Sarah Winckler

M1652 Heligmosomoides Polygyrus Exposure Improves Established Inflammation in NSAID-Synchronized CD25-Depleted Transfer Colitis

Melanin-concentrating hormone (MCH) is localized mainly in the brain, but it is also found in the intestine. MCH plays a clear role in feeding behavior. Recent data suggests that MCH and the MCH receptor 1 mediate intestinal inflammation in mice. Two diverse MCH receptor 1 antagonists (DABA-821 and DABA-822) have been developed for pharmacological testing. DABA-821 penetrates the blood-brain barrier, while DABA-822 does not. Study Aim: Our goal was to test DABA-821 and DABA-822 in a murine model of TNBS colitis. The efficacy of these compounds was compared to Dexamethasone (Dex).Methods: Female BALB/c mice (n=37) were randomized to receive an intracolonic enema of PBS, 50% ethanol/PBS, or 20 mg/kg TNBS in 50% ethanol. On day 1 (five hours after the enemas) mice were given Vehicle, 30 mg/kg of DABA-821, 30 mg/kg of DABA-822, or 1 mg/kg of Dex by orogastric gavage. Test drugs were also administered bid on days 2 through 6. On day 7, mice were euthanized. The colon was analyzed for overall macroscopic damage, ulceration, total length, distal segment weight, MPO activity and histological pathology. Body and spleen weights were also determined. Results:Macroscopic colonic scores (combination of lesions, diarrhea, adhesions, thickness) were significantly increased in TNBS treated mice, as compared to PBS or 50% ethanol treated animals. These scores were attenuated by 53 to 75% in mice treated with the MCH-1 receptor antagonists. Similar efficacy was found with Dex. Colonic ulceration was clearly apparent in the Vehicle/TNBS treatment group, but ulcer severity was significantly reduced in mice treated with the MCH receptor 1 antagonists. Mean colonic ulcer scores were: 4.7 (Vehicle/TNBS), 2.4 (DABA-821), 2.9 (DABA-822) and 2.8 (Dex). As compared to their vehicle treated cohorts, colons were significantly longer, but weighed less, in drug treated mice. Colonic MPO activity was nearly normalized in mice treated with Dex, but only partially attenuated by treatment with the MCH receptor 1 antagonists. Colonic histology scores were significantly reduced (p < 0.05 vs. Vehicle) in all drug treatment groups. Vehicle/TNBS treated mice lost significant weight throughout the study, as compared to PBS treated animals. None of the treatments normalized this weight loss. Reduced spleen weights were observed in Dex treated mice, as well as in mice treated with DABA-821 and 822. Summary: Two diverse MCH receptor 1 antagonists attenuated various parameters of TNBS-induced colitis in mice. The anti-colitis profile of these antagonists was similar to Dex. Conclusion:MCH receptor 1 antagonismmay represent an effective therapeutic approach for intestinal inflammation.

W1185 CpG Oligonucleotide Stimulation of TLR9 Inhibits Il17 Production from Mesenteric Lymph Node Cells

in C.rodentium infection remain to be seen. Method and Result: C. rodentium which express GFP constitutively (CR/GFP) has been established by the transformation with the GFPexpression vector. The expression of GFP was confirmed by flow cytometry and fluorescence microscopy. To examine the localization of C.rodentium, 3-week-old wild-type mice were orally inoculated with CR/GFP. CR/GFP were detected on the cell surface at the colon at day 3 and some were also detected in the CD11b positive cells such as macrophages or dendtiric cells of the colon. The bacterial number in the section of the infected colon and in the feces were increased until day 14. To examine the antigen-specific immune response of CD4 + T cell in C.rodentium infection, OVA-expressing C.rodentium which produce chicken ovalbumin constitutively (CR/OVA) have also been established by the transformation with OVA producing plasmid. The antigenic expression of OVA was confirmed by western blot and T cell-proliferation assay. IgM and IgG response against OVA were detected 2 weeks after oral infection with CR/OVA. Next, to examine the C.rodentium-specific T cell response, CFSE systems were utilized. Wild-type mice were infected with CR/OVA at day 1. CFSE labeled OVA-specific CD4+ T cells (OT-II cells) were adoptively transferred at day 12 and then, MLN were collected at day 15. Proliferation of OVA-specific CD4+ T cells were demonstrated by the detection of multiple cell CD4+ division of CFSE-loaded transgenic T cells. Moreover, enhanced cytokine production of IFN-γ and IL-4 were also detected after In Vitro stimulation with OVA. Conclusion: These studies show the natural course and host immune response in C. rodentium infection. These model systems are useful to examine the mucosal immunity against C.rodentium infection in detail.