Requirements for developing mixed-chimerism in nonmyeloablative total-lymphoid irradiated mice: role of IL-4 and immunoredirection.

Abstract

Adult mice treated with total-lymphoid irradiation (TLI) followed by hematopoietic cell transfer develop stable mixed-chimerism. The purpose of the study is to examine the requirements for the development of mixed-chimerism and characterize the immune responses associated with mixed-chimerism. T-cell number and function were examined in spleens of TLI-treated mice at various times after the completion of TLI by fluorescence-activated cell sorter (FACS) analysis and enzyme-linked immunosorbent assay (ELISA). TLI-treated BALB/c mice were injected with CAF(1) spleen cells between 2 and 28 days after completing TLI, with or without concurrent anti-IL-4. The extent of mixed-chimerism was determined using multiparameter FACS analysis. Enzyme-linked immunosorbent spot (ELISPOT) assays were used to measure anti-donor CD4+ and CD8+ immune responses. TLI treatment results in transient lymphopenia, sparing CD4 cells relative to CD8 cells, followed by gradual, partial recovery over 28 days. Day 2 post-TLI T cells produce more interleukin (IL)-4, but less IL-2, interferon (IFN)-gamma and IL-10, while day 28 post-TLI T cells produce more IFN-gamma relative to IL-4. More than 70% of mice develop mixed-chimerism when injected with CAF(1) cells at 2 days post-TLI, while none become chimeric if injected at 28 days post-TLI. Mixed-chimeric mice contain more anti-donor Th2 CD4 cells and less anti-donor TC1 CD8 cells compared with nonchimeric mice and nonirradiated controls. Treatment with anti-IL-4 inhibits the development of mixed-chimerism ( <0.05), and these nonchimeric mice contain donor-reactive TC1 CD8 cells. IL-4 plays a role in the development of mixed-chimerism, perhaps by inhibition of anti-donor TC1 CD8 cells or enhancement of anti-donor Th2 CD4 cells.