Interleukin-4 (IL-4) plays an important role in several areas of T-cell biology, including the development of Th2 responses (25) and blocking Th1 effector mechanisms mediated by gamma interferon (IFN-␥), such as inducible nitric oxide synthase (iNOS) induction (1,10,28,43,50). Further, IL-4 can inhibit NO production by upregulating arginase expression, thereby providing an alternative pathway for metabolism of L-arginine, the precursor for NO (34).The outcome of schistosome infection in IL-4 Ϫ/Ϫ animals is different from that in wild-type (WT) mice (13,14,18). WT mice normally develop a strong Th2 response and survive infection (14,18). Survival is attributed to the ability of Th2 cells to orchestrate granulomatous lesions which sequester parasite eggs away from surrounding liver tissue while at the same time secreting cytokines that are selectively anti-inflammatory (4,13,15,18,24). Infected IL-4 Ϫ/Ϫ mice, in contrast, fail to develop a Th2 response and produce elevated levels of inflammatory mediators, such as IFN-␥ and NO, and develop severe cachexia and die despite still being able to make adequate granulomas (13,14,18).An interesting feature of schistosomiasis in infected IL-4mice is the failure of these animals to develop the splenomegaly that is characteristic of infection in WT mice (39). Since NO is known to inhibit T-cell proliferation (2,3,8,37,48,50), we hypothesized that increased NO levels in IL-4 Ϫ/Ϫ animals could be contributing to the lack of splenomegaly by inhibiting the proliferation of lymphocytes during infection. Our findings presented here support this view.
MATERIALS AND METHODSParasites and mice. IL-4 Ϫ/Ϫ C57BL/6 (B6) mice (14) were bred at Cornell University. WT B6 mice were obtained from Taconic Farms, Germantown, N.Y. Mice were anesthetized with sodium pentobarbitone intraperitoneally (i.p.), and each was infected percutaneously with approximately 70 Schistosoma mansoni cercariae (NMRI strain). Mice were weighed using a spring balance (Forestry Suppliers, Jackson, Miss.). Experiments were terminated once infected IL-4 Ϫ/Ϫ mice lost ϳ20% of their body weight (14); mice were euthanatized with CO 2 .Cell cultures. Spleens were harvested, and single-cell suspensions of pooled samples were prepared using sterile 70-m-pore-size nylon mesh (Becton Dickinson, Franklin Lakes, N.J.) and Dulbecco minimal essential medium (DMEM). Erythrocytes were lysed using ammonium chloride. Viable cells were counted using trypan blue exclusion and resuspended at 10 7 /ml in DMEM containing 30 mM HEPES, 100 U of penicillin/ml, 100 g of streptomycin/ml, 2 mM glutamine, 5 ϫ 10 Ϫ5 M 2-mercaptoethanol, and 10% hiFCS (complete medium). Cells were incubated alone or were stimulated with 50 g of soluble egg antigen (SEA)/ml (11) or with plate-bound anti-CD3 monoclonal antibody (MAb) (Pharmingen, San Diego, Calif.), 0.5 g/well, with or without aminoguanidine (AMG) (hemisulfate salt, 1 mM), in 96-well flat-bottom plates in 5% CO 2 at 37°C. Cell culture supernatants were collected at 72 h and stored at Ϫ20°C until cyto...
