Changes in cell surface glycosylation occur during the development and differentiation of cells and have been widely correlated with the progression of several diseases. Because of their structural diversity and sensitivity to intra-and extracellular conditions, glycans are an indispensable tool for analyzing cellular transformations. Glycans present on the surface of intestinal epithelial cells (IEC) mediate interactions with billions of native microorganisms, which continuously populate the mammalian gut. A distinct feature of IECs is that they differentiate as they migrate upwards from the crypt base to the villus tip. In this study, nano-LC/ESI QTOF MS profiling was used to characterize the changes in glycosylation that correspond to Caco-2 cell differentiation. As Caco-2 cells differentiate to form a brush border membrane, a decrease in high mannose type glycans and a concurrent increase in fucosylated and sialylated complex/hybrid type glycans were observed. At day 21, when cells appear to be completely differentiated, remodeling of the cell surface glycome ceases. Differential expression of glycans during IEC maturation appears to play a key functional role in regulating the membrane-associated hydrolases and contributes to the mucosal surface innate defense mechanisms. Developing methodologies to rapidly identify changes in IEC surface glycans may lead to a rapid screen-

The phorbol ester 12-O-tetradecanoylphorbol 13-acetate induces tumour promotion, inflammation, cell proliferation and prostaglandin release. Recent reports suggest that the prostaglandins released by 12-O-tetradecanoylphorbol 13-acetate (TPA) initiate a cascade of events leading to polyamine synthesis and cell proliferation. In experiments designed to test this contention, it was found that addition of TPA (1 microM to 1 nM) to confluent mouse 3T3 fibroblasts successively caused the release of prostaglandins E2 and I2, induction of the enzyme ornithine decarboxylase (EC 4.1.1.17), stimulation of [3H]thymidine incorporation into DNA, and cell proliferation. Pretreatment of the cells with the anti-inflammatory steroid dexamethasone (1 microM) or the non-steroidal anti-inflammatory drug indomethacin (1 microM) inhibited TPA-induced prostaglandin release. However, dexamethasone enhanced the other effects of TPA, whereas indomethacin was ineffective. Addition of prostaglandin E2 to the cultures did not induce ornithine decarboxylase activity and cell proliferation. Pretreatment of the cells with 1,3-diaminopropane (1 mM) or alpha-methylornithine (5 mM), inhibitors of polyamine synthesis, decreased TPA-induced ornithine decarboxylase activity without affecting DNA synthesis. TPA stimulated [3H]thymidine incorporation into DNA, even when the ornithine decarboxylase activity was completely blocked. These data suggest that the proliferative effect of TPA on 3T3 cells is independent of prostaglandin release and polyamine synthesis.