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Entry of B lymphocytes into the persistent cell pool in non-immunized mice is not accompanied by somatic mutation of
J A Chies1, M P Lembezat, A A Freitas
1Unité d'Immunobiologie, CNRS, URA 359, Institut Pasteur, Paris, France.
In this study we compare VH-gene repertoires of short-lived and persistent B lymphocytes in normal nonimmunized mice. Enriched populations of persistent peripheral B cells were obtained in vivo either by (i) repeated injections with hydroxyurea or (ii) maintained ganciclovir administration to herpes simplex virus-1 thymidine kinase transgenic mice. Both approaches have previously been shown to deplete newly formed, short-lived B cells. VH genes expressed by persistent or unselected B cell populations were amplified by polymerase chain reaction, cloned using the lambda-ImmunoZAP system (Stratagene) and sequenced. The results presented here concern a total of 116 complete VH sequences from two VH gene families of established germ-line composition: VH7183 and VHX24. No differences were found between the two cell populations as to usage of D or JH segments and to the presence of N sequence additions at D/JH or VH/DJH junctions and CDR3 length. Over 90% of the sequenced VH genes were of germ-line arrangement with no evidence of somatic mutation. These results show that persistent B cells in normal mice are not of embryonic origin and that somatic hypermutation is not necessary for B cell survival. They also suggest that a significant fraction of persistent IgM+ B cells in normal mice are not generated by conventional antigenic stimulation and could represent a novel class of "memory" cells expressing germ-line repertoires.
In this study we compare VH-gene repertoires of short-lived and persistent B lymphocytes in normal nonimmunized mice. Enriched populations of persistent peripheral B cells were obtained in vivo either by (i) repeated injections with hydroxyurea or (ii) maintained ganciclovir administration to herpes simplex virus-1 thymidine kinase transgenic mice. Both approaches have previously been shown to deplete newly formed, short-lived B cells. VH genes expressed by persistent or unselected B cell populations were amplified by polymerase chain reaction, cloned using the lambda-ImmunoZAP system (Stratagene) and sequenced. The results presented here concern a total of 116 complete VH sequences from two VH gene families of established germ-line composition: VH7183 and VHX24. No differences were found between the two cell populations as to usage of D or JH segments and to the presence of N sequence additions at D/JH or VH/DJH junctions and CDR3 length. Over 90% of the sequenced VH genes were of germ-line arrangement with no evidence of somatic mutation. These results show that persistent B cells in normal mice are not of embryonic origin and that somatic hypermutation is not necessary for B cell survival. They also suggest that a significant fraction of persistent IgM+ B cells in normal mice are not generated by conventional antigenic stimulation and could represent a novel class of "memory" cells expressing germ-line repertoires.