Abstract
Correlates of protection (CoP) are invaluable for iterative vaccine design studies, especially in pursuit of complex vaccines such as a universal influenza vaccine (UFV) where a single antigen is optimized to elicit broad protection against many viral antigenic variants. Since broadly protective antibodies against influenza virus often exhibit mutational evidence of prolonged diversification, we studied germinal center (GC) kinetics in hemagglutinin (HA) immunized mice. Here we report that as early as 4 days after secondary immunization, the expansion of HA-specific GC B cells inversely correlated to protection against influenza virus challenge, induced by the antigen. In contrast, follicular T helper (TFH) cells did not expand differently after boost vaccination, suggestive of a B-cell intrinsic difference in activation and differentiation inferred by protective antigen properties. Importantly, differences in antigen dose only affected GC B-cell frequencies after primary immunization. The absence of accompanying differences in total anti-HA or epitope-specific antibody levels induced by vaccines of different efficacy suggests that the GC B-cell response upon revaccination represents an early and unique marker of protection that may significantly accelerate the pre-clinical phase of vaccine development.
Highlights
While antibodies targeting hemagglutinin (HA) protect against influenza virus [1,2], the humoral anti-HA response continuously requires adaptation to newly emerging HA variants
To distinguish between mature B-cell subsets reflecting differences in functionality and maturity, we included a B-cell lineage marker (CD19), an exclusion marker (CD4), markers for germinal center (GC) differentiation (GL7 and CD95/FAS), a marker for plasmablast and plasma cell differentiation (CD138/syndecan-1), and membrane markers upregulated by subsets of memory B cells (CD80 and PD-L2)[24,25]
To verify whether antigen-specific B cells can be detected at increased frequencies in HAimmunized animals, we immunized 8 weeks-old influenza-naive mice with either FL H1#2316 or PBS adjuvanted with alum and subsequently analyzed the B-cell response in draining lymph nodes using the described flow cytometric phenotyping panel
Summary
While antibodies targeting hemagglutinin (HA) protect against influenza virus [1,2], the humoral anti-HA response continuously requires adaptation to newly emerging HA variants. Seasonal vaccines are currently the most effective measure to prevent influenza, the laborious production of yearly reformulated vaccines prohibits immediate tuning of vaccine HA composition to viral antigenic drift or shift[3], with the associated potential for mismatches between circulating virus strains and the strains included in the vaccine[4]. The resulting suboptimal protection by these vaccines has prompted different approaches to design. Recall germinal center response and influenza vaccination study:http://flowrepository.org/id/RvFrbzEPV 0VBPcsZpvUgNHMJABrRK6ZEGfGwqvRVAhFp U0XJgDmrKSFVhc1nPIrb. HA ELISA DATA: doi.
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