Monocytes Acquire the Ability to Prime Tissue-Resident T Cells via IL-10-Mediated TGF-β Release - PubMed (original) (raw)

Monocytes Acquire the Ability to Prime Tissue-Resident T Cells via IL-10-Mediated TGF-β Release

Elizabeth A Thompson et al. Cell Rep. 2019.

Abstract

Using non-human primates (NHPs), mice, and human primary cells, we found a role for interleukin-10 (IL-10) in the upregulation of the tissue-resident memory T cell (TRM) marker CD103. In NHPs, intravenous, but not subcutaneous, immunization with peptide antigen and an adjuvant combining an agonistic anti-CD40 antibody plus poly(IC:LC) induced high levels of CD103+ TRMs in the lung, which correlated with early plasma IL-10 levels. Blocking IL-10 reduced CD103 expression on human T cells stimulated in vitro with the adjuvant combination as well as diminished CD103 on lung-resident T cells in vivo in mice. Monocyte-produced IL-10 induced the release of surface-bound transforming growth factor β (TGF-β), which in turn upregulated CD103 on T cells. Early TGF-β imprinted increased sensitivity to TGF-β restimulation, indicating an early commitment of the T cell lineage toward TRMs during the priming stage of activation. IL-10-mediated TGF-β signaling may therefore have a critical role in the generation of TRM following vaccination.

Keywords: IL-10; T cell; TGF-beta; TRM; anti-CD40; monocyte; non-human primate; tissue-resident memory T cells; toll-like receptor; vaccine.

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Conflict of interest statement

DECLARATION OF INTERESTS

R.M.K. is a founder of ImmuRx, a vaccine company for which intellectual property is based on the combined Toll-like receptor (TLR) agonist and anti-CD40 immunization platform. The remaining authors declare no competing interests.

Figures

Figure 1.. Local APC Targeting with Anti-CD40Ab following Subcutaneous Administration

(A) Rhesus macaques were immunized subcutaneously or i.v. with Alexa-Fluor-680-labeled anti-CD40Ab and poly(IC:LC). For s.c. immunization, indicated samples were collected at 24 and 48 h. Shown are representative histograms of the monocyte Alexa Fluor 680 signal from immunized (gray) or unimmunized (black) animals. (B) Proportion of the CD40 Alexa Fluor 680 signal (mean fluorescent intensity, MFI) of APCs in collected tissues. (C–F) CD40 Alexa Fluor 680 MFI on APCs. (G) Monocyte binding following s.c. or i.v. administration. Each data point represents a separate animal (n = 3, mean ± SEM). See also Figure S1.

Figure 2.. Differential TRM Phenotype after i.v. and s.c. Vaccine Administration Correlates with IL-10 Production

Rhesus macaques were immunized with Env peptides, anti-CD40Ab, and poly(IC:LC) (n = 4/group). BAL samples were stimulated overnight with an immunizing peptide pool to recall total memory Ag-specific T cells (CCR7+CD45RA+ naive T cells excluded, producing interferon-γ (IFN-γ), IL-2, or tumor necrosis factor [TNF]). (A–C) Frequency of Ag-specific CD8 or CD4 T cells 3 weeks (A) and 9 months (C) post-immunization. (B) Proportion of multifunctional T cells at 3 weeks. (D) Frequency of CD103+CD69+ TRMs of Ag-specific CD8 T cells. (E) MFI of CD103 expression on Ag-specific TRMs at 3 weeks. (F) IL-10 levels in plasma. (G) IL-10 levels in plasma after i.v. administration of Poly IC:LC, anti-CD40, or the combination (n = 2/group). (H) Correlation of frequency of TRM of Ag-specific CD8 T cells in BAL at 3 weeks (y axis) with IL-10 (pg/mL) in plasma at 6 h (x axis). Each data point represents a separate animal (mean ± SEM). Significance was tested using an unpaired Student’s t test (A–E), one-way ANOVA (F), or Spearman’s test (H). See also Figure S2.

Figure 3.. APC Stimulation Induces TRM Differentiation via IL-10

(A–C) APC-enriched blood, epidermal APCs, or dermal APCs were cultured with CFSE-labeled allogenic naive T cells for 6 days, with or without prestimulation with anti-CD40Ab and poly(I:C) (CD40/PIC) (n = 12). T cells were cultured with anti-CD3-CD28 stimulation beads as a control (n = 6). (A) Representative flow plot of CFSE dilution and CD103 expression. (B) CD8 T cell proliferation as assessed by %CFSE low. (C) Frequency of proliferating CD8 T cells expressing CD103. (D) IL-10 levels in supernatants from overnight stimulated APC cultures (n = 4). (E and F) Co-cultures were supplemented with a neutralizing anti-IL-10 Ab or isotype control (E) or recombinant IL-10 (F). Frequency of proliferating CD8 T cells expressing CD103 is shown. Each data point represents a separate human donor (n = 4–6). (G) Wild-type C57BL6 mice were immunized i.v. with ova, poly(I:C), and anti-CD40, with or without intraperitoneal (i.p.) injection of anti-IL10R blocking Ab 4 h prior to immunization. Lung-resident extravascular T cells were identified by i.v. injection of anti-CD45. Shown are representative flow plots and summary of %CD103+ of CD8 T cells. Each dot represents a separate mouse (n = 4–8, mean ± SEM). Significance was tested using an unpaired Student’s t test (B, C, F, and G) or one-way ANOVA (E). See also Figure S3.

Figure 4.. IL-10 Stimulates the Release of Monocyte TGF-β and TRM Priming

(A and B) TGF-β1 levels in supernatants of stimulated blood APC cultures after 6 h with anti-IL10 (A) or recombinant IL-10 (B) (n = 7). (C and D) Surface expression of TGF-β1, TGF-β2, and TGF-β3 after 6 h in culture unstimulated (C) or stimulated (D) (n = 6). (E) Correlation of secreted TGF-β1 (x axis) and monocyte TGF-β MFI (y axis) (n = 5). (F) Purified CD14+ monocytes were cultured with CFSE-labeled allogenic naive T cells (n = 6). (G) Purified CD14+ monocytes were cultured in a transwell assay with CFSE-labeled naive T cells stimulated with CD3-CD28 beads, or supernatants were transferred from monocytes after overnight stimulation. Frequency of proliferating CD8 T cells expressing CD103 is shown (n = 6). (H–J) Naive CFSE-labeled T cells were stimulated with CD3-CD28 beads (n = 4). (H) Cultures were supplemented with TGF-β1 on day 0, 2, or 5 of the culture and evaluated for CD103 on day 6. (I) Experimental setup. T cells were cultured in the presence or absence of TGF-β1. Cells were washed on day 3 with fresh media and sorted for CD103− cells on day 5. CD103− cells were restimulated with TGF-β1 and evaluated for CD103 expression on day 6. Shown are representative plots of CD103 sorting purity of CD103+ (pink) and CD103− (blue) fractions on day 5 and (J) summary of CD103 expression on day 6 of cultures. Each data point represents a separate human donor (mean ± SEM). (K) Graphical summary of findings. Significance was tested using an unpaired Student’s t test (B, D, F, and G), one-way ANOVA (A, C, H, and J), or Spearman’s test (E). See also Figure S4.

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Monocytes Acquire the Ability to Prime Tissue-Resident T Cells via IL-10-Mediated TGF-β Release - PubMed (original) (raw)