Fig 2: Histamine and Clobenpropit inhibit purified HIV-stimulated pDC activation.(a) pDC were acquired by flow cytometry before and after purification from PBMC of healthy donors. Negative selection was used to obtain purity higher than 90% in all experiments. (b) Chemical structures of histamine (HA) and clobenpropit (CB). (c) Isolated pDC were pretreated with histamine or with CB at the concentration of 10 μM and then stimulated with microvesicles (mock) alone or with HIV overnight. IFN-α was measured in the supernatants by Elisa. (d) IFN-α production overtime was measured in supernatants of purified pDC cultured overnight with microvesicles (mock) alone or with HIV alone or together with CB at the concentration of 10 μM. (e) Levels of membrane TRAIL were assessed by flow cytometry on purified pDC after pre-incubation with CB or the specific TLR7 antagonist, A151, and then overnight HIV stimulation. (f) mRNA levels of TRAIL from purified pDC pre-incubated with CB (10 μM) and stimulated overnight with HIV, were measured by RT-qPCR and normalized to RPL13A. Production of IFN-α by Elisa in supernatants (g) or expression of membrane TRAIL at the surface of the cells by flow cytometry (h) from purified pDC pre-incubated with CB and then cultured overnight with Influenza A or Dengue viruses. Inhibition of IFN-α production (i) and inhibition of TRAIL expression (j) were assessed in supernatants of PBMC and by flow cytometry on PBMC cultured overnight with Influenza A virus alone or together with histamine (HA) or CB at a concentration ranging from 0.5 μM to 1 mM. Data shown as the mean of three independent experiments ±s.e.m. P values (p) were determined using a two-tailed Student's t test. ***P<0.001, **P<0.01, *P<0.05.

Fig 3: Ex vivo pDCs upregulate Type I and III Interferon genes in response to the HCV PAMP.A) Gene expression changes in ex vivo pDCs after HCV PAMP RNA stimulation. Top: subjects with the CC IL28B/IFNλ3 genotype (2 subjects IL28A/IFNλ2, IL28B/IFNλ3, IL29/IFNλ1 and IFNβ1; 1 subject IFNα2; RIG-I SNPs GG and GA); middle: CT IL28B/IFNλ3 genotype (1 subject, RIG-I SNP AA); Bottom: TT IL28B/IFNλ3 genotype (1 subject RIG-I SNP GG). Top graph is combined data from 2 independent experiments while middle and bottom graphs are data from a single independent experiment each. B) Same gene expression data as in A graphed together. Compared to the non-CC genotypes, the CC subjects had significantly more IFN mRNA. p values are the Wilcoxon signed rank result for (A) each gene compared to the X-region stimulation (dashed line) from the same gene, and (B) each gene CC genotype compared to each gene non-CC genotype. * p<0.05 ** p<0.01 *** p<0.001 # p≤0.0001. Bars represent the mean and error bars are +/− SEM.

Fig 4: Patients with acute SARS-CoV-2 infection show a considerable decrease in DC percentages and TLR9-dependent IFN-a production. Bar graphs representing the percentage of total mDCs, CD1c+, CD141+, and CD16+ mDCs (A) and the percentage of pDCs and IFN-a production in response to CpG-A in acute SARS-CoV-2 infected patients (acute) and healthy donors (HD) (B). The median with the interquartile range is shown. Correlation between the percentage of pDCs and IFN-a production in acute patients and HD. Each dot represents an individual (C). *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001. Mann–Whitney U test was used for groups’ comparisons and Spearman test for nonparametric correlations

Fig 5: HA and CB inhibit Influenza-induced production of type I interferons in PBMC and mice.(a–c) mRNA levels of IFN-α (a), IFN-β (b) and IFN-λ2/3 (c) from PBMC pre-incubated with histamine, and CB and stimulated overnight with Influenza, were measured by RT-qPCR and normalized to RPL13A. Data shown as the means of three independent experiments ±s.e.m. (d–f) 29S8 mice were infected with X31 (800 TCID50) and IFN-α (d), IFN-β (e) and IFN-λ2/3 (f) levels in BAL fluid were measured by ELISA (as described in methods). Data shown as the means ±s.e.m. ***P<0.0001, **P<0.001, *P<0.01 as measured by two-way ANOVA with Bonferroni post-tests.