LT-K63 Enhances B Cell Activation and Survival Factors in Neonatal Mice That Translates Into Long-Lived Humoral Immunity

Abstract

Adjuvants enhance magnitude and duration of immune responses induced by vaccines. In this study we assessed in neonatal mice if and how the adjuvant LT-K63 given with a pneumococcal conjugate vaccine, Pnc1-TT, could affect the expression of tumor necrosis factor receptor (TNF-R) superfamily members, known to be involved in the initiation and maintenance of antibody responses; B cell activating factor receptor (BAFF-R) and B cell maturation antigen (BCMA) and their ligands, BAFF, and a proliferation inducing ligand (APRIL). Initially we assessed the maturation status of different B cell populations and their expression of BAFF-R and BCMA. Neonatal mice had dramatically fewer B cells than adult mice and the composition of different subsets within the B cell pool differed greatly. Proportionally newly formed B cells were most abundant, but they had diminished BAFF-R expression which could explain low proportions of marginal zone and follicular B cells observed. Limited BCMA expression was also detected in neonatal pre-plasmablasts/plasmablasts. LT-K63 enhanced vaccine-induced BAFF-R expression in splenic marginal zone, follicular and newly formed B cells, leading to increased plasmablast/plasma cells, and their enhanced expression of BCMA in spleen and bone marrow. Additionally, the induction of BAFF and APRIL expression occurred early in neonatal mice immunized with Pnc1-TT either with or without LT-K63. However, BAFF + and APRIL + cells in spleens were maintained at a higher level in mice that received the adjuvant. Furthermore, the early increase of APRIL + cells in bone marrow was more profound in mice immunized with vaccine and adjuvant. Finally, we assessed, for the first time in neonatal mice, accessory cells of the plasma cell niche in bone marrow and their secretion of APRIL. We found that LT-K63 enhanced the frequency and APRIL expression of eosinophils, macrophages, and megakaryocytes, which likely contributed to plasma cell survival, even though APRIL + cells showed a fast decline. All this was associated with enhanced, sustained Aradottir Pind et al. LT-K63 and B-cell Survival Mechanisms vaccine-specific antibody-secreting cells in bone marrow and persisting vaccine-specific serum antibodies. Our study sheds light on the mechanisms behind the adjuvanticity of LT-K63 and identifies molecular pathways that should be triggered by vaccine adjuvants to induce sustained humoral immunity in early life.

A comparative study of adjuvants effects on neonatal plasma cell survival niche in bone marrow and persistence of humoral immune responses

“…Ab persistence is mediated by long-lived plasma cells that reside in specialized survival niches in the bone marrow ( 34 ) and their survival was recently shown to be dependent on direct contacts with stromal cells as well as APRIL : BCMA binding ( 35 ). In line with that, we demonstrated that LT-K63 enhanced early APRIL expression by bone marrow accessory cells, particularly by eosinophils, macrophages and megakaryocytes after immunization of neonatal mice with Pnc1-TT ( 15 ). Additionally, a higher proportion of plasmablasts and plasma cells of neonatal mice immunized with Pnc1-TT with LT-K63 expressed BCMA ( 15 ).…”

“…In line with that, we demonstrated that LT-K63 enhanced early APRIL expression by bone marrow accessory cells, particularly by eosinophils, macrophages and megakaryocytes after immunization of neonatal mice with Pnc1-TT ( 15 ). Additionally, a higher proportion of plasmablasts and plasma cells of neonatal mice immunized with Pnc1-TT with LT-K63 expressed BCMA ( 15 ). Therefore, we wanted to explore whether the difference we previously observed ( 20 ) in the persistence of humoral immune responses induced by the selected adjuvants could be explained by their different effects on expression of plasma cell survival factors by bone marrow accessory cells and BCMA expression of plasmablasts/plasma cells up to this 6 week time point, where LT-K63 is used as a positive control.…”

“…TT-specific ASC were enumerated by ELISPOT, as previously described ( 15 , 19 , 20 , 38 ). MultiScreen High protein binding immobilon-P membrane plates (Millipore Corporation, Bedford, MA) were coated with 10 μg/ml TT overnight at 37°C, blocked with complete RPMI 1640 (Life Technologies BRL, Life Technologies, Paisley, U.K.).…”

“…We evaluated effects of four adjuvants to overcome limitations of neonatal immunity and induce potent and persistent immune responses following neonatal immunization with the protein vaccine tetanus toxoid (TT) and compared with the previously established effect of LT-K63 ( 15 ). The adjuvants assessed are of various categories and have been reported to employ different mechanisms of action.…”

Understanding Antibody Responses in Early Life: Baby Steps towards Developing an Effective Influenza Vaccine

“…In addition to use of PAMPS that are likely to affect multiple immune populations, small molecules that bind B cell surface receptors involved in survival and proliferation, such as BAFF, APRIL, and soluble CD40L, have shown promise as potential adjuvants in adult models [ 169 ] ( Figure 2 ). However, the utility of these adjuvants in young infants may be limited by differences in expression of the surface receptors for these molecules [ 170 ]. This is exemplified by the diminished expression of TACI by B cells observed in newborn mice, leading to deficits in plasma cell differentiation even in the presence of exogenous BAFF and APRIL [ 47 ].…”