Interleukin-6 is an independent predictor of mortality in patients starting dialysis treatment

Roberto Pecoits-Filho, Peter Bárány, Bengt Lindholm, Olof Heimbürger and Peter Stenvinkel

Divisions of Renal Medicine and Baxter Novum, Department of Clinical Science, Karolinska Institutet, Huddinge University Hospital, Stockholm, Sweden



   Abstract
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
Background. The mortality rate is high among end-stage renal disease (ESRD) patients, and recent evidence suggests that this may be linked to inflammation. The activity of interleukin-6 (IL-6) and its soluble receptor (sIL-6R) are markedly up-regulated in ESRD patients, and plasma IL-6 levels predict outcome in haemodialysis (HD) patients. However, it has not been established whether elevated plasma IL-6 also predicts outcome in ESRD patients treated by peritoneal dialysis (PD), and how it relates to the data on HD patients. The predictive power of sIL-6R levels on outcome is also unknown in this patient population.

Methods. To determine whether or not plasma IL-6 and sIL-6R predict patient survival, we studied 173 ESRD patients (62% males, 53±1 years of age) near the initiation of dialysis treatment (99 PD, 74 HD patients). The patients were followed for a mean period of 3.1±0.1 years (range 0.1–7.1 years) and were stratified at the start of dialysis treatment according to age, gender, presence of cardiovascular disease, malnutrition (determined by subjective global assessment), diabetes mellitus, and IL-6 and sIL-6R plasma levels.

Results. A significantly different (P<0.0001) mortality rate was observed in different groups when patients were divided into quartiles according to IL-6 levels. Furthermore, the same differences were observed, less notably however, for sIL-6R (P<0.05). When patients were stratified according to IL-6 quartiles and analysed separately according to the different initial treatment groups, a similar profile of survival was observed for PD (P<0.01) and HD (P<0.05) patients. In a Cox proportional hazard model adjusting for the impact of age, malnutrition, diabetes mellitus and male gender, log IL-6 values were independently associated with poor outcome (P<0.05).

Conclusions. The present study demonstrates that the strong predictive value of elevated IL-6 levels for poor outcome in ESRD patients is similar in both HD and PD patients starting treatment.

Keywords: haemodialysis; interleukin-6; outcome; peritoneal dialysis; soluble interleukin-6 receptors



   Introduction
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
The annual mortality rate in dialysis patients remains high despite the marked improvement in dialysis technology and patient care. Many haemodialysis (HD) and peritoneal dialysis (PD) patients [14] show serological evidence of an activated inflammatory response, as clearly indicated by increased circulating levels of non-specific markers of inflammation and pro-inflammatory cytokines including interleukin-6 (IL-6). IL-6 plays a pivotal role in inflammation, being the most powerful inducer of the hepatic acute-phase response. The prototypes of this response, C-reactive protein (CRP) (positive phase) and serum albumin (negative phase) are both known as important predictors of mortality in end-stage renal disease (ESRD) [1,4]. Similarly, studies also found plasma IL-6 levels to be a strong predictor of mortality in HD patients [5] and in patients with normal renal function [6]. Elevated concentrations of IL-6 proved to be a predictor of mortality in the elderly population, independent of prevalent vascular disease, smoking and traditional risk factors, and stronger than, yet additive to, that for CRP [7]. Additionally, recent studies have pointed out the important role of IL-6 soluble receptors (sIL-6R) in determining the bioactivity of IL-6 [8], but its importance as a predictor of morbidity and mortality has not been investigated until now.

Hence, for the first time we report the results from a prospective study designed to determine predictors of outcome in ESRD patients starting dialysis treatment. Our results show a strong impact of plasma IL-6 levels identified shortly before the initiation of dialysis treatment on mortality in both HD and PD patients.



   Subjects and methods
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
Patients
One hundred and seventy-three (62% males, 29% diabetics) patients with ESRD were studied shortly before the onset of dialysis therapy. These patients represented a subgroup of patients enrolled in an ongoing prospective study of pre-dialysis patients, part of which has been presented previously [9]. The study exclusion criteria were age >70 years and unwillingness to participate in the study. Thirty-four per cent of the patients had clinical signs of cardiovascular, cerebrovascular and/or peripheral vascular disease and were defined and grouped as cardiovascular disease (CVD) patients. Thirty-five per cent of the patients had signs of malnutrition, as defined by a subjective global assessment score (SGA) of >1.

After an overnight fast, venous blood samples were taken for analysis of serum albumin, serum lipids, high-sensitivity CRP (hsCRP) (n=131), IL-6 and sIL-6R (n=135). Survival was evaluated after a mean follow-up of 3.1±0.1 years (range 0.1–7.1 years). The effects of age, gender, CVD, malnutrition, diabetes and Log serum IL-6 were determined with the Cox proportional hazard model and the relative risk of death was calculated. Survival was measured beginning with the day of sample collection until death or censoring, which was done at the end of the follow-up (7 February 2002). The prescribed dialysis doses were individualized to achieve the recommended current adequacy targets in both PD and HD patients.

Seventy-one patients received a kidney transplant after entering the study, but were not excluded from the follow-up. Additionally, patients who switched from PD to HD and vice versa during the study period were followed in the same manner as those who remained on the initial form of treatment. The Ethics Committee of the Karolinska Institutet approved the study protocol at Huddinge University Hospital, Stockholm, and informed consent was obtained from all patients.

Methods
Determinations of serum albumin (bromcresol purple method), and hsCRP (nephelometric analysis), were carried out in the Department of Clinical Chemistry, Huddinge University Hospital. Plasma IL-6 and sIL-6Rs were measured by a commercially available photometric enzyme-linked immunosorbent assay (ELISA) (Boehringer Mannheim, Mannheim, Germany and R&D Systems, Minneapolis, MN, USA, respectively).

Statistics
Values are presented as mean±SEM or medians with P<0.05 indicating significance. Comparisons between groups of patients were performed using the Mann–Whitney U-test, whereas comparisons between two groups for nominal variables were made with Fisher's exact test. Correlations were determined with Spearman rank analysis. Survival analyses were made with Kaplan–Meier and Cox regression analyses. For the survival analysis, patients were divided according to quartiles based on the plasma levels of IL-6 and sIL-6R.



   Results
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 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
The basal clinical characteristics of the two groups, divided according to initial treatment modality patients are given in Table 1Go. No significant differences were found at basal between the two dialysis-treatment groups. As expected, significant correlations were found between IL-6 and age (Rho=0.55, P<0.0001), serum albumin (Rho=–0.36, P<0.0001) and hsCRP (Rho=0.61, P<0.0001, n=131). In addition, IL-6 correlated to sIL-6R (Rho=0.18, P<0.05), total cholesterol (Rho=–0.15, P<0.05) and HDL cholesterol (Rho=0.21, P<0.005). Body mass index (BMI) was weakly but significantly correlated to both sIL-6R (Rho=0.22, P<0.05) and IL-6 (Rho=0.16, P<0.05) concentrations.


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Table 1.  Baseline clinical characteristics of patients starting dialysis treatment

 
Diabetic patients had similar plasma IL-6 concentrations [6.4 (range 1.3–35.1) pg/ml vs 6.4 (range 0.9–44.9) pg/ml] and sIL-6R (45.8±1.4 vs 43.6±0.9 ng/ml) compared to non-diabetics. Patients with CVD showed significantly higher levels of plasma IL-6 in comparison to the patients without CVD [9.5 (range 1.4–44.9) pg/ml vs 5.4 (range 0.9–25.2) pg/ml, P<0.001]. On the other hand, no difference was observed in sIL-6R (44.6±1.5 vs 44.2±1.0 ng/ml) levels between the two groups of patients. Malnourished patients (SGA>1) presented with significantly higher IL-6 levels than the non-malnourished [9.2 (range 1.1–44.9) pg/ml vs 5.1 (range 0.9–35.1) pg/ml, P<0.001], whereas similar sIL-6R levels were found between the two groups divided according to SGA (45.0±1.3 vs 42.8±1.0 ng/ml).

The Kaplan–Meier survival analysis for the overall population is shown in Figure 1Go. A highly significant difference in survival was observed for patient groups divided according to quartiles of IL-6 levels ({chi}2 20.8, P<0.0001). Furthermore, a significant, but less remarkable difference was observed for patient groups stratified according to sIL-6R levels ({chi}2 7.3, P<0.05). The survival analysis of the patient groups starting PD and HD separately, divided according to percentiles, is shown in Figure 2Go. The predictive impact of elevated IL-6 showed a similar profile in both PD ({chi}2 11.3, P<0.01) and HD ({chi}2 8.3, P<0.05) patients. The Cox proportional hazard model was applied to the overall study population to adjust event-free times for age, CVD, malnutrition, diabetes mellitus, Log IL-6, CVD and male gender (Table 2Go). While age, CVD, malnutrition, diabetes mellitus, Log IL-6 and male gender were associated with poor outcome in the univariate analysis, only CVD, malnutrition, diabetes and Log IL-6 were independently associated with mortality in a multivariate analysis (Table 2Go). The same model was applied to 125 patients with available sIL-6R plasma determination, to test the value of sIL-6R as an independent predictor of mortality. In this case, while malnutrition ({chi}2 5.1, P<0.05) and CVD ({chi}2 12.5, P<0.0005) were independent predictors of mortality, sIL-6R ({chi}2 0.9), gender ({chi}2 0.1), age ({chi}2 1.8) and diabetes mellitus ({chi}2 2.0) were not.



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Fig. 1.  Survival by Kaplan–Meier in patients divided into groups according to quartiles of IL-6 (n=173) and sIL-6R (n=134) concentrations at start of dialysis.

 


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Fig. 2.  Survival by Kaplan–Meier in patients divided into groups according to quartiles of IL-6 at start of PD (n=99) or HD (n=74) treatment.

 

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Table 2.  Adjusted hazards ratio for overall mortality in 164 patients starting dialysis treatmenta

 



   Discussion
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 
Markedly elevated circulating IL-6 levels are found in ESRD patients [10,11], which may be due to impaired removal of cytokines [12], and increased synthesis due to various infectious processes [13], co-morbid conditions such as coronary heart disease [14], chronic heart failure [15], increased body fat mass [16], as well as other as yet unknown factors [17]. The present study shows that elevated plasma IL-6 levels and the plasma concentration of its soluble receptor predicted outcome both in PD and HD patients followed for a mean period of more than 3 years. Importantly, the relationship between IL-6 and outcome was not altered in a multiple regression analysis adjusting for the impact of age, CVD, malnutrition, diabetes and gender. Thus, our results confirm those previously reported by Bologa et al. [5] demonstrating that elevated plasma IL-6 levels predicted outcome in 90 HD patients followed for 900 days. However, our data extend this finding, showing that the predictive value of elevated circulating IL-6 levels is similar in patients starting PD. Moreover, our study is, to the best of our knowledge, the first analysis of sIL-6 receptor, an important factor influencing the IL-6 system bioactivity, as a marker of poor outcome in ESRD.

Elevated circulatory IL-6 concentrations predict cardiovascular morbidity [6] and overall mortality [7] in non-renal as well as renal patients, there are reasons. First, it has been shown that IL-6 mediates malnutrition in dialysis patients (a strong predictor of outcome) [18] and stimulates the breakdown of muscle proteins [19]. IL-6 also seems to promote cancer cachexia [20]. Secondly, experimental evidence suggests an important role of IL-6 in the atherosclerotic process, as injection of recombinant IL-6 exacerbates early atherosclerosis in ApoE-deficient mice [21]. Moreover, increased IL-6 expression is found in the fibrous plaque stage of the atherosclerotic process [22], and we have recently established that IL-6 is an independent predictor of the progression of carotid atherosclerosis in patients on dialysis treatment for 12 months [23]. Thus, high circulatory IL-6 levels may contribute to malnutrition and atherosclerotic cardiovascular disease, both of which are strong predictors of mortality in dialysis patients [17]. The findings that baseline levels of plasma IL-6 predict patient survival many years prior to the events may seem puzzling; however, such associations have also been found for albumin and CRP (other markers of acute-phase response) in various groups of ESRD patients [1,2,6,7,17]. In our patient population, strong correlations between proteins of the acute phase (CRP, albumin) and IL-6 are demonstrated. The association between higher levels of IL-6 in the group of malnourished patients, and the negative correlation between IL-6 levels and total cholesterol and HDL cholesterol (as previously described by Bologa et al. [5]), point to a causal link between inflammation and nutritional status in ESRD.

An important finding in our study was the similar predictive power of IL-6 for outcomes comparing the patients in different renal replacement modalities. This may be of importance, as HD has been shown to induce inflammation to a greater extent than PD [2]. Thus, our findings support the hypothesis that factors associated with the uraemic syndrome per se, and not with the dialysis procedure, are the main determinants of chronic inflammation in ESRD [24].

This report is the first attempt to analyse the relationship between sIL-6R and clinical outcome in ESRD patients. In the present study, preliminary results, including part of our patient population, show that sIL-6R also predicts outcome in ESRD patients, although a correlation to CVD, malnutrition and co-morbidities was not evident. The soluble form of IL-6, most probably arising from the cleavage of the membrane form of the receptor, is an important regulator of IL-6 activity, since it binds to IL-6 in the circulation, expanding the IL-6/sIL-6R complex activity to organs containing the gp130 membrane-binding site [25]. Moreover, it was recently demonstrated in a combination of clinical and experimental settings that sIL-6R provides an important signalling pathway intimately involved in the transition between the early and late phase of the inflammatory response [8]. Thus, future longitudinal studies of inflammatory diseases should include sIL-6R in the analysis of IL-6 bioactivity.

In conclusion, we report that elevated circulating IL-6 predicts mortality similarly in patients starting PD and HD. This finding highlights the importance of inflammation as an unfavourable prognostic factor in ESRD patients.



   Acknowledgments
 
We acknowledge the skilled technical assistance of Ms A. Lif. This study was supported by the Baxter Extramural Grant Program (PS) and Trone Holsts Foundation (PS). During these studies, Roberto Pecoits-Filho was a recipient of a Research Award number 98/13385–6 from the São Paulo Foundation for Research Support (FAPESP).



   Notes
 
Correspondence and offprint requests to: Roberto Pecoits-Filho MD, Divisions of Renal Medicine and Baxter Novum, K-56 Huddinge University Hospital, S-141 86 Stockholm, Sweden. Email: roberto.pecoits\|[hyphen]\|filho{at}klinvet.ki.se Back



   References
 Top
 Abstract
 Introduction
 Subjects and methods
 Results
 Discussion
 References
 

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Received for publication: 17.12.01
Accepted in revised form: 18. 3.02