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The use of biologics has promising potential in the treatment of inflammation. Studies with cultured cells and mouse models of disease have ascribed proinflammatory and anti-inflammatory functions to oncostatin M (OSM) and the related cytokine, interleukin-6 (IL-6). Here, we examined the effect of systemic administration of adenoviral (Ad) vectors encoding either murine OSM (AdMuOSM) or murine IL-6 (AdMuIL-6) in a mouse model of colitis. BALB/c mice were treated with a 5-day course of 4% dextran-sodium sulfate (DSS) water with or without administration of adenoviral vectors (i.p. or i.m. at 10(7) plaque-forming units [pfu]) given as a cotreatment or therapy. The deletion variant of the adenovirus served as a control for adenoviral infection. Colitis was assessed by (1) morphology (damage score, macrophage infiltration, apoptosis) and (2) function (myeloperoxidase activity and Ussing chamber analysis of epithelial ion transport). Infection with adenovirus alone did not affect colonic form or function. AdMuOSM (either i.p. or i.m.) significantly reduced the severity of the DSS-induced colitis. There was less damage, reduced macrophage infiltration, fewer apoptotic bodies, and a significant improvement in stimulated ion transport in colonic tissues from the treated mice. No benefit of AdMuIL-6 treatment was observed in this model system. Thus, systemic administration of AdMuOSM given as a cotreatment and to a lesser extent as a therapy was found to be of benefit in DSS-induced colitis, a murine model of inflammatory bowel disease (IBD).

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is a clinically important bacterial enteropathogen that manipulates a variety of host cell signal transduction cascades to establish infection. However, the effect of EHEC O157:H7 on Jak/Stat signaling is unknown. To define the effect of EHEC infection on epithelial gamma interferon (IFN-gamma)-Stat1 signaling, human T84 and HEp-2 epithelial cells were infected with EHEC O157:H7 and then stimulated with recombinant human IFN-gamma. Cells were also infected with different EHEC strains, heat-killed EHEC, enteropathogenic E. coli (EPEC) O127:H6, and the commensal strain E. coli HB101. Nuclear and whole-cell protein extracts were prepared and were assayed by an electrophoretic mobility shift assay (EMSA) and by Western blotting, respectively. Cells were also processed for immunofluorescence to detect the subcellular localization of Stat1. The EMSA revealed inducible, but not constitutive, Stat1 activation upon IFN-gamma treatment of both cell lines. The EMSA also showed that 6 h of EHEC O157:H7 infection, but not 30 min of EHEC O157:H7 infection, prevented subsequent Stat1 DNA binding induced by IFN-gamma, whereas infection with EPEC did not. Immunoblotting showed that infection with EHEC, but not infection with EPEC, eliminated IFN-gamma-induced Stat1 tyrosine phosphorylation in both dose- and time-dependent fashions and disrupted inducible protein expression of the Stat1-dependent gene interferon regulatory factor 1. Immunofluorescence revealed that EHEC infection did not prevent nuclear accumulation of Stat1 after IFN-gamma treatment. Also, Stat1 tyrosine phosphorylation was suppressed by different EHEC isolates, including intimin-, type III secretion- and plasmid-deficient strains, but not by HB101 and heat-killed EHEC. These findings indicate the novel disruption of host cell signaling caused by EHEC infection but not by EPEC infection.

Infection with helminth parasites typically evokes a T helper 2-cell response in the mammalian host. Interleukin (IL)-4 and IL-13 are important in the rapid expulsion of parasitic nematodes, with the absence of either cytokine being compensated for by the other. However, the IL-4 and IL-13 common signaling molecule, signal transducer and activator of transcription 6 (STAT-6) appears to be mandatory for the spontaneous expulsion of enteric helminths. Mice genetically deficient in IL-4, IL-13, or STAT-6 were infected with the cestode Hymenolepis diminuta and worm infectivity and jejunal goblet cell responses assessed 12-18 days postinfection (PI). Only the STAT-6 knockout (KO) animals harbored adult worms; neither gravid adult nor stunted H. diminuta was obtained from the infected IL-4 KO or IL-13 KO mice > or = 12 days PI. Also, the establishment of worms in the intestine of STAT-6 KO animals was associated with a reduced goblet cell response. These findings support the hypothesis that increased mucin production is an important part of the host response to tapeworm infection and that functional STAT-6 signaling may be an absolute requirement for the rejection of intestinal cestodes and thus, helminth parasites in general.

Growth factors affect a variety of epithelial functions. We examined the ability of TGF-beta to modulate epithelial ion transport and permeability. Filter-grown monolayers of human colonic epithelia, T84 and HT-29 cells, were treated with TGF-beta (0.1-100 ng/ml, 15 min-72 h) or infected with an adenoviral vector encoding TGF-beta (Ad-TGF beta) for 144 h. Ion transport (i.e., short-circuit current, I(sc)) and transepithelial resistance (TER) were assessed in Ussing chambers. Neither recombinant TGF-beta nor Ad-TGF beta infection affected baseline I(sc); however, exposure to > or = 1 ng/ml TGF-beta led to a significant (30-50%) reduction in the I(sc) responses to forskolin, vasoactive intestinal peptide, and cholera toxin (agents that evoke Cl(-) secretion via cAMP mobilization) and to the cell-permeant dibutyryl cAMP. Pharmacological analysis of signaling pathways revealed that the inhibition of cAMP-driven epithelial Cl(-) secretion by TGF-beta was blocked by pretreatment with SB-203580, a specific inhibitor of p38 MAPK, but not by inhibitors of JNK, ERK1/2 MAPK, or phosphatidylinositol 3'-kinase. TGF-beta enhanced the barrier function of the treated monolayers by up to threefold as assessed by TER; however, this event was temporally displaced from the altered I(sc) response, being statistically significant only at 72 h posttreatment. Thus, in addition to TGF-beta promotion of epithelial barrier function, we show that this growth factor also reduces responsiveness to cAMP-dependent secretagogues in a chronic manner and speculate that this serves as a braking mechanism to limit secretory enteropathies.

Inhibition of the inducible form of nitric oxide (NO) synthase prolonged the murine enteropathy evoked by the bacterial superantigen, Staphylococcus aureus enterotoxin B (SEB). We examined the ability of NO to alleviate SEB-induced epithelial dysfunction and immune cell activation. Human peripheral blood mononuclear cells (PBMC) were activated by SEB for 24 h +/- the NO donors, S-nitroso-N-acetylpenicillamine and spermine-NONOate. The conditioned medium (CM) was collected and applied to T84 epithelial monolayers, and permeability [i.e., transepithelial resistance (TER)] and stimulated ion transport (i.e., short-circuit current responses to carbachol and forskolin) were assessed 24 h later. Exposure to CM led to an approximately 40% drop in TER and hyporesponsiveness to both secretagogues. CM made in the presence of NO donors (10(-4) M) had no significant effect on epithelial barrier or ion transport parameters. NO donors alone had no effect on naive epithelia, and addition of the NO donors to previously made CM did not affect the ability of this CM to alter epithelial function. Moreover, the NO donors dose-dependently reduced SEB-evoked PBMC proliferation and cytokine production (i.e., interferon-gamma, tumor necrosis factor alpha) but did not affect viability. These findings suggest a beneficial role for NO in inflammation by reducing immune cell activation and thus ameliorating consequent physiological abnormalities, in this instance, perturbed epithelial permeability and active ion transport.