Fig. 1.

CRF and UcnII inhibition by RNAi in rat ileum. Groups of rats (n = 4–6) were treated with long dsRNA for CRF (dsCRF), UcnII (dsUcnII), or nonspecific control (dsGFP) for 5–7 days. (A) CRF peptide is undetectable by IHC in the nerve fibers (arrows) of the submucosal (SM) plexus of rats treated with dsRNA for CRF, whereas other treatment groups continued to express CRF in the ileum. (B) Results from RT-PCR reveal that mRNA for CRF or UcnII is significantly inhibited by RNAi treatment, with no UcnII cDNA product amplifiable in dsUcnII groups or very low levels of CRF cDNA product in dsCRF group. Cyclophilin, a housekeeping gene, was detectable in all three groups, and its expression was unaffected by RNAi treatment. (A and C) CRF expression is induced 3 h after TxA treatment in the submucosa and lamina propria (LP). This induction is specifically blocked by 6 days of pretreatment with long dsRNA against CRF. CRF, but not UcnII, is proinflammatory. (D and E) TxA treatment markedly destroys tissue morphology in control or UcnII dsRNA groups (n = 4–6) as assessed by epithelial damage, edema, and neutrophil infiltration. Inhibition of CRF by dsRNA resulted in significant protection of tissue damage that appeared similar to buffer-treated naïve animals. ANOVA indicated a global significant effect [F (3, 12) = 5.5; P < 0.02]. (F) Reduced tissue damage also resulted in reduced fluid secretion in dsCRF animals as a measured loop ratio (weight of the loop/length). ANOVA indicated a global effect [F (4, 17) = 8; P < 0.001]. Different letters denote statistically significant differences among groups (P < 0.01). [Magnification: ×10 (A and D) and ×40 (C).]