DNAM1 and TIGIT balance the T cell response, with low T cell TIGIT expression corresponding to inflammation in psoriatic disease

Abstract

Objectives: Signals at the contact site of antigen-presenting cells (APCs) and T cells help orchestrate the adaptive immune response. CD155 on APCs can interact with the stimulatory receptor DNAM1 or inhibitory receptor TIGIT on T cells. The CD155/DNAM1/TIGIT axis is under extensive investigation as immunotherapy target in inflammatory diseases including cancer, chronic infection and autoimmune diseases. We investigated a possible role for CD155/DNAM1/TIGIT signaling in psoriatic disease.

Methods: By flow cytometry, we analyzed peripheral blood mononuclear cells of patients with psoriasis (n = 20) or psoriatic arthritis (n = 21), and healthy individuals (n = 7). We measured CD155, TIGIT, and DNAM1 expression on leukocyte subsets and compared activation-induced cytokine production between CD155-positive and CD155-negative APCs. We assessed the effects of TIGIT and DNAM1 blockade on T cell activation, and related the expression of CD155/DNAM1/TIGIT axis molecules to measures of disease activity.

Results: High CD155 expression associates with tumor necrosis factor (TNF) production in myeloid and plasmacytoid dendritic cells (DC). In CD1c+ myeloid DC, activation-induced CD155 expression associates with increased HLA-DR expression. CD8 T cells - but not CD4 T cells - express high levels of TIGIT. DNAM1 blockade decreases T cell pro-inflammatory cytokine production, while TIGIT blockade increased T cell proliferation. Finally, T cell TIGIT expression shows an inverse correlation with inflammation biomarkers in psoriatic disease.

Conclusion: CD155 is increased on pro-inflammatory APCs, while the receptors DNAM1 and TIGIT expressed on T cells balance the inflammatory response by T cells. In psoriatic disease, low TIGIT expression on T cells is associated with systemic inflammation.

Keywords: CD155; DNAM1; TIGIT; inflammation; psoriatic disease.

Graphical Abstract

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Figure 1.

CD155 surface expression by APCs. Pooled flow cytometry analysis of PBMCs ex vivo, of healthy controls (n = 7, symbol with cross), psoriasis (n = 7, open symbol), and psoriatic arthritis (n = 7, filled symbol) patients. Shown data are from six APC subsets: CD14+CD16- classical monocyte (cM), CD14+CD16+ intermediate monocyte (iM), CD14-CD16+ non-classical monocyte (ncM), CD141+ myeloid DC (mDC), CD1c+ mDC, and CD303+ plasmacytoid DC (pDC). (A) Proportion of APC subset in PBMCs (gate ‘Single Cells’ Supplementary Fig. S1A). (B) Proportion of CD155-positive cells in APC subset. (C–H) Gating strategy of the selection of CD155-positive cells in cM (C), iM (D), ncM (E), CD141+ mDC (F), CD1c+ mDC (G), and pDC (H).

Figure 2.

High TNF production by CD155 expressing APCs. Flow cytometry analysis of psoriatic disease patients PBMCs stimulated for 4 hours with LPS (100 ng/ml) in the presence of Brefeldin A (1:1000). (A) Proportion of TNF producing cells within CD155-positive and CD155-negative APC subsets of psoriasis patients (n = 13, open symbol) and psoriatic arthritis patients (n = 15, filled symbol): CD14+CD16- classical monocyte (cM), CD1c+ myeloid dendritic cell (mDC), and CD123+ plasmacytoid dendritic cell (pDC) (detailed gating strategy shown in Supplementary Fig. S1B). (B–D) Representative flow plots of TNF production by CD1c+ mDC: CD155-positive TNF FMO control (B), CD155-negative CD1c+ mDC (C), and CD155-positive CD1c+ mDC (D). *Significant P-value MWU. NA: not applicable.

Figure 3.

Positive correlation of CD155 and HLA-DR in activated CD1c+ mDC. Flow cytometry analysis of psoriatic disease patients PBMCs stimulated for 4 hours with LPS (100 ng/ml). Shown is the positive correlation of the percentage of CD155-positive CD1c+ mDC and the MFI of HLA-DR expressed by CD1c+ mDC (Spearman’s rank correlation coefficient r_s = 0.664 [95% CI 0.433–0.813], P = < 0.0001).

Figure 4.

Higher TIGIT and comparable DNAM1 expression by CD8 versus CD4 T cells. Flow cytometry analysis of CD4 (square) and CD8 (circle) T cells ex vivo of healthy controls (HC, n = 7, symbol with cross), psoriasis (PsO, n = 7, blank symbol), and psoriatic arthritis (PsA, n = 7, filled symbol) patients. (A–D) Pooled data of all subjects. (E–F) Data of HC, PsO, and PsA patients shown separately. (A) Significantly higher proportion of TIGIT-positive CD8 T cells compared to CD4 T cells. (B) Comparable proportion of DNAM1-positive CD8 and CD4 T cells. (C) Significantly higher TIGIT MFI of CD8 T cell compared to CD4 T cells. (D) Comparable DNAM1 MFI of CD4 and CD8 T cells. (E) Comparable proportion of TIGIT-positive CD4 T cells in HC, PsO, and PsA. (F) Comparable proportion of DNAM1-positive CD4 T cells in HC, PsO and PsA. (G) Comparable proportion of TIGIT-positive CD8 T cells in HC, PsO, and PsA. (G) Comparable proportion of DNAM1-positive CD8 T cells in HC, PsO, and PsA. *Significant P-value MWU.

Figure 5.

TIGIT blockade increases T cell proliferation, and DNAM1 blockade reduces T cell pro-inflammatory cytokine production. Flow cytometry analysis of PBMCs stimulated for 3 days with CD3/CD28 Dynabeads (PBMC:Dynabead 10:1), after either 10 µg/ml DNAM1 blocking antibody, 10 µg/ml TIGIT blocking antibody or 10 µg/ml DNAM1 and TIGIT blocking antibody isotypes. Pooled data of healthy controls, psoriasis, and psoriatic arthritis patients. Shown are percentages of proliferated T cells, stained with 2 µM CellTrace Violet reagent (A, B, G, and H) and percentages of TNF and IFNγ producing T cells after 4 hours re-stimulation with 50 ng/ml PMA, 1 µg/ml ionomycin in the presence of Brefeldin A (1:1000) (C–F, H–K). (A) TIGIT block significantly increases CD4 T cell proliferation. (B) DNAM1 block has no significant effect on CD4 T cell proliferation. (C) No significant difference in CD4 T cell TNF production after TIGIT blockade (44.5% vs. 45.4%, P > 0.05). (D) No significant decrease in TNF production by CD4 T cells after DNAM1 block (44.5% vs. 39.7%, P > 0.05). (E) No significant increase in CD4 T cell IFNγ production after TIGIT blockade (15.6% vs. 17.4%, P > 0.05). (F) Significantly decreased IFNγ production by CD4 T cells after DNAM1 block (15.6% vs. 13.0%, P = 0.0015). (G) TIGIT block significantly increases CD8 T cell proliferation. (H) DNAM1 block has no significant effect on CD8 T cell proliferation. (I) No significant difference in CD8 T cell TNF production after TIGIT blockade (35.8% vs. 36.1%, P > 0.05). (J) Significant decrease in TNF production by CD8 T cells after DNAM1 block (36.1% vs. 29.6%, P = 0.0039). (K) Trend toward increased CD8 T cell IFNγ production after TIGIT blockade (29.4% vs. 32.0%, P > 0.05). (L) Significant decrease in IFNγ production by CD8 T cells after DNAM1 block (29.4% vs. 25.9%, P = 0.0140). *Significant P-value Wilcoxon-signed rank test.

Figure 6.

Reduced T cell TIGIT expression associates with increased level of acute phase reactants. Correlation of acute phase reactants in serum and the percentage of TIGIT-expressing T cells analyzed ex vivo in psoriasis (n = 7, blank symbol) and psoriatic arthritis (n = 7, filled symbol) patients, using flow cytometry. (A) Significant correlation of percentage TIGIT-positive CD4 T cells and ESR (r_s = −0.7918 [95% CI −0.9457 to −0.3476], P = 0.0052). (B) Trend toward correlation of CD4 TIGIT-positive cells and CRP (r_s = −0.792 [95% CI −0.8701 to 0.01175], P = 0.0521). (C) Significant correlation of percentage TIGIT-positive CD8 T cells and erythrocyte sedimentation rate (ESR) (Spearman’s Rank correlation coefficient [r_s] = −0.705 [95% CI −0.920 to −0.162], P = 0.0189). (D) Significant correlation of CD8 TIGIT-positive CD8 T cells and C-reactive protein (CRP) (r_s = −0.663 [95% CI −0.900 to −0.125], P = 0.022).