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The interaction between Fas ligand and Fas, both expressed on activated T cells, is the major pathway in the regulation of activation-induced cell death. However, activated T cells that express membrane Fas are initially resistant to anti-Fas-induced apoptosis and become susceptible only after proliferation in vitro. Since IL-2 is known to regulate activation-induced cell death, we studied the effect of IL-2 on anti-Fas-mediated apoptosis. Interference with the IL-2 pathway was achieved by 1) inhibition of cytokine synthesis using cyclosporin A or FK506, 2) neutralization of IL-2 by anti-IL-2 Ab, 3) inhibition of binding to IL-2R by CD25 mAb, and 4) blocking of IL-2R signaling by rapamycin. We show that Fas expression is independent of the IL-2 pathway, whereas Fas-mediated apoptosis does not develop in the presence of inhibitors of IL-2 production or signaling. While the addition of rIL-2 reversed the inhibitory effect of cyclosporin A and FK506, the addition of rIL-4, rIL-7, or rIFN-gamma did not, although these cytokines induced progression into the S phase of the cell cycle. Aphidicolin-treated activated T cells that do not progress into the S phase were susceptible to Fas-mediated apoptosis. Therefore, Fas-mediated apoptosis is controlled by signals generated by IL-2 in agreement with the reported alteration of apoptosis in mice deficient in IL-2 or IL-2R.

Galectin-3 is a beta-galactoside binding protein expressed by activated macrophages, epithelial cells, and certain other cell types. Galectin-3 has a C-terminal carbohydrate binding domain, an N-terminal part consisting of a proline- and glycine-rich repetitive domain, and a small N-terminal domain. Two independent LPS binding sites on galectin-3 were demonstrated by binding of biotinylated LPS to immobilized recombinant galectin-3. One appears to be the carbohydrate binding site in the C-terminal domain that confers binding of LPS from Klebsiella pneumoniae that has a beta-galactoside-containing polysaccharide chain. This binding is best demonstrated using galectin-3 immunocaptured by a mAb to the N-terminal part (M3/38) and is inhibited by lactose. In contrast, Salmonella minnesota R7 LPS (Rd mutant), which is devoid of beta-galactosides, appears to bind to a site within the N-terminal part of galectin-3. This interaction is best demonstrated using galectin-3 directly immobilized in wells, and it is inhibited by the Ab M3/38, but not by lactose. Binding inhibition by polymyxin B and the profile of inhibition by a panel of LPSs with different amounts of the inner and outer cores present indicate that this second binding site recognizes the lipid A/inner core region of LPSs.