LPS elicits several immediate proinflammatoy responses in peripheral blood leukocytes via a recently described pathway including CD14, Toll-like receptors (TLR), serine-threonine kinases, and NF-κB transcription factor. However, the functional responses of intestinal epithelial cells (IEC) to stimulation with LPS are unknown. Expression of mRNA and protein for CD14 and TLRs were assessed by RT-PCR, immunoblotting, and immunohistochemistry in mouse and human IEC lines. LPS-induced activation of signaling pathways (p42/p44 mitogen-activated protein kinase (MAPK), c-Jun NH2-terminal kinase (JNK), p38, p65, NF-κB) were assessed by immunoblotting and gel shifts. CD14 mRNA and protein expression were not detectable in IEC. However, human TLR2, TLR3, and TLR4 mRNA were present in IEC. TLR4 protein was expressed in all cell lines; however, TLR2 protein was absent in HT29 cells. Immunofluorescent staining of T84 cells demonstrated the cell-surface presence of the TLRs. LPS-stimulation of IEC resulted in activation (>1.5-fold) of the three members of the MAPK family. In contrast, LPS did not significantly induce activation of JNK and p38 in CMT93 cells, p38 in T84 cells and MAPK and JNK in HT29 cells. Downstream, LPS activated NF-κB in IEC in a time-, dose-, and serum-dependent manner. IEC express TLRs that appear to mediate LPS stimulation of specific intracellular signal transduction pathways in IEC. Thus, IEC may play a frontline role in monitoring lumenal bacteria.

SummaryC3H/HeJBir mice are a new substrain that spontaneously develop colitis early in life. This study was done to determine the T cell reactivity of C3H/HeJBir mice to candidate antigens that might be involved in their disease. C3H/HeJBir CD4 ϩ T cells were strongly reactive to antigens of the enteric bacterial flora, but not to epithelial or food antigens. The stimulatory material in the enteric bacteria was trypsin sensitive and restricted by class II major histocompatibility complex molecules, but did not have the properties of a superantigen. The precursor frequency of interleuken (IL)-2-producing, bacterial-reactive CD4 ϩ T cells in colitic mice was 1 out of 2,000 compared to 1 out of 20,000-25,000 in noncolitic control mice. These T cells produced predominately IL-2 and interferon ␥ , consistent with a T helper type 1 cell response and were present at 3-4 wk, the age of onset of the colitis. Adoptive transfer of bacterial-antigen-activated CD4 ϩ T cells from colitic C3H/HeJBir but not from control C3H/HeJ mice into C3H/HeSnJ scid / scid recipients induced colitis. These data represent a direct demonstration that T cells reactive with conventional antigens of the enteric bacterial flora can mediate chronic inflammatory bowel disease.

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Experimental animal models, particularly the newer mouse models, have convincingly demonstrated that CD+ T cells play a central role in chronic intestinal inflammation. Such CD4+ effector T cells are induced by the bacterial flora. In at least one model, it is conventional protein antigens that are stimulating these pathogenic T cells. The antigens driving disease seem to be a selective subset of immunodominant proteins, likely derived from a subset of organisms. Multiple genes contribute to colitis susceptibility and a number of these genes are being localized.

Technically easy, intraductal US is a valuable adjunct to endoscopic retrograde cholangiography/tissue sampling that increases the ability to distinguish malignant from benign strictures. Intraductal US is unsuitable for assessing lymph nodes associated with malignant strictures.

Intestinal epithelial cells may be actively involved in the immunoregulatory pathways leading to intestinal inflammation. The aim of this study was to assess expression by intestinal epithelial cells of cytokines with potential involvement in the development of intestinal inflammation in interleukin (IL)-2-deficient [(−/−)] mice. Wild-type mice, mice heterozygous for the disrupted IL-2 gene, and IL-2(−/−) mice were studied at 6, 16, and 24 wk of age. The mRNA levels of transforming growth factor-β1 (TGF-β1), tumor necrosis factor-α (TNF-α), IL-1β, IL-6, IL-15, KC, JE, and CD14 in colonic and small intestinal epithelial cells were assessed by Northern blot analysis. CD14 was also measured by Western blotting and reverse transcriptase polymerase chain reaction (RT-PCR). TGF-β1 mRNA was constitutively expressed in both colonic and small intestinal epithelial cells with increased expression in the colonic epithelium of colitic mice. CD14 was detected only in colonic epithelial cells, and mRNA levels increased severalfold in IL-2(−/−) mice with colitis. Northern analysis demonstrated increased levels of TGF-β1 and CD14 mRNA in colonic epithelial cells of IL-2(−/−) mice before the development of signs of colitis. CD14 mRNA and protein expression in the epithelial cells of colitic mice were confirmed by RT-PCR and Western blot analysis of isolated cells. In addition, IL-2(−/−) mice also expressed increased levels of IL-15 mRNA in small intestinal and colonic epithelial cells compared with heterozygous control mice. TNF-α, IL-1β, IL-6, KC, and JE mRNAs were only detectable in colonic epithelial cells of mice after the onset of colitis. Enhanced expression of TGF-β1, IL-15, and CD14 by colonic epithelial cells may play a role in the subsequent development of colitis in IL-2(−/−) mice.

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