In senescence, B lymphopoiesis declines dramatically. In the mouse, decreased production of pre-B cells in the bone marrow is accompanied by alterations in the generation of the B cell specificity repertoire. Such disfunction in B cell repertoire expression has been shown to adversely affect protective murine humoral immunity to infectious microorganisms. The molecular mechanisms responsible for the diminished B lymphopoiesis during aging have not been addressed. We hypothesize that I) with senescence, mice express significantly lower levels o the enzymes Rag-1/2 and terminal deoxynucleotidyl transferase (TdT); 2) this results in altered Vh gene usage and diminished N segment diversity and leads to production of mu heavy chain V region sequences which are less effective in combining with surrogate light chains (lambda5; VpreB) and/or fail to produce functional pre-B cell receptors; and 3) as a result, fewer new pre-B cells are recruited into the mitotically active large pre-B cell stage. The consequences of these events would be both diminished numbers of small, resting pre-B cells within the bone marrow and alteration of the Vh repertoire seen at the pre-B cell level. In order to test this hypothesis, we propose in Specific Aim 1 to evaluate expression of Rag- 1/2, and TdT within cell sorted pro-B and pre-B cells at both the protein and mRNA levels as a function of age.
In Specific Aim 2, the expression of mu heavy chain and surrogate light chain expression in pro-B and pre-B cells from young and senescent mice will be compared. Finally, in Specific Aim 3, CDR3 Vh sequences for two gene families, Vh36-60 and Vh7183, will be amplified from genomic DNA by PCR, cloned, and sequenced. The expression of these VII genes as productive and non-productive rearrangements will be determined among nascent (CD43+ CD25+ B220+) and later (CD43- CD25+ B220+) pre-B cells from young and aged mice. These studies will provide insight into the effect of aging on the VII- dependent recruitment/proliferation apparent among pre-B cells. Completion of these Specific Aims will provide a molecular understanding of the events which underlie the progressive loss of B lymphopoiesis and alterations in repertoire expression seen during senescence.
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| Sherwood, E M; Xu, W; King, A M et al. (2000) The reduced expression of surrogate light chains in B cell precursors from senescent BALB/c mice is associated with decreased E2A proteins. Mech Ageing Dev 118:45-59 |
| Sherwood, E M; Blomberg, B B; Xu, W et al. (1998) Senescent BALB/c mice exhibit decreased expression of lambda5 surrogate light chains and reduced development within the pre-B cell compartment. J Immunol 161:4472-5 |
