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. 2001 Mar;48(3):326–332. doi: 10.1136/gut.48.3.326

Increased expression of IL-16 in inflammatory bowel disease

D Seegert 1, P Rosenstiel 1, H Pfahler 1, P Pfefferkorn 1, S Nikolaus 1, S Schreiber 1
PMCID: PMC1760149  PMID: 11171821

Abstract

BACKGROUND—Inflammatory bowel disease (IBD) is characterised by infiltration of inflamed mucosal regions with CD4+ T lymphocytes and other mononuclear cells. Interleukin (IL)-16 exerts a strong chemoattractant activity on CD4+ cells. Moreover, IL-16 activates expression and production of proinflammatory cytokines such as IL-1β, IL-6, IL-15, and tumour necrosis factor α (TNF-α) in human monocytes.
AIM—To examine if IL-16 expression is increased in IBD patients compared with healthy controls.
METHODS—Twenty one patients with IBD (10 with ulcerative colitis (UC), 11 with Crohn's disease (CD)), seven disease specificity controls (DSC), and seven healthy controls were studied. Biopsies were taken during colonoscopies and IL-16 mRNA as well as protein expression were investigated by reverse transcriptase-polymerase chain reaction, ELISA, western blot, and immunohistochemistry.
RESULTS—IL-16 mRNA and protein expression in the colonic mucosa of IBD patients were increased twofold compared with healthy controls, DSC, or IBD patients under steroid treatment. Most of the detected IL-16 protein was in its bioactive 17 kDa form and was predominantly expressed in eosinophils. Increased IL-16 expression in UC patients appeared to be mainly restricted to the inflamed regions of the colonic mucosa. Levels of caspase 3, which processes the 68 kDa IL-16 precursor molecule into the biological active 17 kDa form, were not increased.
CONCLUSIONS—Our results provide evidence that IL-16 expression is significantly increased in the inflamed colonic mucosa of IBD patients but not in control individuals, DSC, or patients under steroid treatment. Therefore, upregulation of IL-16 expression seems to be specific for chronic intestinal inflammation and could lead to increased secretion of other proinflammatory cytokines in IBD.


Keywords: interleukin-16; T lymphocytes; eosinophils; Crohn's disease; ulcerative colitis; inflammatory bowel disease

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Figure 1  .

Figure 1  

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(A) Total RNA was extracted from biopsies from four normal control individuals and 12 patients with IBD (six Crohn's disease and six ulcerative colitis patients). Interleukin 16 (IL-16) mRNA was amplified by reverse transcriptase-polymerase chain reaction (RT-PCR) and analysed on an agarose gel (top panel). The use of equal amounts of total RNA was confirmed in a parallel experiment by amplification of β-actin mRNA (bottom panel). PCR products were analysed on a 1% agarose gel. (B) Densitometric quantification of IL-16 specific PCR products. Values are given as mean (SD) percentage of intensity of the highest detected densitometric strength. *p<0.05 v controls.

Figure 2  .

Figure 2  

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Quantification of interleukin 16 (IL-16) protein in colonic mucosal biopsy specimens from 21 patients with inflammatory bowel disease (10 ulcerative colitis (UC) patients (6/10 under corticosteroid (cs) therapy), 12 Crohn's disease (CD) patients (8/12 under cs treatment)), seven disease specificity controls (DSC), and seven normal individuals. Tissues were lysed and adjusted to a protein concentration of 0.5 mg/ml: 50 µl of each sample were used to detect IL-16 content by IL-16 specific ELISA. Values are mean (SD).

Figure 3  .

Figure 3  

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Detection of CD4 and interleukin 16 (IL-16) by immunofluorescence staining in biopsies from control individuals and patients with inflammatory bowel disease (ulcerative colitis and Crohn's disease). The specificity of the IL-16 antibody was confirmed by an irrelevant isotype antibody (biotin conjugated rabbit IgG). CD4 infiltration was visualised with a Cy3 conjugated antibody (red) while IL-16 protein was detected by an FITC labelled antibody (green).

Figure 4  .

Figure 4  

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Staining of biopsies from Crohn's disease patients with anti-eosinophilic peroxidase antibody (left) and anti-interleukin 16 (IL-16) antibody (panel).

Figure 5  .

Figure 5  

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(A) Interleukin 16 (IL-16) protein expression was analysed by western blotting in biopsies taken from four control individuals (lanes 1-4), four ulcerative colitis (UC) patients (lanes 5-8), and four Crohn's disease (CD) patients (lanes 9-12). The experiment revealed the predominant existence of the bioactive 17 kDa form of IL-16 in IBD patients compared with controls. (B) Biopsies from four UC and three CD patients were taken from non-inflamed (—) or inflamed (+) regions of the colon and tested for IL-16 protein expression in a western blot assay. (C) Extracts of the same biopsies as in (A) were used to detect caspase 3 protein expression. A lysate from unstimulated human A431 cells supplied with the anti-caspase 3 antibody served as a control (WB, western blot).

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