Journal: eBioMedicine

Article Title: Monocytes are the main source of STING-mediated IFN-α production

doi: 10.1016/j.ebiom.2022.104047

Figure Lengend Snippet: Investigating the sexual dimorphism in the TLR7-, TLR9- and STING-mediated innate cytokine responses with aging . (a) demographic data of the study cohort, age and sex distribution. (b) Schematic summary of study design: Blood samples were obtained from 310 healthy community-dwelling individuals (ages 18–97), samples were separated in two for whole blood stimulation with the indicated PRR ligands, and the other part was used to isolate PBMCs for FACS analysis as indicated. (c, d, e) Levels of cytokine release after whole blood stimulation with the three studied PRR ligands: R848 (c), ODN M362 (d) or cGAMP (e). Concentrations of indicated cytokines were quantified at 24 h in plasma. The results are given in log 10 of the concentrations (pg/ml) with background (without ligands) subtracted. (c, d, e, f-h) Sex-differences in IFN-α release in response to R848 (f), ODN M362 (g) and cGAMP (h), before and after the age of 60. Statistical differences between groups were analyzed using the unpaired Student's t -test (c-d) or by a one-way ANOVA test followed by Sidak's multiple comparisons test (f–h).

Article Snippet: Stock solutions of pattern recognition receptor (PRR) ligands (Resiquimod (R848) (Cat# tlrl-r848), ODN M362 (a class C CpG ODN, (Cat# tlrl-m362-5) and Cyclic [G(2’,5’)pA(3’,5’)p] cGAMP, (Cat# tlrl-nacga23-1) all purchased from InvivoGen, Toulouse, France) were prepared in RPMI.

Techniques:

Journal: eBioMedicine

Article Title: Monocytes are the main source of STING-mediated IFN-α production

doi: 10.1016/j.ebiom.2022.104047

Figure Lengend Snippet: The female predominance in TLR7-driven production of IFN-α is robustly maintained in elderly subjects above 80 . (a–c) From the source groups depicted in a (age range >80), the production of IFN-α is shown in response to R848 (a), ODN M362 (b) and cGAMP (c) by sex, female (F), male (M). Statistical differences between groups were analyzed using the unpaired Student's t -test .

Article Snippet: Stock solutions of pattern recognition receptor (PRR) ligands (Resiquimod (R848) (Cat# tlrl-r848), ODN M362 (a class C CpG ODN, (Cat# tlrl-m362-5) and Cyclic [G(2’,5’)pA(3’,5’)p] cGAMP, (Cat# tlrl-nacga23-1) all purchased from InvivoGen, Toulouse, France) were prepared in RPMI.

Techniques:

Journal: eBioMedicine

Article Title: Monocytes are the main source of STING-mediated IFN-α production

doi: 10.1016/j.ebiom.2022.104047

Figure Lengend Snippet: Influence of CMV serological status on the decrease of IFN-α release with aging . IFN-α release expressed as in in response to stimulation with R848 is shown according to the CMV status: CMV positive (CMV pos) and CMV negative (CMV neg) individuals (a) or as a function of age (b). The Pearson's R coefficient and correlation probabilities were calculated by means of XLSTAT. Panel (c) shows the IFN-α production in CMV neg and CMV pos individuals by sex. (d) Production of IFN-α in CMV pos elderly subjects above 80. Differences between groups were analyzed using the Student's t-test (a, d) or using a one-way ANOVA test followed by Sidak's multiple comparisons test (c).

Article Snippet: Stock solutions of pattern recognition receptor (PRR) ligands (Resiquimod (R848) (Cat# tlrl-r848), ODN M362 (a class C CpG ODN, (Cat# tlrl-m362-5) and Cyclic [G(2’,5’)pA(3’,5’)p] cGAMP, (Cat# tlrl-nacga23-1) all purchased from InvivoGen, Toulouse, France) were prepared in RPMI.

Techniques:

Journal: eBioMedicine

Article Title: Monocytes are the main source of STING-mediated IFN-α production

doi: 10.1016/j.ebiom.2022.104047

Figure Lengend Snippet: Aging-related changes in pDCs and monocytes differ between sexes . (a) Absolute cell counts of pDC, mDC and monocytes per mm 3 were calculated from FCM data as described in materials and methods. Absolute counts per mm 3 (given in log 10 ) are compared in males and females for individuals under or above 60. (b, c) Correlation between log 10 IFN-α release in response to R848 (b) or ODN M362 (c) stimulation and log 10 absolute numbers per mm 3 of plasmacytoid dendritic cells. Pearson's correlation coefficients “ R” and probabilities of correlation were calculated by means of Statview software. (d, e) Residual values (difference between the observed IFN-α concentration and the predicted value) were calculated from the linear regression of IFN-α response as a function of pDC number shown in (b) and (c), respectively. Differences between groups were analyzed using a one-way ANOVA test followed by Sidak's multiple comparisons test (a, d, e).

Article Snippet: Stock solutions of pattern recognition receptor (PRR) ligands (Resiquimod (R848) (Cat# tlrl-r848), ODN M362 (a class C CpG ODN, (Cat# tlrl-m362-5) and Cyclic [G(2’,5’)pA(3’,5’)p] cGAMP, (Cat# tlrl-nacga23-1) all purchased from InvivoGen, Toulouse, France) were prepared in RPMI.

Techniques: Software, Concentration Assay

Journal: eBioMedicine

Article Title: Monocytes are the main source of STING-mediated IFN-α production

doi: 10.1016/j.ebiom.2022.104047

Figure Lengend Snippet: Monocytes are the main source of IFN-α in response to cGAMP . Impact of aging on monocyte counts (a) and IFN-α response (b) after cGAMP stimulation for males and females. Correlation coefficients “R” and probabilities of correlation were calculated using Pearson test. (a) Correlation between the absolute number of monocytes/mm 3 (log 10 ) and age (years). (b) Correlation between the cGAMP-stimulated release of IFN-α (log 10 ) and the absolute number of monocytes/mm 3 (log 10 ). (c) PBMC from 5 donors were depleted of CD14 + cells (> 98% depletion) using magnetic particles or mock-depleted and stimulated with cGAMP (20 µg/ml) or R848 (1 µg/ml) for 24 h. IFN-α production was measured by ELISA. Paired Student's t -test was used for statistical analysis. (d, e) Total PBMCs (5 × 10 5 cells/well) or monocyte-enriched cells (% CD14 + cells > 77% at 3 × 10 5 cells/well) were stimulated with cGAMP or left untreated (NS). The production of IFN-α (d) or IP-10/CXCL-10 (e) was then assessed in 24 h-supernatants. Median values in pg/ml for [IFN-α] or [IP-10] concentrations normalized to 3 × 10 5 cells/well are shown in parentheses. (f, g) Intracellular analysis of IFN-α synthesis in 18 h-stimulated monocytes, gated as CD11c pos HLA-DR pos cells, from male or female donors. Brefeldin A was added for the last 3 h of culture. Frequencies of IFN-α pos monocytes are shown from individual female or male donors stimulated or not with cGAMP. Statistical differences between groups were assessed using the Kruskal-Wallis's test corrected for multiple comparisons.

Article Snippet: Stock solutions of pattern recognition receptor (PRR) ligands (Resiquimod (R848) (Cat# tlrl-r848), ODN M362 (a class C CpG ODN, (Cat# tlrl-m362-5) and Cyclic [G(2’,5’)pA(3’,5’)p] cGAMP, (Cat# tlrl-nacga23-1) all purchased from InvivoGen, Toulouse, France) were prepared in RPMI.

Techniques: Enzyme-linked Immunosorbent Assay