Volume 119, Issue 6, Pages 1514-1523 (December 2000) Interleukin 18 is a potent proliferative factor for intestinal mucosal lymphocytes in Crohn's disease  Takanori Kanai, *, Mamoru Watanabe, *, Akira Okazawa, ‡, Koichi Nakamaru, ‡, Makiyo Okamoto, ‡, Makoto Naganuma, ‡, Hiromasa Ishii, ‡, Masao Ikeda, §, Masashi Kurimoto, §, Toshifumi Hibi, *,‡  Gastroenterology  Volume 119, Issue 6, Pages 1514-1523 (December 2000) DOI: 10.1053/gast.2000.20260 Copyright © 2000 American Gastroenterological Association Terms and Conditions

Fig. 1 (A) Serum IL-18 concentrations in normal controls (15 cases), patients with CD (21 cases), and patients with UC (23 cases) were measured using a human IL-18–specific ELISA. Serum IL-18 concentration was significantly higher in CD (P < 0.001 vs. normal controls; P < 0.005 vs. UC). NL, normal control. (B) Correlation between serum IL-18 concentration and disease activity determined by CDAI in CD. To assess the correlation between serum IL-18 concentration and CDAI, Person's correlation coefficient test was used. Significant (R = 0.73, P = 0.02) correlation between serum IL-18 concentration and CDAI in CD patients was observed. Gastroenterology 2000 119, 1514-1523DOI: (10.1053/gast.2000.20260) Copyright © 2000 American Gastroenterological Association Terms and Conditions

Fig. 2 (A) Immunohistochemical analysis of IL-18 protein expression in colonic mucosa using affinity-purified goat anti-human IL-18 antibody and control goat IgG. In normal control colonic mucosa and UC colonic inflamed mucosa, IL-18–positive cells were scarce. However, in CD-inflamed colonic mucosa, many IL-18–positive mononuclear cells were observed in the lamina propria. This figure is representative of samples from 7 separate subjects (original magnification, 200×). (B) Immunohistochemical analysis of IL-18 and CD68 protein expression in CD-inflamed ileal mucosa. IL-18–positive mononuclear cells were localized mainly to CD68-positive macrophages. This figure is representative of samples from 4 separate subjects (original magnification, 200×). (C) Double staining for IL-18 and CD68 detected in CD-inflamed ileal mucosa by the confocal laser scanning microscope. CD68-positive mononuclear cells within the lamina propria possess abundant cytoplasm, extended cytoplasmic processes, and morphologically consistent tissue macrophages or dendritic cells. IL-18 staining showed complete overlap of those CD68-positive cells, indicating that IL-18–positive cells were macrophages or dendritic cells. This figure is representative of samples from 5 separate subjects (original magnification, 500×). Gastroenterology 2000 119, 1514-1523DOI: (10.1053/gast.2000.20260) Copyright © 2000 American Gastroenterological Association Terms and Conditions

Fig. 3 Flow-cytometric analysis of IL-18 receptor cell surface expression in freshly isolated LPLs derived from normal controls and UC and CD patients. Cells were stained with anti–IL-18 receptor mAb or isotype-matched control mAbs (dotted lines) and FITC-conjugated goat anti-mouse IgG1. After washing, cells were incubated with PE-conjugated anti-CD3 mAb. Analysis gates were set on CD3+ cells. A minimum of 10,000 cells were analyzed for each histogram. The results are representative of 3 similar experiments. Gastroenterology 2000 119, 1514-1523DOI: (10.1053/gast.2000.20260) Copyright © 2000 American Gastroenterological Association Terms and Conditions

Fig. 4 (A) Effect of IL-18 in proliferation of freshly isolated LPLs. Recombinant human IL-18 stimulated significant increase in DNA synthesis in freshly isolated LPLs from CD patients (n = 102; 7 from inflamed and 5 from noninflamed mucosa) in a dose-dependent manner, compared with those from 9 normal controls. Proliferation of LPLs from CD was significantly higher than that of LPLs from UC after stimulation with IL-18 at 15.6 and 62.5 ng/mL, but not at 250 ng/mL. In contrast, proliferative responses of freshly isolated LPLs from UC (n = 9; 5 from inflamed and 4 from noninflamed mucosa) did not increase. (B) Proliferative responses of LPLs from inflamed CD mucosa were significantly higher than those of LPLs from noninflamed CD mucosa after stimulation with IL-18 at 15.6 and 62.5 ng/mL, but not at 250 ng/mL. *P < 0.05 vs. NL; †P < 0.05 vs. UC; ‡P < 0.05 vs. noninflamed CD. Gastroenterology 2000 119, 1514-1523DOI: (10.1053/gast.2000.20260) Copyright © 2000 American Gastroenterological Association Terms and Conditions

Fig. 5 The synergistic effect of IL-12 and IL-18 in proliferation of freshly isolated LPLs. Combined stimulation with IL-12 (1 ng/mL) and IL-18 (100 ng/mL) stimulated significant increase in DNA synthesis in freshly isolated LPLs from inflamed mucosa from CD patients (n = 9), NL mucosa (n = 9), and UC (n = 7). *P < 0.05. Gastroenterology 2000 119, 1514-1523DOI: (10.1053/gast.2000.20260) Copyright © 2000 American Gastroenterological Association Terms and Conditions

Fig. 6 Flow-cytometric analysis of the expression of IL-2 receptor α and IL-12 receptor β2 on IL-18–stimulated LPLs. IL-2 receptor expression of IL-18–stimulated LPLs derived from CD was significantly higher than that of normal controls and UC. IL-12 receptor β2 expression did not change after stimulation with IL-18. The results are representative of 5 similar experiments. Gastroenterology 2000 119, 1514-1523DOI: (10.1053/gast.2000.20260) Copyright © 2000 American Gastroenterological Association Terms and Conditions