Antigen-specific IgA B memory cell responses to Shigella antigens elicited in volunteers immunized with live attenuated Shigella flexneri 2a oral vaccine candidates

Abstract

We studied the induction of antigen-specific IgA memory B cells (B(M)) in volunteers who received live attenuated Shigella flexneri 2a vaccines. Subjects ingested a single oral dose of 10(7), 10(8) or 10(9) CFU of S. flexneri 2a with deletions in guaBA (CVD 1204) or in guaBA, set and sen (CVD 1208). Antigen-specific serum and stool antibody responses to LPS and Ipa B were measured on days 0, 7, 14, 28 and 42. IgA B(M) cells specific to LPS, Ipa B and total IgA were assessed on days 0 and 28. We show the induction of significant LPS-specific IgA B(M) cells in anti-LPS IgA seroresponders. Positive correlations were found between anti-LPS IgA B(M) cells and anti-LPS IgA in serum and stool; IgA B(M) cell responses to IpaB were also observed. These B(M) cell responses are likely play an important role in modulating the magnitude and longevity of the humoral response.

Figure 1. Antigen-specific IgA B_M cell responses

Shown are recipients of 10⁷ (triangles), 10⁸ (circles) and 10⁹ (squares) CFU of CVD 1204 (solid lines) and CVD 1208 (broken lines) who mounted a ≥4-fold rise of anti-LPS (n=12) and anti-IpaB (n=7) IgA pre to post vaccination and evidenced appropriate B_M expansion in vitro; LPS (a) and IpaB (b) ELISPOT performed on days 0 and 28; comparisons made by Wilcoxon Signed Rank Test for 1204 and 1208 combined. Results are expressed as the % of specific B_M SFC per total IgA⁺ expanded cell populations; horizontal lines represent the median of the corresponding groups.

Figure 2. Correlation of antigen-specific B_M cells per 10⁶ expanded PBMC with antigen-specific B_M cells/total IgA B_M cells

B_M cells specific for LPS (a) and IpaB (b) /10⁶ expanded cells 28 days after immunization were plotted against the % of specific SFC divided by total IgA-secreting expanded cell populations for all volunteers studied; dotted lines represent the 95% confidence interval.

Figure 3. Correlation of anti-LPS B_M cell responses with serum and stool antibodies

Recipients of CVD 1204 and CVD 1208 who mounted a ≥4-fold rise of anti-LPS/total stool sIgA pre to post vaccination (mucosal responders) and evidenced appropriate B_M expansion in vitro (n=16) were included in the analysis. Peak serum IgA titers to LPS (a), peak stool IgA specific for LPS divided by total IgA (b), and peak LPS specific IgA ASC (c) were plotted against the number of LPS-specific B_M cells /10⁶ expanded cells 28 days after immunization; dotted lines represent the 95% confidence interval.

Figure 4. Pre to post vaccination assessment of B cell subsets expressing the gut homing receptor integrin α₄/β₇

Shown are the % of CD19⁺ integrin α₄/β₇⁺ cells co-expressing CD27 and CD20 (left panels) and CD27 but not CD20 (right panels) pre and day 28 post vaccination for IgA⁺ (upper panels) and IgG⁺ cells (lower panels) among LPS seroresponders and non-responders. *p <0.05 by Wilcoxon Rank Sum comparing net change in % of the indicated subsets post minus pre immunization among LPS seroresponders versus non-responders; LPS seroresponse is defined as ≥4-fold rise of antigen-specific antibody in serum (all responders were both IgA and IgG seroresponders).