Reduced Ig Class Switch in Aged Mice Correlates with Decreased E47 and Activation-Induced Cytidine Deaminase

Abstract

The capacity to class switch the IgH chain is critical to the effectiveness of humoral immune responses. We show that in vitro-stimulated splenic B cells from senescent mice are deficient in production of multiple class switch isotypes (IgG1, G2a, G3, and E), class switch recombination (CSR), and induction of the E2A-encoded transcription factor E47. E47 has previously been shown to be required for CSR, at least in part via expression of the activation-induced cytidine deaminase. Our studies show that impaired induction of E47, and subsequently activation-induced cytidine deaminase, contribute to poor CSR and production of secondary isotypes in senescence.

Mouse CMV infection delays antibody class switch upon an unrelated virus challenge

“…It remains unclear if the effect of CMV infection on Ig class-switch is truly CD4 independent, or if CMV induces a subtle change in the function of CD4 T-cells, which could not be measured by our assay. Alternatively, CMV may impair B-cell responses by disrupting the lymph-node architecture and thus the signaling network that is necessary for the germinal center reaction, or by direct effects on the B-cell compartment, for instance by dysregulating TNFα signaling (Frasca et al, 2012) or by affecting E47 or activation-induced cytidine deaminase, similarly to effects observed in aging mice (Frasca et al, 2008;Frasca et al, 2004). These important questions may be addressed in future experiments.…”

Aging murine B cells have decreased class switch induced by anti-CD40 or BAFF

“…Because in B cells from both young mice (Castigli et al, 2005a) and humans (Litinskiy et al, 2002) an anti-CD40/IL-4 signal gives about twice the amount of CSR compared to BAFF/IL-4, here we hypothesized that aged mice would also show a similar importance of anti-CD40/IL-4 stimulation for CSR. We also hypothesized that E47 would be the major regulator of CSR in old B cells, as suggested from our previous work on the regulation of anti-CD40/IL-4 response (Frasca et al, 2004a), and that NF-kB, the major transcription factor induced by BAFF, which has been shown to be down-regulated by aging in many cell types (Gosselin and Abbadie, 2003), including T cells (Haynes et al, 2004), would be downregulated in old B cells also. Results show that anti-CD40/IL-4 gave better (2-fold) CSR than BAFF/IL-4 in young B cells, as measured by RT-PCR of post-switch transcripts and flow cytometry.…”

“…This concentration of IL-4 was chosen because it gave the optimum response for both the young and the old splenic B cell cultures (not shown); its effects were comparable to those given by a different IL-4 preparation (Biosource Int. PMC0046) which was used at the dose of 1 μg/ml used in our previously published experiments (Frasca et al, 2004a). At the end of the incubation time, cells were harvested, protein extracts prepared (for EMSA experiments), and mRNA extracted (for RT-PCR and real-time PCR experiments).…”

“…Protein extracts were prepared from the same numbers of cultured spleen cells essentially as previously published (Frasca et al, 2004a); briefly, cells were harvested and centrifuged in a 5415C Eppendorf microfuge (2,000 rpm, 5 min.). The pellet was resuspended in 30 μl of solution A containing Hepes 10 mM, pH 7.9, KCl 10 mM, EDTA 1.0 mM, DTT 1 mM, MgCl 2 1.5 mM, PMSF 1 mM, 1 tablet of protease inhibitor cocktail (Boeringer Manheim, Germany) (per 20 ml) and Nonidet P-40 (0.1%), briefly vortexed and centrifuged (8,000 rpm, 5 min., 4°C).…”

“…Overexpression of E47 can directly induce Aicda gene expression both in a B cell line and in splenic B cells activated in vitro. We have recently demonstrated that in vitro stimulated splenic B cells from senescent mice are deficient in production of multiple class switch isotypes (IgG1, IgG2a, IgG3, and IgE), CSR, induction of the E2A-encoded transcription factor E47, and AID (Frasca et al, 2004a).…”