text: High Antigen Doses and STAT4 Are Required for the In Vivo Generation of IL-10-Producing Th1 Cells
  To address the mechanisms regulating IL-10 production by Th1 cells invivo, we transferred DO11.10 cells into BALB/c recipient mice and immunized the recipients with very high doses of OVA-protein with or without added lipopolysacharide (LPS). Tcells were recovered from the inguinal lymph nodes 3 days after priming and restimulated invitro with OVA peptide for 48 hr. This invivo immunization induced IL-10 and IFN-gamma production, and the amount of IL-10 production was enhanced by addition of LPS in the immunization (Figure3A) and with higher doses of OVA (3 muM versus 1 muM, data not shown). To test the role of STAT4 and STAT6 signaling in the invivo development of IL-10-producing Th1 cells, we transferred STAT4- or STAT6-deficient or WT DO11.10 cells into recipient BALB/c mice and immunized with OVA-protein plus LPS as before. In vivo expression of both IL-10 and IFN-gamma was markedly reduced but not completely abrogated in the absence of STAT4 signaling (Figures 3B and 3C), suggesting the existence of compensatory mechanisms that were absent in the invitro system. Signaling through STAT6 had no effect on IL-10 production by Th1 cells as shown by intracellular cytokine staining (ICS) and by immunoassay in STAT6-deficient Tcells (Figures 3B and 3C). 
  