Increased plasma IL-21 and IL-21R expression in CD4 T cells is associated with humoral response to the influenza vaccine.

<p>(<b>A</b>) Circulating levels of IL-21 were measured by ELISA at t0 and wk4 in the plasma of 11 HIV^– seroprotected (HIV^– SP), 12 HIV⁺ seroprotected (HIV⁺ SP) and 4 HIV⁺ non seroprotected (HIV⁺ NSP) post-menopausal women. (<b>B</b>) Correlation between TNFα plasma levels at baseline and the ratio between wk4 and baseline plasma IL-21. (<b>C</b>, <b>D</b>) IL-21R mean fluorescence intensity (MFI) was established in live CD4 (<b>C</b>) and CD8 (<b>D</b>) T cells of 8 HIV^– SP, 11 HIV⁺ SP and 4 HIV⁺ NSP aging women. Frozen cells obtained before and four weeks after vaccine administration were thawed, rested overnight, stained for surface markers and acquired by flow cytometry. IL-21R MFI was established for ViViD^-CD3⁺CD4⁺ or ViViD^-CD3⁺CD8⁺ events. Statistical analysis was performed using paired Student’s <i>t</i>-test for comparisons within groups, unpaired Student’s <i>t</i>-test for comparisons between groups, and Pearson correlation as appropriate.</p

T cell activation and senescence are associated with decreased humoral response to influenza vaccination.

<p>Frozen PBMC obtained before influenza vaccination were thawed, rested over night and stained with monoclonal antibodies for immunophenotyping of CD4 (ViViD^-CD3⁺CD4⁺), pTfh (ViViD^-CD3⁺CD4⁺CD45RO⁺CXCR5⁺) and CD8 (ViViD^-CD3⁺CD8⁺) T cell subsets. Correlations were established between the reciprocal of influenza Ab titer at week 4 and the frequency of activated (CD38⁺HLA-DR⁺), dividing (Ki-67⁺) and senescent (CD28^-CD57⁺) CD4 (<b>A</b>), pTfh (<b>B</b>) and CD8 (<b>C</b>) T cells for 10 HIV^– (open dots) and 15 HIV⁺ (filled dots) post-menopausal women. Pearson analysis was utilized to establish statistical correlation between variables.</p

Impaired Antibody Response to Influenza Vaccine in HIV-Infected and Uninfected Aging Women Is Associated with Immune Activation and Inflammation

<div><p>Background</p><p>Aging and HIV infection are independently associated with excessive immune activation and impaired immune responses to vaccines, but their relationships have not been examined.</p> <p>Methods</p><p>For selecting an aging population we enrolled 28 post-menopausal women including 12 healthy volunteers and 16 HIV-infected women on antiretroviral treatment with <100 HIV RNA copies/ml. Antibody titers to trivalent influenza vaccination given during the 2011-2012 season were determined before and 4 weeks after vaccination.</p> <p>Results</p><p>Seroprotective influenza antibody titers (≥1:40) were observed in 31% HIV⁺ and 58% HIV-uninfected women pre-vaccination. Following vaccination, magnitude of antibody responses and frequency of seroprotection were lower in HIV⁺ (75%) than in HIV^– (91%) women. Plasma IL-21, the signature cytokine of T follicular helper cells (Tfh), and CD4 T cell IL-21R were upregulated with seroconversion (≥4 fold increase in antibody titer). Post-vaccine antibody responses were inversely correlated with pre-vaccination plasma TNFα levels and with activated CD4 T cells, including activated peripheral (p)Tfh. Plasma TNFα levels were correlated with activated pTfh cells (r=0.48, p=0.02), and inversely with the post-vaccination levels of plasma IL-21 (r=-0.53, p=0.02). In vitro TNFα blockade improved the ability of CD4 T cells to produce IL-21 and of B cells to secrete immunoglobulins, and addition of exogenous IL-21 to cell cultures enhanced B cell function. Higher frequencies of activated and exhausted CD8 T and B cells were noted in HIV⁺ women, but these markers did not show a correlation with antibody responses.</p> <p>Conclusions</p><p>In aging HIV-infected and uninfected women, activated CD4 and pTfh cells may compromise influenza vaccine-induced antibody response, for which a mechanism of TNFα-mediated impairment of pTfh-induced IL-21 secretion is postulated. Interventions aimed at reducing chronic inflammation and immune activation in aging, HIV-infected patients may improve their response to vaccines.</p> </div

Phenotypic alterations in B cell subsets in HIV-infected aging women are not associated with antibody response to influenza vaccination.

<p>PBMC were stained with surface MoAbs to CD20, CD21, CD10, CD27 and FcRL4. Lymphocytes were gated based on forward and side scatter. CD3^-CD20⁺ cells were gated into CD21^hi and CD21^lo/neg cells and further divided based on the expression of CD27 and CD10 as: early transitional (CD21^lo/negCD27^-CD10⁺), late transitional (CD21^hiCD27^-CD10⁺), naïve (CD21^hiCD27^-CD10^–), resting memory (CD21^hiCD27⁺CD10^–), activated memory (CD21^lo/negCD27⁺CD10^–), and exhausted tissue-like (CD21^lo/negCD27^-CD10^–) subsets. (<b>A</b>) The percentage of cells in each B cell subpopulation was determined in 10 healthy controls (open dots) and 15 HIV-infected women (filled dots) before vaccination. (<b>B</b>) Expression of the exhaustion marker FcRL4 was evaluated in each B cell subset. (<b>C</b>) Correlations were established between the B cell populations that resulted significantly different between HIV- and HIV+ in A and B and the reciprocal of influenza Ab titer at week 4. Correlation between variables was established with Pearson analysis.</p

TNFα blockade promotes IL-21 secretion and IgG production in CD4-B cell co-cultures.

<p>Memory CD4 T cells obtained from 4 healthy donors were cultured with autologous B cells at a 1:1 ratio in the presence of SEB and/or TNFα, IL-21, anti-anti-TNFα or anti-IL-21R blocking antibodies or the appropriate isotype control. (<b>A</b>) Phenotyping was performed on day 3 of co-culture in gated live CD4 and pTfh cells. (<b>B</b>) IL-21 levels measured by ELISA in day 3 culture supernatants. (<b>C</b>, <b>D</b>) IgM, IgG and IgA levels were evaluated by ELISA after 7 days of co-culture. In (<b>D</b>), IgG levels in samples where IL-21R-Fc was added were not measurable because of the cross-reactivity to the Fc portion of IL-21R-Fc. P values were calculated using paired Student’s <i>t</i>-test for comparisons between two groups, and repeated measures one-way ANOVA for comparisons between larger groups.</p

Antibody response to influenza vaccination.

<p>Antibody titers specific for the whole 2011-2012 seasonal influenza vaccine were determined before (t0) and 4 weeks (wk4) after vaccination by hemagglutination inhibition assay in the plasma of 12 HIV^– and 16 HIV⁺ elderly women. Reverse cumulative distribution curves for the antibody titers prior to (<b>A</b>) and 4 weeks after (<b>B</b>) vaccination for HIV-uninfected (open circles) and HIV-infected (filled circles) donors were derived to illustrate immune responses. (<b>C</b>) Reciprocal of Ab titers before and after vaccination. (<b>D</b>) Ratio between the wk4 and the baseline reciprocal of Ab titer. P values were calculated with Student’s <i>t</i>-test or Mann-Whitney test as appropriate.</p

Other measured extrinsic and intrinsic health factors among the first booster vaccination sub-cohort.

Other measured extrinsic and intrinsic health factors among the first booster vaccination sub-cohort.</p

Linear mixed effects model (LMM) evaluating the relationship between post-primary vaccination antibody titers and time, COVID-19 vaccine manufacturer, prior COVID-19 infection status, and biological sex.

Linear mixed effects model (LMM) evaluating the relationship between post-primary vaccination antibody titers and time, COVID-19 vaccine manufacturer, prior COVID-19 infection status, and biological sex.</p

Post-primary vaccination mean Log₂ Ab across COVID vaccine groups.

There was no significant difference (T = 1.26, p = 0.211) between mean log2 Ab between participants who received Moderna mRNA-1273 (13.50 ± 1.71) or Pfizer BNT162b2 vaccines (13.14 ± 1.64) for their primary vaccination after 2 weeks of vaccination onset.</p

Post-primary vaccination mean Log₂ Ab across biological sex groups.

The mean log2 Ab in male participants (12.87 ± 12.87) was statistically lower (T = 2.47, p = 0.015) than that of female (13.56 ± 1.46) participants after primary vaccination.</p