10.3389/fimmu.2018.02483.s001 Anne-Margarete Brennecke Anne-Margarete Brennecke Sandra Düber Sandra Düber Bishnudeo Roy Bishnudeo Roy Irene Thomsen Irene Thomsen Annette I. Garbe Annette I. Garbe Frank Klawonn Frank Klawonn Oliver Pabst Oliver Pabst Karsten Kretschmer Karsten Kretschmer Siegfried Weiss Siegfried Weiss Image_1_Induced B Cell Development in Adult Mice.JPEG Frontiers 2018 bone marrow RAG B cell development B-2/B-1a/B-1b antibodies CSR T-dependent/-independent VH usage 2018-10-31 13:45:24 Figure https://frontiersin.figshare.com/articles/figure/Image_1_Induced_B_Cell_Development_in_Adult_Mice_JPEG/7276262 <p>We employed the B-Indu-Rag1 model in which the coding exon of recombination-activating gene 1 (Rag1) is inactivated by inversion. It is flanked by inverted loxP sites. Accordingly, B cell development is stopped at the pro/pre B-I cell precursor stage. A B cell-specific Cre recombinase fused to a mutated estrogen receptor allows the induction of RAG1 function and B cell development by application of Tamoxifen. Since Rag1 function is recovered in a non-self-renewing precursor cell, only single waves of development can be induced. Using this system, we could determine that B cells minimally require 5 days to undergo development from pro/preB-I cells to the large and 6 days to the small preB-II cell stage. First immature transitional (T) 1 and T2 B cells could be detected in the bone marrow at day 6 and day 7, respectively, while their appearance in the spleen took one additional day. We also tested a contribution of adult bone marrow to the pool of B-1 cells. Sublethally irradiated syngeneic WT mice were adoptively transferred with bone marrow of B-Indu-Rag1 mice and B cell development was induced after 6 weeks. A significant portion of donor derived B-1 cells could be detected in such adult mice. Finally, early VH gene usage was tested after induction of B cell development. During the earliest time points the VH genes proximal to D/J were found to be predominantly rearranged. At later time points, the large family of the most distal VH prevailed.</p>