Abstract
In this study, we describe a novel murine model of chronic intestinal inflammation induced by the hapten reagent 2,4,6-trinitrobenzene sulfonic acid (TNBS). Rectal application of low doses of TNBS in BALB/c and SJL/J mice resulted in a chronic transmural colitis with severe diarrhea, weight loss, and rectal prolapse, an illness that mimics some characteristics of Crohn's disease in humans. The colon of TNBS-treated mice on day 7 was marked by infiltration of CD4+ T cells; furthermore, in situ polymerase chain reaction studies revealed high levels of interferon (IFN)-gamma mRNA in diseased colons. Isolated lamina propria (LP) CD4+ T cells from TNBS-treated mice stimulated with anti-CD3 and anti-CD28 antibodies exhibited a Th1 pattern of cytokine secretion: a 20-50-fold increase in IL-2 and IFN-gamma levels and a 5-fold decrease in IL-4 levels as compared with those of stimulated LP CD4+ T cells from control BALB/c mice. Administration of monoclonal anti-IL-12 antibodies to the TNBS-treated mice both early (at 5 d) and late (at 20 d) after induction of colitis led to a striking improvement in both the clinical and histopathological aspects of the disease and frequently abrogated the established colitis completely. Furthermore, LP CD4+ T cells isolated from anti-IL-12-treated mice failed to secrete IFN-gamma upon in vitro stimulation. In summary, the data demonstrate the pivotal role of IL-12 and IFN-gamma in a TNBS-induced murine model of chronic intestinal inflammation. Furthermore, they suggest the potential utility of anti-IL-12 antibodies in patients with Crohn's disease.
Highlights
Based on previous studies showing that rectal administration of the hapten reagent trinitrobenzene sulfonic acid (TNBS) induces colitis in rats [22, 23], we explored the possibility that administration of TNBS can induce a chronic inflammation of the murine colon
We found that BALB/c and SJL/J mice subjected to intrarectal administration of TNBS in 50% ethanol reproducibly developed pancolitis with severe diarrhea and rectal prolapse tensity usually peaked between 2 and 4 wk after administration of TNBS
FACS | analysis o f spleen lymphocytes revealed a twofold increase in the percentage of CD4 § and CD8 + T cells in TNBS-treated mice compared with control ethanol-treated mice and normal BALB/c mice, along with a reduction in B220 + B cells
Summary
Specific pathogen-free 2-4-too-old female BALB/c or SJL/J mice were obtained from the National Cancer Institute (Bethesda, MD) and maintained in the Building 10A Animal Facility at the National Institutes of Health. The mice were lightly anesthetized with metofane (methoxyflurane; Pitman-Moore, Mundelein, IL). A 3.5F catheter was carefully inserted into the colon such that the tip was 4 cm proximal to the anus. 0.5 mg of the hapten reagent TNBS (Sigma Chemical Co., St. Louis, MO) in 50% ethanol (to break the intestinal epithelial barrier) was slowly administered into the lumen of the colon via the catheter fitted onto a l-ml syringe. The total injection volume was 100 ~1 in both groups allowing TNBS or ethanol to reach the entire colon, including the caecum and appendix. Animals were kept in a vertical position for 30 s and returned to their cages
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