Synthesis of DNA by the spleens of germ-free mice during the primary response to sheep red cells

The mitotic activity of the spleens of non-immunized germ-free (GF) mice was less than in similar specific pathogen-free (SPF) animals. No evidence of germinal center activity was noted in these GF mouse spleens before challenge with sheep erythrocytes (SRC). These centers only began to develop 4 days after immunization and were not fully developed before 8 days. In control SPF mice, the spleens showed very few germinal centers which were small in size, but they showed the same pattern of evolution as in similarly immunized GF mice. The changes in the red pulp, characterized by the development of clusters of nuclei labeled with [³H]thymidine, followed closely the development of plaqueforming cells (PFC). The numbers of direct PFC reached the same peak level in GF and SPF mice on day 4 of the response to SRC, but were not so well sustained in the former animals after this time. Indirect PFC were much lower in the spleens of GF mice than in SPF animals. The pattern and degree of increase of DNA synthesis in the spleen of GF and SPF mice following immunization with SRC differed from and was less than that of mice reared in less clean conditions. Increased DNA synthesis occurred very soon after injection of SRC (6 to 24 h) and the increase was sustained for 4 days without further significant rise and then declined. Autoradiographs of the spleen of immunized GF mice given [³H]thymidine showed that the first increase of labeled nuclei in the white pulp occurred around the central arterioles as early as 6 h after SRC. This was followed by increased labeling in the mantle layer of the white pulp and the characteristic pattern of germinal center labeling developed after 4 days. Increased labeling of nuclei developed in the red pulp as early as in the white pulp, while the subcapsular and trabecular areas showed high labeling indices even in the spleens of non-immunized controls. The ratio of labeling index/mitotic index which is governed by the respective durations of DNA synthesis and mitosis in those cells in division cycle, varied between 10 and 280 in different areas of the spleen. This indicated a vast excess of cells synthesising DNA in relation to the numbers of dividing cells actually present in the spleens of these mice.