A molecular mechanism for TNF-α-mediated downregulation of B cell responses.

B cell function with age is decreased in class switch recombination (CSR), activation-induced cytidine deaminase (AID), and stability of E47 mRNA. The latter is regulated, at least in part, by tristetraprolin (TTP), which is increased in aged B cells and also negatively regulates TNF-α. In this study, we investigated whether B cells produce TNF-α, whether this changes with age, and how this affects their function upon stimulation. Our hypothesis is that in aging there is a feedback mechanism of autocrine inflammatory cytokines (TNF-α) that lowers the expression of AID and CSR. Our results showed that unstimulated B cells from old BALB/c mice make significantly more TNF-α mRNA and protein than do B cells from young mice, but after stimulation the old make less than the young; thus, they are refractory to stimulation. The increase in TNF-α made by old B cells is primarily due to follicular, but not minor, subsets of B cells. Incubation of B cells with TNF-α before LPS stimulation decreased both young and old B cell responses. Importantly, B cell function was restored by adding anti-TNF-α Ab to cultured B cells. To address a molecular mechanism, we found that incubation of B cells with TNF-α before LPS stimulation induced TTP, a physiological regulator of mRNA stability of the transcription factor E47, which is crucial for CSR. Finally, anti-TNF-α given in vivo increased B cell function in old, but not in young, follicular B cells. These results suggest new molecular mechanisms that contribute to reduced Ab responses in aging.

Deviation of the B cell pathway in senescent mice is associated with reduced surrogate light chain expression and altered immature B cell generation, phenotype, and light chain expression.

B lymphopoiesis in aged mice is characterized by reduced B cell precursors and an altered Ab repertoire. This likely results, in part, from reduced surrogate L chains in senescent B cell precursors and compromised pre-BCR checkpoints. Herein, we show that aged mice maintain an ordinarily minor pool of early c-kit(+) pre-B cells, indicative of poor pre-BCR expression, even as pre-BCR competent early pre-B cells are significantly reduced. Therefore, in aged mice, B2 B lymphopoiesis shifts from dependency on pre-BCR expansion and selection to more pre-BCR-deficient pathways. B2 c-kit(+) B cell precursors, from either young or aged mice, generate new B cells in vitro that are biased to larger size, higher levels of CD43, and decreased kappa L chain expression. Notably, immature B cells in aged bone marrow exhibit a similar phenotype in vivo. We hypothesize that reduced surrogate L chain expression contributes to decreased pre-B cells in aged mice. The B2 pathway is partially blocked with limited B cell development and reduced pre-BCR expression and signaling. In old age, B2 pathways have limited surrogate L chain and increasingly generate new B cells with altered phenotype and L chain expression.

Old mice retain bone marrow B1 progenitors, but lose B2 precursors, and exhibit altered immature B cell phenotype and light chain usage.

The bone marrow of old adult mice ( approximately 2 years old) has reduced B lymphopoiesis; however, whether the B1 pathway in adult bone marrow is also compromised in senescence is not known. Herein, we show that phenotypic (IgM(-)Lin(-)CD93(+)[AA4.1(+)] CD19(+)B220(low/-)) B1 progenitors are retained in old bone marrow even as B2 B cell precursors are reduced. Moreover, B1 progenitors from both young adult and old mice generated new B cells in vitro enriched for CD43 expression, likely due to their activation, and exhibited increased lambda light chain usage and diminished levels of kappa light chain expression. B1 progenitors were shown to have lower surrogate light chain (lambda5) protein levels than did B2 pro-B cells in young mice and these levels decreased in both B1 and B2 precursor pools in old age. These results indicate that the B1 B cell pathway persists during old age in contrast to the B2 pathway. Moreover, B1 B cell progenitors generated new B cells in the adult bone marrow that have distinct surface phenotype and light chain usage. This is associated with decreased surrogate light chain expression, a characteristic held in common by B1 progenitors as well as B2 precursors in old mice.