Interleukins IL-33 and IL-17/IL-17A in Patients with Ulcerative Colitis
Jasna Ajduković, MD, MSc,* Ante Tonkić, MD, PhD,† Ilza Salamunić, PhD,‡ Izet Hozo,
MD, PhD,† Miroslav Šimunić, MD, PhD,† Damir Bonacin, MD†
*General Practice, Sinj, Croatia
†Department of Internal Medicine, Split University Hospital Center, Split, Croatia
‡Deparment of Medical Laboratory Diagnosis, Split University Hospital Center, Split, Croatia
Short title: IL-33 and IL-17/IL-17A in Ulcerative Colitis
Fratarski prolaz 1
21230 Sinj, Croatia
Background: Disturbance of immune homeostasis in ulcerative colitis (UC) is related to the
predominance of T-helper-2 (Th2) immune response. Interleukin (IL)-33 stimulates Th
lymphocytes to produce Th2 cytokines, such as IL-4, IL-5, and IL-13, which are believed to
induce pathological changes in the intestinal mucosa. The pro-inflammatory role of IL-17 in
UC is still unclear. Our aim was to determine serum concentrations of IL-33 and IL-17 in
patients with UC and healthy controls.
Methods: Serum concentrations of IL-33 and IL-17 were measured in 18 patients (10 men)
with UC and 16 control subjects (10 men) by using two-layer immunoenzyme procedure
Results: Median serum concentrations of IL-33 in patients with UC and controls were 140
pg/mL (interquartile range [IQR], 72.5 pg/mL) and 165 pg/mL (IQR, 140.0 pg/mL),
respectively, but the difference was not statistically significant (Mann-Whitney U = 112, p =
0.281). The median serum concentration of IL-17/IL-17A in patients with UC was
significantly higher (100 pg/mL, IQR 35.75 pg/mL) than that in controls (65 pg/mL, IQR
32.25 pg/mL) (Mann-Whitney U = 55, p = 0.002).
Conclusion. Serum concentration of IL-33 in patients with UC was not increased in
comparison with that in controls, which is in accordance with current evidence that its
primary biological effect is transcriptional rather than cytokinal. Further research is needed to
explain whether increased concentration of IL-17 in UC is protective or harmful and to
elucidate its immunological and pathogenic role.
Key words: ulcerative colitis; IL-33 human; immunity, mucosal; interleukins; interleukin-
Ulcerative colitis (UC) is a chronic, disabling inflammatory bowel disease (IBD) of unknown
cause, whose pathogenesis is believed to involve aberrant immune response of the intestinal
mucosa. Various studies in IBD indicate that aberrant mucosal immune response in
genetically predisposed individuals may be induced by different environmental and
endogenous factors.[2-5] Increased secretion of cytokines and increased expression of HLA-
DR II molecules on epithelial cells have been found in IBD, as well as increased expression of
adhesion receptors on endothelial cells, which facilitates neutrophil entrance into the intestinal
lamina propria. The total number of T lymphocytes in the intestinal lamina propria of
patients with UC is also increased, although the ratio of CD4+ to CD8+ remains unchanged
(2:1). The number of T cells with expression of HLA-DR II is not increased, but the
number of those expressing the IL-2 receptor (CD25) may be increased, as well as of those
with memory phenotype CD45RO+. Furthermore, intestinal lamina propria in patients with
IBD contains 2-5 times more lymphocytes than the normal mucosa.
The disturbance of immune homeostasis in UC is related to the predominance of Th2 immune
response. Mucosal T lymphocytes in patients with UC secrete more Th2 cytokines IL-4, IL-5,
and IL-13,. The cytokine that stimulates this increased production is IL-33.. Thus, it
is believed that IL-33 induces pathologic changes in the intestinal mucosa via these Th2
IL-17, also called IL-17A, is produced by Th17 cells, dendritic cells (DCs) and γδ T cells.
It induces the release of different chemokines (e.g., IL-8), growth factors from mesenchymal
cells (e.g., granulocyte-macrophage colony-stimulating factor), proinflammatory cytokines
(e.g., IL-6), and co-stimulatory molecules (e.g., intercellular adhesion molecule-1) from
stromal/mesenchymal cells. The release of these molecules leads to the recruitment and
activation of neutrophils and increase in tissue inflammation. The role of IL-17 in the
development of intestinal inflammation has been disputed since it did not contribute
significantly to CD4+ T cell adoptive transfer colitis model and it played a protective role in
dextran sulfate sodium (DSS)-induced colitis.[15-17]
To the best of our knowledge, serum concentrations of IL-33 and IL-17 have not been
measured in patients with UC. Therefore, the aim of the study was to determine the
concentration of cytokines IL-33 and IL-17 in the serum of patients with UC and healthy
MATERIALS AND METHODS
The study included patients with previously confirmed UC who visited or were invited to visit
our Gastroenterology Outpatient Clinic or Department of Internal Medicine for follow-up
examination. Diagnosis of UC in all patients was based on patient history, physical findings,
colonoscopy, and histological findings including cryptitis or crypt abscesses, neutrofil
accumulation in the lamina propria, and reduced mucosa thickness. Of 21 eligible patients, 3
were excluded due to previous immunomodulatory or immunosuppressive therapy with
corticosteroids. The patient group included 18 patients (10 men and 8 women) aged between
20 and 65 years (median, 52 years), with the mean clinical activity index (CAI) of 4.5.
The control group consisted of 16 healthy subjects (10 men and 6 women) without history of
gastrointestinal symptoms. The control subjects were recruited from asymptomatic blood
donors and their median age was 46.5 years (range, 19-59 years).
Ethics Committee of the Split University Hospital Center approved the study and all study
subjects gave their informed consent to participate.
Blood samples were obtained between September 18, 2008 and April 01, 2009. After
centrifugation, serum samples were stored at -20 °C until analysis. For the quantitative
determination of IL-33 and IL-17 serum levels, we used the two-layer (sandwich)
immunoenzyme procedure (ELISA) on the semiautomatic analyzer mini-Boss (BIOMEDICA
ges. MbH, Vienna, Austria).
Quantitative assay for IL-33 was performed using Human IL-33 ELISA Quantitation Kit
(GenWay Biotech, Inc., San Diego, CA, USA), with the reference standard curve prepared by
using known amounts of IL-33 (0 - 500 pg/mL). The test sensitivity was 7 pg/mL.
The concentration of IL-17 was determined using Quantikine Human IL-17 Immunoassay
ELISA test (R&D Systems, Minneapolis, MN, USA) following the manufacturer’s
instructions. The reference standard curve was prepared using known amounts of IL-17 (0 -
2000 pg/mL). The test sensitivity was 15 pg/mL.
Data were presented as medians with interquartile range (IQR) and analyzed with Mann-
Whitney U- test. The level of statistical significance was set at p < 0.05. The statistical
analysis was performed with the Statistical Package for the Social Sciences 17.0 (SPSS Inc.,
Chicago, IL, USA).
The median serum concentration of IL-33 was 140 pg/mL (IQR, 72.5 pg/mL) in patients with
UC and 165 pg/mL (IQR, 140.0 pg/mL) in healthy control subjects (Figure 1). Although the
concentration of IL-33 was lower in patients with UC than healthy controls, the difference
was not statistically significant (Mann-Whitney U = 112, p = 0.281). On the other hand, the
significant difference was found in the serum concentration of IL-17, which was significantly
higher in patients with UC than healthy controls (Mann-Whitney U = 55, p = 0.002). The
median serum concentration of IL-17 measured in patients with UC was 100 pg/mL, whereas
its concentration in control subjects was 65 pg/mL (IQR, 32.3 pg/mL; Figure 2).
The serum values of IL-33 in patients with UC were not significantly different from those in
healthy controls, but the serum values of IL-17 in patients with UC were significantly
It is well known that soluble IL-33 stimulates Th lymphocytes that have ST2 receptor to
produce Th2 lymphokines.[8-12] Its action on the activation of monocytes, basophils and
eosinophils has also been confirmed.[8-12] IL-33 is highly expressed in the cell nuclei of
many tissues, including gastrointestinal organs, although its role there has not been fully
elucidated. In laboratory conditions, mice treated with IL-33 showed inflammatory intestinal
changes. It was the reason why we decided to measure the concentration of IL-33 in the
serum of patients with UC and compare it with that in healthy controls. The serum
concentration of IL-33 in patients with UC in our study was not significantly increased in
comparison with healthy controls. This finding is in accordance with the knowledge that the
primary biological effect of IL-33 is transcriptional rather than cytokinal. Therefore, the
serum concentration of IL-33 does not necessarily have to reflect the intensity of its real
activity in UC. Although cytokines play an important role in UC, the role of IL-33 could be
dual – that of a cytokine and a gene promoter.
We found that patients with UC had increased values IL-17 in comparison with healthy
controls, which is in accordance with the findings reported by Fujino et al . However, in
their study, IL-17 in the sera of healthy Japanese subjects was below detection limit.
Potential therapeutic benefit of blocking IL-17 by using a soluble IL-17-receptor-Ig fusion
protein or an auto-vaccine targeting mechanisms upstream of IL-17, such as co-inhibition of
IL-12 and IL-23 or inhibition of TNF-a, has not been confirmed in humans.
The main limitation of our study is a small sample size, which should be taken into account
when interpreting our results. However, to the best of our knowledge, this is the first study to
determine IL-33 and IL-17 serum concentrations in patients with UC. The immunological and
pathogenic role of IL-17 in UC still needs to be elucidated and further research is needed to
determine whether increased concentration of IL-17 in UC has protective or harmful effects.
Figure 1. Serum levels of IL-33 in patients with ulcerative colitis and control group.
Squares – median; error bars – interquartile range.
Figure 2. Serum levels of IL-17 in patients with ulcerative colitis and control group.
Squares – median; error bars – interquartile range.
1 Rutgeerts P, Vermeire S, Van Assche G. Biological therapies for inflammatory bowel
diseases. Gastroenterology. 2009;136:1182-1197.
2 Tsianos EV, Katsanos K. Do we really understand what the immunological disturbances in
inflammatory bowel disease mean? World J Gastroenterol. 2009;15:521-525.
3 Mayer L. Current concept of inflammatory bowel disease: etiology and patogenesis. In:
Kirsner JB, ed. Inflammatory Bowel Disease. Philadelphia, PA: W. B. Saunders
4 Macfarlane S, Steed H, Macfarlane GT. Intestinal bacteria and inflammatory bowel disease.
Crit Rev Clin Lab Sci. 2009;46:25-54.
5 Boden EK, Snapper SB. Regulatory T cells in inflammatory bowel disease. Curr Opin
6 Elson CO. The immunology of inflammatory bowel disease. In: Kirsner JB, ed.
Inflammatory Bowel Disease. Philadelphia, PA: W.B. Saunders Company; 2000:208-231.
7. Podolsky DK, Fiocchi C. cytokines, chemokines, growth factors, eicosanoids and other
bioactive molecules in inflammatory bowel disease. In: Kirsner JB, ed. Inflammatory
Bowel Disease. Philadelphia, PA: W.B. Saunders Company; 2000:191-202.
8 Gadina M, Jefferies CA. IL-33: a sheep in wolf's clothing? Sci STKE. 2007(390):pe31.
9 Hayakawa H, Hayakawa M, Kume A, et al. Soluble ST2 blocks interleukin-33 signaling in
allergic airway inflammation. J Biol Chem. 2007;282:26369-26380.
10 Allakhverdi Z, Smith DE, Comeau MR, et al. Cutting edge: the ST2 ligand IL-33 potently
activates and drives maturation of human mast cells. J Immunol. 2007;179:2051-2054.
11 Iikura M, Suto H, Kajiwara N, et al. IL-33 can promote survival, adhesion and cytokine
production in human mast cells. Lab Invest. 2007;87:971-978.
12 Moussion C, Ortega N, Girard JP. The IL-1-like cytokine IL-33 is constitutively expressed
in the nucleus of endothelial cells and epithelial cells in vivo: a novel 'alarmin'? PLoS
13 Zhou L, Ivanov II, Spolski R, et al. IL-6 programs T(H)-17 cell differentiation by
promoting sequential engagement of the IL-21 and IL-23 pathways. Nat Immunol.
14 Ivanov S, Lindén A. Interleukin-17 as a drug target in human disease. Trends Pharmacol
15 Chang SH, Dong C. IL-17F: regulation, signaling and function in inflammation. Cytokine.
16 Louten J, Boniface K, de Waal Malefyt R. Development and function of TH17 cells in
health and disease. J Allergy Clin Immunol. 2009;123:1004-11.
17 Cheung PF, Wong CK, Lam CW. Molecular mechanisms of cytokine and chemokine
release from eosinophils activated by IL-17A, IL-17F, and IL-23: implication for Th17
lymphocytes-mediated allergic inflammation. J Immunol. 2008;180:5625-5635.
18 Carriere V, Roussel L, Ortega N, et al. IL-33, the IL-1-like cytokine ligand for ST2
receptor, is a chromatin-associated nuclear factor in vivo. Proc Natl Acad Sci U S A.
19 Fujino S, Andoh A, Bamba S, et al. Increased expression of interleukin 17 in inflammatory
bowel disease. Gut. 2003;52:65-70.
Figure 2.Download full-text