang le mefuta e mengata ea boko, ho akarelletsa le li-synapses, methapo ea mali, myelination, ho ruruha le ho se sebetse ha litsela tsa metabolic. Re atlehile ho netefatsa liphanele tsena tsa biomarker ka litlhahlobo tse ngata tsa ho ikatisa, ho kenyelletsa le lisampole tse fetang 500 tsa CSF tse tsoang mafu a fapaneng a neurodegenerative. Litlhahlobo tsena tsa netefatso li kenyelletsa ho hlahloba lipheo tsa sehlopha ho CSF ea bakuli ba nang le matšoao a AD (AsymAD) kapa ho bonts'a bopaki ba ho bokellana ha amyloid e sa tloaelehang tikolohong e tloaelehileng ea kelello. Litlhahlobo tsena li totobatsa phapang ea bohlokoa ea baeloji ho baahi ba AsymAD le ho supa matšoao a ka khonang ho theola batho maemong a pele a lefu lena. Ka kakaretso, liphetho tsena li emela mohato oa bohlokoa ho nts'etsopele ea lisebelisoa tsa protheine biomarker tse thehiloeng ho mekhoa e mengata e ka atlehang ho rarolla mathata a mangata a kliniki a tobaneng le AD.
Morero o ka sehloohong oa phuputso ena ke ho tsebahatsa li-biomarker tse ncha tsa cerebrospinal fluid tse bonts'ang mafu a fapaneng a thehiloeng bokong a lebisang ho AD. Setšoantšo sa S1 se hlalosa mokhoa oa rona oa ho etsa lipatlisiso, o kenyelletsang (i) tlhahlobo e pharaletseng e tsamaisoang ke liphuputso tsa pele tsa AD CSF le proteome ea boko ba marang-rang ho khetholla li-biomarker tse ngata tse amanang le boko ba CSF, le (ii) phetiso e latelang: Li-biomarker tsena li ka har'a cerebrospinal lihlopha tsa metsi. Patlisiso e shebaneng le ho sibolloa e qalile ka tlhahlobo ea polelo e fapaneng ea CSF ho batho ba 20 ba tloaetseng ho tseba le bakuli ba 20 ba AD Setsing sa Patlisiso sa Mafu a Alzheimer's Emory Goizueta (ADRC). Ho hlahlojoa ha AD ho hlalosoa e le ho senyeha ho hoholo ha kutloisiso ka pel'a Aβ1-42 e tlaase le maemo a phahameng a tau le p-tau ka har'a cerebrospinal fluid [Mean Montreal Cognitive Assessment (MoCA), 13.8 ± 7.0] [ELISA (ELISA) )]] (Lethathamo la S1A). Taolo (e bolelang MoCA, 26.7 ± 2.2) e ne e na le maemo a tloaelehileng a li-biomarker tsa CSF.
CSF ea motho e tšoauoa ka mefuta e mengata ea liprotheine tse ngata, moo albumin le liprotheine tse ling tse ngata haholo li ka thibelang ho fumanoa ha liprotheine tse khahlisang (24). Ho eketsa botebo ba ho sibolloa ha protheine, re tlositse liprotheine tsa pele tse 14 tse ngata haholo sampoleng e 'ngoe le e 'ngoe ea CSF pele ho tlhahlobo ea mass spectrometry (MS) (24). Kakaretso ea li-peptide tse 39,805 li ile tsa khetholloa ke MS, tse ileng tsa etsoa 'mapa oa li-proteome tse 3691 ka lisampole tse 40. Palo ea liprotheine e etsoa ka litlhaku tse ngata tsa tandem mass (TMT) (18, 25). E le ho rarolla lintlha tse sieo, re kenyelelitse feela liprotheine tse neng li lekantsoe bonyane 50% ea lisampole tlhahlobisong e latelang, kahoo qetellong ra lekanya liprotheine tse 2875. Ka lebaka la phapang e kholo maemong a bongata ba protheine e felletseng, sampole ea taolo e ile ea nkuoa e le kantle ho naha (13) mme ha ea ka ea kenyelletsoa tlhahlobong e ileng ea latela. Litekanyetso tse ngata tsa lisampole tse 39 tse setseng li ile tsa fetoloa ho latela lilemo, bong, le batch covariance (13-15, 17, 18, 20, 26).
Ho sebelisa lipalo-palo tsa tlhahlobo ea t-test ho lekola polelo e fapaneng mabapi le sete ea data ea regression, tlhahlobo ena e ile ea supa liprotheine tseo maemo a tsona a mangata a ileng a fetoloa haholo (P <0.05) lipakeng tsa taolo le linyeoe tsa AD (Letlapa la S2A). Joalokaha ho bontšitsoe setšoantšong sa 1A, bongata ba liprotheine tsa 225 ho AD bo fokotsehile haholo, 'me bongata ba liprotheine tsa 303 bo ile ba eketseha haholo. Liprotheine tsena tse hlalositsoeng ka tsela e fapaneng li kenyelletsa matšoao a 'maloa a AD a cerebrospinal fluid, a kang microtubule-associated protein tau (MAPT; P = 3.52 × 10−8), neurofilament (NEFL; P = 6.56 × 10−3), Protein e amanang le kholo ea 43 (GAP43; P = 1.46 × 10-5), Mafura a Tlamang Protheine ea 3 (FABP3; P = 2.00 × 10−5), Chitinase 3 joaloka 1 (CHI3L1; P = 4.44 × 10−6), Neural Granulin (NRGN; P = 3.43 × 10-4) le VGF methapo ea ho hōla ha methapo (VGF; P = 4.83 × 10-3) (4-6). Leha ho le joalo, re ile ra boela ra khetholla lipheo tse ling tsa bohlokoa haholo, tse kang GDP dissociation inhibitor 1 (GDI1; P = 1.54 × 10-10) le SPARC e amanang le modular calcium e tlamang 1 (SMOC1; P = 6.93 × 10-9) . Tlhahlobo ea Gene Ontology (GO) ea liprotheine tse 225 tse fokolitsoeng haholo li senotse likamano tse haufi le lits'ebetso tsa mokelikeli oa 'mele joalo ka steroid metabolism, mali a coagulation, le tšebetso ea lihomone (Setšoantšo sa 1B le Lethathamo la S2B). Ka lehlakoreng le leng, protheine e eketsehang haholo ea 303 e amana haufi-ufi le sebopeho sa lisele le metabolism ea matla.
(A) Sebaka sa seretse se chesang se bontša phetoho ea log2 fold (x-axis) e amanang le -log10 statistical P value (y-axis) e fumanoeng ke teko ea t, e sebelisetsoang ho lemoha phapang pakeng tsa taolo (CT) le Maemo a AD a CSF proteome Of all proteins. Liprotheine tse nang le maemo a fokotsehileng haholo (P <0.05) ho AD li bonts'oa ka boputsoa, ha liprotheine tse nang le maemo a eketsehileng a mafu li bontšoa ka bofubelu. Protheine e khethiloeng e ngotsoe. (B) Mantsoe a holimo a GO a amanang le protheine a fokotsehile haholo (putsoa) mme a eketseha (e khubelu) ho AD. E bonts'a mantsoe a mararo a GO a nang le lintlha tse phahameng ka ho fetisisa tsa z mafapheng a ts'ebetso ea baeloji, mesebetsi ea limolek'hule, le likarolo tsa cellular. (C) MS e ile ea lekanya boemo ba MAPT ho sampuli ea CSF (ka ho le letšehali) le ho hokahana ha eona le sampole ELISA boemo ba tau (ka ho le letona). Coefficient ea khokahano ea Pearson e nang le P value e amehang e ea hlaha. Ka lebaka la khaello ea lintlha tsa ELISA bakeng sa nyeoe e le 'ngoe ea AD, lipalo tsena li kenyelletsa boleng ba linyeoe tse 38 ho tse 39 tse hlahlobiloeng. (D) Tlhahlobo ea lihlopha tse laoloang (P <0.0001, Benjamini-Hochberg (BH) e fetotsoe P <0.01) ho laola 'me AD CSF e fumane mehlala e sebelisang liprotheine tsa 65 tse fetotsoeng ka ho fetisisa ho data. Standardize, normalize.
Boemo ba proteomic ba MAPT bo amana haufi-ufi le tekanyo ea ELISA tau e lekantsoeng e ikemetseng (r = 0.78, P = 7.8 × 10-9; Setšoantšo sa 1C), e tšehetsang bonnete ba tekanyo ea rona ea MS. Kamora tšilo ea trypsin boemong ba protheine ea amyloid precursor (APP), li-peptide tse khethehileng tsa isoform tse entsoeng 'mapeng oa C-terminus ea Aβ1-40 le Aβ1-42 li ke ke tsa etsoa ionized hantle (27, 28). Ka hona, li-peptide tsa APP tseo re li tšoaileng ha li amane le maemo a ELISA Aβ1-42. Bakeng sa ho lekola polelo e fapaneng ea nyeoe e 'ngoe le e' ngoe, re sebelisitse liprotheine tse hlahisitsoeng ka mokhoa o fapaneng le P <0.0001 [false discovery rate (FDR) e lokisitsoeng P <0.01] ho etsa tlhahlobo ea lihlopha tse hlokometsoeng tsa lisampole (Letlapa la S2A). Joalokaha ho bontšitsoe setšoantšong sa 1D, liprotheine tsena tse 65 tsa bohlokoa haholo li ka kopanya lisampole ka nepo ho latela boemo ba lefu, ntle le nyeoe e le 'ngoe ea AD e nang le litšobotsi tse kang taolo. Har'a liprotheine tsena tse 65, tse 63 li eketsehile ka AD, ha tse peli feela (CD74 le ISLR) li fokotsehile. Ka kakaretso, litlhahlobo tsena tsa cerebrospinal fluid li bonts'itse makholo a liprotheine ho AD tse ka sebetsang e le li-biomarker tsa mafu.
Eaba re etsa tlhahlobo e ikemetseng ea marang-rang ea AD boko proteome. Sehlopha sa boko ba ho sibolloa ha sena se ne se kenyelletsa dorsolateral prefrontal cortex (DLPFC) ho tloha taolong (n = 10), lefu la Parkinson (PD; n = 10), linyeoe tsa AD / PD (n = 10) le AD (n = 10) tse tsoakiloeng. ) Mohlala. Emery Goizueta ADRC. Palo ea batho ba linyeoe tsena tse 40 esale e hlalositsoe (25) mme e akaretsoa ho Lethathamo la S1B. Re sebelisitse TMT-MS ho sekaseka liphatsa tsena tse 40 tsa boko le sehlopha sa ho pheta-pheta sa linyeoe tse 27. Ka kakaretso, lisebelisoa tsena tse peli tsa data tsa boko li hlahisitse li-peptide tse ikhethang tsa 227,121, tse neng li entsoe 'mapa ho li-proteoms tse 12,943 (25). Ke feela liprotheine tse neng li lekantsoe bonyane 50% ea linyeoe li kenyelelitsoe liphuputsong tse latelang. Setsi sa ho qetela sa data se sibolotsoeng se na le liprotheine tse 8817 tse lekantsoeng. Fetola maemo a bongata ba protheine ho ipapisitse le lilemo, bong, le nako ea ho shoa ha lefu (PMI). Tlhaloso e fapaneng ea tlhaloso ea lintlha tse behiloeng ka mor'a ho fokotseha e bontšitse hore> maemo a protheine a 2000 a fetotsoe haholo [P <0.05, tlhahlobo ea phapang (ANOVA)] ho lihlopha tse peli kapa ho feta tsa mafu. Ka mor'a moo, re ile ra etsa tlhahlobo ea lihlopha tse laoloang ho latela liprotheine tse hlalositsoeng ka mokhoa o fapaneng, le P <0.0001 ho AD / control le / kapa AD / PD papiso (Figure S2, A le B, Table S2C). Liprotheine tsena tse 165 tse fetotsoeng haholo li bontša ka ho hlaka linyeoe tse nang le lefu la AD ho tloha taolong le disampole tsa PD, tse tiisang liphetoho tse matla tse khethehileng tsa AD ho proteome eohle.
Ka mor'a moo re ile ra sebelisa algorithm e bitsoang Weighted Gene Co-expression Network Analysis (WGCNA) ho etsa tlhahlobo ea marang-rang ho proteome ea boko e sibollotsoeng, e hlophisang lintlha tse behiloeng ka li-module tsa protheine tse nang le mekhoa e tšoanang ea polelo (11-13). Tlhahlobo e ile ea khetholla li-module tsa 44 (M) li-protheine tse kopanetsoeng, tse hlophisitsoeng le ho baloa ho tloha ho tse kholo ka ho fetisisa (M1, n = 1821 liprotheine) ho ea ho tse nyenyane ka ho fetisisa (M44, n = 34 liprotheine) (Setšoantšo sa 2A le Letlapa la S2D)). Joalokaha ho boletsoe ka holimo (13) Bala profil ea polelo ea moemeli kapa protheine ea sebopeho sa mojule ka mong, 'me u e kopanye le boemo ba lefu le lefu la AD, ke hore, theha selekane sa Alzheimer's Disease Registry (CERAD) le Braak Score (Figure 2B). Ka kakaretso, li-module tsa 17 li ne li amana haholo le AD neuropathology (P <0.05). Bongata ba limmojule tsena tse amanang le lefu le tsona li ruile ka matšoao a ikhethileng a mofuta oa lisele (Setšoantšo sa 2B). Joalokaha ho boletsoe ka holimo (13), ntlafatso ea mofuta oa sele e khethoa ka ho hlahloba ho kopana ha mojule le lenane la litšupiso tsa liphatsa tsa lefutso tse khethehileng tsa mofuta oa sele. Liphatsa tsa lefutso tsena li tsoa ho data e phatlalalitsoeng ho li-neuron tsa litoeba tse ikhethileng, lisele tsa endothelial le glial. Teko ea tatellano ea RNA (RNA-seq) (29).
(A) Fumana WGCNA ea proteome ea boko. (B) Biweight midcorrelation (BiCor) tlhahlobo ea protheine ea modular signature (karolo ea pele e kholo ea polelo ea protheine ea modular) e nang le litšobotsi tsa AD neuropathological (holimo), ho kenyelletsa le CERAD (Aβ plaque) le Braak (tau tangles) lintlha. Matla a likamano tse ntle (tse khubelu) le tse mpe (tse putsoa) li bontšoa ke 'mapa oa mocheso oa mebala e' meli, 'me linaleli li bontša bohlokoa ba lipalo (P <0.05). Sebelisa Hypergeometric Fisher's Exact Test (FET) (tlase) ho lekola kamano ea mofuta oa sele ea mojule ka mong oa protheine. Matla a moriti o mofubelu a bontša tekanyo ea ho ntlafatsa mofuta oa lisele, 'me linaleli li bontša bohlokoa ba lipalo (P <0.05). Sebelisa mokhoa oa BH ho lokisa boleng ba P bo nkiloeng ho FET. (C) Tlhahlobo ea GO ea liprotheine tsa modular. Mekhoa e amanang haufi-ufi ea baeloji e bontšoa bakeng sa mojule ka mong kapa sehlopha se amanang le sona. oligo, oligodendrocyte.
Sehlopha sa li-modules tse hlano tse amanang haufi-ufi tsa astrocyte le microglia-rich (M30, M29, M18, M24, le M5) li bontšitse kamano e matla e ntle le AD neuropathology (Setšoantšo sa 2B). Tlhahlobo ea Ontology e hokahanya likarolo tsena tsa glial le kholo ea lisele, ho ata, le ho itšireletsa mafung (Setšoantšo sa 2C le Lethathamo la S2E). Li-module tse ling tse peli tsa glial, M8 le M22, le tsona li laoloa ka matla ho mafu. M8 e amana haholo le tsela ea ho amohela li-Toll-like receptor, e leng pontšo ea cascade e phethang karolo ea bohlokoa karabelong ea tlhaho ea 'mele ea ho itšireletsa mafung (30). Ka nako e ts'oanang, M22 e amana haufi-ufi le phetoho ea morao-rao ea phetolelo. M2, e nang le li-oligodendrocyte tse ngata, e bonts'a kamano e matla e ntle le lefu la AD le kamano ea ontological le nucleoside synthesis le DNA replication, e bontšang ho eketseha ho matla ha lisele ho mafu. Ka kakaretso, liphuputso tsena li tšehetsa ho phahama ha li-module tsa glial tseo re li boneng pele ho AD network proteome (13, 17). Hona joale ho fumanoa hore li-modules tse ngata tse amanang le AD tse amanang le glial marang-rang li bonts'a maemo a tlaase a ho bua ka maemo a taolo le linyeoe tsa PD, ho totobatsa boemo ba bona ba lefu le phahameng ka AD (Figure S2C).
Ke li-module tse 'nè feela tsa proteome ea marang-rang ea rona (M1, M3, M10, le M32) tse amanang ka matla le lefu la AD (P <0.05) (Setšoantšo sa 2, B le C). Ka bobeli M1 le M3 li na le matšoao a mangata a neuronal. M1 e amana haholo le matšoao a synaptic, ha M3 e amana haufi-ufi le mosebetsi oa mitochondrial. Ha ho na bopaki ba ho ntlafatsa mofuta oa lisele bakeng sa M10 le M32. M32 e bonahatsa kamano pakeng tsa M3 le metabolism ea lisele, ha M10 e amana haholo le kholo ea lisele le mosebetsi oa microtubule. Ha ho bapisoa le AD, li-module tse 'nè kaofela li eketsoa taolong le PD, ho ba fa liphetoho tsa AD tse khethehileng (Figure S2C). Ka kakaretso, liphetho tsena li tšehetsa palo e fokotsehileng ea li-module tse ruileng tsa neuron tseo re li boneng pele ho AD (13, 17). Ka kakaretso, tlhahlobo ea marang-rang ea proteome ea boko eo re e fumaneng e hlahisitse li-module tse fetotsoeng ka ho khetheha tsa AD tse tsamaellanang le liphuputso tsa rona tse fetileng.
AD e tšoauoa ka mohato oa pele oa asymptomatic (AsymAD), moo batho ba bonts'ang ho bokellana ha li-amyloid ntle le ho fokotseha ha kelello ea bongaka (5, 31). Mokhahlelo ona oa asymptomatic ke fensetere ea bohlokoa bakeng sa ho lemoha le ho kenella kapele. Re kile ra bonts'a ts'ireletso e matla ea modular ea AsymAD le AD brain network proteome ho pholletsa le li-data tse ikemetseng (13, 17). E le ho etsa bonnete ba hore marang-rang a boko ao re a fumaneng hona joale a lumellana le liphuputso tsena tse fetileng, re ile ra hlahloba ho boloka li-module tsa 44 boitsebisong bo phetoang bo tsoang mekhatlong ea 27 DLPFC. Mekhatlo ena e kenyelletsa taolo (n = 10), AsymAD (n = 8) Le linyeoe tsa AD (n = 9). Taolo le lisampole tsa AD li kenyellelitsoe tlhahlobisong ea sehlopha sa rona sa ho sibolla boko (Letlapa la S1B), athe linyeoe tsa AsymAD li ne li ikhethile feela sehlopheng sa boikaketsi. Linyeoe tsena tsa AsymAD le tsona li tsoa bankeng ea boko ea Emory Goizueta ADRC. Le hoja temoho e ne e tloaelehile nakong ea lefu, maemo a amyloid a ne a phahame ka mokhoa o sa tloaelehang (ho bolela CERAD, 2.8 ± 0.5) (Letlapa la S1B).
Tlhahlobo ea TMT-MS ea lisele tsena tse 27 tsa boko e entse hore ho be le palo ea liprotheine tsa 11,244. Palo ena ea ho qetela e kenyelletsa feela liprotheine tse lekantsoeng bonyane 50% ea lisampole. Setsi sena se phetoang sa data se na le 8638 (98.0%) ea liprotheine tsa 8817 tse fumanoeng tlhahlobong ea rona ea boko, 'me e batla e le 3000 e fetotse liprotheine tse pakeng tsa taolo le AD cohorts (P <0.05, ka mor'a tlhahlobo ea Tukey e kopantsoeng t bakeng sa tlhahlobo ea phapang) ( Tafole S2F). Har'a liprotheine tsena tse hlahisitsoeng ka mokhoa o fapaneng, 910 e boetse e bonts'itse liphetoho tse kholo lipakeng tsa AD le linyeoe tsa taolo ea proteome ea boko (P <0.05, kamora tlhahlobo ea t-teko ea ANOVA Tukey). Ke habohlokoa ho hlokomela hore matšoao ana a 910 a lumellana haholo ka tsela ea phetoho pakeng tsa proteome (r = 0.94, P <1.0 × 10-200) (Setšoantšo sa S3A). Har'a liprotheine tse ntseng li eketseha, liprotheine tse nang le liphetoho tse tsitsitseng ka ho fetisisa pakeng tsa lisebelisoa tsa data ke haholo-holo litho tsa glial-rich M5 le M18 modules (MDK, COL25A1, MAPT, NTN1, SMOC1, le GFAP). Har'a liprotheine tse fokotsehileng, tse nang le liphetoho tse tsitsitseng ka ho fetisisa li ne li batla li le litho tsa M1 module (NPTX2, VGF, le RPH3A) tse amanang le synapse. Re ile ra boela ra netefatsa liphetoho tse amanang le AD tsa midkine (MDK), CD44, protheine e amanang le frizzled 1 (SFRP1) le VGF by western blotting (Figure S3B). Tlhahlobo ea ho boloka mojule e bontšitse hore hoo e ka bang 80% ea li-module tsa protheine (34/44) bokong ba proteome li ne li bolokiloe haholo ho sete ea data ea ho ikatisa (z-score> 1.96, FDR e lokisitsoe P <0.05) (Figure S3C). Li-module tse leshome le metso e mene li ne li bolokiloe ka ho khetheha pakeng tsa li-proteome tse peli (z-score> 10, FDR e lokisitsoe P <1.0 × 10-23). Ka kakaretso, ho sibolloa le ho pheta-pheta ha boemo bo phahameng ba ho lumellana ha polelo e fapaneng le sebopeho sa modular pakeng tsa proteome ea boko e totobatsa ho ikatisa ha liphetoho ho liprotheine tsa AD frontal cortex. Ntle le moo, e boetse e netefalitse hore AsymAD le mafu a tsoetseng pele haholo a na le sebopeho sa marang-rang a boko bo ts'oanang haholo.
Tlhahlobo e qaqileng haholoanyane ea polelo e fapaneng ka har'a data ea ho pheta-pheta boko e totobatsa tekanyo e kholo ea liphetoho tsa protheine ea AsymAD, ho kenyelletsa le kakaretso ea liprotheine tsa 151 tse fetotsoeng haholo pakeng tsa AsymAD le taolo (P <0.05) (Figure S3D). Tumellanong le mojaro oa amyloid, APP bokong ba AsymAD le AD e eketsehile haholo. MAPT e fetoha haholo feela ho AD, e lumellanang le maemo a ntseng a eketseha a li-tangles le kamano ea eona e tsejoang le ho fokotseha ha kelello (5, 7). Li-modules tse nang le glial-rich (M5 le M18) li bonahala haholo ka liprotheine tse ntseng li eketseha ho AsymAD, ha mochine oa M1 o amanang le neuron ke oona o emelang liprotheine tse fokotsehileng ho AsymAD. Bongata ba matšoao ana a AsymAD a bonts'a liphetoho tse kholo ho mafu a matšoao. Har'a matšoao ana ke SMOC1, protheine ea glial ea M18, e amanang le lihlahala tsa boko le kholo ea mahlo le maoto le matsoho (32). MDK ke ntlha ea kholo e tlamang ea heparin e amanang le kholo ea lisele le angiogenesis (33), setho se seng sa M18. Ha ho bapisoa le sehlopha sa taolo, AsymAD e ile ea eketseha haholo, e lateloa ke keketseho e kholo ea AD. Ka lehlakoreng le leng, synaptic protein neuropentraxin 2 (NPTX2) e ile ea fokotseha haholo bokong ba AsymAD. NPTX2 e ne e kile ea amahanngoa le neurodegeneration mme e na le karolo e tsebahalang ho kopanyelletsa li-synapse tse thabisang (34). Ka kakaretso, liphetho tsena li senola mefuta e fapaneng ea liphetoho tse fapaneng tsa protheine ea pele ho AD tse bonahalang li tsoela pele ka ho teba ha lefu lena.
Ka lebaka la hore re fihletse botebo bo boholo ba tšireletso ea protheine ha ho sibolloa proteome ea boko, re leka ho utloisisa ka botlalo ho fetana ha eona le transcriptome ea boemo ba marang-rang ea AD. Ka hona, re ile ra bapisa proteome ea boko eo re e fumaneng le module eo re neng re e hlahisitse pele ho tekanyo ea microarray ea liphatsa tsa lefutso tsa 18,204 AD (n = 308) le taolo (n = 157) DLPFC (13). pharallana. Ka kakaretso, re khethile li-module tse fapaneng tsa RNA tse 20, tse ngata tsa tsona li bonts'itse ho ntlafatsa mefuta e itseng ea lisele, ho kenyelletsa le li-neuron, oligodendrocyte, astrocyte le microglia (Setšoantšo sa 3A). Liphetoho tse ngata tsa li-module tsena ho AD li bontšoa setšoantšong sa 3B. Ho lumellana le tlhahlobo ea rona e fetileng ea protheine-RNA e sebetsanang le ho sebelisa MS proteome e tebileng e sa ngolisoang (e ka bang 3000 protheine) (13), boholo ba li-module tsa 44 bokong ba proteome network eo re e fumaneng e le marang-rang a transcriptome Ha ho na ho kopana ho bohlokoa. ho sibolloa ha rona le ho pheta-pheta ha li-module tsa liprotheine tse 34 tse bolokiloeng haholo bokong ba proteome, ke ba 14 feela (~40%) ba ileng ba feta tekong e nepahetseng ea Fisher (FET) e ipakile e na le kamano ea bohlokoa ea lipalo le transcriptome (Figure 3A). E lumellana le tokiso ea tšenyo ea DNA (P-M25 le P-M19), phetolelo ea protheine (P-M7 le P-M20), RNA binding/splicing (P-M16 le P-M21) le targeting ea protheine (P-M13 le P- M23) ha e kopane le li-module ho transcriptome. Ka hona, le hoja sete ea data e tebileng ea proteome e sebelisoa tlhahlobisong ea hona joale ea ho kopana (13), boholo ba proteome ea marang-rang ea AD ha e na 'mapa oa marang-rang a transcriptome.
(A) Hypergeometric FET e bonts'a ntlafatso ea matšoao a ikhethileng a mofuta oa sele mojuleng oa RNA oa AD transcriptome (holimo) le tekanyo ea ho fetana lipakeng tsa RNA (x-axis) le li-protein (y-axis) tsa boko ba AD. (ka tlase). Matla a moriti o mofubelu a bontša tekanyo ea ho ruisa mefuta ea lisele karolong e ka holimo le matla a ho kopana ha li-module karolong e ka tlaase. Linaleli li bontša bohlokoa ba lipalo (P <0.05). (B) Tekanyo ea khokahano lipakeng tsa liphatsa tsa lefutso tsa mojule ka mong oa transcriptome le boemo ba AD. Li-module tse ka letsohong le letšehali ke tsona tse amanang le AD (tse putsoa), 'me tse ka ho le letona ke tsona tse amanang le AD (e khubelu). Palo e fetotsoeng ea BH-e lokisitsoeng ea P e bonts'a tekanyo ea bohlokoa ba lipalo tsa khokahano ka 'ngoe. (C) Li-module tsa bohlokoa tse tsamaisanang tse nang le mofuta oa lisele tse arolelanoang. (D) Tlhahlobo ea kamano ea phetoho ea log2 fold ea protheine e ngotsoeng (x-axis) le RNA (y-axis) mojulung o fetang. Coefficient ea khokahano ea Pearson e nang le P value e amehang e ea hlaha. likokoana-hloko, microglia; lihloliloeng tsa leholimo, li-astrocyte. CT, taolo.
Bongata ba liprotheine le li-module tsa RNA li arolelana lintlha tse tšoanang tsa mofuta oa lisele le litsela tse sa fetoheng tsa AD (Setšoantšo sa 3, B le C). Ka mantsoe a mang, mojule oa M1 o amanang le synapse oa boko ba proteome (PM1) o entsoe ka 'mapa oa li-module tsa RNA tse tharo tsa neuronal-rich homologous (R-M1, R-M9 le R-M16), tse ho AD Ka bobeli li bonts'itsoe. boemo bo fokotsehileng. Ka mokhoa o ts'oanang, li-protein tsa glial-rich M5 le M18 li kopana le li-module tsa RNA tse ruileng ka li-astrocyte le li-microglial markers (R-M3, R-M7, le R-M10) 'me li ameha haholo mafu Ho eketseha. Likarolo tsena tse arolelanoang tsa modular lipakeng tsa li-data tse peli li ts'ehetsa ntlafatso ea mofuta oa sele le liphetoho tse amanang le mafu tseo re li boneng ho proteome ea boko. Leha ho le joalo, re hlokometse liphapang tse ngata tsa bohlokoa lipakeng tsa RNA le maemo a protheine ea matšoao a motho ka mong ho li-module tsena tse arolelanoang. Tlhahlobo ea kamano ea polelo e fapaneng ea liprotheine le li-transcriptomics tsa limolek'hule tse ka har'a li-module tsena tse fetang (Setšoantšo sa 3D) se totobatsa ho se lumellane hona. Ka mohlala, APP le liprotheine tse ling tse ngata tsa glial module (NTN1, MDK, COL25A1, ICAM1, le SFRP1) li bontšitse keketseho e kholo ea proteome ea AD, empa ho ne ho batla ho se na phetoho ho AD transcriptome. Liphetoho tsena tse khethehileng tsa protheine li ka 'na tsa amana haufi-ufi le li-amyloid plaques (23, 35), tse totobatsang proteome e le mohloli oa liphetoho tsa pathological,' me liphetoho tsena li ka 'na tsa se ke tsa bonahala ho transcriptome.
Kamora ho itlhahloba ka boikemelo ba boko le li-proteome tsa CSF tseo re li fumaneng, re ile ra etsa tlhahlobo e felletseng ea lisebelisoa tse peli tsa data ho khetholla li-biomarker tsa AD CSF tse amanang le pathophysiology ea marang-rang a boko. Re tlameha ho qala ka ho hlalosa ho kopana ha liprotheine tse peli. Le hoja ho amohetsoe hohle hore CSF e bonahatsa liphetoho tsa neurochemical bokong ba AD (4), tekanyo e nepahetseng ea ho kopana pakeng tsa boko ba AD le CSF proteome ha e hlake. Ka ho bapisa palo ea lihlahisoa tse arolelanoang tsa liphatsa tsa lefutso tse fumanoeng liprotheine tsa rona tse peli, re fumane hore hoo e batlang e le 70% (n = 1936) ea liprotheine tse khetholloang ka cerebrospinal fluid le tsona li ne li lekanngoa bokong (Setšoantšo sa 4A). Bongata ba liprotheine tsena tse fetang (n = 1721) li entsoe 'mapa ho e' ngoe ea li-module tsa 44 tsa co-expression ho tsoa ho sete ea data ea boko (Setšoantšo sa 4B). Joalokaha ho lebeletsoe, li-module tse tšeletseng tse kholo tsa boko (M1 ho M6) li bonts'itse palo e kholo ka ho fetisisa ea ho kopana ha CSF. Leha ho le joalo, ho na le li-modules tse nyenyane tsa boko (mohlala, M15 le M29) tse finyellang tekanyo e phahameng e sa lebelloang ea ho kopana, e kholo ho feta mojule oa boko ka makhetlo a mabeli boholo ba eona. Sena se re susumelletsa ho sebelisa mokhoa o qaqileng haholoanyane, o tsamaisoang ke lipalo-palo oa ho bala ho kopana pakeng tsa boko le cerebrospinal fluid.
(A le B) Liprotheine tse fumanoeng bokong ba ho sibolloa le lisebelisoa tsa data tsa CSF lia kopana. Bongata ba liprotheine tsena tse pota-potileng li amahanngoa le e 'ngoe ea li-module tsa 44 tsa co-expression network ea boko ba co-expression network. (C) Fumana ho kopana lipakeng tsa cerebrospinal fluid proteome le proteome network network. Mola o mong le o mong oa 'mapa oa mocheso o emela tlhahlobo e arohaneng ea hypergeometric FET. Mola o ka holimo o bontša ho kopana (motho o moputsoa / o motšo) pakeng tsa mojule oa boko le proteome eohle ea CSF. Mohala oa bobeli o bontša hore ho kopana pakeng tsa li-modules tsa boko le protheine ea CSF (e nang le moriti o mofubelu) e laoloa haholo ho AD (P <0.05). Mola oa boraro o bontša hore ho kopana pakeng tsa li-modules tsa boko le protheine ea CSF (blue shading) e theohile haholo-e laoloa ho AD (P <0.05). Sebelisa mokhoa oa BH ho lokisa boleng ba P bo nkiloeng ho FET. (D) Phanele ea mojule ea ho mena e ipapisitse le kamano ea mofuta oa sele le mantsoe a amanang le GO. Liphanele tsena li na le kakaretso ea liprotheine tse amanang le boko tse 271, tse nang le polelo e fapaneng e fapaneng ho proteome ea CSF.
Re sebelisa li-FET tse nang le mohatla o le mong, re ile ra lekola bohlokoa ba ho kopana ha protheine lipakeng tsa CSF proteome le li-module tsa boko bo bong. Tlhahlobo e senoletse hore kakaretso ea li-modules tsa boko ba 14 ho CSF data set na statistically overlaps (FDR e fetotsoe P <0.05), le module e eketsehileng (M18) eo ho kopana ha eona ho leng haufi le bohlokoa (FDR e fetotsoe P = 0.06) (Setšoantšo sa 4C , mola o ka hodimo). Re boetse re na le thahasello ho li-module tse sebetsanang haholo le liprotheine tsa CSF tse hlalositsoeng ka tsela e fapaneng. Ka hona, re sebelisitse litlhahlobo tse ling tse peli tsa FET ho fumana hore na (i) protheine ea CSF e eketsehile haholo ho AD le (ii) protheine ea CSF e fokotsehile haholo ho AD (P <0.05, paired t test AD/control) li-module tsa boko tse nang le kamano e utloahalang. pakeng tsa tsona. Joalokaha ho bontšitsoe meleng e bohareng le e ka tlaase ea Setšoantšo sa 4C, litlhahlobo tsena tse eketsehileng li bontša hore 8 ea li-module tsa boko ba 44 li kopana haholo le protheine e kentsoeng ho AD CSF (M12, M1, M2, M18, M5, M44, M33, le M38) . ), ha li-modules tse peli feela (M6 le M15) li bontšitse ho kopana ho nang le morero le protheine e fokotsehileng ho AD CSF. Joalo ka ha ho lebelletsoe, li-module tse 10 kaofela li ka har'a li-module tse 15 tse nang le kamano e phahameng ka ho fetisisa le CSF proteome. Ka hona, re nka hore li-module tsena tsa 15 ke mehloli e phahameng ea lihlahisoa tsa li-biomarker tsa CSF tse nkiloeng bokong tsa AD.
Re ile ra mena limmojule tsena tse 15 tse tobaneng ho etsa liphanele tse hlano tse kholo tsa protheine ho ipapisitsoe le bohaufi ba tsona ho setšoantšo sa sefate sa WGCNA le kamano ea tsona le mefuta ea lisele le liphatsa tsa lefutso (Figure 4D). Phanele ea pele e na le li-module tse ruileng ka li-neuron marker le liprotheine tse amanang le synapse (M1 le M12). Sehlopha sa synaptic se na le kakaretso ea liprotheine tsa 94, 'me maemo a CSF proteome a fetohile haholo, a etsa hore e be mohloli o moholo ka ho fetisisa oa matšoao a CSF a amanang le boko har'a liphanele tse hlano. Sehlopha sa bobeli (M6 le M15) se bontšitse kamano e haufi le lisele tsa endothelial le 'mele oa vascular, o kang "pholiso ea maqeba" (M6) le "taolo ea ho arabela ha humoral immune immune" (M15). M15 e boetse e amana haholo le lipoprotein metabolism, e amanang haufi-ufi le endothelium (36). Phanele ea methapo e na le matšoao a 34 CSF a amanang le boko. Sehlopha sa boraro se kenyelletsa li-modules (M2 le M4) tse amanang haholo le li-marker tsa oligodendrocyte le ho ata ha lisele. Mohlala, mantsoe a boemo bo holimo a ontology a M2 a kenyelletsa "taolo e ntle ea ho ikatisa ha DNA" le "purine biosynthesis process". Ho sa le joalo, tsa M4 li kenyelletsa "phapang ea lisele tsa glial" le "karohano ea chromosome". Phanele ea myelination e na le matšoao a 49 CSF a amanang le boko.
Sehlopha sa bone se na le li-modules tse ngata (M30, M29, M18, M24, le M5), 'me hoo e batlang e le li-module tsohle li ruile haholo ka li-microglia le li-marker tsa astrocyte. Ho tšoana le phanele ea myelination, karolo ea bone e boetse e na le li-modules (M30, M29, le M18) tse amanang haufi-ufi le ho ata ha lisele. Li-module tse ling tsa sehlopha sena li amana haholo le mantsoe a immunology, a kang "mokhoa oa ho itšireletsa mafung" (M5) le "taolo ea karabo ea 'mele" (M24). Sehlopha sa 'mele sa ho itšireletsa mafung sa glial se na le matšoao a 42 a CSF a amanang le boko. Qetellong, karolo ea ho qetela e kenyelletsa matšoao a 52 a amanang le boko ho li-module tse 'nè (M44, M3, M33, le M38), tseo kaofela li leng' meleng o amanang le ho boloka matla le metabolism. E kholo ka ho fetisisa ho li-module tsena (M3) e amana haufi-ufi le mitochondria 'me e ruile ka matšoao a khethehileng a neuron. M38 ke e 'ngoe ea litho tse nyane tsa mojule metabolome ena hape e bonts'a boits'oaro bo itekanetseng ba neuron.
Ka kakaretso, liphanele tsena tse hlano li bonahatsa mefuta e mengata ea lisele le mesebetsi ho AD cortex, 'me ka kakaretso li na le matšoao a 271 a amanang le boko a CSF (Letlapa la S2G). Bakeng sa ho lekola bonnete ba liphetho tsena tsa MS, re sebelisitse proximity extension assay (PEA), thekenoloji ea orthogonal antibody e nang le bokhoni ba ho pheta-pheta, kutloisiso e phahameng le boits'oaro, 'me ra hlahlobisisa lisampole tsa cerebrospinal fluid re fumane karoloana ea li-biomarker tsena tse 271. (n = 36). Lipheo tsena tsa 36 li bonts'a phetoho ea AD multiple ea PEA, e amanang haufi-ufi le liphuputso tsa rona tse thehiloeng ho MS (r = 0.87, P = 5.6 × 10-12), E tiisitseng ka matla liphello tsa tlhahlobo ea rona e feletseng ea MS (Figure S4 ).
Lihlooho tsa likokoana-hloko tse hatisoang ke lihlopha tsa rona tse hlano, ho tloha ho synaptic signaling ho metabolism ea matla, kaofela li amana le pathogenesis ea AD (1-3). Ka hona, li-module tsohle tse 15 tse nang le liphanele tsena li amana le lefu la AD ho proteome ea boko eo re e fumaneng (Setšoantšo sa 2B). Ntho e hlokomelehang ka ho fetisisa ke kamano e phahameng e ntle ea pathological pakeng tsa li-module tsa rona tsa glial le kamano e matla e mpe ea pathological pakeng tsa li-module tsa rona tse kholo ka ho fetisisa tsa neuronal (M1 le M3). Tlhahlobo ea polelo e fapaneng ea proteome ea rona ea boko e phetoang (Setšoantšo sa S3D) e boetse e totobatsa liprotheine tsa glial tse tsoang ho M5 le M18. Ho AsymAD le matšoao a AD, liprotheine tsa glial tse ntseng li eketseha ka ho fetisisa le li-synapse tse amanang le M1 Protheine e fokotsehile ka ho fetisisa. Litlhokomeliso tsena li bontša hore matšoao a 271 cerebrospinal fluid ao re a hlokometseng lihlopheng tse hlano a amana le mekhoa ea mafu ho AD cortex, ho kenyeletsa le tse hlahang methating ea pele ea asymptomatic.
E le ho hlahloba hantle tsela ea phetoho ea liprotheine tse bokong le mokelikeli oa mokokotlo, re ile ra hula tse latelang bakeng sa e 'ngoe le e' ngoe ea li-module tse 15 tse holimo: (i) re fumane boemo ba bongata ba mojule sebakeng sa data ea boko le (ii) module. protheine Phapang e hlahisoa ka cerebrospinal fluid (Setšoantšo sa S5). Joalokaha ho boletsoe pejana, WGCNA e sebelisetsoa ho tseba bongata ba module kapa boleng ba protheine bokong (13). 'Mapa oa seretse se chesang o sebelisoa ho hlalosa polelo e fapaneng ea liprotheine tsa modular ho cerebrospinal fluid (AD/control). Lipalo-palo tsena li bontša hore liphanele tse tharo ho tse hlano li bontša mekhoa e fapaneng ea ho bua bokong le mokelikeli oa mokokotlo. Li-modules tse peli tsa phanele ea synapse (M1 le M12) li bontša ho fokotseha ha tekanyo ea bongata bokong ba AD, empa haholo li kopana le protheine e eketsehileng ho AD CSF (Figure S5A). Li-modules tse amanang le neuron tse nang le metabolome (M3 le M38) li bontšitse boko bo tšoanang le cerebrospinal fluid expression mekhoa e sa lumellaneng (Figure S5E). Sehlopha sa methapo se ile sa boela sa bontša mekhoa e fapaneng ea ho hlalosa, le hoja li-modules tsa eona (M6 le M15) li ne li eketseha ka mokhoa o itekanetseng bokong ba AD 'me li fokotsehile ho CSF e kulang (Figure S5B). Liphanele tse peli tse setseng li na le marang-rang a maholo a glial ao liprotheine tsa tsona li lulang li laoloa ka mokhoa o tsitsitseng likarolong tse peli (Setšoantšo sa S5, C le D).
Ka kopo elelloa hore litloaelo tsena ha lia tloaeleha ho matšoao ohle a liphanele tsena. Ka mohlala, phanele ea synaptic e kenyelletsa liprotheine tse 'maloa tse fokolitsoeng haholo bokong ba AD le CSF (Figure S5A). Har'a matšoao ana a laoloang fatše a cerebrospinal fluid ke NPTX2 le VGF ea M1, le chromogranin B ea M12. Leha ho le joalo, ho sa tsotellehe mekhelo ena, boholo ba matšoao a rona a synaptic a phahame ho AD spinal fluid. Ka kakaretso, litlhahlobo tsena li khonne ho khetholla mekhoa ea lipalo-palo ea maemo a boko le cerebrospinal fluid ho e 'ngoe le e' ngoe ea liphanele tsa rona tse hlano. Mekhoa ena e totobatsa kamano e rarahaneng le hangata e fapaneng pakeng tsa boko le polelo ea protheine ea CSF ho AD.
Ka mor'a moo, re sebelisitse tlhahlobo e phahameng ea ho pheta-pheta ea MS (CSF replication 1) ho fokotsa lihlopha tsa rona tse 271 tsa li-biomarker ho lipheo tse tšepisang ka ho fetisisa le tse ka hlahisoang hape (Setšoantšo sa 5A). CSF khopi 1 e na le kakaretso ea lisampole tse 96 ho tsoa ho Emory Goizueta ADRC, ho kenyeletsoa taolo, AsymAD, le sehlopha sa AD (Tafole S1A). Linyeoe tsena tsa AD li khetholloa ke ho fokotseha ho fokolang ha kelello (ho bolela MoCA, 20.0 ± 3.8), le liphetoho ho li-biomarker tsa AD tse tiisitsoeng ka cerebrospinal fluid (Letlapa la S1A). Ho fapana le tlhahlobo ea CSF eo re e fumaneng, phetisetso ena e etsoa ho sebelisoa mokhoa oa MS o sebetsang hantle le o phahameng oa "single-shot" (ntle le karoloana ea kantle ho marang-rang), ho kenyeletsoa le protocol e nolofalitsoeng ea ho itokisa e felisang tlhokeho ea immunodepletion ea lisampole ka bomong. . Sebakeng seo, "seteishene sa ntlafatso" se felisitsoeng ke 'mele oa ho itšireletsa mafung se sebelisoa ho holisa lets'oao la liprotheine tse ngata haholo (37). Le hoja e fokotsa kakaretso ea proteome, mokhoa ona oa ho thunngoa o le mong o fokotsa nako ea mochine haholo 'me o eketsa palo ea lisampole tse ngotsoeng ka TMT tse ka hlahlojoang tse sebetsang (17, 38). Ka kakaretso, tlhahlobo e bonts'itse li-peptide tse 6,487, tse bonts'itseng liprotheine tse 1,183 maemong a 96. Joalo ka tlhahlobo ea CSF eo re e fumaneng, ke liprotheine feela tse lekantsoeng bonyane 50% ea lisampole tse kenyellelitsoeng lipalong tse latelang, mme data e ile ea khutlisoa bakeng sa litlamorao tsa lilemo le bong. Sena se ile sa etsa hore ho be le palo ea ho qetela ea liprotheine tse 792, tseo 95% ea tsona le tsona li ileng tsa khetholloa ho sete ea data ea CSF e fumanoeng.
(A) Lipheo tsa protheine tsa CSF tse amanang le boko li tiisitsoe sehlopheng sa pele sa CSF se phetoang 'me se kenyelelitsoe karolong ea ho qetela (n = 60). (B ho isa ho E) Maemo a liphanele tsa biomarker (lintlha tse kopantsoeng tsa z) tse lekantsoeng ka lihlopha tse 'ne tsa ho ikatisa tsa CSF. Liteko tsa t tse kopantsoeng kapa ANOVA e nang le tokiso ea morao-rao ea Tukey e ile ea sebelisoa ho lekola bohlokoa ba lipalo-palo ba liphetoho tse ngata tlhahlobong e 'ngoe le e 'ngoe ea phetisetso. CT, taolo.
Kaha re thahasella haholo ho netefatsa lipheo tsa rona tsa CSF tse 271 tse amanang le boko ka tlhahlobo e felletseng, re tla fokotsa tlhahlobo e eketsehileng ea proteome ena e phetoang ho matšoao ana. Har'a liprotheine tsena tsa 271, 100 e ile ea fumanoa ka ho pheta-pheta ha CSF 1. Setšoantšo sa S6A se bontša tlhaloso e fapaneng ea matšoao ana a 100 a fetang pakeng tsa taolo le li-sampuli tsa ho pheta-pheta AD. Synaptic le metabolite histones li eketseha ka ho fetisisa ho AD, ha liprotheine tsa vascular li fokotseha haholo mafung. Boholo ba matšoao a 100 a fetang (n = 70) a bolokile tataiso e tšoanang ea phetoho ho lihlopha tse peli tsa data (Figure S6B). Lits'oants'o tsena tse 70 tse netefalitsoeng tsa CSF tse amanang le boko (Letlapa la S2H) li bonts'a haholo mekhoa ea polelo ea phanele e kileng ea bonoa, ke hore, taolo e tlase ea liprotheine tsa methapo le taolo ea liphanele tse ling kaofela. Ke 10 feela ea liprotheine tsena tse 70 tse netefalitsoeng tse bonts'itseng liphetoho ho bongata ba AD tse hananang le mekhoa ena ea phanele. E le ho hlahisa phanele e bonts'ang hantle mokhoa o akaretsang oa boko le cerebrospinal fluid, re qheletse ka thoko liprotheine tsena tse 10 ho phanele ea thahasello eo qetellong re e netefalitseng (Setšoantšo sa 5A). Ka hona, sehlopha sa rona se qetella se kenyelletsa kakaretso ea liprotheine tse 60 tse netefalitsoeng ka lihlopha tse peli tse ikemetseng tsa CSF AD tse sebelisang litokiso tse fapaneng tsa sampole le tlhahlobo ea sethala sa MS. Merero ea polelo ea z-score ea liphanele tsena tsa ho qetela ho CSF kopi 1 taolo le linyeoe tsa AD li netefalitse mokhoa oa phanele o hlokometsoeng sehlopheng sa CSF seo re se fumaneng (Setšoantšo sa 5B).
Har'a liprotheine tsena tse 60, ho na le limolek'hule tse tsejoang hore li amahanngoa le AD, tse kang osteopontin (SPP1), e leng pro-inflammatory cytokine e 'nileng ea amahanngoa le AD lithutong tse ngata (39-41), le GAP43, A synaptic protein. seo se hokahane ka ho hlaka le neurodegeneration (42). Liprotheine tse netefalitsoeng ka ho fetisisa ke matšoao a amanang le mafu a mang a neurodegenerative, a kang amyotrophic lateral sclerosis (ALS) e amanang le superoxide dismutase 1 (SOD1) le Parkinson's disease e amanang le desaccharase (PARK7) . Re boetse re netefalitse hore matšoao a mang a mangata, joalo ka SMOC1 le boko bo nang le boko bo nang le "membrane attachment attachment 1" (BASP1), li na le lihokelo tse fokolang tsa nakong e fetileng tsa neurodegeneration. Ke habohlokoa ho hlokomela hore ka lebaka la bongata ba bona bo fokolang ka kakaretso ho CSF proteome, ho thata ho rona ho sebelisa mokhoa ona oa ho lemoha o le mong oa boemo bo phahameng ho fumana ka botšepehi MAPT le liprotheine tse ling tse amanang le AD (mohlala, NEFL le NRGN ) ( 43, 44 ).
Ka mor'a moo re ile ra hlahloba matšoao ana a pele a 60 litlhahlobong tse ling tse tharo. Ho CSF Copy 2, re sebelisitse TMT-MS e le 'ngoe ho sekaseka sehlopha se ikemetseng sa taolo ea 297 le lisampole tsa AD ho tsoa ho Emory Goizueta ADRC (17). CSF replication 3 e kenyelelitse ho hlahlojoa bocha ha data e fumanehang ea TMT-MS ho tsoa ho taolo ea 120 le bakuli ba AD ba tsoang Lausanne, Switzerland (45). Re fumane likarolo tse fetang peli ho tse tharo tsa matšoao a mantlha a 60 pokellong ka 'ngoe. Le hoja thuto ea Switzerland e sebelisitse li-platform tse fapaneng tsa MS le mekhoa ea ho lekanya TMT (45, 46), re ile ra hlahisa ka matla mekhoa ea rona ea liphanele ka litlhahlobo tse peli tse pheta-phetoang (Setšoantšo sa 5, C le D, le Tables S2, I, le J) . E le ho hlahloba boemo ba lefu la sehlopha sa rona, re sebelisitse TMT-MS ho hlahloba sete ea bone ea replication data (CSF replication 4), e neng e sa akarelletse feela linyeoe tsa taolo (n = 18) le AD (n = 17), empa hape le PD ( n = 14)), ALS (n = 18) le disampole tsa 'dementia' ea frontotemporal (FTD) (n = 11) (Letlapa la S1A). Re atlehile ho lekanya hoo e ka bang karolo ea bobeli ho tse tharo ea liprotheine tsa phanele sehlopheng sena (38 ho tse 60). Liphetho tsena li totobatsa liphetoho tse ikhethileng tsa AD ho liphanele tsohle tse hlano tsa biomarker (Setšoantšo sa 5E le Lethathamo la S2K). Keketseho ea sehlopha sa metabolite e bontšitse matla a matla ka ho fetisisa a AD, a lateloa ke sehlopha sa myelination le glial. Ka tekanyo e fokolang, FTD e boetse e bontša keketseho pakeng tsa lihlopha tsena, tse ka 'nang tsa bontša liphetoho tse tšoanang tsa marang-rang (17). Ka lehlakoreng le leng, ALS le PD li bontšitse hoo e batlang e le li-profile tsa myelination, glial, le metabolome e le sehlopha sa taolo. Ka kakaretso, ho sa tsotellehe liphapang tsa ho lokisetsa mehlala, sethaleng sa MS, le mekhoa ea ho lekanya TMT, litlhahlobo tsena tse pheta-phetoang li bontša hore matšoao a rona a pele a na le liphetoho tse khethehileng tsa AD ho feta lisampole tse ikhethang tsa 500 tsa CSF.
AD neurodegeneration e 'nile ea tsejoa ka bophara lilemo tse' maloa pele matšoao a kelello a qala, kahoo ho na le tlhokahalo e potlakileng ea li-biomarker tsa AsymAD (5, 31). Leha ho le joalo, bopaki bo ntseng bo eketseha bo bontša hore biology ea AsymAD e hole le ho homogeneous, 'me tšebelisano e rarahaneng ea kotsi le botsitso e lebisa ho phapano e kholo ea batho ka mor'a tsoelo-pele ea mafu (47). Leha e ne e sebelisetsoa ho khetholla linyeoe tsa AsymAD, maemo a li-biomarker tsa CSF tsa mantlha (Aβ1-42, tau kaofela le p-tau) ha a so ka a paka hore a ka tseba esale pele hore na ke mang ea tla hatela pele ho 'dementia' (4, 7), a bonts'a ho feta. ho hlokahala ho kenyelletsa lisebelisoa tse akaretsang tsa biomarker tse ipapisitseng le likarolo tse ngata tsa fisioloji ea boko ho beha ka nepo kotsi ea baahi bana. Ka hona, ka mor'a moo re ile ra hlahloba phanele ea rona ea AD-valided biomarker sebakeng sa AsymAD ea CSF kopi ea 1. Linyeoe tsena tsa 31 AsymAD li bontšitse maemo a sa tloaelehang a mantlha a biomarker (Aβ1-42 / kakaretso ea tau ELISA, <5.5) le kutloisiso e feletseng (e bolela MoCA, 27.1). ± 2.2) (Letlapa la S1A). Ntle le moo, batho bohle ba nang le AsymAD ba na le lintlha tsa 'dementia' tsa kliniki tsa 0, tse bontšang hore ha ho na bopaki ba ho theoha ha ts'ebetso ea letsatsi le letsatsi ea kelello kapa ea ts'ebetso.
Re ile ra qala ho sekaseka maemo a liphanele tse netefalitsoeng ho likopi tsohle tsa 96 CSF 1, ho kenyeletsoa sehlopha sa AsymAD. Re fumane hore liphanele tse 'maloa tsa sehlopha sa AsymAD li ne li e-na le liphetoho tse ngata tsa AD-joaloka bongata, phanele ea vascular e bontšitse mokhoa oa ho theoha ho AsymAD, ha likarolo tse ling kaofela li bontša mokhoa o holimo (Setšoantšo sa 6A). Ka hona, liphanele tsohle li bontšitse kamano ea bohlokoa haholo le ELISA Aβ1-42 le litekanyetso tsa tau kaofela (Setšoantšo sa 6B). Ka lehlakoreng le leng, kamano lipakeng tsa sehlopha le lintlha tsa MoCA e batla e fokola. E 'ngoe ea liphuputso tse makatsang ho tsoa litlhahlobong tsena ke palo e kholo ea bongata ba sehlopha sa AsymAD. Joalokaha ho bontšitsoe setšoantšong sa 6A, boemo ba phanele ea sehlopha sa AsymAD hangata bo tšela boemo ba sehlopha sa taolo le sehlopha sa AD, se bonts'a phapang e batlang e phahame. Ho tsoela pele ho hlahloba phapang ena ea AsymAD, re sebelisitse tlhahlobo ea Multidimensional Scaling (MDS) ho linyeoe tsa 96 CSF replication 1. Tlhahlobo ea MDS e lumella ho bona ka mahlo ho tšoana pakeng tsa linyeoe tse thehiloeng ho mefuta e itseng ea data. Bakeng sa tlhahlobo ena ea lihlopha, re sebelisa feela matšoao a phanele a tiisitsoeng a nang le phetoho e kholo ea lipalo (P <0.05, AD/control) boemong ba ho sibolloa ha CSF le ho pheta-pheta 1 proteome (n = 29) (Table S2L). Tlhahlobo ena e hlahisitse lihlopha tse hlakileng tsa sebaka pakeng tsa taolo ea rona le linyeoe tsa AD (Setšoantšo sa 6C). Ka lehlakoreng le leng, linyeoe tse ling tsa AsymAD li kopantsoe ka ho hlaka sehlopheng sa taolo, ha tse ling li le maemong a AD. Ho tsoela pele ho hlahloba AsymAD heterogeneity, re sebelisitse 'mapa oa rona oa MDS ho hlalosa lihlopha tse peli tsa linyeoe tsena tsa AsymAD. Sehlopha sa pele se ne se kenyelletsa linyeoe tsa AsymAD tse kopantsoeng haufi le taolo (n = 19), ha sehlopha sa bobeli se ne se khetholloa ke linyeoe tsa AsymAD tse nang le letšoao le haufi le AD (n = 12).
(A) Boemo ba polelo (z-score) ea sehlopha sa biomarker sa CSF ho disampole tsohle tse 96 sehlopheng sa CSF replication 1, ho kenyeletsoa AsymAD. Tlhahlobo ea phapang le tokiso ea morao-rao ea Tukey e ile ea sebelisoa ho lekola bohlokoa ba lipalo-palo ba liphetoho tse ngata tsa phanele. (B) Tlhahlobo ea kamano ea phanele ea protheine abundance level (z-score) e nang le lintlha tsa MoCA le kakaretso ea tau ho ELISA Aβ1-42 le CSF copy 1 sampuli. Coefficient ea khokahano ea Pearson e nang le P value e amehang e ea hlaha. (C) MDS ea 96 CSF kopi ea linyeoe tsa 1 e ne e itšetlehile ka maemo a mangata a 29 a tiisitsoeng a li-panel markers, tse fetotsoeng haholo ka bobeli ho sibolloa le CSF copy 1 data sets [P <0.05 AD / control (CT)]. Tlhahlobo ena e ne e sebelisetsoa ho arola sehlopha sa AsymAD ho laola (n = 19) le AD (n = 12) lihlopha tse nyenyane. (D) Morero oa ho foqoha ha seretse se chesang o bonts'a polelo e fapaneng ea liprotheine tsohle tsa CSF tse pheta-phetoang 1 tse nang le phetoho ea log2 fold (x-axis) e amanang le boleng ba -log10 lipalo pakeng tsa lihlopha tse peli tsa AsymAD. Li-biomarker tsa phanele li na le mebala. (E) CSF replication 1 boemo ba bongata ba li-biomarker tsa sehlopha sa khetho li hlahisoa ka tsela e fapaneng lipakeng tsa lihlopha tse nyane tsa AsymAD. Tlhahlobo e fetotsoeng ea Tukey ea ho fapana e ile ea sebelisoa ho lekola bohlokoa ba lipalo.
Re hlahlobile polelo ea phapang ea protheine lipakeng tsa taolo ena le linyeoe tsa AD-joaloka AsymAD (Setšoantšo sa 6D le Lethathamo la S2L). 'Mapa oa seretse se chesang o bonts'a hore matšoao a 14 a fetohile haholo lipakeng tsa lihlopha tsena tse peli. Boholo ba matšoao ana ke litho tsa synapse le metabolome. Leha ho le joalo, SOD1 le myristoylated alanine-rich protein kinase C substrate (MARCKS), e leng litho tsa lihlopha tsa 'mele tsa ho itšireletsa mafung tsa myelin le glial, ka ho latellana, le tsona ke tsa sehlopha sena (Setšoantšo sa 6, D le E) . Sehlopha sa methapo se ile sa boela sa kenya letsoho matšoao a mabeli a fokolitsoeng haholo sehlopheng sa AD-joaloka AsymAD, ho kenyelletsa le AE e tlamang protheine 1 (AEBP1) le ho tlatsa setho sa lelapa sa C9. Ho ne ho se na phapang e khōlō pakeng tsa taolo le lihlopha tse nyenyane tsa AD-joaloka AsymAD ho ELISA AB1-42 (P = 0.38) le p-tau (P = 0.28), empa ho ne ho hlile ho e-na le phapang e khōlō boemong ba tau kaofela (P = 0.0031 ) (Setšoantšo sa S7). Ho na le matšoao a 'maloa a liphanele a bontšang hore liphetoho pakeng tsa lihlopha tse peli tsa AsymAD li bohlokoa ho feta kakaretso ea maemo a tau (mohlala, YWHAZ, SOD1, le MDH1) (Figure 6E). Ka kakaretso, liphetho tsena li bonts'a hore karolo ea rona e netefalitsoeng e kanna ea ba le li-biomarker tse ka fokotsang kotsi ea bakuli ba nang le lefu la asymptomatic.
Ho na le tlhokahalo e potlakileng ea lisebelisoa tsa biomarker tse thehiloeng ho sistimi ho lekanya hamolemo le ho shebisa mafu a fapaneng a amanang le AD. Lisebelisoa tsena li lebeletsoe hore li se ke tsa fetola feela moralo oa rona oa ho hlahloba AD, empa hape li khothalletsa ho amoheloa ha mekhoa ea phekolo e sebetsang, e tobileng ea mokuli (1, 2). Ho fihlela sena, re sebelisitse mokhoa o felletseng oa proteomics ho AD bokong le CSF ho tsebahatsa li-biomarker tsa CSF tse thehiloeng marang-rang tse bonts'ang mefuta e mengata e fapaneng ea pathophysiology e thehiloeng bokong. Tlhahlobo ea rona e hlahisitse liphanele tse hlano tsa li-biomarker tsa CSF, tseo (i) li bonts'ang li-synapses, methapo ea mali, myelin, ho se sebetse ha 'mele le metabolism; (ii) bonts'a ho ikatisa ho matla ho li-platform tse fapaneng tsa MS; ( iii) Bontša liphetoho tse ntseng li tsoela pele tsa lefu ho pholletsa le mekhahlelo ea pele le ea morao ea AD. Ka kakaretso, liphetho tsena li emela mohato o ts'episang ho nts'etsopele ea lisebelisoa tse fapaneng, tse tšepahalang, tse shebaneng le webo bakeng sa lipatlisiso tsa AD le lits'ebetso tsa bongaka.
Liphetho tsa rona li bonts'a mokhatlo o sirelelitsoeng haholo oa proteome ea marang-rang ea boko 'me e ts'ehetsa ts'ebeliso ea eona e le ankora bakeng sa nts'etsopele ea biomarker e thehiloeng ho sistimi. Tlhahlobo ea rona e bonts'a hore li-dataset tse peli tse ikemetseng tsa TMT-MS tse nang le boko ba AD le AsymAD li na le modularity e matla. Liphuputso tsena li eketsa mosebetsi oa rona oa pele, ho bontša ho boloka li-module tse matla tsa lisele tsa boko tse fetang 2,000 tse tsoang ho lihlopha tse ngata tse ikemetseng tse ka pele, parietal le temporal cortex (17). Marang-rang ana a tumellano a bonts'a liphetoho tse fapaneng tse amanang le mafu tse hlokometsoeng liphuputsong tsa hajoale, ho kenyelletsa ho eketseha ha li-module tsa glial-rich inflammatory le ho fokotseha ha li-module tse ruileng tsa neuron. Joalo ka lipatlisiso tsa hajoale, marang-rang ana a maholo a boetse a na le liphetoho tse kholo tsa modular ho AsymAD, e bonts'ang mefuta e fapaneng ea li-preclinical pathophysiology (17).
Leha ho le joalo, ka har'a moralo ona o itšetlehileng ka mokhoa o ts'oarellang haholo, ho na le liphatsa tsa lefutso tse ntle haholo, haholo-holo har'a batho ba mekhahlelo ea pele ea AD. Sehlopha sa rona sa biomarker se khona ho hlahisa lihlopha tse peli ho AsymAD, tse bonts'ang phapang e kholo ea matšoao a mangata a CSF. Sehlopha sa rona se khonne ho totobatsa phapang ea tlhaho pakeng tsa lihlopha tsena tse peli tse nyenyane, tse neng li sa totobala boemong ba li-biomarker tsa AD tsa mantlha. Ha ho bapisoa le sehlopha sa taolo, Aβ1-42/total tau ratios ea batho bana ba AsymAD e ne e le tlase ka mokhoa o sa tloaelehang. Leha ho le joalo, ke maemo a kakaretso a tau feela a neng a fapane haholo pakeng tsa lihlopha tse peli tsa AsymAD, ha maemo a Aβ1-42 le p-tau a ntse a batla a bapisoa. Kaha tau e phahameng ea CSF e bonahala e le selelekela se betere sa matšoao a kelello ho feta maemo a Aβ1-42 (7), re belaela hore lihlopha tse peli tsa AsymAD li ka ba le likotsi tse fapaneng tsa tsoelo-pele ea mafu. Ka lebaka la sampole e lekantsoeng ea AsymAD ea rona le khaello ea data ea longitudinal, lipatlisiso tse ling lia hlokahala ho fihlela liqeto tsena ka kholiseho. Leha ho le joalo, liphetho tsena li bonts'a hore phanele ea CSF e thehiloeng ho sistimi e ka ntlafatsa bokhoni ba rona ba ho qhekella batho ka nepo nakong ea boemo ba lefu lena.
Ka kakaretso, liphuputso tsa rona li tšehetsa karolo ea mesebetsi e mengata ea likokoana-hloko ho pathogenesis ea AD. Leha ho le joalo, dysregulated energy metabolism e bile sehlooho se hlahelletseng sa liphanele tsa rona tse hlano tse netefalitsoeng tsa ho ngola. Liprotheine tsa Metabolic, tse kang hypoxanthine-guanine phosphoribosyltransferase 1 (HPRT1) le lactate dehydrogenase A (LDHA), ke li-synaptic biomarkers tse tiisitsoeng ka ho fetisisa, tse bontšang hore keketseho ea AD CSF ke thobalano e ka hlahisoang haholo. Methapo ea rona ea mali le liphanele tsa glial le tsona li na le matšoao a 'maloa a amehang metabolism ea lintho tse oxidative. Liphuputso tsena li tsamaisana le karolo ea bohlokoa eo lits'ebetso tsa metabolic li e bapalang bokong kaofela, eseng feela ho fihlela tlhoko e phahameng ea matla a li-neurone, empa hape le ho fihlela tlhoko e phahameng ea matla ea bonohe ba linaleli le lisele tse ling tsa glial (17, 48). Liphetho tsa rona li ts'ehetsa bopaki bo ntseng bo hola ba hore liphetoho tse ka bang teng ho redox le ho sitisoa ha litsela tsa matla e kanna ea ba sehokelo sa mantlha lipakeng tsa lits'ebetso tse 'maloa tsa bohlokoa tse amehang ho pathogenesis ea AD, ho kenyeletsoa mathata a mitochondrial, ho ruruha ha glial, le tšenyo ea Vascular (49). Ntle le moo, li-biomarker tsa metabolic cerebrospinal fluid li na le palo e kholo ea liprotheine tse ruileng ka mokhoa o fapaneng lipakeng tsa taolo ea rona le li-subgroups tse kang AD-joaloka AsymAD, tse fanang ka maikutlo a hore tšitiso ea litsela tsena tsa matla le redox e kanna ea ba bohlokoa mothating oa pele oa lefu lena.
Mekhoa e fapaneng ea boko le cerebrospinal fluid panel eo re e boneng le eona e na le litlamorao tse khahlisang tsa biology. Li-synapses le metabolomes tse ruileng ka li-neurone li bonts'a maemo a fokotsehileng bokong ba AD le ho eketseha ha mokelikeli oa cerebrospinal. Ka lebaka la hore li-neurone li na le li-mitochondria tse hlahisang matla ho li-synapse ho fana ka matla bakeng sa matšoao a tsona a mangata a ikhethileng (50), ho lebelletsoe ho ts'oana ha litlhaloso tsa lihlopha tsena tse peli tsa neuron. Tahlehelo ea li-neurone le extrusion ea lisele tse senyehileng li ka hlalosa mekhoa ena ea boko le CSF ka lefu la morao-rao, empa li ke ke tsa hlalosa liphetoho tsa pele tseo re li bonang (13). Tlhaloso e 'ngoe e ka bang teng bakeng sa liphuputso tsena tsa lefu la pele la asymptomatic ke ho faola ho sa tloaelehang ha synaptic. Bopaki bo bocha mefuteng ea litoeba bo fana ka maikutlo a hore microglia-mediated synaptic phagocytosis e kanna ea kengoa tšebetsong ka mokhoa o sa tloaelehang ho AD mme ea lebisa tahlehelong ea pele ea synapse bokong (51). Lintho tsena tse lahliloeng tsa synaptic li ka 'na tsa bokellana ho CSF, ke ka lebaka leo re bonang keketseho ea CSF phanele ea neuron. Ho faola li-synaptic ka mokhoa oa ho itšireletsa mafung ho ka boela ha hlalosa ho eketseha ha liprotheine tsa glial tseo re li bonang bokong le cerebrospinal fluid nakong eohle ea lefu lena. Ntle le ho pruning ea synaptic, ho se tloaelehe ka kakaretso tseleng ea exocytic ho ka boela ha lebisa bokong bo fapaneng le lipolelo tsa CSF tsa matšoao a neuronal. Liphuputso tse 'maloa li bontšitse hore litaba tsa li-exosomes ho pathogenesis ea boko ba AD li fetohile (52). Tsela ea extracellular e boetse e ameha ho ata ha Aβ (53, 54). Ho bohlokoa ho elelloa hore khatello ea secretion ea exosomal e ka fokotsa lefu la AD ho mefuta ea litoeba tsa AD transgenic (55).
Ka nako e ts'oanang, protheine e ka har'a phanele ea methapo e bontšitse keketseho e itekanetseng ea boko ba AD, empa e fokotsehile haholo ho CSF. Ho se sebetse hantle ha "blood-brain barrier" (BBB) ho ka hlalosa liphetho tsena. Lithuto tse ngata tse ikemetseng tsa postmortem tsa batho li bontšitse ho senyeha ha BBB ho AD (56, 57). Lithuto tsena li netefalitse liketsahalo tse fapaneng tse sa tloaelehang tse potileng lera lena le tiisitsoeng ka thata la lisele tsa endothelial, ho kenyeletsoa ho lutla ha capillary le ho bokellana ha liprotheine tse fetisitsoeng maling (57). Sena se ka fana ka tlhaloso e bonolo bakeng sa liprotheine tse phahameng tsa vascular bokong, empa ha li khone ho hlalosa ka botlalo ho fokotseha ha liprotheine tsena ka har'a cerebrospinal fluid. Monyetla o mong ke hore tsamaiso ea methapo e bohareng e ikakhela ka setotsoana limolek'hule tsena ho rarolla bothata ba ho ruruha ho eketsehileng le khatello ea oxidative. Ho fokotseha ho tse ling tsa liprotheine tse matla ka ho fetisisa tsa CSF phanele ena, haholo-holo tse amehang taolong ea lipoprotein, li amana le ho thibela maemo a kotsi a ho ruruha le ts'ebetso ea methapo ea methapo ea mefuta ea oksijene e sebetsang. Sena ke 'nete bakeng sa Paroxonase 1 (PON1), enzyme e tlamang lipoprotein e ikarabellang bakeng sa ho fokotsa maemo a khatello ea oxidative ho potoloha (58, 59). Alpha-1-microglobulin/bikunin precursor (AMBP) ke letšoao le leng le laoloang haholo la sehlopha sa methapo ea mali. Ke selelekela sa bikunin ea lipid transporter, e amehang hape ho hatelleng ho ruruha le Ts'ireletso ea methapo (60, 61).
Ho sa tsotellehe likhopolo-taba tse fapa-fapaneng tse thahasellisang, ho se khone ho lemoha ka ho toba mekhoa ea lefu la biochemical ke moeli o tsebahalang oa tlhahlobo ea proteomics e entsoeng ke ho sibolloa. Ka hona, lipatlisiso tse ling lia hlokahala ho hlalosa ka kholiseho mekhoa ea liphanele tsena tsa biomarker. E le ho fetela molemong oa nts'etsopele ea tlhahlobo ea kliniki e thehiloeng ho MS, tataiso ea nako e tlang e boetse e hloka hore ho sebelisoe mekhoa ea palo e lebisitsoeng bakeng sa netefatso e kholo ea biomarker, joalo ka tlhahlobo e ikhethileng kapa e ts'oanang ea karabelo (62). Haufinyane tjena re sebelisitse tlhahlobo ea karabelo e tšoanang (63) ho netefatsa liphetoho tse ngata tsa protheine ea CSF tse hlalositsoeng mona. Lipheo tse 'maloa tsa bohlokoa li lekantsoe ka nepo e kholo, ho kenyeletsoa YWHAZ, ALDOA, le SMOC1, e leng 'mapa oa liphanele tsa rona tsa synapse, metabolism le ho ruruha, ka ho latellana (63). Independent Data Acquisition (DIA) le maano a mang a thehiloeng ho MS le ona a ka ba molemo bakeng sa netefatso ea sepheo. Bud le ba bang. (64) Haufinyane tjena ho ile ha bontšoa hore ho na le ho kopana ho hoholo pakeng tsa li-biomarker tsa AD tse khethiloeng ho sete ea rona ea data ea CSF ea ho sibolla le data e ikemetseng ea DIA-MS, e nang le hoo e ka bang 200 ea lisampole tsa CSF tse tsoang lihlopha tse tharo tse fapaneng tsa Europe. Liphuputso tsena tsa morao-rao li tšehetsa bokhoni ba liphanele tsa rona ho fetoha ho lemoha ho thehiloeng ho MS. Tšebeliso ea li-antibody ea setso le aptamer e boetse e bohlokoa bakeng sa nts'etsopele ea li-biomarker tsa AD. Ka lebaka la bongata bo fokolang ba CSF, ho thata haholo ho lemoha li-biomarker tsena ho sebelisa mekhoa e phahameng ea MS. NEFL le NRGN ke mehlala e 'meli e joalo ea li-biomarker tsa CSF tse nang le bongata bo tlase, tse kentsoeng sethaleng tlhahlobong ea rona e felletseng, empa li ke ke tsa bonoa ka ts'epo ho sebelisa leano la rona le le leng la MS. Maano a ho lebela a ipapisitseng le li-antibodies tse ngata, joalo ka PEA, a ka khothaletsa phetoho ea kliniki ea matšoao ana.
Ka kakaretso, thuto ena e fana ka mokhoa o ikhethileng oa proteomics bakeng sa ho tsebahatsa le ho netefatsa li-biomarker tsa CSF AD tse ipapisitseng le litsamaiso tse fapaneng. Ho ntlafatsa liphanele tsena tsa matšoao ho li-cohorts tse ling tsa AD le li-platform tsa MS ho ka ts'episa ho ntšetsa pele stratification ea kotsi ea AD le kalafo. Lithuto tse lekanyang boemo ba nako e telele ba liphanele tsena ha nako e ntse e ea li bohlokoa hape ho fumana hore na ke motsoako ofe oa matšoao o behang hantle kotsi ea lefu la pele le liphetoho ho matla a lefu.
Ntle le lisampole tse 3 tse kopilitsoeng ke CSF, lisampole tsohle tsa CSF tse sebelisitsoeng phuputsong ena li ile tsa bokelloa tlas'a tšehetso ea Emory ADRC kapa mekhatlo ea lipatlisiso e amanang haufi-ufi. Kakaretso ea lihlopha tse 'nè tsa lisampole tsa Emory CSF li sebelisitsoe lithutong tsena tsa proteomics. Sehlopha sa CSF se fumanoe se na le lisampole ho tsoa ho litsamaiso tse 20 tse phetseng hantle le bakuli ba 20 ba AD. CSF kopi 1 e kenyelletsa disampole ho tsoa ho li-control tse 32 tse phetseng hantle, batho ba 31 ba AsymAD, le batho ba 33 ba AD. CSF kopi 2 e na le 147 controls le 150 AD disampole. Sehlopha sa 4 sa CSF replication 4 se kenyelletsa li-control tse 18, 17 AD, 19 ALS, 13 PD, le disampole tse 11 tsa FTD. Ho ea ka tumellano e amohetsoeng ke Boto ea Tlhahlobo ea Setsi sa Emory University, bankakarolo bohle ba thuto ea Emory ba fumane tumello ka tsebo. Ho ea ka 2014 National Institute of Aging Best Practice Guidelines for Alzheimer's Centers (https://alz.washington.edu/BiospecimenTaskForce.html), cerebrospinal fluid e ile ea bokelloa 'me ea bolokoa ka ho phunngoa ha lumbar. Bakuli ba Control le AsymAD le AD ba fumane tlhahlobo e tloaelehileng ea kelello ho Emory Cognitive Neurology Clinic kapa Goizueta ADRC. Mehlala ea bona ea cerebrospinal fluid e ile ea lekoa ke INNO-BIA AlzBio3 Luminex bakeng sa ELISA Aβ1-42, kakaretso ea tau le p-tau (65). Litekanyetso tsa ELISA li sebelisoa ho ts'ehetsa likarolo tsa tlhahlobo ea lithuto tse ipapisitseng le litekanyetso tse khaotsoeng tsa AD biomarker (66, 67). Lintlha tsa motheo tsa palo ea batho le tsa tlhahlobo ea mafu a mang a CSF (FTD, ALS, le PD) le tsona li fumanoa ho Emory ADRC kapa mekhatlong ea lipatlisiso e amanang le eona. Kakaretso ea metadata ea linyeoe tsena tsa Emory CSF e ka fumanoa ho Lethathamo la S1A. Litšobotsi tsa sehlopha sa Swiss CSF replication 3 cohort li se li phatlalalitsoe pele (45).
CSF e fumane mohlala. E le ho eketsa botebo ba ho sibolloa ha data ea CSF, tšebeliso ea 'mele ea ho itšireletsa mafung ea liprotheine tse ngata e ile ea etsoa pele ho trypsinization. Ka bokhuts'oane, 130 μl ea CSF ho tsoa ho lisampole tse 40 tsa CSF le bophahamo bo lekanang (130 μl) ea High Select Top14 Abundance Protein Depletion Resin (Thermo Fisher Scientific, A36372) li ile tsa beoa kholomong ea spin (Thermo Fisher Scientific, A89868) ka phaposing. mocheso Koamisa). Kamora ho ohla metsotso e 15, centrifuge sampole ho 1000g metsotso e 2. Sesebelisoa sa filtara sa 3K ultracentrifugal (Millipore, UFC500396) se sebelisitsoe ho tsepamisa sampole ea litšila ka centrifuging ho 14,000g metsotso e 30. Hlakola mefuta eohle ea sampole ho 75 μl ka letsoai la phosphate buffered. Tekanyo ea protheine e ile ea hlahlojoa ka mokhoa oa bicinchoninic acid (BCA) ho latela protocol ea moetsi (Thermo Fisher Scientific). The immunodepleted CSF (60 μl) ho tsoa ho lisampole tsohle tse 40 e ile ea chekoa ka lysyl endopeptidase (LysC) le trypsin. Ka bokhutšoanyane, sampuli e ile ea fokotseha 'me ea alkylated ka 1.2 μl 0.5 M tris-2 (-carboxyethyl) -phosphine le 3 μl 0.8 M chloroacetamide ka 90 ° C ka metsotso ea 10, ebe e sonicated ka ho hlapa metsi ka metsotso e 15. Mohlala o ile oa hlapolloa ka 193 μl 8 M urea buffer [8 M urea le 100 mM NaHPO4 (pH 8.5)] ho ea ho motsoako oa ho qetela oa 6 M urea. LysC (4.5 μg; Wako) e sebelisoa bakeng sa tšilo ea bosiu ka mocheso oa kamore. Joale sampole e ile ea hlapolloa ho 1 M urea ka 50 mM ammonium bicarbonate (ABC) (68). Eketsa palo e lekanang (4.5 μg) ea trypsin (Promega), ebe u kenya sampole ka lihora tse 12. Eketsa tharollo ea peptide e chesitsoeng ho 1% formic acid (FA) le 0.1% trifluoroacetic acid (TFA) (66), 'me e be letsoai ka kholomo ea 50 mg Sep-Pak C18 (Metsi) joalo ka ha ho hlalositsoe kaholimo (25) . Joale peptide e ile ea hlakoloa ka 1 ml ea 50% acetonitrile (ACN). Ho leka-lekanya palo ea liprotheine ho pholletsa le lihlopha (25), li-aliquots tse 100 μl ho tsoa lisampoleng tsohle tse 40 tsa CSF li ile tsa kopanngoa ho hlahisa sampole e tsoakiloeng, e ileng ea aroloa ka lisampole tse hlano tsa maemo a ka hare tsa lefatše (GIS) (48). Lisampole tsohle tsa motho ka mong le litekanyetso tse kopantsoeng li omisitsoe ke vacuum ea lebelo le phahameng (Labconco).
CSF e kopitsa mohlala. Dayon le basebetsi-'moho ba kile ba hlalosa ho fokotseha ha 'mele oa ho itšireletsa mafung le tšilo ea lijo tsa CSF sampuli ea 3 (45, 46). Lisampole tse setseng tse replicate ha lia ka tsa felloa ke matla ka bonngoe. Cheka lisampole tsena tse sa tlosoeng ho trypsin joalo ka ha ho hlalositsoe pejana (17). Bakeng sa tlhahlobo e 'ngoe le e' ngoe e pheta-phetoang, li-aliquots tse 120 μl tsa peptide e hlahisitsoeng ho tloha sampoleng e 'ngoe le e' ngoe li ile tsa kopanngoa 'me tsa aroloa ka li-aliquots tse lekanang ho sebelisoa e le tekanyetso ea ka hare ea lefatše ea TMT (48). Lisampole tsohle tsa motho ka mong le litekanyetso tse kopantsoeng li omisitsoe ke vacuum ea lebelo le phahameng (Labconco). Bakeng sa ho matlafatsa lets'oao la protheine ea CSF e ngata haholo, ka ho kopanya 125 μl ho tsoa sampoleng e 'ngoe le e' ngoe, ho lokiselitsoe sampole e "ntlafalitsoeng" bakeng sa tlhahlobo e 'ngoe le e 'ngoe [ke hore, sampole ea baeloji e etsisang sampole ea lipatlisiso, empa chelete e teng ke. e kholoanyane haholo (37, 69)] e kopantsoeng ho ba sampole e tsoakiloeng ea CSF (17). Joale sampole e tsoakiloeng e ile ea tlosoa ho sebelisoa 12 ml ea High Select Top14 Abundance Protein Removal Resin (Thermo Fisher Scientific, A36372), e silehileng joalokaha e hlalositsoe ka holimo, 'me e kenyelelitsoe ho tse ngata tse latelang tsa TMT tse ngotsoeng.
Nako ea poso: Aug-27-2021