Interleukin 37 expression protects mice from colitis

Abstract

IL-37, a newly described member of the IL-1 family, functions as a fundamental inhibitor of innate inflammation and immunity. In the present study, we examined a role for IL-37 during experimental colitis. A transgenic mouse strain was generated to express human IL-37 (hIL-37tg), and these mice were subjected to dextran sulfate sodium (DSS)-induced colitis. Despite the presence of a CMV promoter to drive expression of IL-37, mRNA transcripts were not present in colons at the resting state. Expression was observed only upon disruption of the epithelial barrier, with a six- to sevenfold increase (P = 0.02) on days 3 and 5 after continuous exposure to DSS. During the development of colitis, clinical disease scores were reduced by 50% (P < 0.001), and histological indices of colitis were one-third less in hIL-37tg mice compared with WT counterparts (P < 0.001). Reduced inflammation was associated with decreased leukocyte recruitment into the colonic lamina propria. In addition, release of IL-1β and TNFα from ex vivo colonic explant tissue was decreased 5- and 13-fold, respectively, compared with WT (P ≤ 0.005), whereas IL-10 was increased sixfold (P < 0.001). However, IL-10 was not required for the anti-inflammatory effects of IL-37 because IL-10-receptor antibody blockade did not reverse IL-37-mediated protection. Mechanistically, IL-37 originating from hematopoietic cells was sufficient to exert anti-inflammatory effects because WT mice reconstituted with hIL-37tg bone marrow were protected from colitis. Thus, IL-37 emerges as key modulator of intestinal inflammation.

Fig. 1.

Reduced severity of DSS colitis in hIL-37tg mice. DSS or water vehicle was administered to WT and IL-37tg mice ad libitum in drinking water for 7 d. (A) Weight change during treatment was expressed as percentage change from day 0. (B) Clinical DAI (SI Methods). (C) Macroscopic images of colons from indicated treatment cohorts. (D) Colon lengths were assessed at necropsy on day 7. Colon lengths were assessed at necropsy on day 7. Data are expressed as mean ± SEM. (A) **P < 0.01, ***P < 0.001 vs. IL-37tg. (B) ***P < 0.001 vs. WT-H₂O, ⁺⁺⁺P < 0.001 vs. WT-DSS (n = 19–27 mice from four independent experiments).

Fig. 2.

Amelioration of histological indices in hIL-37tg mice subjected to DSS colitis. (A and B) Histological assessment of colitis severity demonstrated amelioration of all assessed parameters in IL-37tg mice compared with WT mice. Data are expressed as mean ± SEM. ***P < 0.001 vs. WT (n = 27 mice per cohort from four independent experiments). (C) Representative micrograph images of colons from indicated treatment cohorts. (Scale bars: 200 μm.)

Fig. 3.

Induction of IL-37 mRNA correlates with intestinal barrier breakdown. (A) Time course of IL-37 mRNA transcripts within colonic tissues during DSS colitis. (B) Colonic tissues from DSS-treated WT and hIL-37tg mice were harvested on indicated days, and the severity of colitis was assessed as described in Methods. (C) Representative micrographs of the progression of colitis at the indicated time points in WT and IL-37tg mice. Data are expressed as mean ± SEM (n = 4 mice per time point from two independent experiments). *P < 0.05, ***P < 0.001 vs. day 0 (A) or WT (B). (Scale bars: 200 μm.)

Fig. 4.

Levels of colonic IL-10, TNFα, and IL-1β. Colonic tissue explants were harvested at 7 d after continuous DSS exposure and cultured ex vivo at 37 °C for 24 h. Secreted cytokines were assayed from supernatants as described in Methods and expressed as picogram of cytokine per milligram of tissue (pg/μg). Data are expressed as mean ± SEM. *P < 0.05, **P < 0.01 vs. WT-H₂O vehicle, ⁺⁺P < 0.001, ⁺⁺⁺P < 0.001 vs. WT-DSS (n = 19–27 mice from four independent experiments).

Fig. 5.

Effect of hIL-37tg expression on colonic leukocyte infiltrates. Assessment of colonic leukocyte populations from WT and IL-37tg mice after DSS treatment. (A) Major leukocyte subsets were expressed as absolute numbers. (B and C) Representative flow cytometry plots of dendritic cells (CD11c^high/MHC^high), macrophages (CD11c^low/MHCII^high), eosinophils (Siglec F^high/GR1^neg), and neutrophils (SiglecF^neg/GR1^high). Data are expressed as mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001 vs. WT-DSS (n = 6 mice per strain from two independent experiments).

Fig. 6.

Hematopoietic-derived IL-37 protects mice from colitis. BM chimeric mice were generated after irradiation of WT(CD45.1) or hIL-37tg(CD45.2) mice as described in Methods. (A) Flow cytometry analyses of donor and recipient CD45⁺ splenocytes demonstrated >95% reconstitution. DSS or water vehicle was administered to indicated chimeric mice ad libitum in drinking water for 7 d. (B) Weight changes during treatment were expressed as percentage change from day 0. (C) Clinical DAI (SI Methods). Data are expressed as mean ± SEM. **P < 0.01, ***P < 0.001 vs. WT→WT-DSS (n = 6–11 mice per strain from two independent experiments).

Fig. 7.

Effect of hematopoietic-derived IL-37 on colitis and cytokine production. DSS or water vehicle was administered ad libitum in drinking water for 7 d to WT, IL-37tg, and indicated BM chimeric mice. (A) Colon lengths were assessed at necropsy on day 7. (B) Histological evaluation of colons was performed as described. (C) Representative micrographs of colons of indicated BM chimeric mice. (Scale bars: 200 μm.) (D) Ex vivo colonic tissue explants of indicated BM chimeric mice treated with DSS were cultured at 37 °C for 24 h. Data are expressed as mean ± SEM. (A) ***P < 0.001 vs. WT-H₂O, ⁺⁺⁺P < 0.001 vs. WT→WT-DSS. (B) ***P < 0.001 vs. WT→WT-DSS, ⁺⁺⁺P < 0.001 vs. IL-37tg→WT-DSS. (D) *P < 0.05, **P < 0.001, IL-37tg→WT vs. WT→WT-DSS (n = 6–11 mice per cohort from two independent experiments).