Expansion of memory Th2 cells over Th1 cells in neonatal primed mice

BALB/c mice primed with CAF1 splenocytes during the neonatal stage developed A/J-specific tolerance with prolonged survival (> 60 days) of A/J skin grafts. Mice failed to develop A/J-specific cytotoxicity, but rejected third-party skin grafts and generated appropriate third-party cytotoxic T cell responses. We demonstrated previously that graft acceptance was associated with enhanced interleukin (IL)-4 and diminished interferon [IFN]-gamma tolerogen-specific cytokine production, whereas third-party graft rejection was associated with the opposite pattern of cytokine production. We now report that neonatal mice do not mount mixed lymphocyte reaction responses against A/J, but the mice contain a higher percentage of IL-4-producing cells that were characterized as CD4+Mel-14lo cells. Although alloantigen priming of both neonatal and adult control mice expands the CD4+Mel-14lo subset, CD4+Mel-14lo cells from neonatal primed mice produce significantly higher levels of IL-4 and IL-10 and lower IFN-gamma, whereas CD4+Mel-14lo cells from adult primed mice produce mainly IFN-gamma. Moreover, enzyme-linked spot immunosorption analysis demonstrates that, compared with adult primed mice, neonatal primed mice contain more IL-4-producing CD4 cells and less IFN-gamma-producing cells, which indicates that neonatal antigen exposure induces and expands alloreactive Th2 memory CD4 cells. The addition of neutralizing antibodies against IL-4 and IL-10 to primary MLR failed to recover IFN-gamma by CD4+Mel-14lo cells, but cells secreted IFN-gamma after a second in vitro restimulation with tolerogen, which indicates that CD4 cells from neonatal tolerant mice have the capacity to differentiate into Th1 cells. In summary, neonatal tolerant mice contain higher ratios of Th2/Th1 CD4 cells, and the Th2 cytokines function to maintain the ratio by inhibiting Th1 differentiation.