immunosenescence in humans: deterioration or remodelling?

ISSN 0883-0185 IRIMEH 12(1) 1-84 (1995) Volume 12 Number 1, (1995)

INTERNATIONAL REVIEWS OF

IMMUNOLOGYEditar in Chìef

Constantin A. BonaMount Sinai School of Medicineof thè City University o/New York

The Reshaping of Gucst Editore

. Claudio Franceschithè Immune System Wlth Age Andrea Cossarizza

Introduction: The Reshaping of thè Immune System with AgeClaudio Franceschi and Andrea Cossarizza 1

Aging Associated Changes in thè Generation and Development of B CeliMemory

Norman R. Klinman and G. Jeanette Thorbecke 5

Changes in thè Intestìnal Lymphoid Compartment Throughout Life:Implications for thè Locai Generation of Intestina! T Cells

J. Rozing and B. de Geus 13*"i:

V(ì Gene Repertoire in thè Aging Mouse: A Developmental PerspectiveRosana Gonzalez-Quintial, Roberto Baccalà, Robert S. Balderas,and Argyrìos N. Theofilopouios 27

T Celi Repertoire Usage hi Hurnans, from Newborns to CentenariansAndrea Cossarizza, Daniela Barbieri, and Marco Londei 41

Immunosenescence in Humans: Deterioration or Remodellìng?Claudio Franceschi, Daniela Monti, Daniela Barbieri, Emanuela GrassUU,Leonardo Troiano, Stefano Salvioli, Paolo Negro, Miriam Capri, Marcella Guido,Roberto Azzi, Paolo Sansoni, Roberto Paganelli, Umberto Fagiolo,Giovannella Saggio, Sabrina Donazzan, Stefano Mariotti, Sergio D'Addato,Antonio Caddi, Claudio Ortolani, and Andrea Cossarizza 57

Immunoendocrìne Reshaping with AgeAlberto E. Panerai and Enzo Ottaviani 75

harwood academic publishers

Intera. Rev. Immurici. Voi. 12. 1995, pp. 57-74Reprims available directly from thè publisherPhotocopying permitted by license only

«1995 Harwood Academic Publishers GmbHPrìnted in Malaysia

Immunosenescence in Humans:Deterioration or Remodelling?CLAUDIO FRANCESCHI/ DANIELA MONTI/ DANIELA BARBIERI/EMANUELA GRASSILLI/ LEONARDA TROIANO/ STEFANO SALVIGLI/PAOLO NEGRO/ MIRIAM CAPRI/ MARCELLO GUIDO/ ROBERTO AZZI/PAOLO SANSONI/ ROBERTO PAG ANELLI,» UMBERTO FAGIOLO,1'GIOVANNELLA BAGGIO,' SABRINA DONAZZAN," STEFANO MARIOTTI/SERGIO D'ADDATO,0 ANTONIO CADDI,0 CLAUDIO ORTOLANI,*and ANDREA COSSARIZZAf

T Dipartì mento di Scienze Biomediche, Sezione di Patologia Generale, Università di Modena,Modena, ItalyIstituto dì Clinica Medica Generale e Terapia Medica, Università di Parma, Parma, Italy^Cattedra di Immunologia Clinica e Allergologia, Università di Roma "La Sapienza," Roma,Italy''Istituto di Medicina Interna, Università di Padova, Padova, Italy^Istituto di Endocrinologia, Metodologia Clinica e Medicina del Lavoro, Tirrenia-Pisa, Italy°Centro per lo Studio dell'Atcrosclerosi, Servizio di Gerontologia, Policlinico S- OrsolaMalpighi, Bologna, Italy'Ospedale SS. Giovanni e Paolo, Venezia, Italy

KEYWORDS: aging. centenarians, immunosenescence, autotmmunìly, NK cells, cytokines

AGING AND LONGEVITY: A THEORETICAL APPROACH

Aging has been studied extensively. Conversely, longevity, and particularly human longev-ity, has been neglected [I], Hundreds of theories are available on aging, indicating thatscientists are stili far from understanding thè biological and cultural basis of this process. Tothis long lisi of theories, we bave added a new one, based on thè consideration that thèmaintenance of soma integrity is thè consequence of a continuous activity of a limitednumber of cellular defense mechanisms [2-5]. We have hypothesized that DNA repair,enzymatic and non-enzymatic antioxidants, production of heat shock and stress proteins,and activity of po!y(ADP-ribose)polymerase form a network of interconnected cellulardefense mechanisms, whose global efficìency has been evolutionary set at different levels indifferent species and in different individuai of thè same species. We have also speculatedthat apoptosis is a fundamental biological process which canjoin thè listof cellular defensemechanisms, being an ancestra! process used to eliminate damaged, mutated, viral-infectedor transformed cells [6, 7]. On thè whole, thè above mentioned cellular defense mecha-nisms can be considered as thè basic molecular and cellular anti-aging systems.

Addressfor correspondence: Prof. Claudio Franceschi, Dipartimento di Scienze Biomediche, Sezionedi Patologia Generale, Università di Modena. via Campi 287, 41100 Modena, Italy. Tei.: +39 59360044; Fax: +3959362206.

57

58 C. FRANCESCHI et al.

However, it can be argued that aging is not simply a cellular mechanism but that itrepresents thè failure of more integrated systems whose purpose is to preserve bodyintegrity. The nervous, thè endocrine and thè immune systems are ali devoted to thèmaintenance of body homeostasis. On thè basis of phylogenetic studies, we suggested that aco-evolutìon of these systems probably occurred [8]. Accordingly, we and others arguedthat these three systems nave to be studied as a whole, and that thè more appropriateapproach is to consider immunoneuroendocrine cells and organs as part of a unique Systemdevoted to cope with ali kinds of internai and external stimuli and damaging agents [9, IO].In previous papers we have suggested that thè immunoneuroendocrine System relies uponthè above mentìoned molecular and cellular defense mechanisms for its continuous activity[3]. This point of view represents a tentative to combine molecular and cellular withsystemic theories of aging. It can be predicted that thè optimal functioning of thisimmunoneuroendocrine System is of major importance for survival, aging and longevity.Accordingly, as far as human longevity is concerned, we hypothesized that people whosurvived in good conditions for long periods, dose to thè maximum life span of our species,should be equipped with an optimal immunoneuroendocrine System [3]. For this reason, aresearch started some years ago on thè biological basis of human longevity, in a selectedgroup of healthy centenanans. We will report here some of thè data collected in thè last fewyears on thè immune System of centenarians. These data are thè first part of a broaderinvestigation in which immune and neuroendocrine parameters will be analyzed in order tounderstand some of thè molecular and cellular basis of human longevity. A genetic searchfor longevity assurance genes is also in progress.

AN OLD TENET: IMMUNOSENESCENCE = LMMUNODETERIORATION

The aging of thè immune System has been thoroughly studied in a variety of models and,particularly, in rodents and humans.

The result of this intense investigation can be summarized with thè equation immuno-senescence — immunodeterioration [11]. Increased sensitivity to infectious diseases andcancer, decreased antibody production to non-self antigens and increased levels of autoanti-bodies, defective NK activity and decreased T lymphocyte proliferation have been consid-ered a paradigm of a defective immune responsi vene ss with age. Moreover, it was alsoassumed that most of this age-related immune deterioration has to be ascribed to profoundand early changes of thymus, whose involution starts immediately after puberty [12]. As aconsequence, another popular tenet in immunogerontology is that thè cellular, or Tlymphocyte branch, suffers because of age more than thè humoral, or B lymphocyte,branch. This scenario is mainly based on data obtained in rodents. In this review, we will tryto demonstrate that in humans thè deterioration of thè immune system with age is not asdramatic as that reported in experimental animals, Moreover, we will review data suggest-ing that thè lime is mature to challenge thè above mendoned tenet.

IMMUNOGERONTOLOGICAL BIAS

We and others have been interested in human immunosenescence. Some years ago, Dutchimmunologists suggested that strict biochemical and clinical inclusion and exclusioncriteria, known as SENIEUR Protocol, should be adopted in studies of immunosenescence

(13, 14].thè confemergedinvestiganot as aimedicinebut alsoimmuneatheroscK

In correlderly hiyears. Thand thatparticularThus, thèlogically.

Anothepeople areof "old" fage is o verthat age, i ipossible a;age-relatei-newbornsage-relatetthyroid [1?first and ti

Humancriteria formany repoabsolute niwith age isignificane

CENTEN,

Forali thes<-thè last dee,cai immunewill refer temental (e.eGenerally, e

First, thèsurviving leItaly, prelinLuciano Methan 4,000.

bui that it;rve body[ed to thè•sted that a^rs arguedppropriate[uè systemits[9,10].elies upon>us activityllular withng of this[ongevity.

eople whourspecies,s reason, aa selected

:he last few" a broader. in order toletìc search

NON

nodels and,

m immuno-iiseases andof autoahtì->een consid-

it was alsoto profoundty[ I2] .Asallular, or T.ymphocyte,/, we will tryjge is not asJata suggest-

IMMUNOSENESCENCE IN HUMANS 59

rs ago, Dutchnd exclusionnosenescence

[13,14]. This proposai was aimed at avoiding a classica! bias in gerontological studies, i.e.thè confusion between aging and age-related diseases. Indeed, an entirely new scenarioemerged when thè immune system of carefully selected healthy, elderly people wasinvestigated. A variety of immune parameters and responses measured in those people werenot as altered as previously thought [15]. This approach is a great challenge for bio-medicine, owing to thè importance of thè immune system for immunological pathologiesbut also for other diseases not traditìonally included among immune diseases, in whichimmune and inflammatory responses can play an important or even cruciai role (e.g.,atherosclerosis, dementia, cancer).

In comparison with thè enormous literature on immunosenescence in rodents and inelderly humans, only scanty and anecdotal data are available on people older than 80-85years. This is strange ifweconsider that human Ufespan is much longer, i.e. 110-120 years,and that thè number of old people is dramatically increasing in ali countries, andparticularly in those that are economically (and immunologically!) developed [16, 17].Thus, thè last three to four decades of human lìfe have been left unexplored immuno-logically.

Another possible bias in human immunogerontology is that in most studies two groups ofpeople are compared, i.e. agroupof "young" people.usually 20-30 years old, andagroupof "old" people. This second group is usually composed of subjects whose chronologicalage is over 60-65 years. Apart that not ali gerontologists would agree that aging starts afterthat age, it appears that it is difficult, with such an approach, to fully appreciate thè trend ofpossible age-related modifications. Indeed, we found that, in order to follow and understandage-related changes in immune parameters, one has to study human subjects fromnewborns to centenarians. We found that centenarians are extremely useful to appreciateage-related changes in immune system and other organs and functions suchas, forexample,thyroid [18, 19]. This approach avoids thè bias of assessing aging without considering thèfirst and thè last 2-3 decades of human life, for a total of about half a century.

Human aging is a slow process, and it is difficult to choose appropriate and reliablecriteria for assessing it. Finally, thè literature on immunosenescence is confused since, inmany reports, data referring to lymphocyte subsets are presented as percentages but not asabsolute numbers [20]. These inconsistencies can create thè illusion that nothing changeswith age (e.g., thè percentage of CD3 + cells) whereas upon closer examination, asignificant decrease in these cells does occur [21].

CENTENARIANS ARE EXCEPTIONAL INDIVIDUAI^

Fora!! these reasons (1. selection and healthy status of thè subjects; 2. necessity of exploringthè last decades of human life; 3. importance of successful aging to appreciate physiotogi-cal immunosenescence) we began to investigate thè immune system of centenarians. Wewill refer to centenarians as people who are not only older than 100 years, but also in goodmental (e.g., practicing pharmacist) and physìcal condition (able to chop firewood!).Generally. centenarians are considered to be a rare curiosity. However, this is not thè case.

First, their number is increasing dramatically, and, according to recent predictions, thosesurviving longer than 95 or ÌOO years will soon represent a consistent group [16, 17]. InItaly, preliminary data based on a nationwide investigation coordinated by ProfessorLuciano Motta (University of Catania) indicate that thè cenienarians are nowadays morethan 4,000. About one thìrd of them are in relatively good mental and physical condition,


and can be considered "healthy" centenarians. We think that thè SENIEUR Protocol,proposed by Ligthart et al. for elderly people [13, 14], should be revisited to be applied topeople over 100 years. We are elaborating such a protocol, to be adapted to healthycentenarians, within thè framework of thè Concerted Action Programme on MolecularGerontology of thè EU BioMed Research Programme.

Second, healthy centenarians are thè best example of successful aging, narnely peoplewho have escaped major age-related diseases and reached thè extreme limit of human life ingood clinical conditions [22]. In most cases, histories of these exceptional individualareveal them to be free of cancer, dementia, diabetes, cardiovascular diseases, cataract.Moreover, as discussed above, in orderto reach such an advanced age, centenarians shouldbe equipped with well preserved and effìcient immuno- and defense mechanisms andoptimal combinations of an appropriate Hfestyle and genetic background [3, 23]. Thus, thèstudy of centenarians, and particularly that of healthy centenarians is not only of broadbiologica! and medicai interest, but can help in identifying genes that prevent thè abovementioned age-related diseases.

C<ntEcc

HUMORAL LMMUNITY AND SUCCESSFUL IMMUNOSENESCENCE:INCREASE IN IMMUNOGLOBULINS, DECREASE IN B CELLS,LACK OF ORGAN-SPECIFIC AUTOANTIBODIES

As far as humoral immunity ìs concerned, thè tenet that aging = deterioration is based onthè observation that there is an increased frequency with age of pathological processesinvolving B cells and antibody production, such as B chronic lymphocytic leukemia(B-CLL), presence of autoantibodies or monoclonal gammopathies, amyloidogenesis.Moreover, thè decreased antibody response may also result in a propensity for infectiousdiseases, particularly pneumonia, or recurrent infections, as well as poor responses tovaccinations against thè causative agents, resulting on thè whole in an increased morbidityand mortality in elderly subjects. Most of thè data on this topic have been collected beforethè "revolution" of immunosenescence caused by thè use of SENIEUR Protocol inimmunogerontological studìes. For this reason, we began thè analysis of humoral immunityin healthy elderly people, including centenarians. First of ali, we faced an unpredictedparadox concerning humoral immunity with age, i.e. an increase of immunoglobulin (Ig)serum level and a concomitant decrease in peripheral blood B lymphocytes (Fig. I)[18,21,24].

Most studies in thè past two decades have addressed thè assay of Ig classes and subclassesin sera from human aged subjects, in attempts to establish thè norma! range for laboratorypurposes, as well as to investigate thè physiology of this classìcal parameter of humoralimmunity with aging. By analyzing 87 sera of healthy subjects belonging to severa! agegroups carefully selected according to thè established criteria of thè SENIEUR Protocol,and including a group of healthy centenarians, we found that both IgG and IgA serum levelssignificantly rise in correlation with age, whereas IgM levels remain unchanged [24].Moreover, among IgG subclasses, we observed that IgG 1, 2, and 3 showed a significantincrease, whereas IgG4 did not (Fig. 2) [24]. An increased in vitro Ig production by B cellsfrom aged people has been previously reported [25]. It is interesting to remind that IgGl and3 are mainly involved in thè humoral responses to virai and bacterial antigens, IgG2-~together with IgM—are responsive for responses to polysaccharides (mainly outer walJantigens of capsulated bacteria), and IgG4, with IgE, are increased in response .to

FIGUR.subclas.-reportec

parasi!exposiagainsnotewt.except

Dee25]. DdecreaincreasCDI9-of thesgroupsmoleciilymphccytic leunreco-

The:WÌlh 3.Ì.

particu!detecta1

with agThis

practicathis is r

iUR Protocol,3 be applied toted to healthyon Molecular

namely peopleif humanlifeinlai individualsases, cataract.narians shouldxhanisms and23].Thus, thèonly of broad

vent thè above


40

E 30

D

cc<octmcc

20

10

IMMUNOGLOBULINS

B LYMPHOCYTES

O 10 20 30 40 50 60 70 80 90 100 110

4CE:

ion is based on;ical processes;ytic leukemia.yloidogenesis.' for infectiousr responses toased morbidityollected beforeR Protocol inioral immunìtyin unpredictedloglobulìn (Ig)icytes (Fig. 1)

and subciassesfor laboratory

-•ter of humoralto several age

EUR Protocol,:A scruni levelsichanged [24].ed a significantction by B cellsid that IgGl andtigens, IgG2—tinly outer wallin re spense to

AGE (YEARS)FIGURE 1 The first paradox of ìmmunosenescence is shown: serum leve! of immunoglobulin ciasscs andsubciasses increases with age, whik thè absolute numberof circulating B iymphocytes decreases (based on datareported in Paganelli et al., 1992; Sansoni et al., 1953).

parasite antigens, as well as beìng thè "memory" isotype in conditionsof chronic high doseexposure [26]. This increase of IgG and Ig A antibodies may afford greater protectionagainst virai and bacterial infections in healthy old aged people and centenarians. It is alsonoteworthy that very few IgG subclass defects were found in elderly subjects, with thèexception of IgG4 deficiency [24].

Decreased numbers of Iymphocytes are found in selected healthy elderly subjects [21,25]. Despite small changes in percentages of lymphocyte subsets, ali are significantlydecreased as absolute numbers [21]. B Iymphocytes in centenarians were reported to beìncreased, although not significantly [27]. In centrasi, we observed a striking decrease ofCD19+ B cells both in thè 70-83-year age group, and in centenarians (24). The percentageof these cells also decreased, from 13.5% in young controls, to 9.4% and 3.2% in thè twogroups of aged people. Further, we found that thè CD19+ cells coexpressing thè CD5moìecule also decreased with age [18, 24]. These cells represent a distinct subset of BIymphocytes able to produce polyreactive autoantibodies, and originate chronic lympho-cytic leukemia of thè B celi type. This finding confirmed that we did not select subjects withunrecognized B-CLL, which is common in elderly subjects.

The scenario regarding thè changes in humoral immunity which occur in elderly subjectswith age is even more complex if we consider thè problem of autoimmunity, and, inparticular, that of autoantibodies. It has been reported that thè frequency of subjects withdetectable serum levels of organ-specific or non-organ-specific autoantibodies increaseswith age.

This tenet was challenged by our observation that organ-specific autoaniibodies arepractically absent in thè plasma of healthy centenarians [18]. We subsequently showed thatthis is not a peculiar characteristic of centenarians, as thè absence of autoantibodies also


3000

|2000-

1000-

coHz

CCbmcc

O 10 20 30 40 50 60 70 80 90 100 110

|

••• ~~m i9—- _ . -* . mm °*.

10 20 30 40 50 60 70 80 90 1 00 110

AGE (YEARS)FIGURE 2 A detailed analysis of thè data conccrning Ig plasma levels reveals that IgG, IgA, and IgGl. IgG2,IgG3 significanti increase with agc (based on data reported in Paganelli et al., 1992).

occurs in healthy old people [19]. In centrasi, unselecied, eldedy people presented an age-related increase in ihese autoantibodies (Fig. 3) [18,19]. Non organ-specific auloantibodiesfollow a different trend, increasing also in healthy aged donors, including centenarians(manuscript in preparation). - - ' • ' '

These data on humoral immunily of elderly subjects raise severa! questions, and differentpossibilities can be envisaged to explaìn these age-related changes. In particular, il is

FIGURE 3semm levelunselected aon data rcp-

possible torgans othgerminai ihas to be rhoming alaboratonprocessespropensi!;damaging

CELLULPRESENSUBJECSINCE F

Unexpect'what wesubjects.others ineCD8+T'were mar!destiny.

Anothe

O 100 110

'O 100 110

gA.andIgGl.IgG2.

presented an age-iìc autoantibodiesiing centenanans

ons, anddifferenla particular. il is


OT

ZD>CC<CC

mcc

10 20 30 40 50 60 70 80 90 100 110

AGE (YEARS)FIGURE 3 The importance for immunogerontology of studying strictly selecied heaJrhy subjects is shown: thèsenim level of organ-specific autoantibodies (antì-cnyroid, anti-pancreas, etc.) increases with age, but only inunselected aged people. Heaithy clderly and centenarians do not show any increase of these autoantibodies (basedon data reported in Mariotti et al., 1992 and 1993).

possible to hypothesize that increased number of B lymphocytes and plasma cells occurs inorgans other than periphera! blood, or that thè lifespan of B lymphocyles and plasma cells ingerminai centers is increased in aged people. Finally, an increased production of Ig per celihas to be ruled out. According to first and second hypotheses, alterations with age of B celihomìng and propensity to apoptosis can be predicted. Indeed, recent data from ourlaboratory suggest that thè membrane expression of certain molecules involved in homìngprocesses and of certain celi adhesion molecules changes with age [28]. A differentpropensity to apoptosis has been observed in PBL from centenarians after exposure todamaging virai or chemical agents (manuscript in preparation).

CELLULAR IMMUNITY AND SUCCESSFUL IMMUNOSENESCENCE:PRESENCE OF A WELL-PRESERVED NUMBER OF T CELLS IN ELDERLYSUBJECTS AND CENTENARIANS, DESPITE A THYMUS INVOLUTINGSINCE PUBERTY

Unexpectedly, age-related changes in thè T celi compartment were much less dramatic thanwhat we would have predicted according to thè data in rodents and in unselected agedsubjects. Moreover, it is noteworthy that some parameters were found decreased, whileothers increased. In particular, we found that thè absolute number of CD3 + , CD4+ andCD8+ T cells decreased with age [21, 29], while activated peripheral T cells [HLA-DR+)were markedly augmented (Figs. 4 and 5). In both cases centenarians did not escape thisdestiny.

Another interesting trend has been recorded as far as percentage and absolute number of


4000

3000

UJO

i 2000-

DCUJffl

D 1000

o C03+• CD4+

p<0.01

O 10 20 30 40 50 60 70 80 90 100 110

AGE (YEARS)

FIGURE 4 Age-re!ated changes in thè absolute number of circulating T lymphocytes (CD3 -I-), and of one majorTcellsubset (CD4+). When a large groupof subjects ofdifferent age. from newborn tocentenanans. ìs smdied,thè age-related Ircnd of an immunologicai parameter emerges. In this figure, it appears that thè number of Tlymphocytes significanlly decrease with age. However, it ìs important to note that thè number of T lymphocytes insome centenarians Ìs higher than that found in some young subjects. Because of this individuai variability mostimmune paramcters canno! be laken as reliable biomarker of immunosenescence (based on data reported inSansoni et al., 1993).

1500

UJO

U.occUJm2DZ

1200

900

600"

300'

0 C08+• CD19+

jxO.01

p<0-01

90 ioo no;

AGE (YEARS) ';

FIGURE 5 Age-rclated changes in thè absotuie number of circutatingT (CDS+) and B lymphocyta (CJJj?Ì>The age-related decrease of B cells is more dramatic than thal of T cells. Indeed, B cells ire tboutJOOjWfechildrcn and young subjects. and about 30-40/u.l in centenarians, while, for «ampie. CDS + T cèlb tre wcnt900/M-I in young subjects and about 450/^1 in centenarians (based on data reported in Sansoni et al., 1993).

"virgin/unprircerned:theseIndeed, a dra.years, with a estili showed a

Thus, thè fcsharp changesdecades of liftthat take piacelifespan of seunderstand or;thè thymus hadifficult toexfpossible new iother have beeis required forthymus becorrtaken over bynegative anddemonstratedremnants or sucells every da.

CELL DIVISBUI LONGE

Aging is chartLossof prolif;Hayflick pheiundergo. It icapability ancproliferatìve emay be consi<

In a recensenescence, tprogrammedequìpped witfin order and eresponsible fsenescence nfunction is u.cellular senesthè continuoineoplastic gnMoreover, w.between celi irecently founing age of ti


o CD3+• CD4+

100 110

CD3 + ), and of one major) centenarians, is studied,rars that thè number of Tmber of T lymphocyies inndividual vahability most>ased on data reported in

o CD8+• CD19+

o o

io fxO.01

p<0.01

)0 100 110

J B lymphocytes (CD19+).! cclls are about 300/jil in:. CD8 + T cells are iboutin Sansoni et al., 1993).

"virgin/unprimed" (CD45RA+) and "memory/activated" T cells (CD45R04-) was con-cerned: these parameters did not change signìficantly after thè fourth decade of iife [30].Indeed, a dramatic decrease in CD45RA-H T cells occurred from birth to thè age of 30years, with aconcomitant increase in CD45RO+ lymphocytes. Old people and centenariansstili showed a consistent number of either CD4-H or CD8+ virgin T cells (Fig. 6) [30].

Thus, thè following questions arise: Where do these T lymphocytes come from? Why aresharp changes in expression of CD45 isoforms mainly observed in thè first two to threedecades of Iife? These modifications seem to mirror thè invotutive changes of thè thymus,that take piace immediately after puberty [31], i.e. at least 80 years ago. Assuming that aHfespan of several decades is highly improbable for most memory T cells, we need tounderstand origin and continuous renewal of virgin and memory T celi compartments whenthè thymus has likely undergone a profound involution, as in centenarians. In any case, it isdifficult to explain why a consistent number of T cells shows a virgin phenotype, waiting forpossiblenew antigens, even in far advancedage. Indeed, reversions from one isoform to thèother have been described [32-35], and it has been suggested that thè presence of antigensis requìred for "memory" cells to survive [36-38], Thus, thè possibility exists that thèthymus becomes progressively less important as thè T lymphocyte producer, and its rote istaken over by other "peripheral" lymphoid organ(s) [39]. We must assume therefore thatnegative and positive selection are occumng successfully in this (these) organ(s), asdemonstrated by a lack of autoimmune responses in healthy centenarians. Thus, thymicremnants or substituting organ(s) are probably able to produce and select high numbers of Tcells every day, until thè extreme limit of human Iife.

CELL DIVISION IN CENTENARIANS: SLIGHTLY DELAYEDBUI LONGER LASTING

Aging is characterized by a variety of alterations which occur in most organs and celi types.Loss of proliferadve vigor is considered a marker of thè aging process and is related to thèHayflick phenomenon, i.e. thè limited number of replications that normal cells canundergo. It has been demonstrated that mere is a correlation between proliferativecapabìlity and maximum Iife span in different species, and an inverse correlation betweenproliferative capability and donor age [40-42]. Thus, a decreased capability to proliferatemay be considered a characteristic of cellular senescence.

In a recent paper we argued that an intriguìng relationship exists among cellularsenescence, tumor growth and longevity [7]. AH these phenomena are deeply related toprogrammed celi death or apoptosis [2]. A possible scenario is thè following: cells may beequipped with genes which actively promote cellular senescence thus controlling celi deathin order and escaping trans formai io rt [6, 43, 44]. This situation is balanced by other genesresponsive for survival and viability. Circumstanttal evidence suggests that cellularsenescence may be considered a peculiar type of celi differentiation whose biologica!function is to counteract uncontrolled celi proliferation [45]. From this point of view,cellular senescence can be considered one of thè most important mechanisms in avoidingthè continuous onset of tumors. The most effective evolutive way for a celi to controlneoplasie growth is likely to set up genes which promote apoptotic celi death [46-60].Moreover, we were able to show that an intrìguing relationship at thè molecular levelbetween celi proliferation and apoptosis [61-66]. Interestingly, and unexpectedly, we haverecently found that lymphocytes become progressively resistant to apoptosis with increas-ing age of thè donor, and centenarians follow this trend (manuscript in preparation).


-I_JLUOtu>E55oCL

IL.O

100-fìo oo CD45RA

CD45RO

2 80-tfjLLJU

60

* ULM3HU «• ' . • • •• .»•

* ' • f -—• • • • • * • *s» *•i ? '/; 'o. i%> "• ': " !-

O 10 20 30 40 50 60 70 80 90 HO

100

so

> 60

40

20- • • * •»^ — V

L« * '

o^'V .

o CD45RA• CD45RO

10 20 30 40 50 60 70 80 90 100 110

AGE (YEARS)FIGURE 6 Age-related changes in thè absolute number of circulating "virgin/unprimed" (CD45RA + ) lod"memory/activated" (CD45RO+), among CD4+ (upper pane!) or CD8+ (lower panel) T cells. The mostdramatic changes occur during thè first 2-3 decades of life. It is important to note that there is an unbalance, wbosemeaning ìs unknown, between CEM+ and CD8 -t- T cclls. as far as thè expression of CD45 isofonrn Ìs cooceroed.Moreovcr, it is also interesting to note that people of far advanced age and centcnarians stili bave a high number ofvirgin/unprimed T cells in their peripheral blood. An open question is where do these virgin T ceDs come Eroro,taking into account that thè organ in which these cclls should be produced, i.e. thyraus, started io imohrtiOQimmcdiatcly after puberty. severa! decades ago (partially based on data from Cossarizza et al., 1992). ̂ /V^

At present, it ibased on thè copotentially trancellular longevi

It has beencapability of itsexposure to antiwere fully capa(20-30 years oproliferative ca]notion that thejpreparation). Thloss of prolifera

SOLUBLE RE(THE UNBALA

In recent years itnetwork of hundifferentiation, \

Q.£

zoH

CCOaetoo2

ir•oHXco

140000].

120000-.

100000-

80000-

60000-

40000-'.

20000-

o-

FIGURE 7 Prolifcr.and 17 health cenien;thyrnidìne incorporairespond to this mitog


At present, it is difficult to reconcile this findings with thè above mentioned hypothesisbased on thè consideration that apoptosis is thè main mechanism to get rid of mutated andpotentially transformed cells. In any case, resistance to apoptosis could contribute tocellular longevity, and, possibly, to organistnic longevity.

It has been suggested that a cruciai change in thè immune System is thè reducedcapability of its component cells to proliferate, a problem related to donai expansion afterexposure to antigenic stimulì [67]. We found that T lymphocytes from healthy centenarianswere fully capable of proliferating, and that thè only difference vis a vis young people(20-30 years old) is a delay in peak responsi ve ness (Fig. 7) [23]. Recent data on thèproliferative capability of fibroblasts from centenarians are in full agreement with thènotion that they experience no major change in proliferative capability (manuscript inpreparation). These data cast some doubt on thè proposed relationship between aging andloss of proliferative vigor [68].

100 110

SOLUBLE REGULATORY MECHANISMS:THE UNBALANCE OF CYTOKINE NETWORK

In recent years it became clear that thè immune orchestra depends on a subtle and well tunednetwork of humoral mediators, collectively called cytokines, that are responsive fordifferentiation, proliferation and survival of lymphoid cells. They include interleukins,

sJ 8.

0*•08

I-• ta

1 08

D•

100

0

•

•110

:d" (CD45RA+) and•I) T cells. The mosts an unbalance, whose

^ 1400001E

; a£ 120000-z9. 100000-H

£ 80000on•*•

O 60000-02- 40000-CCTJH- 20000-I

o-l

sofonns is concerned. (havcahighnuraberof \n T cells come from,

, started its involutionet al., 1992).

FIGURE 7 Proliferaand 17 health centenaJÈiumii-lin* tnmmstf^t

• CENTENARIANS0 YOUNG SUBJECTS

DAYS OF CULTUREFIGURE 7 Proliferale capability of peripheral blood mononuclcar cells from 12 young (mean age 26 ± 4 years)and 17 health centenarians stimulated with an optimal dose of phytohemoagglutinin (PHA) as assessed by JH-thymidine incorporation (6 hour pulse) after different days of cultures. It is evident that thè overall capability torespond to this tnitogen is well preserved in centenarians {based on data reported in Franceschi et al., 1991).


colony stìmulating factors, interferons, and others, such as tumor necrosis factors (TNF).These molecules, most of which bave been characterized and cloned, constitute a complexnetwork, and act by thè interaction with, and binding to, specific membrane receptors,which must be considered as an integrai part of thè cytokine network. There are cytokines,such as interleukin-2 (IL-2), which have a particular importance for thè proliferation anddifferentiation of T, B, and NK cells. IL-2 and IL-10 lead to increased production ofIgM, IgG and IgA, whereas IL-4 and IL-13 induce IgE and IgG4 synthesis [69-71],

Other cytokines, suchasIL-I, IL-6 and TNF-a are considered prò-in fi ammatory agents,and play an ìmportant role not only in thè immune responses bui also in inflammation. IL-6also amplifies Ig synthesis by committed B cells.

It has been reported that thè production and utilization of one of thè most importantcytokines, IL-2, declines with age [72]. However, we have shown that thè altered produc-tion and utilization of thìs cytokine by cells from aged donors were rescued by thè expo-sure of cells to low frequency pulsed electromagnetic fields, suggesting that thè abovementioned alterations are not irreversible, and can be positively modulated [73].

What about other cytokines, whose production and utilization has not been criticallyanalyzed during human immunosenescence? We reported that thè capability of mono-nuclear cells from healthy, aged subjects [74], as well as from centenarians [manuscript inpreparation], to produce pro-ìnflammatory cytokines such as IL-1, IL-6 and TNF-aincreases with age (Fig. 8). These data suggest that thè cytokine network undergo profoundbui complex changes with age. Indeed, thè production and/or utilization of some cytokinesdecreases with agef while thè production of other cytokines increases. This field is far frombeing clear, as thè data on changes of other cytokines are lacking, especially in humans.Moreover, no data are available on possible changes in cytokine receptor on target

plasma rrreceptor £receptorspathologù

INNATE

T and B etcapable ofmillions ofvìruses anoof recogni:problem: tias chemot;different laprese rving '•did not dis^effective incollaborateagainst a v;

On thè bimmune res

ZD>CC<tetCQCC

401

30-

20

10-

TNF-et

IL-6

IL-2

O 10 20 30 40 50 60 70 60 90 100 110

AGE (YEARS)

FIGURE 8 Age-rclated changes in thè production of cytolunes from peripheral blood mononuclcar cells fromKealihy subjects ofdifferemages. including centenarians. A compiei scenario is evident, wbere thè productwn ofsome cytokines increases with age while that of others dccreases (based on dau reported in Fagiolo et al., 1993,artd manuscript in preparation). ->' - - •:•-''•

-I-JUJo

CCUJffl£

900

750

600

450-

300-

150-

Q+-O

FIGURE 9 Agvariancc with T I.newborns Io cen:


ctors (TNF)..te a complexne receptors,re cytokines,iferation andToduction of[69-71].

atory agents,imation. IL-6

)St importantered produc-by thè expo-at thè above[73].:en critically.ty of mono-nanuscript inand TNF-a

rgo profoundme cytokinesId is far fromy in humans.:or on target

'-a

^~

^v^^^^^

00 110

plasma membrane (number per celi, affinity, etc.), soluble cytokine receptors, as well asreceptor antagonists. These data are urgently needed, considering that cytokines and theirreceptors are heavily involved in thè mechanisms responsible for many age-associatedpathologies (atherosclerosis, dementia, autoimmune dìseases, etc.).

INNATE ÌMMUNITY: THE FIRST TO COME, THE LAST TO GO

T and B cells are classical examples of adaptive Ìmmunity, being clonally distributed andcapable of specifically reacting with sìngle epitopes of a given antigen. However, formillions of years lower creatures had to survive in environments full of pathogens such asviruses and bacteria, despite thè absence of an immune System functioning at a clonal levelof recognition. Mechanisms have been used, and evolved, in order to overcome such aproblem: they are cotlectively called "innate Ìmmunity," and comprehend responses suchas chemotaxis, phagocytosis, naturai cytotoxicity, among others. Evolutive studies fromdifferent laboratories, including ours, suggest that these mechanisms are fully capable ofpreserving body integrity even in thè absence of adaptive Ìmmunity [75-85] . However, theydid not disappear with thè onset of adaptive Ìmmunity. Indeed, they are stili present andeffective in more evolved animals such as mammals, where they are intermìxed with andcollaborate with clonally distributed T and B cells, and stili represent a first line of defenseagainst a variety of pathogens.

On thè basis of these considerations, it can be predicted that thè most sophisticatedimmune responses are also thè most fragile, and prone to age-related alterations. On thè

900-

7^n-• ' wU

Ì

W

J 600UJO

è 45°ocum 3002s

Z150-

n-

° CD16+• • CD57+

o0 --

• • _

* *• e

• *

• * * • Q *°* p<0.01* ^ *" """ _

^ _^ _^ — — '* ~~ *g °>° J *T 0 ° * J

— - — *~~ a o* O n O A • • « ^1 g ° Ih* 8 . • . 2 a o«« n WS* - w> PI A *̂ ' m a ° - K"1*3

84 n **°a R • 0 01

*•* A 9 ^ ° 0 ° O<b 9 °. %0 ° i • ° *° 0° 8° °°«

o o°

luclcar celk fromthè production of

yoloetal., 1993,

O 10 20 30 40 50 60 70 80 90 100 110

AGE (YEARS)

FIGURE 9 Age-rclated changes in thè absolute numbcr of circulaling cells with markcrs of NK activity. Atvariance with T lymphocytes, these cells eitherincrcasc (CD57 + ) or remain unchanged (CD16+) with age, fromnewborns to centenarians (based on data reported in Sansoni ei al., 1993).


contrary, ancestral innate immune responses should be more resistant to age-relatedchanges, being "simpler," more economical, and conserved throughout evolution.

Accordingly, innate naturai immunity was studied in healthy elderly subjects andcentenarians. In particular, we focussed on naturai killer (NK) celi activity and chemotaxis.NK cells and activity during aging bave been studied extensively by severa! groups.Different results (decrease, increase, no change) have been reported, probably because of apoor selection and insufficient inclusion and exclusion criteria [15, 21]. In large groups ofhealthy centenarians, middle aged (40-50 years old) and young (20-30 years oldj subjectsa detailed cytofluorimetric analysis allowed us io demonstrate an age-related increase incells with high NK activity (CD16+,CD57~) [21]. On thè contrary, cells with intermediate(CD16+,CD57+) or low (CD57+,CD16-) NK activity showed only minor modifications[21]. In centenarians, an increase in this high activity NK subset is mirrored by wellpreserved cytotoxicity, as measured by both NK and redirected killing assays. In Down'ssyndrome, an example of precocious aging in humans [31, 86-98], an expansion of NKcells occurs, suggesting that this is peculiar to immunosenescence [95, 99]. However, inthis syndrome, NK cells were, functionally, highly inefficient (Fìg. IO) [99]. Recent datasuggest that a persistently low NK activity is a predictor of impending morbidity [100].Conversely, it can be speculateti that well preserved NK activity can help in becoming acentenarian. The age-related increase of cells bearing NK markers, and of non-MHC-restricted T lymphocytes [21], could be interpreted as a compensatory mechanism to copewith decreases in T cells.

Recent preliminary data on chemotaxis suggest that thè capabilìty of peripheral blood

20

Z3

tetcoir

10

PHYSIOLOGICAL AGING

ACCELERATED AGING

O 10 20 30 40 50 60 70 80 90 100 110

AGE (YEARS)FIGURE IO Naturai killer (NK) celi activity iti physiological aging (from newborns lo centenarians) ind inacceleratcd aging (Down's syndrome). When healthy subjects are studied, thè NK activity of peripheral bloodeffeciors vs K562 tumor target cells in a 4 hour JICr release assay docs not undcrgo sìgnificant changes with igè.On thè contrary. a marked and significant decrease of N*K activity is observcd in young and aduli subjects aflectedby asyndromeof accelerated aging suchas Down's syndrome (based on data reported in Cossarizza et al.,-1991,and Sansoni et al., 1993). • ' ••'"

mononucle(manuscrip

On thè unot unde rgexplain wh;of coping vElderly sublosis, etc.) ;The hypothmajor risk t

CONCLUS

Taking intoimmune sysreshapìng arcomplexity egrasp its subpriate to deswhereas stili

In other wof thecontinoxygen free rdynamic poicellular defe,

Investigatiand directionsince thè mo;do have an ir

Acknowledgi

This paper. carneEU BioMed RcsC.N.R.(Progctuterapeutiche") a;nella Sindrome e

References

1. Franceschi,2. Franceschi.3. Kirkwood,4. Franceschi,

Sa)violi. S-,5. Franceschi,

3-58,19936. Monti, D., '

M. G-, and


to age-relatedolution.subjects and

id chemotaxis.;veral groups.[ybecauseofairge groups ofs old) subjectsed increase inh intermediatemodificationsrored by wellys. In Down's•ansion of NK;. However, in]. Recent datairbidity [100].n becoming aof non-MHC-anism to cope

ripheral blood

1NG

ìING

mononuclear cells to respond to chemotactic stimali is well preserved in centenarians(manuscript in preparatkm).

On thè whole, thè above mentioned data indicate that, as predicted, innate immunity doesnot undergo a significarti deterioration with age. This is probably one of thè reasons toexplain why healthy aged subjects such as healthy centenarians are apparently fully capableof coping with infectious agents, and bave no increased frequency of infectious diseases.Elderly subjects who show an increased susceptibility to infections (influenza, tubercu-losis, etc.) are probably those in whom pathological changes in immune System occurred.The hypothesis can be put forward that physiological aging per se does not represent amajor risk factor for most infectious pathologìes.

CONCLUSIONS AND PERSPECTIVES

Taking into account thè complex (positive and negative) changes which occur in thèimmune System with age, in comparison with young subjects, we prefer to use thè wordsreshapingandretuning, insteadof "alteration," "deterioration," "decline," todescribe thècomplexìty of immunosenescence. It is our opinion that these pejorative descriptions do notgrasp its substance, and that terms such as continuous remodelling are clearly more appro-priate to describe a situation where some immune parameters increase, others decrease,whereas stili others remain unchanged.

In other words, we think that thè body undergo a continuous adaptation as a consequenceof thè continuous exposure to low levels of internai and external damaging agents, such asoxygen free radicals, glucose and other reducìng sugars, radiations, among others. This is adynamic point of view, that considers centenarians as thè end-product of very effectivecellular defense mechanisms selected throughout phylogenesis and ontogenesis.

Investigations on centenarians, thè best example of successfulaging, can clarify thè trendand direction of thè immunosenescence, and go far beyond immunology sensu strictu,since thè most important age-related pathologies (e.g., atherosclerosis, dementia, cancer)do have an immunological component.

Acknowledgments

This paper, carried out within thè framework of thè Concerted Action Programme on Molecular Gerontology of thèEU BioMed Research Programme, has been partially supported by granls from M.U.R.S.T. (40% and 60%),C.N.R. (Progetto Finalizzato "Invecchiamento" and Progetto Strategico "Cìtochine: aree di intervento e strategieterapeutìche") and Regione Emilia-Romagna (Progetto di ricerca: "Prevenzione dell'invecchiamento precocenella Sindrome di Down: studio longitudinale e trasversale") to C. Franceschi.

100 110

•menarians) and in>f peripheral bloodchanges with age.

,lt subjects affected.arizza età!., 1991.

References

1. Franceschi. C. and Fabris, N. Aging C/m. Exp. Res. 5, 333-336. 1993.2. Franceschi. C. Aging Clin. Exp. Res. 1, 3-15, 1989.3. Kirkwood. T. B. L. and Franceschi. C. Ann. N.Y. Acad. Sci. 663, 412-417. 1992.4. Franceschi, C.. Monti, D.,Scarfi, M. R.,Temperani, R, Emilia, G., Sansoni. R, Troiano, L., Agnesini, C.,

Salvioli, S., and Cossarizza, A. Ann. N.Y. Acad- Sci. 663, 4-16, 1992.5. Franceschi. C. In: Trattalo di Gerontologia e Geriatria, ediied by G. Crepaldi, UTET. Torino, cap. 1, pp,

3-58. 1993.6. Monti, D., Troiano. L.. Tropea, F, Grassilli, E., Cossarizza. A., Barozzi, D.. Felloni. M. C., Tamassia.

M. G.. and Franceschi. C. Am. J. Clin. Nutr. 55. 1208S-12I4S. 1992.


1. Monti, D., Grassi 11 i, E., Troiano, L., Cossarizza, A., Salvioli, S., Barbieri, D., Agnesini, C-, Bettuzzi, S-,Ingtetti. M. C-, Corti, A., and Franceschi, C. Mn. N.Y. Acad. Sci. 673, 70-82, 1992.

8. Ottaviani, E., Ferraglia, F, Genedani, S., Bernardi, M., Bertolini. A., Cossarizza, A., Monti, D., andFranceschi, C. Ami. N.Y. Acad. Sci. 594. 454-157, 1990.

9. Ottaviani, E., Caselgrandi, E., Bondi, M., Cossarizza, A, Monti, D., and Franceschi, C. Adv. Neuro-immunol. 1, 27-39. 1991.

10. Ottaviani. E.. Caselgrandi, E., Fontanili. ?.. and Franceschi. C- Acta Biol. Hangar. 43, 293-298, 1992.11. Makinodan. T. and Kay. M. M. B. Adv. Immunol. 29, 287-330, 1980.12. Miller. J. F P A. Lance! ii. 748-749, 1962.13. Ligthart. G. I, Corberand, J. X., Four.nier, C., Garanaud, P, Hijmans, W., Kenncs, B., Mailer-Hermelink,

H. K., and Steinmann, G. G. Mech. Ageing Dev. 28, 47-55. 1984.14. Ligthart. G. J.. Corberand, J. X., Geertzen. H. G. M., Meinders. A. E., Knook, D. L.. and Hijmans. W.

Mech. Ageing Dev. 55, 89-97, 1990.15. Ligthart, G. J.. Schuit. H. R. E., and Hijmans. W. Immunotogy 68, 396-402. 1989.16. Olshansky, S. I, Carnes, B. A., and Casse!, C. Science 250. 634-640. 1990.17. Barinaga. M. Science 254. 936-938. 1991.18. Mariotti, S., Sansoni. P. Barbestno. G.. Caturegli. R, Monti, D., Cossarizza. A., Giacomelli, T,, Passeri.

G., Fagiolo. U., Pinchera. A., and Franceschi. C. Lancet 339. 1506-1508. 1992.19. Mariotti. S.. Barbesino, G., Caturegli, P, Bartalena, L., Sansoni, R. Fagnoni, F. Monti, D., Fagiolo, U.,

Franceschi, C., and Pinchera. A. /. din. Endocrinol. Metab. 177, 1130-1134, 1993.20. Goldstein. G. Blood 82. 2601-2604, 1993.21. Sansoni, R, Cossarizza, A., Brianti, V., Fagnoni, F, Snelli, G-, Monti, D., Marcato, A., Passeri, G.,

Ortolani, C., Forti, E., Fagiolo. U. Passeri, M., and Franceschi. C. Blood 80. 2767-2773. 1993.22. Rowe. J. W. and Kahn. R- L. Science 237. 143-149, 1987.23. Franceschi, C., Monti, D., Cossarizza, A., Fagnoni, F, Passeri, G., and Sansoni, P Ann. N.Y. Acad. Sci.

621. 428-441. 1991.24. Paganelli. R.. Quinti, !.. Fagiolo, U, Cossarizza, A., Ortolani. C., Guerra. E., Sansoni. P, Pucillo. L. P,

Cozzi, E., Bertollo. L.. Monti. D., and Franceschi, C. Clin. Exp. Immunol. 90. 351-354, 1992.25. Lehtonen, L., Eskola, 1, Vainio, O., and Lehtonen. A. /. Gerontol. 45, M108-112, 1990.26. Aaìberse, R. C., Van der Gaag, R., and Ven Leewen, J. J. Immunol. 130, 722-726, 1983.27. Thompson, J. S.. Wekstein, D. R.. Rhoades. J. L., Kirkpatrick, C., Brown. S. A., Roszman, T, Straus, R.,

and Tietz, N. / Am. Ceroni. Soc. 32. 274-281, 1984.28 Cossarizza, A..Ortolani, C..Sansoni, P,Donazzan, S., Saggio, G-, Paganelli, R.,Zambruno,G., Balde!!!,

M., Capri, M., Barbieri, D., Monti, D., and Franceschi, C. Eur. J. Histochem. 37 (suppl), 26, 1993.29. Cossarizza, A, Kahan, M., Ortolani, C., Franceschi, C., and Londei, M. Eur. J. Immunol. 21, 1571-1574,

1991.30. Cossarizza. A., Ortolani, C., Paganelli, R.. Monti, D., Barbieri, D.. Sansoni, P, Fagiolo, U., Forti, E.,

Londei, M., and Franceschi, C. / Immunol. Res. 4, 118-126, 1992.31. Fabris,N.,Mocchegiani,E., Amadio, L., Zannotti. M., Licastro, E, and Franceschi, C. Lanceti, 983-986,

1984.32. Bell. E. B- and Sparshott, S. M. Nature 348. 163-166, 1990.33. Michie, C.. McLean, A., Alcock, C., and Beverley. P C. L. Nature 360, 264-265, 1992.34. Brod, S. A.. Rudd. C. E., Purvee, M., and Hafler, D. A. / Exp. Med. 170. 2147-2152, 1989.35. Rothsteiri. D. M.. Yamada. A., Schlossman. S. F. and Morimoto. C. / Immunol. 146. 1175-1183, 1991.36. Gray. D. and Skarvall. H. dature 336, 70-73, 1988.37. Gray. D. and Marzinger. P / Exp. Med. 174. 969-974. 1991.38. Gray. D. Anna. Rev. Immunol. 11, 49-77. 1993.39. Rocha. B., Vassalli. P. and Guy-Grand. D. Immunol. Todav 13. 449-154. 1992. .40. Martin. G. M.. Sprague. C. A., and Epstein. C. J. Lab. Inveii. 23, 86-92, 1970.4L Rohme. D. Proc. Nati. Acad. Sci. USA 78. 5009-5013, 1981.42. Witkowski. J. A. Exp. Cerantol- 22. 231-248, 1987.43. Apoptosis. The molecular basis of celi dealh. In: Currenl Communications in Celi and Molecular Biology,

Voi. 3, edited b> Tornei, L. D. and Cope. F O., Cold Spring Harbor Uboratory Press. 1991, pp 1-321.44. Cohen. J. J. Adv. Immunol. 50. 55-85, 1991.45. Pereira-Smith. O. M. and Smilh. J. R. Science 221, 964-966. 1983.46. Pereira-Smith, O. M. and Smith. J. R. Proc. Nati. Acad. Sci. USA 85, 6042-6046, 1988.47. Smith, J. R. and Pcreira-Smith, O. M. Adv. Cancer Res. 54, 63-71, 1990.48. Holliday. R. / Gerontol. Biol. Sci. 45. B36-41. 1990.49. Knudson Jr. A. G. Anna. Rev. Genet. 20. 231-251. 1986.50. Sager. R. Science 246. 1406-1412. 1989.51. Levine. A. J. BioEisays 12. 60-67. 1990.52. Levine, A. J. and Momand, J. Biochim. Biophys. Acta 1032, 119-136, 1990.53. O'Brten. W.. Stenman. G., and Sager. R. Proc. Nati. Acad. Sci. USA 83, 8659-8663, 1986.54. Hockenbery. D.. Nunez. G., MUlirnan. C., Schreiber, R. D.. and Korsmeyer. S. i Nature 348. 334-336. 1990.

55. Shay, ;56. Yonish

1991.57. Gregor

Nature58. Hende:

Rickin59. Strasse60. SentmL

1991.61. Bcttuzz

France;-62. Grassi]

Commi.63. Grassi).

Acad. i64. Grassi1.:

and Suz65. Sikora.

192. 38,'66. Sikora.

Biophys67. Muraski68. Cristofa:69. Snapper70. Sutton. ;7!. Defranct

179, 13572. Gillis, S73. Cossariz

Francese74. Fagiolo,

C., and75. Francese

1991.76. Francese77. Ottavian

B, 245,78. Francese79. Ottavian:

1992.80. Ottaviani8 1 . Ottaviani

446-15282. Ottaviani83. Onaviani84. Genedan

107C, 7'85. Genedan

203-20686. Francese;

179-181.87. Francese;

M. fiumi88. Francesi

M. P. Pa89. Licastro,

75. 111-190. Chiricolo

145-152.91. Francescr

Meni. De92. Cossarizz

Thymus 1


ni, C-, Bettuzzi, S.,'2.A., Monti. D., and

hi, C. Adv. Neuro-

ìì. 293-298, 1992.

Miiller-Hermclink,

., and Hijmans, W.

omelli, T., Passeri,

i t i , D., Fagiolo, U.,

o, A., Passeri, G.,-2773. 1993.

nn. N.Y. Acad. Sci.

li, P, Pucillo, L. P,-354. 1992..990.1983-man.T, Straus.R.,

bruno, G., Baldelli,uppl), 26, 1993.noi. 21, 1571-1574,

jiolo, U., Forti, E.,

lance: i, 983-986,

1992.152, 1989.6, 1175-1183. 1991.

/ Motecuiar Biology,•sì, 1991, pp 1-321.

1988.

Ì3. 1986.348. 334-336,1990.

55. Shay, J. W., Wright, W. E., and Werbin. H. Biochim. Biophys. Acta 1072, 1-7, 1991.56. Yonish-Rouach, E., Resnilzki, D., Lotem, J., Sachs, L., Kimchil, A., and Oren, M. Nature 352. 345-347,

1991.57. Gregory, C. D.. Dive, C, Henderson, S.,Smith,C. A., Williams, G. T., Gordon, J.. and Rickinson. A. B.

Nature 349, 612-614, 1991.58. Henderson. S., Rowe, M., Gregory, C., Groom-Carter, D., Wang. E, Longnecker, R., Kieff, E., and

Rickinson, A. Celi 65, 1107-1115, 1991.59. Strasser. A.. Harris, A. W., and Cory. S. Celi 67. 889-899, 1991.60. Semman, C. L.. Shutter, J. R., Hockenbery, D., Kanagawa, O., and Korsmeier, S. G. Ceti 67, 879-888,

1991.61. Bertuzzi, S., Troiano, L., Davalli, P, Tropea, E. Ingletti, M. C., Grassilli, E., Monti, D., Corti, A., and

Franceschi. C. Biochem. Biophys. Res. Commuti. 175, 810-815, 1991.62. Grassilli, E., Bettuzzi, S., Monti, D.. Ingletti, M. C., Franceschi, C, and Corti, A. Biochem. Biophys. Res.

Commun. 180, 59-63, 1991.63. Grassilli, E.. Bettuzzi. S., Troiano, L., Ingletti, M. C., Monti, D., Corti, A., and Franceschi, C. Ann. N.Y.

Acad. Sci. 663, 471-474, 1992.64. Grassilli, E., Carcereri de Prati, A., Monti, D., Troiano, L., Menegazzi, M., Barbieri, D., Franceschi, C.,

and Suzuki, H. Biochem. Biophys. Res. Commun. 188, 1261-1266, 1992.65. Sikora, E., Grassilli. E., Bellesia, E.. Troiano, L., and Franceschi, C. Biochem. Biophys. Res. Commun.

192, 386-391, 1993.66. Sikora. E., Grassilli, E., Radziszewska, E., Bellesia, E., Barbieri, D., and Franceschi, C. Biochem.

Biophys. Res. Commun. 197, 709-715, 1993.67. Muras'ko, D. M., Weiner, P, and Kaye, D. Ctin. Exp. Immunol. 70, 440^(48, 1987.68. Cristofalo. V J. and Pignolo, R. J. Physiol. Rev. 73, 617-638, 1993.69. Snapper, C. M. and Mond, J. J. Immunol. Today 14, 15-17, 1993.70. Sunoo, B. J. and Gould, H. J. Nature 366, 421-428, 1993.71. Defrance, T., Carayon, P, Billian, G., Guillcmot, J.-C-, Minty, A., Caput, D., and Ferrara, P J. Exp. Med.

179, 135-143, 1994.72. Gillis, S., Kozak. R.. Durante. M., and Weksler, M. / C/in. Inveii. 67, 937-942, 1981.73. Cossarizza, A., Monti, D., Cantini, M., Paganelli, R., Monlagnani, G., Cadessi, R., Bersani, E, and

Franceschi, C. FEBS Len. 248, 141-144, 1989.74. Fagiolo, U, Cossarizza, A., Scala, E., Fanales-Belasio, E., Ortolani, C., Cozzi, E., Monti, D., Franceschi,

C., and Paganellì, R. Eur. J. Immunol. 23, 2375-2378, 1993.75. Franceschi, C., Cossarizza, A-, Ortolani, C., Monti, D., and Ottaviani, E. Adv. Neuroimmunol. 1, 99-113,

1991.76. Franceschi, C., Cossarizza, A., Monti, D., and Ottaviani, E. Eur. J. Immunol. 21, 489-493, 1991.77. Ottaviani, E., Cossarizza, A., Ortolani, C., Monti, D., and Franceschi, C. Proc. RoyatSoc. London Series

fi. 245, 215-218, 1991.78. Franceschi, C. and Ottaviani, E. J. Immunol. Res. 4. 53-55, 1992.79. Ottaviani, E., Caselgrandi, E., Petraglia, E, and Franceschi, C. Cen. Compar. Endocrinol. 87, 354-360,

1992.80. Ottaviani. E., Franchìni, A., Cossarizza, A., and Franceschi, C. Neuropeptides 23, 215-219, 1992.81. Otiavìani, E., Caselgrandi, E., Franchinì, A., and Franceschi, C. Biochem. Biophys. Res. Commun. 193,

446-452. 1993.82. Ottaviani, E., Franchini. A., and Franceschi. C. Biochem. Biophys. RES. Commun. 195, 984-988. 1993.83. Oltaviani, E. and Franceschi, C. Trends Compar. Biochem. Physiol. 1, 109-117, 1993.84. Genedani, S., Bernardi, M., Fontanili, R, Ottaviani. E., and Franceschi, C. Compar. Biochem. Physiol. B.

107C, 79-81, 1994.85. Genedani, S., Bernardi. M., Ottaviani, E., Franceschi, C., Leung, M. L-, and Stefano. G. B. Peptìdeì 15.

203-206, 1994.86. Franceschi. C.. Licastro, F, Paolucci, P, Masi, M., Cavicchi, S., and Zannotti, M. J. Meni. Defic. Res. 22.

179-181, 1978.87. Franceschi, C., Licastro. E, Chiricolo, M., Fantini, M. P. Paolucci, P, Fabris, N.. Zannotii, M., and Masi,

M. Hìanan Genetìcs 2 (suppl.), 238-239, 1981.88. Franceschi, C., Licastro, E. Chiricolo, M.. Bonetti. F, Zannotti, M., Fabris, N., Mocchegiani, E., Fantini.

M. R, Paolucci, P, and Masi. M. / Immunol. 126, 2161-2164. 1981.89. Licastro. F, Chiricolo, M., Tabacchi, P. L., Barboni, E, Zannotti, M., and Franceschi, C. Celi. Immunol.

75, 111-121, 1983.90. Chiricoìo.M., Minetlì, L., Licastro, E, Tabacchi, P L., Zannotti, M., and Franceschi. C. CeroniologyìO,

145-152, 1984.91. Franceschi, C.. Chiricolo, M.. Licastro, F, Zannotti, M.. Masi, M., Mocchegiani, E., and Fabris. N. /

Meni. Dejic. Res. 32, 169-181, 1988.92. Cossarizza, A., Monti, D., Montagnani, G.. Dagna-Bricarelli, F, Forabosco, A., and Franceschi, C.

TTiymus 14, 163-170. 1989.


93. Franceschi,C..Monti, D.,Cossarizza, A.,Tornasi, A..Sola, R, and Zannotti, M.Adv.Exp.Med. Biol. 264.499-502. 1990.

94. Cossarizza, A., Monti, D., Dagna-BricarelH. E, Montagnani, G., and Franceschi, C. Immunat. Lett. 24,137-140, 1990.

95. Cossarizza. A.. Monti, D., Montagnani, G., Ortolani, C., Masi, M., Zannotti, M., and Franceschi, C.Am. 3. Med. Gtnet. 7 (suppl.). 213-218, 1990.

96. Scarfi, M. R..Cossarizza. A..Monti, D., Bersani, E, Zannotti, M., Lici, M. B-, and Franceschi. C. Mutar,Res. 237, 217-222, 1990.

97. Licastro, F, Chiricolo. M., Mocchegiani, E., Fabris. N., Cossarizza, A., Masi, M., Arena, G., Zannotti, \. Monti, D., Bcltrandi, E.. Mancini, R.. Casadeì-Maldini. M., and Franceschi, C. J. Immunol. Res, 2,

95-100. 1990. i98. Cossarizza, A., Monti, D.. Bersani, E. Scarfi, M. R., Zannotli, M.. Cadessi, R., and Franceschi, C. Aging

din. Exp. Res. 3, 241-246, 1991.99. Cossarizza, A. .Ortolani, C. .Forti, E. .Montagnani, G.. Paganelli, R. Zannotti, M. .Marini, M. .Monti, D.,

and Franceschi, C. BW 77, 1263-1270. 1991.100. Levy, S. M., Herberman, R. B.. Lee, J., Whiteside, T., Beadle. M.. Heiden, L., and Simons, A. Nat.

Immun. Celi Growth Reg. 10, 289-307, 1991.

immunosenescence in humans: deterioration or remodelling?

Documents