This Article Abstract Full Text (PDF) Services Download to citation manager PubMed PubMed Citation

Interleukin-1 Receptor Antagonist, Interleukin-6, and C-Reactive Protein as Predictors of Mortality in Nonagenarians: The Vitality 90+ Study

¹ Tampere School of Public Health, University of Tampere, Finland.
Departments of ² Clinical Chemistry, ³ Forensic Medicine, ⁴ Microbiology and Immunology, University of Tampere Medical School, Finland.
⁵ Research Unit of the Laboratory Center, Tampere University Hospital, Finland.
⁶ Department of Social Welfare and Health Services, State Provincial Office of Western Finland.

Address correspondence to Marja Jylhä, MD, PhD, Tampere School of Public Health, FIN-33014 University of Tampere, Finland. E-mail: marja.jylha{at}uta.fi

	Abstract

Top Abstract Materials and Methods Results Discussion References

 
Background. Inflammation plays a major role in both aging and chronic disease. Longitudinal studies in very old people can improve our understanding of these processes. We investigated blood levels of C-reactive protein (CRP), interleukin-6 (IL-6), interleukin-1 receptor antagonist (IL-1ra), and their combinations as predictors of mortality in nonagenarians.

Methods. This is a prospective population-based study including both community-dwelling and institutionalized nonagenarians enrolled in the Vitality 90+ Study. Altogether 285 persons participated in the baseline interview and gave blood. Information on chronic disease was drawn from health center registers. Data on mortality over 4 years were obtained from the Population Register Center. In Cox proportional hazards models, chronic disease and major risk factors were adjusted for.

Results. Plasma levels of IL-1ra, IL-6, and CRP were higher in persons who died during the follow-up than in those who survived. When sex, education, cardiovascular disease, diabetes, cancer, history of infections, high density lipoprotein cholesterol, Mini-Mental State Examination, body mass index, smoking status, and exercise were adjusted for, only IL-1ra was a significant predictor of mortality (hazard ratio [HR] 2.12; 95% confidence interval [CI], 1.24–3.62). Persons in the upper tertiles of both CRP and IL-1ra (HR 2.72; 95% CI, 1.25–6.00), or in the upper tertile of all three markers (HR 2.34; 95% CI, 1.23–4.61), had higher mortality than those who were not in the upper tertile in any of the markers.

Conclusions. IL-1ra is a powerful prognostic marker in very old people. Our results implicate its role in the complex interaction between inflammatory markers in aging and disease.

The present study focuses on the prognostic role of three inflammatory markers, interleukin-1 receptor antagonist (IL-1ra), interleukin-6 (IL-6), and C-reactive protein (CRP), in long-lived people. IL-1ra is a specific, high-affinity antagonist to the other two members of the IL-1 family, IL-1 and IL-1ß (7), which belong to the earliest mediators of the acute phase response in inflammation. They are inductors of IL-6, which in turn induces production of CRP (5), but also suppresses IL-1 production and increases the synthesis of IL-1ra (8). IL-1ra in turn inhibits IL-6 release (9). IL-6, IL-1ra, and CRP also have several other functions, some of them synergistic.

Chronic, low-grade inflammatory activity is involved in the pathogenesis and course of several age-related disorders (5,8,10). CRP and IL-6 have been found to predict cardiovascular disease and mortality (11–13), all-cause mortality (14–16), disability (17), and loss of muscle strength (18). The role of IL-1ra has been examined mainly in patient samples and in in vitro studies. Early animal models suggested that it prevents death from septic shock, but recent studies stress the importance of the IL-1ra/IL-1 ratio and suggest that higher levels of IL-1ra may be predictive of mortality (10,19). In one study, high levels of CRP, IL-6, and IL-1ra were all independently associated with poor physical performance in old people (20).

The results from the few population studies with very old people vary. In 126 centenarians, elevated CRP was associated with 5-year all-cause mortality, but IL-6 was not (2), whereas in a sample of 333 octogenarians IL-6 did predict mortality during 6 years (21). To our knowledge, there are no previous data on the association of IL-1ra with mortality in randomly selected old populations, and in general, data on inflammatory markers in the very old population are scarce.

We investigated the value of CRP, IL-6, IL-1ra, and their combinations as predictors of mortality in a population sample of people aged 90 years in the Vitality 90+ Study. Major chronic conditions and risk factors that are known to be associated with both mortality and inflammation were taken into account as possible confounders. We hypothesized that (i) CRP, IL-6, and IL-1ra predict mortality in nonagenarians, (ii) the association between these markers and mortality can partly be explained by comorbidity and major risk factors, and (iii) joint elevations in the inflammatory markers have a stronger association with mortality than do any of the markers alone.

	MATERIALS AND METHODS

Top Abstract Materials and Methods Results Discussion References

 
Participants and Study Design
This investigation is part of Vitality 90+, a prospective multidisciplinary population-based study of people aged 90 or older in Tampere, Finland (22,23). The study population in this analysis consisted of all people born in 1909–1910 who, according to the local population register as of January 2000, resided in the city of Tampere (n = 535). Both community-dwelling and institutionalized persons were invited to participate in home interviews and blood tests. According to the national Population Register Center, 66 persons had died before the beginning of data collection, leaving 496 persons eligible for the study. An additional 42 persons died during the study but before being examined, 86 persons refused to participate, mostly due to poor physical or mental condition, and 7 persons could not be reached. Another 45 persons refused blood tests and only took part in the interviews. The final sample for the present study thus consisted of 285 (66% of the eligible population) participants who agreed to have blood tests taken and most of whom (92%; n = 262) also participated in home interviews. Most of the participants (240, 84%) lived in the community; 45 (16%) lived in institutions.

Interviews, performance measurements, and blood tests were carried out at homes for community-dwelling participants or in institutions for institutionalized participants. Medical diagnoses were collected from health center records.

The study protocol was approved by the Ethics Committee of the Pirkanmaa Hospital District and the Ethics Committee of Tampere Health Center. All participants gave their written informed consent.

Biochemical Measurements
Blood samples were obtained in the morning after an overnight fast using EDTA tubes in ice. After separation of plasma by low-speed centrifugation (15 minutes at 700 g), the plasma was divided into aliquots and stored at –80°C until analyzed. Concentrations of IL-1ra and IL-6 were determined using commercially available enzyme-linked immunosorbent assay (ELISA) kits (Quantikine; R&D Systems, Minneapolis, MN for IL-1ra; Pelikine Compact human IL-6 ELISA kit; CLB, Amsterdam, The Netherlands for IL-6). The optical density of individual wells was determined with a Multiscan Biochromatic 348 (Labsystems, Helsinki, Finland) spectrophotometer.

High-sensitivity CRP and high-density lipoprotein (HDL) cholesterol concentrations were analyzed using a Cobas Integra 700 automatic analyzer with reagents and calibrators as recommended by the manufacturer (Hoffmann-La Roche Ltd., Basel, Switzerland; COBAS Integra C-Reactive Protein, Latex).

Covariates
Medical diagnoses were available for 257 participants. They were collected from records maintained by public health care physicians, including diagnoses made in hospitals, and were coded according to the International Classification of Diseases, 10th Revision (ICD-10) (24). Total cardiovascular morbidity (ICD codes 0–50) included hypertension, coronary heart disease, chronic heart failure, myocardial infarction, and atrial fibrillation. Infectious disease indicates a history of any infectious disease treated in the hospital, including gastroenteritis, erysipelas, hepatitis, pneumonia, urinary infections, and others. This variable, therefore, measures not only infections at the time of baseline examination, but also previous infections, if severe enough to require hospital treatment. Other diseases included were diabetes and cancer (other than basal cell carcinoma).

Cognitive status, as assessed by the Mini-Mental State Examination (MMSE), was used as a dichotomized variable, using the median value (25) as the cut point. Body mass index (BMI, computed as weight in kilograms divided by height in meters squared) was categorized as < 20, 20–24.99, 25–29.99, or 30.

Educational level was classified as (i) less than primary, (ii) primary, or (iii) secondary or more. Physical exercise was categorized as (i) daily, (ii) at least once a week, or (iii) not very often, and smoking status was categorized as (i) current or former or (ii) never.

Mortality Follow-Up
Dates of deaths were drawn from the Population Register Center. There were no losses to mortality follow-up. Follow-up time was calculated from February 21, 2000 to the date of death or to April 5, 2004 for survivors.

Statistical Analysis
Differences between the groups were analyzed using Pearson's chi-square test for categorized variables and the Mann–Whitney U test for medians. Because CRP and IL-6 were not normally distributed, the analyses were also performed by using their log values. For clarity, original values are used in describing the results. Variables are reported as medians and interquartile range (q1-q3), or percentages. As predictors of mortality, inflammatory markers are used as tertiles (< 0.9, 0.9–3.6, and > 3.6 mg/L for CRP; < 1.97, 1.97–3.8, and > 3.8 pg/mL for IL-6; < 312, 312–454, and > 454 pg/mL for IL-1ra). In addition, an inflammation score was constructed. First, a measure was created to describe the different combinations of high levels of inflammatory markers (11). In this variable, the first category consists of (i) participants whose values were below the upper tertile in all of the three markers. The other categories—(ii) upper tertile in any one of the three markers, (iii) CRP and IL-6 in the upper tertile, (iv) IL-6 and IL-1ra in the upper tertile, (v) CRP and IL-1ra in the upper tertile, and (vi) all three values in the upper tertile—were each contrasted to category (i). In the inflammation score, the categories of any two markers in the upper tertile (categories iii–v) were combined.

Cox proportional hazards models were used to analyze associations between inflammatory markers and all-cause mortality over the 4-year follow-up. The analyses were adjusted for (i) sex, and (ii) then also for cardiovascular diseases, (iii) then also for diabetes, cancer, and history of infectious disease, and (iv) finally also for education, MMSE score, BMI, physical exercise, smoking status, and HDL cholesterol. The results between (ii) and (iii) were essentially similar, and only the latter are shown here.

Statistical analyses were performed using SPSS versions 13.0 and 14.0. A p value <.05 was considered significant.

	RESULTS

Top Abstract Materials and Methods Results Discussion References

 
The three inflammatory markers were correlated: The correlation between CRP and IL-6 was.31 (p <.01), between CRP and IL-1ra it was.35 (p <.01), and between IL-6 and IL-1ra it was.13 (p <.05).

Altogether, 171 (60.0%) participants died during the 4-year follow-up. The characteristics of the sample according to survival status are presented in Table 1. Mortality was higher among men and participants who had lower MMSE scores, either low or high BMI, who exercised less often, and for participants who had cardiovascular disease, a history of infectious disease, or cancer. The deceased participants had significantly higher levels of CRP, IL-6, and IL-1ra than did those who survived. The deceased participants were also more often in the upper tertile in combinations that included high IL-1ra and high CRP.

When, instead of the inflammation score, the combination measure was used that separated the three different combinations of two inflammatory markers in the upper tertile (data not shown, see Table 1 for the variable), the HR for the combination of high CRP and high IL-1ra was significant (HR 2.72; 95% CI, 1.25–6.00). For the two other combinations, mortality was not significantly elevated.

When men and women were analyzed separately and without adjustments, none of the individual inflammatory markers was significantly associated with mortality in the rather small subgroup of men (n = 67). Still, mortality was significantly elevated for men who were in the upper tertile of all three markers in the inflammation score. In women, the associations found were essentially the same as those for the total sample (data not shown).

To better understand the impact of major disease on the results, we performed separate analyses in certain subgroups. All these analyses were adjusted for sex (data not shown). Among participants with cardiovascular disease (n = 200), both the upper CRP tertile (HR 1.63; 95% CI, 1.07–2.47), upper IL-1ra tertile (HR 1.17; 95% CI, 1.14–2.69), and the upper tertiles of all three markers (HR 2.71; 95% CI, 1.52–4.84) were associated with mortality. In participants without a diagnosis of cardiovascular disease (n = 57), none of the inflammatory markers was significant, although the direction of the coefficients was the same. Among participants with a history of infectious disease (n = 115), no significant association was observed between the inflammatory markers and mortality, whereas in participants without a history of infectious disease (n = 142) both the upper IL-1ra tertile (HR 2.15; 95% CI, 1.21–3.81) and the combined upper tertile in all three markers (HR 6.60; 95% CI, 2.87–15.44) were associated with mortality. In the community-dwelling group (n = 240), higher mortality was associated with the upper CRP tertile (HR 1.59; 95% CI, 1.04–2.41), the upper IL-1ra tertile (HR 1.58; 95% CI, 1.03–2.42), and with the combined upper tertile of all inflammatory markers (HR 3.72; 95% CI, 1.99–6.99). In the small group of institutionalized persons (n = 45), none of the individual markers was significant, but mortality was elevated in participants in the upper tertile of two or all three inflammatory markers.

When those 35 deaths that occurred within 1 year of baseline data collection were left out of the analysis, the results were still similar to the main analyses: The upper tertile of CRP, the upper tertile of IL-1ra, the combined upper tertiles of two markers, and the upper tertile in all three markers were still associated with higher mortality.

	DISCUSSION

Top Abstract Materials and Methods Results Discussion References

 
We investigated the role of blood levels of three inflammatory markers (CRP, IL-6, and IL-1ra) and their combinations, as predictors of mortality during 4 years in a population sample of 90-year-old people. Baseline blood concentrations of all three markers were significantly higher in participants who died during the 4 years than in those who survived. A novel finding was that IL-1ra showed a strong graded association with mortality even when major chronic disease and risk factors were adjusted for. CRP also predicted mortality independently of major diseases, but there was no significant association between IL-6 and mortality. The inflammation score indicated a higher mortality for combined high levels of CRP and IL-1ra, both with and without a high level of IL-6.

This research is one of only a few longitudinal studies that have investigated inflammatory markers in a population-based sample of very old people, using a variety of data from clinical diagnoses to lifestyles. We invited the whole cohort born in 1909–1910 and living in the city of Tampere to participate, and 66% of those who were alive at the beginning of the study agreed. The gender distribution did not differ between the participants and nonparticipants. One strength of the study is that both community-dwelling (participation rate 69%) and institutionalized (participation rate 57%) persons were included. However, the loss was reasonably high, and there were missing data in several variables. Thus, the sample is too small to allow for adjusted stratified analyses in different clinical subgroups. This is a clear weakness of this study.

In our data, levels of CRP, IL-6, and IL-1ra were linearly correlated, but their associations with mortality were different. Several earlier studies with old populations have identified CRP as a predictor of mortality (2,11), and it has been suggested that this association mainly reflects activities of tumor necrosis factor- and IL-6 (2). Here, however, IL-6 was not associated with mortality when sex or other confounders were adjusted for. Instead, IL-1ra was a strong independent predictor both alone and in combination with CRP. Also, the baseline characteristics of the sample differed between the tertiles of IL-1ra more than between the tertiles of the two other indicators. Levels of IL-1ra were significantly elevated in participants with low MMSE scores, either a very low or high BMI, cardiovascular disease, diabetes, or higher levels of HDL cholesterol.

Earlier studies have shown that IL-1ra predicts mortality in acute infectious conditions (19,25). In our study, the association was maintained when the deaths that occurred within 1 year of baseline measurements were left out of the analysis. Furthermore, IL-1ra was not significantly associated with mortality in the subgroup with a history of infectious disease, although the blood levels were slightly, albeit not significantly, higher in the group with the disease. Thus, it is not likely that the higher levels in participants who died reflect response to acute infectious or inflammatory exposure. Our earlier work (26) and other studies (27) have shown that the genotype or allele frequency of IL-1ra VNTR did not differ between long-lived people and younger controls. Neither was mortality during 4 years in our sample associated with genotype (Hurme M, Jylhä M, Lehtimäki T, Karhunen PJ, Hervonen A, unpublished observations). In Italian centenarians without acute pathological conditions, blood levels of IL-1ra showed no relationship with genotype (27). These results indicate that the elevated levels of IL-1ra that are associated with shorter remaining life expectancy are neither directly genetically regulated nor a sign of acute inflammatory exposure, but rather a response to the low-grade proinflammatory activity in aging, or chronic disease and its severity, or both. The stronger association of IL-1ra with mortality in the group suffering from cardiovascular disease compared to that in the group without cardiovascular disease implicates the role of this cytokine in vascular pathologies. However, immunosenescence is best understood as a continuum of changes related to age-associated pathology (3), and it is difficult to make a strict separation between aging and pathological conditions. Still, the role of IL-1ra in major pathological conditions deserves further research.

The exact mechanisms that associate IL-1ra levels with mortality in very old persons remain unclear. Recent research indicates that it is not the level of IL-1ra as such, but the balance between IL-1 and IL-1ra and the timing of the response that is critical to the course and outcome of different conditions (10,19). Furthermore, the roles of different cytokines can be understood only in their mutual interaction. The IL-1 family likely acts in a close network—particularly with CRP, IL-6, and tumor necrosis factor-—but also with other cytokines and inflammatory indicators. In these networks, the functions of one cytokine are modified, modulated, and substituted by another one (6). Thus, rather than being in a direct causal relationship with mortality, IL-1ra should be considered as a prognostic marker in the very old population, and as part of the dynamic reshaping of the cytokine network in the state of "inflamm-aging"(6).

	Acknowledgments

Top Abstract Materials and Methods Results Discussion References

 
This study was funded by Competitive Research Funding from Pirkanmaa Hospital District (AH), the Academy of Finland (MJ), and the Emil Aaltonen Foundation (TL).

We thank our collaborators working in health and social care in the city of Tampere for their help and support during the data collection.

	Footnotes

Top Abstract Materials and Methods Results Discussion References

 
Decision Editor: Luigi Ferrucci, MD, PhD

Received October 12, 2006

Accepted December 12, 2006

	References

Top Abstract Materials and Methods Results Discussion References

 

1. Ershler WB. Interleukin-6: a cytokine for gerontologists. J Am Geriatr Soc. 1993;41:176-181.[Medline]
2. Bruunsgaard H, Pedersen BK. Age-related inflammatory cytokines and disease. Immunol Allergy Clin North Am. 2003;23:15-39.[Medline]
3. Krabbe KS, Pedersen M, Bruunsgaard H. Inflammatory mediators in the elderly. Exp Gerontol. 2004;39:687-699.[Medline]
4. Ferrucci L, Corsi A, Lauretani F, et al. The origins of age-related proinflammatory state. Blood. 2005;105:2294-2299.
5. Bruunsgaard H, Pedersen M, Pedersen BK. Aging and proinflammatory cytokines. Curr Opin Hematol. 2001;8:131-136.[Medline]
6. De Martinis M, Franceschi C, Monti D, Ginaldi L. Inflammation markers predicting frailty and mortality in the elderly. Exp Mol Pathol. 2006;80:219-227.[Medline]
7. Dinarello CA. The interleukin-1 family: 10 years of discovery. FASEB J. 1994;8:1314-1325.[Abstract]
8. Maggio M, Guralnik JM, Longo DL, Ferrucci L. Interleukin-6 in aging and chronic disease: a magnificent pathway. J Gerontol A Biol Sci Med Sci. 2006;61:575-584.[Abstract/Free Full Text]
9. Tilg H, Trehu E, Atkins MB, Dinarello CA, Mier JW. Interleukin-6 (IL-6) as an anti-inflammatory cytokine: induction of circulating IL-1 receptor antagonist and soluble tumor necrosis factor receptor p55. Blood. 1994;83:113-118.[Abstract/Free Full Text]
10. Arend WP. The balance between IL-1 and IL-1Ra in disease. Cytokine Growth Factor Rev. 2002;13:323-340.[Medline]
11. Harris TB, Ferrucci L, Tracy RP, et al. Associations of elevated interleukin-6 and C-reactive protein levels with mortality in the elderly. Am J Med. 1999;106:506-512.[Medline]
12. Pepys MB, Hirschfield GM. C-reactive protein: a critical update. J Clin Invest. 2003;111:1805-1812.[Medline]
13. Volpato S, Guralnik JM, Ferrucci L, et al. Cardiovascular disease, interleukin-6, and risk of mortality in older women: the Women's Health and Aging Study. Circulation. 2001;103:947-953.[Abstract/Free Full Text]
14. Reuben DB, Cheh AI, Harris TB, et al. Peripheral blood markers of inflammation predict mortality and functional decline in high-functioning community-dwelling older persons. J Am Geriatr Soc. 2002;50:638-644.[Medline]
15. Roubenoff R, Parise H, Payette HA, et al. Cytokines, insulin-like growth factor 1, sarcopenia, and mortality in very old community-dwelling men and women: the Framingham Heart Study. Am J Med. 2003;115:429-435.[Medline]
16. Stork S, Feelders RA, van den Beld AW, et al. Prediction of mortality risk in the elderly. Am J Med. 2006;119:519-525.[Medline]
17. Ferrucci L, Harris TB, Guralnik JM, et al. Serum IL-6 level and the development of disability in older persons. J Am Geriatr Soc. 1999;47:639-646.[Medline]
18. Schaap LA, Pluijm SM, Deeg DJ, Visser M. Inflammatory markers and loss of muscle mass (sarcopenia) and strength. Am J Med. 2006;119:526.e9–526.17.
19. Ashare A, Powers LS, Butler NS, Doerschug KC, Monick MM, Hunninghake GW. Anti-inflammatory response is associated with mortality and severity of infection in sepsis. Am J Physiol Lung Cell Mol Physiol. 2005;288:L633-L640.[Abstract/Free Full Text]
20. Cesari M, Penninx BW, Newman AB, et al. Inflammatory markers and cardiovascular disease (The Health, Aging and Body Composition [Health ABC] Study). Am J Cardiol. 2003;92:522-528.[Medline]
21. Bruunsgaard H, Ladelund S, Pedersen AN, Schroll M, Jorgensen T, Pedersen BK. Predicting death from tumour necrosis factor-alpha and interleukin-6 in 80-year-old people. Clin Exp Immunol. 2003;132:24-31.[Medline]
22. Jylha M, Hervonen A. Functional status and need of help among people aged 90 or over: a mailed survey with a total home-dwelling population. Scand J Public Health. 1999;27:106-111.[Medline]
23. Lehtimaki T, Ojala P, Rontu R, et al. Interleukin-6 modulates plasma cholesterol and C-reactive protein concentrations in nonagenarians. J Am Geriatr Soc. 2005;53:1552-1558.[Medline]
24. Goebeler S, Jylha M, Hervonen A. Medical history, cognitive status and mobility at the age of 90. A population-based study in Tampere, Finland. Aging Clin Exp Res. 2003;15:154-161.[Medline]
25. Suharti C, van Gorp EC, Dolmans WM, et al. Cytokine patterns during dengue shock syndrome. Eur Cytokine Netw. 2003;14:172-177.[Medline]
26. Wang XY, Hurme M, Jylha M, Hervonen A. Lack of association between human longevity and polymorphisms of IL-1 cluster, IL-6, IL-10 and TNF-alpha genes in Finnish nonagenarians. Mech Ageing Dev. 2001;123:29-38.[Medline]
27. Cavallone L, Bonafe M, Olivieri F, et al. The role of IL-1 gene cluster in longevity: a study in Italian population. Mech Ageing Dev. 2003;124:533-538.[Medline]

This Article Abstract Full Text (PDF) Services Download to citation manager PubMed PubMed Citation