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. 2004 Nov;138(2):323–329. doi: 10.1111/j.1365-2249.2004.02599.x

Imbalance between interleukin-1 agonists and antagonists: relationship to severity of inflammatory bowel disease

O LUDWICZEK *,§, E VANNIER †,§, I BORGGRAEFE †, A KASER *, B SIEGMUND ‡, C A DINARELLO ‡, H TILG *
PMCID: PMC1809217  PMID: 15498044

Abstract

Interleukin (IL)-1 is a key mediator in the pathogenesis of inflammatory bowel disease (IBD). Naturally occurring IL-1 modulators include IL-1 receptor antagonist (IL-1Ra), IL-1 soluble receptor Type I (IL-1sRI), IL-1sRII and IL-1 receptor accessory protein (AcP). Systemic and mucosal levels of IL-1 soluble receptors remain unknown in IBD. Plasma or colonic tissues were obtained from 185 consecutive unselected patients with Crohn's disease (CD) or ulcerative colitis (UC) and from 52 control subjects. Plasma and colonic explant culture supernatants were assessed for IL-1α, IL-1β, IL-1Ra, IL-1sRI and IL-1sRII. Plasma IL-1Ra levels were higher in UC (+93%) than in healthy subjects. IL-1α and IL-1β were not detected. IL-1sRII levels were marginally lower in CD (−10%) and UC (−9%), whereas IL-1sRI levels were elevated in CD (+28%) only. Plasma IL-1sRI levels correlated positively (P < 0·01) with Crohn's disease activity index (r = 0·53), C-reactive protein (r = 0·46) and α1-acid glycoprotein (r = 0·42). In colonic explant cultures, IL-1α and IL-1Ra levels were elevated in non-lesional (+233% and +185% respectively) and lesional CD (+353% and +1069%), lesional UC (+604% and +1138%), but not in non-lesional UC. IL-1β was elevated in lesional UC (+152%) and CD (+128%). In contrast, IL-1sRII levels were elevated in non-lesional CD (+65%), but remained unchanged in lesional CD, non-lesional and lesional UC. IL-1sRI levels did not differ between patient and control groups. These results indicate that (i) the proinflammatory moiety IL-1sRI is a systemic marker of inflammation and activity in CD and (ii) local shedding of the functional antagonist IL-1sRII may dampen colonic inflammation in CD, but not in UC.

Keywords: cytokine receptors, cytokines, human, inflammation

INTRODUCTION

Ulcerative colitis (UC) and Crohn's disease (CD) are chronic relapsing inflammatory bowel diseases (IBD) of unknown etiology [1]. Several exogenous factors, such as bacterial cell wall components, viruses or dietary products, are thought to initiate and/or perpetuate a sequence of chronic immune processes that are not down-regulated appropriately in genetically predisposed individuals and that may lead to intestinal mucosal injury [1–3]. Cytokines, such as tumour necrosis factor (TNF)-α and interleukin (IL)-1, play a major role in the development of IBD [4–11]. TNF-α and IL-1β induce synthesis of chemokines, including IL-8, a potent neutrophil chemoattractant [12,13]. Activation of neutrophils recruited to the intestinal mucosa results in the synthesis of proinflammatory cytokines [1] and the release of neutrophil granule enzymes involved in oxidative burst and tissue damage, such as myeloperoxidase [14]. Accordingly, tissue levels of IL-1 correlate closely with the degree of mucosal inflammation and necrosis [15]. These observations suggest that IL-1 is one of the critical mediators of intestinal inflammation in IBD.

Activities of IL-1 are, in part, regulated by the naturally occurring inhibitor IL-1 receptor antagonist (IL-1Ra) [5,16–18]. IL-1Ra specifically inhibits IL-1 activities by binding to IL-1 receptors, but does not display agonist activity [5,12,16,19–22]. In the intestinal mucosa, epithelial cells and lamina propria mononuclear cells are the major sources of IL-1Ra [23]. An imbalance between the production of IL-1 and IL-1Ra has been described in freshly isolated intestinal mucosal cells [24] and in colonic mucosal biopsies obtained from inflamed intestinal tissue of IBD patients [25]. Administration of recombinant IL-1Ra prevents mucosal inflammation and necrosis in a rabbit model of dextran-induced colitis [19,26]. Conversely, neutralization of endogenous IL-1Ra increases the severity of intestinal inflammation [22], indicating that endogenous IL-1Ra plays an anti-inflammatory role. The importance of IL-1 and IL-1Ra in the pathogenesis of IBD has been corroborated by the association between carriage of IL-1RN allele 2, low production of IL-1Ra and severity of disease in UC patients of Jewish or Hispanic genetic background [27].

All activities of IL-1 are mediated via the IL-1 receptor type I (IL-1RI) [5]. Myeloid cells also express IL-1 receptor type II (IL-1RII), a decoy receptor that presents a high degree of homology with the extracellular domain of IL-1RI [5]. However, IL-1RII has a short (29 residue) intracytoplasmic domain that does not bind MyD88 and IRAK, signalling units otherwise recruited to members of the Toll-like receptor/IL-1R family [28]. IL-1RII is therefore a naturally occurring functional antagonist of IL-1. IL-1 receptors play an additional role in the control of IL-1 activities through the proteolytic cleavage of their extracellular domains. Shedding of IL-1RII yields the IL-1 soluble receptor type II (IL-1sRII) which binds IL-1β avidly, but IL-1α and IL-1Ra with low affinity [17,29–31]. Thus, IL-1sRII contributes to IL-1 antagonism through the preferential neutralization of IL-1β activity [29,32,33]. In contrast, IL-1 soluble receptor type I (IL-1sRI) retains the ability of membrane-bound IL-1RI to bind IL-1Ra and IL-1α with greater affinity than IL-1β, and is therefore regarded as a proinflammatory moiety [30,33–35]. To date, the release and functional importance of IL-1 soluble receptors in intestinal inflammation remain poorly understood. Strong et al. reported that intestinal mucosal mesenchymal cells contain detectable levels of IL-1RII transcripts [36], but undetectable levels of IL-1RI transcripts. On the other hand, Panja et al. [37] observed IL-1β binding to intestinal epithelial cells freshly isolated from normal subjects and IBD patients. However, there has been no report on surface expression or shedding of IL-1RI and IL-1RII by intestinal epithelial cells or lamina propria mononuclear cells from patients with CD or UC.

Following binding of IL-1α or IL-1β to the type I IL-1R, a second protein, the IL-1 receptor accessory protein (AcP), is recruited to the complex [38]. Both proteins, IL-1R and AcP, are required for signalling. In addition to the transmembrane form of AcP another form exists, encoded by an alternatively spliced mRNA [38,39]. This form contains only the extracellular domain of AcP. Recently, it has been shown that soluble human AcP can associate with ligand-bound soluble IL1R–II. Association of sIL1R-II with AcP increases the affinity of binding to both human IL-1α and human IL-1β by approximately two orders of magnitude, which renders human sIL1R-II an effective inhibitor of the former as well as the latter [40].

The aim of the present study was to assess whether the balance between the proinflammatory moieties (IL-1α, IL-1β and IL-1sRI) and the naturally occurring IL-1 antagonists (IL-1Ra and IL-1sRII) is altered in patients with IBD, at the systemic and intestinal mucosal levels. We also investigated whether circulating and mucosal levels of IL-1 modulators correlate with severity of disease in CD and UC.

MATERIALS AND METHODS

Study subjects for plasma samples

The study enrolled 141 patients with IBD. Ninety-eight patients were diagnosed with CD, 43 with UC. Diagnosis was made by radiology, endoscopy and histology examinations. Thirty-nine healthy volunteers were recruited as control subjects. Patient characteristics are reported in Tables 1 and 2. Note that CD patients had a median age of 34 years (range: 17–76) whereas UC patients had a median age of 48 years (range: 18–64). None of the studied CD patients had received infliximab before blood sampling. Healthy controls had a median age of 33 years (range: 24–59), 21 were female, 18 male. Informed consent was obtained from each subject. Approval from the local ethical committee was obtained. If patients underwent colonoscopy, blood was sampled before the endoscopic procedure. From the 32 CD patients with fistulas or abscesses, 25 were receiving metronidazole.

Table 1.

Clinical features of patients with Crohn's disease

Plasma Colonic biopsies
Female/male 55/43 13/7
Age (years) 34 (range: 17–76) 36 (range: 16–69)
Disease duration (months) 32 (range: 1–372) 13 (range: 1–146)
Therapy
 No specific therapy 19 7
 Aminosalicylates 38 3
 Aminosalicylates + steroids 13 4
 Steroids 11 0
 Azathioprine 17 6
Site of inflammation
 Small bowel 30 4
 Colon 31 10
 Small bowel + colon 31 6
 Small bowel + colon + stomach 6 0
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Table 2.

Clinical features of patients with ulcerative colitis

Plasma Colonic biopsies
Female/male 16/27 12/12
Age (years) 48 (range: 18–64) 36 (range: 16–57)
Disease duration (months) 36 (range: 1–276) 358 (range: 0–2163)
Therapy
 No specific therapy 7 5
 Aminosalicylates 23 7
 Aminosalicylates + steroids 6 5
 Steroids 2 3
 Azathioprine 5 4
Site of inflammation
 Proctitis 11 4
 Left-sided colon 19 11
 Pancolitis 13 9
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Study subjects for colonic biopsies

We enrolled 44 unselected patients with IBD. Patient characteristics are reported in Tables 1 and 2. There were 20 patients with CD (median age: 36 years, range: 16–69) and 24 patients with UC (median age: 36 years, range: 16–57). For some IBD patients, colonic explants were obtained from both inflamed (lesional) and non-inflamed (non-lesional) areas. Inflammation was assessed macroscopically using commonly accepted criteria, such as vulnerability, ulceration, colour and vascular pattern of the mucosa. The control cohort was made of 13 healthy subjects with no known concomitant disease who underwent colonoscopy for tumour screening (six female, seven male). The control population had a median age of 57 years (range: 45–70). Approval from the local ethical committee was obtained.

Plasma collection

For each subject a 9-ml sample of venous blood was collected on EDTA, and 1-ml aliquots of plasma were stored at −70°C. A consecutive code number was assigned to each sample prior to measurement in a blinded manner.

Clinical laboratory parameters

Serum samples were analysed for C-reactive protein (CRP) and α1-acid glycoprotein (α1-AGP) at the clinical laboratory of the University Hospital, Innsbruck, Austria. Plasma CRP was measured in an immuno-turbidimetric assay (BM/Hitachi 717, Tokyo, Japan). Minimum detection limit was 0·7 mg/dl. Serum α1-AGP was measured by an immuno-turbidimetric assay (AAG-Roche, Cobas Integra, Basel, Switzerland). Minimum detection limit was 8 mg/dl. Erythrocyte sedimentation rate (ESR) was determined by standard laboratory technique.

Colonic explant cultures

Upon endoscopic removal, colonic biopsies were immediately placed in phosphate-buffered saline (PBS) (Biochrom KG, Berlin, Germany) at room temperature. Biopsies were weighed (average weight: 7·65 mg, range: 3·03–16·65 mg), washed three times in PBS at room temperature and placed in 48-well culture plates. Each biopsy was placed in a well containing 1 ml RPMI-1640 supplemented with 10% fetal calf serum (FCS) (Life Technologies, Schoeller Pharma, Vienna, Austria), 100 U/ml penicillin and 100 µg/ml streptomycin (Schoeller Pharma). Cultures were maintained for 24 h at 37°C in a humidified atmosphere containing 5% CO2. Cultures were harvested, centrifuged at 1000 g for 10 min and cell-free supernatants aliquoted for storage at −70°C. Levels of IL-1, IL-1Ra and soluble IL-1R were normalized to biopsy weight.

Measurement of IL-1 soluble receptors

Radioimmunoassays (RIA) for IL-1sRI and IL-1sRII have been described elsewhere [41].

IL-1sRI was not recognized by the polyclonal antibody raised against IL-1sRII, and vice-versa. IL-1α, IL-1β or IL-1Ra (10 ng/ml) added to the RIAs did not affect the detection of IL-1sRI or IL-1sRII by their specific polyclonal antibodies. These results indicate that our polyclonal antisera detect IL-1sR whether unbound or complexed to their preferential ligands, i.e. IL-1α and IL-1Ra for IL-1sRI, and IL-1β for IL-1sRII.

Measurement of IL-1Ra

The RIA specific for human IL-1Ra has been described previously [42]. Briefly, samples or standards were incubated for 24 h at room temperature with diluted (1 : 25 000) polyclonal anti-IL-1Ra antiserum. Bolton–Hunter-labelled [125I]-IL-1Ra [10 000 counts per minute (cpm) per condition] was added. After 24 h at room temperature, immunoprecipitates were formed by addition of goat antirabbit IgG and polyethylene glycol at respective final concentrations of 1% and 3%. After 60 min at room temperature, immunoprecipitates were recovered by centrifugation at 1500 g for 15 min.

Measurement of IL-1

IL-1α and IL-1β were detected using commercially available enzyme-linked immunosorbent assays (ELISAs (R&D Systems, Minneapolis, MN, USA)). The detection limit of the IL-1α assay was 1 pg/ml, the detection limit of the IL-1β assay was 2 pg/ml.

Statistical analysis

Results are expressed as mean ± standard error of the mean (s.e.m.). Mean IL-1 agonist (or antagonist) levels in plasma (or colonic biopsies) from healthy subjects were compared with those from patients with CD or UC by Kruskal–Wallis testing (SPSS, SPSS Inc., Chicago, IL, USA). Subgroups were compared using Mann–Whitney testing. Significance of the correlations was assessed by calculating the Pearson's correlation coefficient. To analyse influences of differences in gender and age between the patient and control groups, univariate analysis of variance was performed. P-values less than 0·05 were considered significant.

RESULTS

Elevated circulating levels of IL-1sRI, but decreased levels of IL-1sRII in IBD patients

As shown in Fig. 1a, plasma levels of IL-1sRI in UC patients (1·86 ± 0·11 ng/ml, n = 43) did not differ (P = 0·17) from those in healthy subjects (1·63 ± 0·05 ng/ml, n = 39). On the other hand, plasma levels of IL-1sRI were significantly higher (P < 0·01) in patients with uncomplicated CD (no fistulas/no abscesses) (2·03 ± 0·10 ng/ml, n = 66, P < 0·05) than in healthy subjects. CD patients with fistulas and/or abscesses had significantly higher IL-1sRI plasma levels (2·46 ± 0·14 ng/ml, n = 32) than CD patients with no complication (2·03 ± 0·10 ng/ml, n = 66, P < 0·05, data not shown). For all measured variables, no statistically significant influence of differences in gender and age distribution between compared groups were found.

Fig. 1.

Fig. 1

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Plasma levels of IL-1 agonists and antagonists in ulcerative colitis and Crohn's disease. One hundred and nine unselected, consecutive patients with IBD were enrolled in the study, 43 with ulcerative colitis (UC) and 66 with Crohn's disease (CD). Thirty-nine healthy subjects served as controls. Blood was collected on EDTA and plasma obtained. IL-1α and IL-1β were undetectable by ELISA. Levels of IL-1sRI (a), IL-1sRII (b) and IL-1Ra (c) were measured using specific RIAs. Results are expressed as mean ± s.e.m. Statistically significant differences between two groups are shown by brackets.

In contrast to IL-1sRI levels, IL-1sRII levels were decreased in patients with uncomplicated CD (4·74 ± 0·11 ng/ml, n = 66, P < 0·01) and in patients with UC (4·83 ± 0·10 ng/ml, n = 43, P < 0·05), when compared to healthy subjects (5·31 ± 0·13 ng/ml, n = 39) (Fig. 1b). IL-1sRII levels were slightly lower in uncomplicated CD (4·74 ± 0·11 ng/ml compared to complicated CD (4·97 ± 0·10 ng/ml, P = 0·05, data not shown). IL-1sRII levels did not differ significantly between CD and UC patients.

Circulating levels of IL-1 family members in IBD patients

IL-1α and IL-1β were neither detected in the plasma from healthy controls nor in the plasma from IBD patients. Plasma levels of IL-1Ra were significantly elevated in patients with UC (0·54 ± 0·09 ng/ml, P < 0·01) (Fig. 1c). In patients with uncomplicated CD, IL-1Ra levels were 0·45 ± 0·03 ng/ml compared to 0·28 ± 0·01 ng/ml in controls, but this difference was not statistically significant (P = 0·191). There was no significant difference between IL-1Ra levels in uncomplicated and complicated CD patients (uncomplicated CD: 0·45 ± 0·03 ng/ml; complicated CD: 0·52 ± 0·04 ng/ml, P = 0·22, data not shown). However, the difference between controls and complicated CD was statistically significant (P < 0·05).

Correlations between markers of disease activity and plasma levels of IL-1 modulators

Disease activity was monitored using the ESR, plasma levels of CRP and α1-AGP, and circulating leucocyte counts. In addition to these laboratory parameters, disease activity was assessed using the Crohn's disease activity index (CDAI) for patients with CD and the Rachmilewitz score [43] for patients with UC.

In CD patients (Table 3), CDAI correlated positively with plasma levels of IL-1sRI (r = 0·53, P < 0·01, n = 40), but not with those of IL-1sRII. IL-1sRI levels further correlated positively with ESR (r = 0·40, P < 0·01, n = 60), CRP (r = 0·46, P < 0·001, n = 59), and α1-AGP levels (r = 0·42, P < 0·01, n = 58) whereas IL-1sRII levels correlated positively with leucocyte counts (r = 0·29, P < 0·05, n = 72).

Table 3.

Correlations between plasma levels of IL-1 soluble receptors and IL-1 receptor antagonist and laboratory parameters in Crohn's disease

Laboratory parameter IL-1sRI IL-1sRII IL-1Ra
ESR r = 0·40, P < 0·01 n.s. n.s.
Leucocytes n.s. r = 0·29, P < 0·05 n.s.
CRP r = 0·46, P < 0·001 n.s. n.s.
α1-AGP r = 0·42, P < 0·01 n.s. n.s.
CDAI r = 0·53, P < 0·01 n.s. n.s.
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α1-AGP: α1-acid glycoprotein; CDAI: Crohn's disease activity index; CRP: C-reactive protein; ESR: erythrocyte sedimentation rate; n.s. = P > 0·05.

In UC patients (Table 4), the Rachmilewitz index did not correlate with plasma levels of IL-1sRI or IL-1sRII. Similarly, there was no correlation between plasma levels of IL-1 soluble receptors, on one hand, and ESR, CRP or α1-AGP levels, on the other hand. IL-1Ra levels correlated positively with leucocyte counts (r = 0·39, P < 0·05, n = 34).

Table 4.

Correlations between plasma levels of IL-1 soluble receptors and IL-1 receptor antagonist and laboratory parameters in ulcerative colitis

Laboratory parameter IL-1sRI IL-1sRII IL-1Ra
ESR n.s. n.s. n.s.
Leucocytes n.s. n.s. r = 0·39, P < 0·05
CRP n.s. n.s. n.s.
α1-AGP n.s. n.s. n.s.
Rachmilewitz Index n.s. n.s. n.s.
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α1-AGP: α1-acid glycoprotein; CRP: C-reactive protein; ESR: erythrocyte sedimentation rate; n.s. = P > 0·05.

Release of IL-1sRI and IL-1sRII from colonic explants

As reported in Fig. 2a, there was no difference in levels of IL-1sRI between lesional and non-lesional biopsies from IBD patients, and between biopsies from control subjects and biopsies from IBD patients.

Fig. 2.

Fig. 2

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Release of IL-1 agonists and antagonists by colonic explants from patients with ulcerative colitis or Crohn's disease. Biopsies were obtained by endoscopy of 44 unselected, consecutive patients with ulcerative colitis (UC, n = 24) or Crohn's disease (CD, n = 20). The control group was made of 13 patients who underwent colonoscopy for tumour screening. Biopsies were left unstimulated, and cultured for 24 h. Culture supernatants were assessed for IL-1sRI (a), IL-1sRII (b), IL-1α (c), IL-1β (d), and IL-1Ra (e). Levels of IL-1 agonists and antagonists are expressed as pg/ml/mg biopsy weight. Results are expressed as mean ± s.e.m. Statistically significant differences between two groups are shown in brackets.

In contrast, levels of IL-1sRII in cultures of non-lesional biopsies from CD patients (99·07 ± 11·12 pg/ml/mg, n = 11) were higher than those of lesional biopsies (62·33 ± 5·88 pg/ml/mg, n = 8, P < 0·01) and control subjects (60·14 ± 5·23 pg/ml/mg, P < 0·01) (Fig. 2b). For UC patients, levels of IL-1sRII in cultures of non-lesional biopsies (68·96 ± 5·24 pg/ml/mg) did not differ from those in cultures of lesional biopsies (60·88 ± 6·50 pg/ml/mg) or biopsies from control subjects (Fig. 2b).

Production of IL-1α and IL-1β by colonic explants

In UC patients levels of IL-1α in cultures of non-lesional biopsies (1·04 ± 0·35 pg/ml/mg, n = 9) did not differ from those from control subjects (0·99 ± 0·47 pg/ml/mg, P < 0·01) However, cultures of lesional biopsies contained more IL-1α (6·97 ± 2·20 pg/ml/mg, n = 10) than those of non-lesional biopsies (P < 0·01) and those from control subjects (P < 0·001).

In CD patients, the levels of IL-1α in cultures of lesional biopsies (4·49 ± 1·00 pg/ml/mg, n = 8) did not differ significantly from those in cultures of non-lesional biopsies (3·31 ± 1·44 pg/ml/mg, n = 8, P = 0·065), but were higher than those in control subjects (P < 0·01). Non-lesional biopsies were also significantly higher than controls (P < 0·05) (Fig. 2c).

As shown in Fig. 2d, colonic explants from healthy controls secreted 25·22 ± 9·49 pg/ml/mg IL-1β. There was a significant difference between controls and lesional areas in UC (63·62 ± 19·70, P = 0·01), but not between controls and non-lesional UC (22·08 ± 12·53 pg/ml/mg).

In CD patients, both non-lesional (49·32 ± 21·30 pg/ml/mg) and lesional (57·52 ± 6·03 pg/ml/mg) biopsies were higher than controls, but only in lesional CD was the difference statistically significant (P < 0·01). The difference between non-lesional and lesional was not significant.

Production of IL-1Ra by colonic explants

In UC patients, colonic tissues from lesional areas (322 ± 58 pg/ml/mg, n = 11, P < 0·001) produced highly significantly more IL-1Ra than tissues from non-lesional areas (32 ± 5 pg/ml/mg, n = 9). There was no difference between non-lesional biopsies and biopsies from control subjects (26 ± 7 pg/ml/mg, P < 0·001). However, the difference between lesional biopsies from UC patients and biopsies from control subjects was highly significant (P < 0·001) (Fig. 2e).

There was a significant difference between non-lesional biopsies from CD patients (74 ± 25 pg/ml/mg, n = 7) and biopsies from control subjects (P < 0·05). Lesional biopsies from CD patients secreted more IL-1Ra (304 ± 115 pg/ml/mg, n = 9) than biopsies from control subjects (P = 0·001). The difference between non-lesional and lesional biopsies was also significant (P = 0·05).

DISCUSSION

One of the factors participating in the initiation and perpetuation of inflammation in IBD may be an inappropriate production of anti-inflammatory cytokines, resulting in a disturbed balance of proinflammatory versus anti-inflammatory cytokines. Although increased levels of IL-1 and an imbalance between IL-1 and IL-1Ra have been documented [12], the role of other IL-1 modulators in IBD has not yet been elucidated. The aim of the present study was to gain a more complete understanding of the IL-1 family members in IBD. For this purpose, plasma levels of IL-1α, IL-1β, IL-1Ra, IL-1sRI and IL-1sRII in patients with CD and UC were compared to those measured in a cohort of healthy subjects. Additionally, we monitored the release of IL-1α, IL-1β, IL-1Ra, IL-1sRI and IL-1sRII by colonic biopsies from patients with CD or UC, and from subjects with no inflammatory intestinal condition.

Patients with CD and UC were characterized by elevated plasma levels of IL-1Ra, but decreased plasma levels of IL-1sRII. In addition, plasma IL-1sRI levels in CD were elevated. Thus, the balance between the proinflammatory IL-1sRI and the anti-inflammatory IL-1sRII is altered in IBD patients. This shift towards an inflammatory state was supported further by the correlation between IL-1sRI levels and biological markers of disease activity (ESR, CRP, α1-AGP) or the CDAI in CD patients. The absence of correlation between these parameters and IL-1sRII (except for leucocyte counts in CD patients) underlines the lack of significant increase of the anti-inflammatory IL-1sRII. In UC patients, neither the parameters of inflammation nor the Rachmilewitz index correlated with IL-1sRI or IL-1sRII. This could be explained by differences in the kinetics of the appearance of these agents in the systemic circulation.

The increase in plasma levels of IL-1Ra in IBD patients is corroborated by the results from whole tissue cultures from colonic biopsies. However, the parallel between local and systemic compartments is not seen with IL-1sRI. In contrast to plasma levels, there was no difference in release of IL-1sRI by biopsies from IBD patients and healthy controls. This may be explained by the more ubiquitous expression of the IL-1RI gene. Thus, the contribution of the gut mucosa to systemic IL-1RI levels is likely to be marginal when compared to the many tissues expressing the IL-1RI gene and releasing IL-1sRI under normal and pathological conditions.

Our results underline the important role of IL-1α/β in gut inflammation. Both IL-1β and IL-1α were elevated in UC patients, compared to healthy controls. Overproduction was restricted to biopsies taken from inflamed areas, whereas biopsies from non-inflamed areas did not differ from healthy controls. In cultures of lesional biopsies from CD patients, there were higher levels of IL-1α and IL-1β. Differently to UC, however, in CD patients IL-1α was also elevated in non-inflamed areas. IL-1β was also elevated in non-lesional areas from CD patients, but this difference failed to reach statistical significance. These results suggest differences in the course of inflammation between UC and CD. Whereas in non-inflamed gut mucosa in UC IL-1 levels are not different to gut mucosa from healthy individuals, there is a state of inflammatory activity − although compensated − in macroscopically apparently normal mucosa in CD patients. This hypothesis is also strengthened by the results of analysing IL-1Ra and IL-1sRII. While there is still no difference between healthy controls and non-lesional UC, both IL-1 antagonists are elevated in non-lesional CD. These findings suggest that, in UC, inflammation is a more local event, restricted to apparently affected mucosa, whereas in CD even in macroscopically normal mucosa there is an overexpression of proinflammtory cytokines which is, however, compensated by a parallel up-regulation of endogenous anti-inflammatory agents. In lesional areas this compensation breaks down, leading to an overwhelming inflammatory reaction.

Although up-regulation of IL-1Ra seems to be an appropriate response to control IL-1-mediated inflammation, this response is not sufficient. Several mechanisms could explain the failure of the naturally occurring anti-inflammatory mediators to set a new balance. First, a large excess of IL-1Ra is required to block the binding of IL-1 to the IL-1 type I receptor [44]. It is generally thought that a 10–100-fold excess of IL-1Ra is required to achieve efficient antagonism. This means that although an increase in IL-1Ra levels is an adequate response to contain an otherwise overwhelming inflammatory reaction, the extent of this reaction might not be sufficient to circumvent the pathological process. This hypothesis is supported by the observation that IBD is characterized by a specific decrease in the IL-1Ra/IL-1 ratio [12]. This decrease might not only result from an inadequate increase in IL-1Ra, but also from an enhanced IL-1Ra-mediated IL-1 secretion. Indeed, studies with IL-1Ra-deficient mice and IL-1Ra transgenic mice have shed some light on the relationship between IL-1Ra levels and IL-1 synthesis. Unexpectedly, serum IL-1α levels following an intraperitoneal challenge with endotoxin were decreased in IL-1Ra-deficient mice but were increased in IL-1Ra-overproducing mice, indicating that IL-1Ra is a positive regulator of IL-1α synthesis in endotoxaemia [45]. This seems to be supported by our results, where increased IL-1α levels coincide with enhanced secretion of IL-1Ra.

In conclusion, our results suggest that IBD is associated with an imbalance of IL-1 modulators that may lead to the perpetuation of IL-1-mediated inflammation. Several mechanisms may be responsible: (i) the lack of increased release of the functional IL-1 antagonist IL-1sRII by inflamed colonic tissue from IBD patients. Plasma levels of IL-1sRII are even decreased in IBD patients. (ii) IL-1Ra might play an ineffective role, although an inducible IL-1 antagonist, elevated IL-1Ra levels might not be sufficient to counteract IL-1-mediated inflammation. Increased IL-1Ra secretion might be compensated by (iii) elevated IL-1sRI levels that, in turn, may contribute to (iiii) increased IL-1α secretion, hence, an imbalance favouring chronic proinflammatory conditions, such as IBD.

Acknowledgments

Drs Edouard Vannier and Ingo Borggraefe were supported by the Alfond Family Research Fund. These studies were supported by the Tufts-NEMC Research Fund (to E. V.) and by the Austrian Science Fund P14641 and P15783 (to H. T.).

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