Establishment and Analysis of Germfree T Cell Receptor Transgenic Mice

Abstract

Recent studies have shown that intestinal bacteria affect the intestinal immune system. In order to elucidate the effects of intestinal microflora on T-cell mediated immune responses of the intestine, we established germfree (GF) T cell receptor transgenic (TCR-Tg) mice. GF ovalbumin (OVA)-specific TCR-Tg mice were obtained from conventional (CV) TCR-Tg mice by hysterectomy. Cells from spleen, Peyer’s patch (PP) and lamina propria (LP) were isolated from GF TCR-Tg mice, and the ratio of CD4⁺ T cells was assessed by flow cytometry and the production of cytokines, in response to in vitro OVA stimulation, was measured by ELISA. The numbers of PPs were significantly lower in GF TCR-Tg mice compared with CV TCR-Tg mice and the ratios of CD4⁺ T cells in PP and LP cells from GF TCR-Tg mice were decreased. When stimulated with OVA, PP cells from GF TCR-Tg mice secreted higher levels of interferon (IFN)-γ, interleukin (IL)-5 and IL-6, and LP cells from GF mice secreted higher levels of IFN-γ and IL-6 compared with CV mice. These results suggested that although the gut-associated lymphoid tissue is poorly developed in germfree mice, intestinal T cells developing under these conditions possessed an enhanced ability to secrete cytokines in response to antigenic stimulation. Our TCR-Tg system should be an informative system to evaluate the effect of intestinal microflora on antigen-specific T cell responses.