Engagement of monocytes, NK cells, and CD4+ Th1 cells by ALVAC-SIV vaccination results in a decreased risk of SIVmac251 vaginal acquisition
Fig 2
Monocytes and NKG2A cells in risk of SIVmac251 acquisition.
Pearson correlation of myeloid cell subsets with SIVmac251 acquisition in the ALVAC-SIV group. The levels of myeloid cells were analyzed only from animals in Part 2 (Table 1). (A) The percentage of total monocytes and (B) CD14+CD16- classical monocytes were associated with decreased rate of SIV acquisition. (C) GSEA analysis of the transcriptomic profile of the ALVAC-SIV/gp120 animals at week 26 revealed an enrichment of classical monocyte markers among genes associated with lower risk of SIV acquisition. The SLEA method was used to summarize classical monocyte markers for each subject. A scatterplot shows average classical monocyte markers as a function of the frequency of classical monocytes measured by FCM in ALVAC-SIV treated animals at week 26. The grey region indicates the 95% confidence-interval of this correlation. (D) Relative frequency of NKG2A cells (defined as CD45+, CD3-, CD20-, and CD14- cells) at week 13 for the ALVAC-SIV/gp120, NYVAC-SIV/gp120, and control groups from Part 2 (horizontal line: mean). (E) Correlation between the frequency of intermediate monocytes in blood at week 26 and the cytotoxic function of mucosal NKG2A cells (week 13). (F) Frequency of vaginal NKG2A CD107+ cells in the ALVAC-SIV vaccinated, NYVAC-SIV vaccinated, and control groups from Part 2 of the study at week 26 (horizontal line: mean). (G) Scatterplot of the average of cytotoxic NK cell markers as a function of the average expression of classical monocyte markers. The gene expression of FAS and TNF, two canonical cytotoxic NKs, as function of the expression of classical monocyte markers is indicated by lines. (H) Heatmaps showing the level of expression of transcriptomic markers of classical monocytes in three NHP studies. In the current study (ALVAC/gp120 [Ivag]; left), animals were primed with ALVAC-SIV and boosted with ALVAC-SIV+gp120 formulated in alum. Blood samples were taken 24h after the first boost (week 12) and after the 2nd boost (week 24). In a prior study (DNA/ALVAC/gp120 [IR]; center), animals were primed with DNA and boosted with ALVAC-SIV/gp120 formulated in alum [16]. Blood samples were taken 24h after the first (week 12) and second (week 24) boosts. In another of our prior studies (ALVAC/gp120 [IR]; right), animals were primed with ALVAC-SIV, boosted with ALVAC-SIV+gp120 formulated in alum, and challenged intrarectally [15]. Blood samples were taken 24 h after the first ALVAC-SIV immunization. GSEA enrichment analysis was used to test for enrichment of transcriptomic markers of classical monocytes [80] among genes correlated with challenges in each study (ALVAC/gp120 [Ivag]: 1st boost, NES = 2.70, FDR < 0.001; 2nd boost, NES = 2.33, FDR < 0.001; DNA/ALVAC/gp120 [IR]: 1st boost: NES = 1.23, FDR = 0.104; 2nd boost: NES = 0.888, FDR = 0.779; ALVAC-SIV/g120 [IR]: NES = 1.54, FDR = 0.146). The markers of classical monocytes correlating with challenges (i.e. leading edge genes) overlapping between the three studies are shown in the heatmap. A blue-to-red color gradient represents the log2 fold-change between post-vaccination and pre-vaccination gene expression. A Spearman correlation and t-test were used. The Spearman correlations were transformed to t statistics and compared to the Student distribution (t-test) using the formula below. In this formula, rho is Spearman’s rho for samples, t is taken to be (1 –[a/2]) of the t-distribution (with n– 2 degrees of freedom), and null hypothesis significance testing rejects the null if |t*| is greater than or equal to t. This was done to statistically assess the correlation between the ordering of the samples by the levels of gene expression and challenge (ALVAC/gp120 [Ivag]: 1st boost, R = 0.48, p = 0.0433; 2nd boost, R = 0.15, p = 0.5464; DNA/ALVAC/gp120 [IR]: 1st boost, R = 0.71, p = 0.0089; 2nd boost, R = 0.37, p = 0.2411; ALVAC-SIV/gp120 [IR]: R = 0.30, p = 0.1305). (I) Scatterplot of average IL18 measured by Luminex assay following envelope stimulation of blood cells at week 13 as a function of the number of SIV challenges to infection in the ALVAC-SIV/gp120 treated animals. The grey region indicates the 95% confidence-interval of this correlation.
