MARK-AGE population: From the human model to new insights

Mechanisms o Ageing and De elopmen 151 (2015) 13–17 Con en s lis s a ailable a ScienceDi ec Mechanisms o Ageing and De elopmen jou nal homepage: www.else ie .com/loca e/mechagede MARK-AGE popula ion: F om he human model o new insigh s Mi iam Cap ia,b,∗,1, Ma ía Mo eno-Villanue ac,1, Elisa Ce eninia, Elisa Pinia, Ma ia Scu ia, Vincenzo Bo ellia, Ma ia Gius ina Palmasb, Ma co Zolid, Ch is iane Schöne, Anne Siepelmeye e, Jü gen Be nha d e, Simone Fiegl , Ge ben Zondagg, An on J.M. de C aenh, An i He oneni, Mikko Hu mei, Ewa Siko aj, E s a hios S. Gonosk, Kons an inos Vou e akisk, Oli ie Toussain l, Flo ence Debacq-Chainiauxl, Bea ix G ubeck-Loebens einm, Alexande Bü klec, Claudio F anceschia,b aDIMES-Depa men o Expe imen al, Diagnos ic and Special y Medicine, Alma Ma e S udio um, Uni e si y o Bologna, 40126 Bologna, I aly bCIG-In e depa men al Cen e “L.Gal ani”, Alma Ma e S udio um, Uni e si y o Bologna, 40126 Bologna, I aly cMolecula Toxicology G oup, Depa men o Biology, Uni e si y o Kons anz, 78457 Kons anz, Ge many dDepa men o Medical and Su gical Sciences, Uni e si y o Bologna and S. O sola-Malpighi Hospi al, Bologna, I aly eBioTeSys GmbH, 73728 Esslingen, Ge many UMIT P i a e Uni e si ä ü Gesundhei swissenscha en, Medizinische In o ma ik und Technik GmbH, 6060 Hall in Ti ol, Aus ia gDNage BV, Leiden, The Ne he lands hDepa men o Ge on ology and Ge ia ics, Leiden Uni e si y Medical Cen e, Leiden, The Ne he lands iSchool o Medicine, Uni e si y o Tampe e, 33014 Tampe e, Finland jLabo a o y o he Molecula Bases o Ageing, Nencki Ins i u e o Expe imen al Biology, Polish Academy o Sciences, 3 Pas eu S ee , 02-093 Wa saw, Poland kIns i u e o Biology, Medicinal Chemis y and Bio echnology, Na ional Hellenic Resea ch Founda ion, 48 Vas. Cons an inou A e. 116 35 A hens, G eece lNARILIS URBC, Uni e si y o Namu (FUNDP), 61, ue de B uxelles, 5000 Namu , Belgium mImmunology Di ision, Ins i u e o Biomedical Aging Resea ch, Uni e si ä Innsb uck, Rennweg 10, 6020 Innsb uck, Aus ia a i c l e i n o A icle his o y: A ailable online 2 Ap il 2015 Keywo ds: Human ageing Bioma ke s Rec ui men MARK-AGE popula ion Many ele an EU p ojec s we e unded du ing las decade and MARK-AGE (Eu opean S udy o Es ablish Bioma ke s o Human Ageing) was he fi s ocused on he iden ifica ion o bioma k- e s connec ing i sel wi h a p e ious unded p ojec , i.e. GEHA (Gene ic o Heal hy Ageing) ha iden ified Eu opean amilies wi h longe i y componen (F anceschi e al., 2007). MARK-AGE was mainly a c oss sec ional s udy, based on defini e assump ions, as Abb e ia ions: CS, Cockayne synd ome; DS, Down synd ome; GO, GEHA o - sp ing; RASIG, andomly ec ui ed age-s a ified indi iduals om he gene al popula ion; SGO, spouses o GO; WS, We ne synd ome. ∗Co esponding au ho a : Depa men o Expe imen al, Diagnos ic and Special y Medicine, Alma Ma e S udio um, Uni e si y o Bologna, Via S. Giacomo, 12 40126 Bologna, I aly. E-mail add ess: [email p o ec ed] (M. Cap i). 1These au ho s con ibu ed equally o his wo k i will be desc ibed below, and ocused on an age ange be ween 34 and 75 yea s, in o de o iden i y ea ly bioma ke s o biolog- ical s. ch onological age, po en ially capable o p edic ing he a e o ageing la e in li e. Complemen a ily, many an i-ageing s a egies ha e been p oposed, such as hose ela ed o immune sys em emodelling (Cap i e al., 2006b), bu a new e a begun on di e en issues-specific epigenomics (Ho a h, 2013; Romanoski e al., 2015), ood/nu i ion science, die in e en ion (San o o e al., 2014; Bacalini e al., 2014; Me cken e al., 2013; Ce enini e al., 2013;Be endsene al.,2014)andi sin e ac ion wi h genomic back- g ound (Co ella and O do ás, 2014) and gu mic obio a emodeling (O a iani e al., 2011; Biagi e al., 2013; Collino e al., 2013). These ecen findings oge he wi h MARKAGE bioma ke s no only gi e a new pe spec i e in e m o ageing a e measu emen , bu also new insigh s o molecula - a ge ed in e en ions o slow down human ageing p ocess and likely, age- ela ed pa hologies onse . h p://dx.doi.o g/10.1016/j.mad.2015.03.010 0047-6374/© 2015 The Au ho s. Published by Else ie I eland L d. This is an open access a icle unde he CC BY-NC-ND license (h p://c ea i ecommons.o g/licenses/ by-nc-nd/4.0/). 14 M. Cap i e al. / Mechanisms o Ageing and De elopmen 151 (2015) 13–17 1. Ageing o Eu opean Coun ies The p opo ion o elde ly people o e 65 yea s in Eu ope (EU- 28Coun ies) is p edic ed o inc ease om 18.2% in 2013 o 28.7% by 2080 (Eu os a sou ce). The sha e o hose aged 80 yea s o abo e in he EU-28’s popula ion is p ojec ed o almos iple be ween 2013 and 2080. Thus, o e he coming decades, Eu ope’s demog aphic makeup will change ha shly. Ou popula ions a e becoming olde han e e be o e due o h ee majo ends. Fi s , he baby-boom gene a ion app oaches e i emen age hus olde people will ise apidly; second, bi h a es ha e emained low o se e al decades; and hi d, Eu opean people, bu no only, a e li ing longe li es, e en i como bidi es and disabili ies a e inc easing, especially a e 62 yea s (Eu os a sou ce). In 2011 he numbe o heal hy li e yea s a bi h was es ima ed a a ound 62 yea s o men and 61 yea s o women in EU-28, meaning ha app oxima ely, 16% and 21% o unheal hy li e o men and women, espec i ely, a e expec ed. This demog aphic emodelling emphasises he c i ical impo ance o iden i ying new s a egies able o coun e ac o slow down age- ing and he onse o age- ela ed diseases and disabili ies. These new s a egies can con ibu e o inc ease he numbe o elde ly ci izens in good heal h, and educing age- ela ed medical and social cos s. The e o e he iden ifica ion o bioma ke s o heal hy o unheal hy ageing (see Bü kle e al., his issue) and he adop ion o heal hy li e s yles be o e he use o d ug-based he apies a e necessa y. This si ua ion has pushed EU go e nance o iden i y scien ific p ojec s able o find solu ions o moni o and coun e ac unheal hy ageing, hus p omo ing a heal hy and ac i e ageing. 2. F om popula ion o indi idual le el On he o he side, he a e o ageing in humans is no uni o m due o he complexi y o gene ics and epigene ics (Cap i e al., 2006a, 2014), which in e ac wi h en i onmen (Biagi e al., 2013; Ga m e al., 2013) and s ochas ici y wi h di e se weigh s a di - e en phases o li e aking in o accoun also emb yonic and oe al de elopmen (Ce enini e al., 2010). Fu he , hese main ac o s could di e en ly a ec he a e o ageing a he le els o cells, is- sues o body sys ems wi hin he same o ganism acco ding o he hypo hesis o he “mosaic o ageing” (Ce enini e al., 2008). Tissues and o gans migh age a di e en a e and u he mo e, indi idu- als o same ch onological age migh ha e di e en biological age. This complexi y makes mo e di ficul he iden ifica ion o a unique comp ehensi e mechanism o ageing and ela ed bioma ke s (Deelen e al., 2013). 3. The human model in MARK-AGE: inno a i e concep s MARK-AGE aimed a he iden ifica ion o bioma ke s o age- ing capable o dis inguishing be ween ch onological and biological ageing (see Giampie i e al., his issue) looking a sys emic pa am- e e s assessed in he blood/u ine and buccal mucosa cells (BMC) o olun ee s, likely mi o ing he en i e o ganism. To achie e his objec i e a obus human model wi h defini e assump ions was concep ualised acco dingly (see Bü kle e al., his issue), as b iefly desc ibed: 1. Subjec s ep esen ing he “no mal” aging: andomly ec ui ed age-s a ified indi iduals om he gene al popula ion o RASIG, co e ing he age ange 35–74 yea s; 2. Subjec s ep esen ing he success ul o “decele a e” aging: sub- jec s bo n om a long-li ing pa en belonging o a amily wi h long li ing sibling(s) al eady ec ui ed in he amewo k o he GEHA -Gene ic o Heal hy Ageing- p ojec (Sky he e al., 2011). These indi iduals (“GEHA o sp ing” o GO) we e ec ui ed oge he wi h hei spouses o SGO ep esen ing he bes con ol o e alua e possible li e s yle e ec s, since hey ha e sha ed he same en i onmen al ac o s o many yea s wi h hei pa ne s. 3. Subjec s ep esen ing accele a ed aging: pa ien s wi h p oge oid synd omes (Cockayne, We ne and Down synd omes), cha ac- e ised by accele a ed “segmen al” ageing. 4. Independen a iables in MARK-AGE MARK-AGE model ook in o accoun essen ial independen a i- ables, i.e. gende , age, geog aphy and popula ions. The selec ion o hese a iables was done on he basis o specific assump ions such as he majo ole o gende in he ageing p ocess, he need o iden i y bioma ke s be o e he la e phase o li e and he ole o en i onmen /geog aphy/cul u e in he popula ion ageing wi h a consis en numbe o indi iduals.). Abou 5% o ec ui ed ol- un ee s we e no classified as “success ully ec ui ed” MARK-AGE indi idual, ei he because exclusion c i e ia such as posi i e esul s o HBV, HCV i uses sc eening o due o e o s in da a en y in o he pheno ypic da abase. The a GENDER. Males and emales we e ec ui ed close o 50% in each age class, gi ing he possibili y o analyse age-gende e ec s. Li e a u e s ongly sugges s a gende e ec on mo ali y a e. Females li e longe e en wi h a highe equency o disabili- ies and como bidi ies (Oksuzyan e al., 2008). This di e ence be ween males and emales seems o be a pa adox and he bio- logical causes a e no ye unde s ood, bu many da a sugges he in ol emen o sexual ch omosomes and hei age-dependen me hyla ion pa e n (Cap i e al., 2014; Gen ilini e al., 2012, 2013). Fu he mo e, p ospec demog aphic s udies show ha he gap o li e expec ancy be ween gende s will become smalle in he nex 40 yea s, e en i an analysis compa ing heal hy li e yea s be ween he gende a he age o 65 shows he e we e 9 EU Mem- be S a es in 2011 whe e men could expec mo e heal h li e yea s han women (Eu os a sou ce). b AGE. An age ange be ween 34 and 75 yea s was hough o be c ucial o iden i y ea ly bioma ke s o ageing. Humans unde go di e en phases o li e and 15 yea s beyond 50–60 yea s, i.e. he ansi ion be ween adul age and he beginning o he consis en ageing p ocess appea o be a c i ical empo al window. Indeed, Table 1 Indi iduals classified as “success ully ec ui ed” RASIG, GO, SGO and DS olun ee s finally inse ed in he da abase and conside ed o MARK-AGE analysis. Cen es RASIGM RASIGF GOM GOF SGOM SGOF DSM DSF MissedCodes Sub o al BioTeSys 162 188 8 358 FUNDP 121 135 33 42 17 21 8 377 LUMC 52 68 58 44 0 222 NENCKI 192 189 25 51 30 17 2 506 NHRF 200 203 5 17 4 2 1 432 UIBK 200 190 0 390 UNIBO 193 199 52 42 22 33 27 20 0 588 UTA 27 63 53 88 24 33 8 296 To al 1095 1167 220 308 155 150 27 20 27 3169 M. Cap i e al. / Mechanisms o Ageing and De elopmen 151 (2015) 13–17 15 Table 2 S a egies o MARKAGE dissemina ion applied by all he cen es in ol ed in he ec ui men . Cen es Popula ion Dissemina ion and s a egy o ec ui men BioTeSys GmbH (DE) RASIG Newspape a icles, in o ma ion e ening a he own hall oge he wi h he go e ning mayo ; egis a ion o fice (le e /flye ), olun ee s known om o he ials conduc ed a BioTeSys a li le wo d-o -mou h ecommenda ion. FUNDP/S a iCELL (BE) RASIG/GO/SGO Con ac ed an open-uni e si y o pe sons belonging o he 3 d age and all socie al ho izons; he Se ices o he Human Resou ces o he Ci y o Namu , he Uni . o Namu , Uni . Clinics o Mon -Godinne, and Regional Hospi al Cen e, dealing wi h all so s o pe sonnel; o ganised p ess con e ence a Uni . o Namu (many p ess a icles, in e iews on na ional adios, local TV news, na ional TV p og amme on ageing). GEHA e e ence o GO lis LUMC (NL) GO/SGO D a ed a lis wi h a numbe o picked nomina i es (GEHA e e ence). NHRF (GR) RASIG/GO/SGO Con ac by email all he pe sonnel o Resea ch Ins i u es o A hens. i. Con ac by phone, call all he GEHA siblings gi ing in o ma ion o GO/SGO on MARK-AGE, sending hem he in o ma i e shee by pos o ax. ii. Con ac by phone, call all ou pe sonal acquain ances gi ing in o ma ion on MARK-AGE NENCKI (PL) RASIG/GO/SGO Ob ained add esses o 3200 RASIG om Minis y o In e io and Adminis a ion (p esen ly Minis y o In e io ) acco ding o PESEL (Na ional Elec onic Census Numbe Sys em); sen 1700 le e s o in i a ion, he esponde s eedback by phone o e-mail (22%). GEHA e e ence o GO lis . UIBK (AT) RASIG A icles in a e y common Ty olean daily newspape ; dissemina ion on local TV news and a icles in o he newspape s and magazines. UNIBO (IT) RASIG/GO/SGO/DS GO/SGO we e ec ui ed be o e RASIG: d a ed a lis wi h a numbe o picked nomina i es. RASIG: popula ion o PIANORO (17,000 inhabi an s) nea Bologna; con ac s wi h he Mayo Ci izen and he Dis ic o Public Heal h. A p esen a ion o he p ojec was pe o med o all he popula ion wi h local Go e nmen and gene al p ac i ione s; local pape , in o ma ion flye s; magazine associa ed o ood discoun s) and TV, ec ui ed also pe sons om he Uni e si y o Bologna. DS and hei amily we e con ac ed by specific associa ion in Bologna UTA (FL) RASIG/GO/SGO Newspape s, local TV announcemen s his pe iod includes he menopause in women and he pe iod whe e majo age ela ed diseases and he a e o mo ali y s a o inc ease (Rause e al., 2006). Acco dingly, one o he majo cha ac e is ic o MARK-AGE is o ocus on he abo e men ioned age ange in o de o iden i y ea ly bioma ke s o biological s. ch onological age, po en ially capable o p edic ing he a e o ageing la e in li e. c GEOGRAPHY. 11 di e en Eu opean beneficia ies, dis ibu ed om No h o Sou h, we e esponsible o he ec ui men . This dis ibu ion has gi en he possibili y o s udy en i onmen- al/geog aphical/cul u al popula ion e ec s. Thus, geog aphical o en i onmen al ac o s could influence pa ame e s, such as me hyla ion pa e ns (Pi azzini e al., 2012), bu di e ences among di e se la i udes can be e ealed and ad hoc analysed. d POPULATIONS. One o he main goal o MARK-AGE was o ec ui consis en g oups o subjec s assumed o ha e di e se a es o ageing, as desc ibed abo e. On he whole 3337 subjec s we e ini ially ec ui ed and he ec ui men o GO/SGO was he mos challenging due o la ge dis ances be ween subjec s esidence and labo a o y place, hus i was mo e ime consuming, mo e expensi e and mo e e o needed o ulfil equi emen s o anspo o biological samples. Fu he mo e, he numbe o SGO was abou hal o GO since somebody declined, di o ced o passed away. 53 Down Synd ome indi iduals we e ec ui ed by UNIBO wi h a specific s a egy commi ed o amily in ol emen and he employmen o specialized human esou ces, such as psychologis s and psychia is s. Fu he a specific ba e y o es s ocused on beha iou , neu opsychological, neu opsychia ic and cogni i e assessmen was se up (see Mo eno-Villanue a, Cap i e al., his issue) in o de o e alua e he unc ional and cogni i e s a us o hese people. DNage B.V. and la e on Uni e si y o Con- s ance (UKON, Ge many) we e esponsible o collec ion o abou 59 exis ing blood samples om WS pa ien s and a mino num- be o CS pa ien s ec ui ed by ex e nal collabo a o s. In iew o he complexi y and he size o he MARK-AGE p ojec , d opou s we e expec ed. Ne e heless, abou 95% o ec ui ed subjec s (i.e. 3169) could be included in he MARK-AGE popula ion o anal- ysis (Table 1 final numbe s o success ully ec ui ed olun ee s and also inse ed in he MARK-AGE da abase we e he ollowing: 2262 RASIG; 528 GO; 305 SGO and 47 DS as shown in Table 1 (59 WS pa ien s samples a e no included). Technical aspec s a e b iefly epo ed in Box I and Table 2. All p ocedu es o da a base managemen and cleaning a e desc ibed (see Bau e al., his issue). Box I: P ac ical aspec s o MARK-AGE. - MARK-AGE RASIG popula ion was ec ui ed by BioTeSys GmbH (Ge many), Uni e si y o Namu (FUNDP, Belgium), Ös e e- ichische Akademie de Wissenscha en (UIBK, Aus ia), Ins i u e o Expe imen al Biology (NENCKI, Poland), Na ional Hellenic Resea ch Founda ion Cen e (NHRF, G eece), Uni e si y o Tam- pe e (UTA, Finland) and Uni e si y o Bologna ALMA MATER STUDIORUM (UNIBO, I aly). Subjec s om GO/SGO coho s we e ec ui ed by Uni e si y o Leiden (LUMC, Ne he land), Uni e si y o Tampe e (UTA, Finland), NENCKI, UNIBO, FUNDP and NHRF. UNIBO addi ionally ec ui ed DS pa ien s, DNage B.V. (Ne he land) and Uni e si y o Cons ance (UKON, Ge many) we e esponsi- ble o collec ion o exis ing blood samples om CS and WS pa ien s ec ui ed by ex e nal collabo a o s. A specific ques- ionnai e was de eloped o RASIG/GO/SGO popula ion, which was adap ed by UNIBO o he DS pa ien s. All he ques ion- nai es and biological samples ha e been desc ibed in he a icle dedica ed o MARK-AGE s anda d ope a ing p ocedu es (see Mo eno-Villanue a, Cap i e al., his issue). Rec ui men s a egies a e de ailed in Table 1. Reasonably, GO/SGO ec ui men was ca - ied ou by Beneficia ies who we e in ol ed in GEHA p ojec since hey had al eady con ac ed hese amilies be o e (F anceschi e al., 2007). - Inclusion and exclusion c i e ia RASIG we e andomly ec ui ed age-s a ified indi iduals om he gene al popula ion (bo h sexes) aged 35–74 and able o gi e in o med consen . Conce ning GO sub- jec s, hey we e sons o daugh e s o GEHA indi iduals and SGO we e GO spouses in age- ange 55–74 yea s. As well as RASIG, all o hem should be able o gi e hei in o med consen . Exclu- sion c i e ia we e he ollowing: i. sel - epo ed se oposi i i y o HIV, HBV (excep se oposi i i y by accina ion) and HCV (HBV and HCV se oposi i i y assessed a e blood collec ion). ii. p esence o ac ual cance and cu en use o an i-cance d ugs o glucoco i- coids; iii. less han 50% o li e ime spen in coun y o esidence. i . inabili y o gi e in o med consen . 5. Robus da a, longi udinal componen and p og ess MARK-AGE was a g ea oppo uni y o ad ancemen in knowl- edge. To asce ain he biological and analy ical obus ness o he measu emen s o candida e bioma ke s, 100 dono s om he whole s udy popula ion we e e-sampled wi hin 3–6 mon hs. Fu - he , wi hin he li e ime o he p ojec (66 mon hs), a limi ed (12%) andom sample o p obands we e e- es ed in o de o es ablish 16 M. Cap i e al. / Mechanisms o Ageing and De elopmen 151 (2015) 13–17 Fig. 1. Ch onological s biological age. Dis ances om bisec o (Y = X) a e only indica i e. Basic assump ion o MARK-AGE highligh s he possibili y o iden i y sys emic bioma ke s which co ela e wi h age and significan ly sepa a e RASIG, GO and DS popula ions o di e en age a es. RASIG: Randomly ec ui ed Age-S a ified Indi iduals om he gene al popula ion (age ange: 34–75 yea s); GO: GEHA o sp ing (age ange: 54–75 yea s); DS: Down synd ome (age ange: 19–68 yea s) a longi udinal componen wi hin he s udy. To make s onges he esul s eme ging om MARK-AGE i would be impo an o pe o m a mo e obus longi udinal s udy on all he ec ui ed subjec s. Ne - e heless, hose pa ame e s, which co ela e wi h age and hose, which significan ly sepa a e coho s o di e en age a es a e hose able o moni o he sys emic age- ela ed changes o he body. Fig. 1 depic s he concep s which MARK-AGE aced and he basic ma he- ma ical app oach om which algo i hms ha e been de eloped. The s onges MARK-AGE bioma ke s and hose ela ed o he epigene ic clock (353CpG si es shaping an ageing clock in e ms o ch oma in s a es and issue a iance), ecen ly aised up om s udies on di e en ly aged human issues, human obesi y and Down Synd ome (Ho a h, 2013; Ho a h e al., 2014, 2015), e eal powe ul issue -specific bioma ke s mi o ing he sys emic con- di ion. These findings open new ques ions on he u u e ole o he bioma ke s ne wo k which suppo s in o ma ion o heal hy s a us and likely could also “ anspo communica ion” among di e en cell, o gans o issues. Thus, new insigh s a e expec ed by bioma k- e s ne wo k, no only highligh ing issue-specific and/o sys emic molecula pa hways, bu also moni o ing he issue-specific and/o sys emic a e o ageing, bo h a indi idual and popula ion le els, and e en ually a ge ing genes o hei p oduc s o age- ela ed pa hology in e en ions. Acknowledgemen s We wish o hank he Eu opean Commission o financial sup- po h ough he FP7 la ge-scale in eg a ing p ojec “Eu opean S udy o Es ablish Bioma ke s o Human Ageing” (MARK-AGE; g an ag eemen no.: 200880). We a e g a e ul o D . Lau a Celani, D . Anna Lau a Ba bie i, D . Claudia Be a elli, D . C is ina Fabb i, Massimo Izzi oge he wi h all he UNIBO eam o hei essen ial con ibu ion on olun ee s’ ec ui men and labo a o y p ocedu es in Bologna (I aly). We would like o hank he BioTeSys eam Jen- ni e Schweika , Simone Sie e , Annabel S ie lin, Ka ha ina Halle and Am ei Ge ull o hei e o in ec ui men managemen and documen a ion as well as sample p ocessing. O. Toussain and F. Debacq-Chainiaux a e, espec i ely Senio Resea ch Associa e and Resea ch Associa e o he FNRS, Belgium and acknowledge he Région Wallonne o he Ne wo k II Senegene I and Senegene p ojec s II, he Pole o Excellence Nano oxico p ojec # 516252, and Complemen ‘QNano’ p ojec # 1117448. We a e g a e ul o D . Do o a Janiszewska, D . Magdalena Dudkowska, Ms. Anna Ka pa, Ms. Ma a Wincenciak om he Nencki eam and all nu ses o hei con ibu ion on olun ee s’ ec ui men and labo a o y p oce- du es in Wa saw (Poland). We would like also o hank D . G azyna Mosieniak o he aluable inpu in he p ojec o ganisa ion. Las ly, we would like o hank all s udy pa icipan s who olun a ily o e ed hei ime and suppo o he benefi o he MARK-AGE p ojec . Re e ences Bacalini, M.G., F iso, S., Oli ie i, F., Pi azzini, C., Giuliani, C., Cap i, M., San o o, A., F anceschi, C., Ga agnani, P., 2014. P esen and u u e o an i-ageing epigene ic die s. Mech. Ageing De . 136–137, 101–115. Be endsen, A., San o o, A., Pini, E., Ce enini, E., Os an, R., Pie uszka, B., Rol , K., Cano, N., Caille, A., Lyon-Belgy, N., Fai wea he , S., Feskens, E., F anceschi, C., 2014. A pa allel andomised ial on he e ec o a heal h ul die on inflammageing and i s consequences in Eu opean elde ly people: design o he NU-AGE die a y in e en ion s udy. Mech. 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