The other major advantage of ZFN is the speed of the procedure since KO rats can be generated

Midostaurin in about 4 months in both inbred and outbred strains 8, 9, 23. Finally, mutations are definitive and transmitted to the progeny. Our characterization of IgM KO and JH KO rats confirm the previous findings in μMT or J KO mice 11, 12 and immunodeficient human patients 24, 25 that the absence of membrane Ig expression results in the absence of B cells. On the contrary, IgM deletion 26 or truncation 13 in mice permitted expression of other heavy chains and allowed B-cell development and maturation due to replacement of IgM by IgD. Similarly to humans 24, 27, IgM KO rats showed only 5% of normal levels of BM pro–pre B cells, whereas μMT mice showed normal levels of BM pro–pre B cells 11. In this regard, deletion in mice of the Ig JH region resulted in a block of Ig gene expression and B-cell development this website at the pro-B-cell stage 12 as for JH KO rats. Thus, like μMT mice in which transcription and translation of μ-chain occurred but did not result in expression

of membrane-bound IgM and like JH KO mice, IgM KO rats showed a shortened μ transcript and the absence of Ig polypeptide production and therefore a very early B-cell block. As for mice and human cells, an enigma still persists

on how B-cell levels can be suppressed early or potentially, after rearrangement at the pre-BCR stage but before a fully functional μ polypeptide is expressed. An answer to this may be dependent on the level of early control of the IgH locus when chromatin is opening and antisense transcription will be initiated before D to J recombination 28. It is possible that strain-specific parameters as well as size and position of the removed or targeted region may determine the B-cell block. Another difference with IgM KO mice 11 is that these mice showed normal levels of IgA and absence of all other Ig isotypes 29, whereas IgM KO rats showed complete deficiency Edoxaban of all isotypes including IgA. Analogously to IgM KO rats, patients with deletion of the μ locus also result in the absence of Ig production for all isotypes including IgA 25. Since in contrast to mice, only 1% of cells recovered from the peritoneal cavity of rats are B cells 17, we did not analyze this compartment. In IgM or JH KO rats’ T-cell numbers in spleen but not in lymph nodes were decreased, as described for μMT mice 14, 15. In μMT mice, this was due to the lack of production of lymphotoxin α1β2 by B cells, required for CCL21 and stromal cell development, and as yet to be defined mechanism(s) for the promotion of T-cell numbers 14.