This case-cohort study of a middle-aged metabolically healthy population at baseline has three main findings. First, circulating concentrations of IL-1Ra were elevated in cases of incident type 2 diabetes 13 years in advance of diagnosis compared with individuals who remained diabetes-free. Second, in control subjects, longitudinal changes could be described by a linear trajectory with only a slight increase over time, whereas in case subjects, IL-1Ra increased rapidly starting 6 (95% CI 4.5–7.5) years before diagnosis. Third, changes in obesity explained the slight linear increase in control subjects throughout the observation period and in case subjects up to 6 years before diabetes diagnosis, but did not account for the steep increase in the IL-1Ra trajectory among case subjects in the last 6 years.
Our findings extend current knowledge on the association between inflammation and type 2 diabetes development, as this is the first study to characterize cytokine trajectories before type 2 diabetes. Our previous report on IL-1Ra and incident diabetes in a nested case-control study was limited to a single IL-1Ra measurement at study baseline (phase 3) (15
) and therefore did not provide information on the time course of IL-1Ra levels preceding diabetes diagnosis.
Our findings are also novel from a pathophysiological perspective because they support the hypothesis that the pre-diabetic stage is characterized not only by proinflammatory alterations, but also by the presence of an anti-inflammatory response. Several mechanisms could explain the upregulation of IL-1Ra before the diagnosis of type 2 diabetes. The steep increase of IL-1Ra in pre-diabetic individuals occurs within the same time window in which indicators of insulin sensitivity, β-cell function, and glycemia deteriorate (supplementary Fig. 1). This may indicate that these unfavorable changes in glucose metabolism and the increase of IL-1Ra production are closely connected. In vitro studies show that both human islets and monocytes respond to high glucose concentrations with an upregulation of IL-1β (22
), so that increased IL-1Ra concentrations before type 2 diabetes may represent a response to glucose-mediated IL-1β upregulation. The balance between IL-1β and IL-1Ra has been postulated to be a major determinant of the time course and severity of inflammatory diseases (25
), and it is conceivable that the local and/or systemic ratio of these cytokines could also be relevant in the pathogenesis of type 2 diabetes. We could not test this hypothesis here because the physiological concentrations of IL-1β in individuals without severe inflammatory diseases are so low that they are mostly undetectable with currently available assays (23
). However, it has been suggested that elevation of only IL-1β may not be sufficient to increase risk of type 2 diabetes; instead, increased IL-1β in combination with elevated levels of other proinflammatory cytokines may be required (28
), thus limiting the predictive relevance of the IL-1Ra/IL-1β ratio without consideration of other risk factors.
The finding that the difference in trajectories between case and control subjects could not be explained by obesity is important because adipose tissue is a major producer of IL-1Ra (9
). We replicated strong correlations of IL-1Ra levels with BMI and waist circumference (30
). However, inclusion of BMI or waist circumference as time-varying covariates had no major effect on the shape of IL-1Ra trajectories, suggesting that the upregulation of IL-1Ra cannot directly be attributed to weight gain. Differences in secretion of leptin, an adipokine strongly upregulated in obesity that can stimulate IL-1β production and lead to increased IL-1Ra release, may be one of the mechanisms involved (34
We did not adjust for glycemia, because our stratifying variable (incident diabetes caseness) already includes fasting and/or postload glucose in its definition, which means that we have already adjusted for glucose values at time 0. Because fasting insulin is not included in the diagnostic criteria, we performed an analysis adjusted for fasting insulin, which substantially attenuated the slope difference between case and control subjects preceding the diagnosis of diabetes. However, a significant acceleration of the slope from years −6 to 0 remains among case subjects, indicating that fasting insulin explains or mediates some, but not all, of the late increase in IL-1Ra. It should be kept in mind that with our study design, we cannot directly establish whether this attenuation is due to confounding, mediation, or shared causation.
Our findings point to the possibility that shape of biomarker trajectories is informative in terms of assessing their predictive value for different time windows. Sequential measurements of cytokines and other biomarkers and the characterization of individual trajectories may improve the estimation of type 2 diabetes risk. Differences in IL-1Ra levels between case and control subjects varied considerably over the lead time such that IL-1Ra may be more useful in predicting short-term diabetes risk, whereas other cytokines may be more strongly associated with long-term risk of type 2 diabetes. To date, attempts to improve diabetes risk prediction in the general population based on biomarker panels measured at a single time point have produced only marginal improvements compared with conventional risk models. It remains to be determined whether it will be possible to improve risk prediction at different pre-diabetic stages in the general population by considering serial biomarker measurements over time.
From a therapeutic point of view, it is noteworthy that recombinant IL-1Ra has been shown to improve metabolic control in patients with type 2 diabetes (13
). In this light, an upregulation of IL-1Ra may be expected to be protective rather than associated with increased risk. In our study, it appears that the steep increase of IL-1Ra levels by approximately one-third was by far not sufficient to prevent the onset of type 2 diabetes. In the anakinra trial, an improvement of glycemia and β-cell function was accompanied by supraphysiological IL-1Ra peak levels in serum that were >1,000-fold higher in the intervention group than in the placebo group (13
Our study has some limitations and strengths that should be acknowledged. First, the study design does not enable us to exclude the alternative interpretation that the increase of IL-1Ra before type 2 diabetes contributes to the disease risk and represents a causal factor rather than an anti-inflammatory response. However, data from the anakinra trial (13
) as well as preclinical data (12
) argue strongly against diabetogenic effects of IL-1Ra. Second, we did not determine IL-1β levels and therefore cannot know whether IL-1β increases before type 2 diabetes and whether such an increase precedes the increase in IL-1Ra. In addition, we did not have data from all three study phases for other proinflammatory markers, such as CRP and IL-6, for a comparison of trajectories. We did not adjust for the available CRP or IL-6 levels at baseline, because this adjustment has the capacity to influence only the intercept of the model, not the shape of the trajectories, and therefore would not help to answer the question of temporal sequences. Third, the Whitehall II study is an occupational cohort and as such is not population based. Although the cohort was healthy at recruitment, as in a community-based sample, the cohort overrepresents white, male, and middle-aged individuals. Fourth, due to missing data, approximately two-thirds of the source population at phase 3 was excluded from the present analysis. However, more than 300 incident cases and a total of almost 6,000 measurement points for the trajectories led to sufficient statistical power to detect differences between case and control subjects. In addition, our dropout analysis (supplementary Table 1) indicates that exclusions are unlikely to have affected internal validity.
The study has notable strengths. It is based on a well-phenotyped cohort, and more than half of incident diabetes cases were diagnosed based on the gold standard oral glucose tolerance test. We applied a sophisticated methodology that considered the interrelation among repeated measurements from the same individual at different time points during the follow-up and based our analysis on a large sample, as noted above.
In conclusion, we characterized cytokine trajectories to understand the evolution of the association between IL-1Ra levels in the circulation and type 2 diabetes before the diagnosis of diabetes. IL-1Ra levels showed an accelerated increase during the last 6 years preceding diagnosis. We showed that crude measures of adiposity did not explain the IL-1Ra trajectory, potentially implicating other factors in disease susceptibility. These data support the hypothesis that an anti-inflammatory response counterbalances metabolic and immunologic disturbances preceding type 2 diabetes. Moreover, these results suggest that multiple measurements of inflammation-related and other biomarkers can markedly improve our understanding of the pathogenesis of type 2 diabetes. The clear temporal characterization shown by our data may help define the optimal clinical use of these biomarkers.