Gliotoxin, an Inhibitor of Nuclear Factor-Kappa B, Attenuates Peptidoglycan–Polysaccharide-Induced Colitis in Rats Leo R. Fitzpatrick, Jian Wang, and Truc Le Gastrointestinal Research Laboratory, Maryland Research Laboratories, Otsuka Maryland Research Institute, Rockville, Maryland, U.S.A. Summary: Gliotoxin is a fungal metabolite that has immuno- suppressive properties. First, we determined if gliotoxin could inhibit bacterial peptidoglycan–polysaccharide-stimulated tu- mor necrosis factor- production, as well as nuclear factor- kappa B (NF-B), in a rat macrophage (NR8383) cell line. Next, the apoptosis-inducing potential of gliotoxin was also evaluated in this cell line. Finally, we evaluated whether glio- toxin could reduce peptidoglycan–polysaccharide-induced co- litis in rats. Gliotoxin (2 mg/kg/day) was dosed from day 14 after the initial intramural colonic injection of peptidoglycan– polysaccharide until day 21. A gross colonic injury score, my- eloperoxidase activity, and cytokine levels were all evaluated on day 21. Gliotoxin dose dependently inhibited cytokine pro- duction, as well as NF-B, and also induced apoptosis in the NR8383 cell line. On day 21, gliotoxin significantly reduced gross colonic injury (adhesions, nodules, mucosal lesions) in rats. Gliotoxin-treated rats also had partially normalized bio- chemical indices of colitis, such as colonic cytokine levels. The colonic level of NF-B was also partially normalized in glio- toxin treated rats. Gliotoxin also exhibited an antiarthritis effect in peptidoglycan–polysaccharide-treated rats. In summary, gliotoxin effectively attenuated the chronic reactivation phase of peptidoglycan–polysaccharide-induced colitis. This antico- litis effect may be related to the inhibition of NF-B in Lewis rats. Key Words: Gliotoxin—Nuclear Factor-kappa B— Colitis—Rats. INTRODUCTION Gliotoxin is a fungal metabolite, which is classified as an epipolythiodioxopiperazine analog (1). This fungal metabolite exhibits potent immunosuppressive effects both in vitro and in vivo (2–6). Specifically, gliotoxin has been shown to affect the function of various immune cells, including lymphocytes, neutrophils, and macro- phages (2–6). More recently, other investigators demonstrated that gliotoxin potently and selectively inhibited the activation of the transcription factor nuclear factor–kappa B [NF- B] in T and B cells (5), as well as in hepatocytes (7). In this regard, gliotoxin prevented the degradation of IB, which is an endogenous inhibitor of NF-B (5). In fact, gliotoxin may prevent such degradation by acting as a specific proteasome inhibitor (8). Of note, NF-B is believed to play a pivotal patho- genic role in both human inflammatory bowel disease (IBD), as well as in animal models of IBD (9–13). Her- farth et al. reported that treatment of mice with gliotoxin could effectively treat the acute phase of dextran sodium sulfate [DSS]-induced colitis in mice (14). They also found that intraperitoneal (i.p.) treatment with gliotoxin down-regulated the enhanced colonic NF-B activity as- sociated with DSS administration to mice (14). We have shown that gliotoxin is effective against two different phases of DSS-induced colitis in rats, but this agent is particularly effective during the acute phase of this co- litis (15). DSS-induced colitis is reported to mimic more closely human ulcerative colitis (UC), as opposed to Crohn’s disease (CD) (16). Therefore, a major goal of our study was to evaluate the efficacy of gliotoxin against pepti- doglycan-polysaccharide (PG–PS)-induced colitis, which more closely mimics CD (17,18). Macrophages have clearly been implicated in the pathogenesis of PG– PS-induced colitis, as well as human CD (17–20). There- fore, using a rat macrophage cell line, we also evaluated the in vitro effects of gliotoxin on PG–PS-induced NF- Received February 21, 2001; accepted November 21, 2001. Address correspondence to Dr. L. R. Fitzpatrick, GI Research Labo- ratory, Maryland Research Laboratories, Otsuka Maryland Research Institute, 9900 Medical Center Drive, Rockville, MD 20850, U.S.A. Inflammatory Bowel Diseases 8(3):159–167 © 2002 Crohn’s & Colitis Foundation of America, Inc. 159