T-cell immunoglobulin and mucin domain 3 is upregulated in rheumatoid arthritis, but insufficient in controlling inflammation

Abstract

Objectives: Rheumatoid arthritis (RA) is a chronic autoimmune disease, that involves both pro- and anti-inflammatory mechanisms. The purpose of the present study is to investigate T-cell immunoglobulin and mucin domain 3 (Tim-3) in RA.

Methods: Plasma levels of soluble (s) Tim-3 in early RA (n=98), were followed, to evaluate association with treatment and disease activity, acquired from a prospective collected biobank (clinicaltrials.gov (NCT00660647)). We also investigate the influence of Tim-3 on spontaneous cytokine production in synovial fluid mononuclear cells (SFMC) from RA patients after addition of neutralizing anti-Tim-3's antibodies, either alone or in combination with neutralizing anti-Programmed Cell death protein 1 (PD-1) antibodies.

Results: Long-time stimulated CD4 T-cells expressed high levels of Tim-3, but tended to decrease their PD-1 expression. Tim-3 expression was exclusively seen co-expressed with PD-1 by CD3, CD4, CD45RO positive cells in the inflamed RA joint. Addition of neutralizing Tim-3 antibodies increased the secretion of IFNγ and MCP-1, in SFMC cultures from RA. Whereas neutralizing anti-PD-1 antibodies showed a broader impact on cytokine production. Finally, we observed that soluble Tim-3 is increased in plasma and is associated with disease activity in early RA.

Conclusion: Taken together, our findings indicate disease-suppressive functions of Tim-3 in RA.

Keywords: DAS28CRP; Rheumatoid arthritis; Tim-3; anti-TNF-alfa.

Conflict of interest statement

None.

AJCEI Copyright © 2022.

Figures

Figure 1

Increased Tim-3 expression after persistent T cell activation. A. Transient and persistent activation of CD4+ T cells stimulation of T cells through the CD3/CD28 axis, results in sustained Tim-3 expression. B. Heat-map showing the frequency of CD3+CD4+ cells with increased expression of the indicated surface receptors at different time-points. D = days. C. Median fluorescence intensity (MFI) of Tim-3 and PD-1 respectively, in either transiently or persistently activated CD4+ T cell cultures at the indicated time-points. Data are normalized to the MFI at D0 and show mean + SD.

Figure 2

Increased percentage of Tim-3+ and PD-1+ cells among CD3+CD4+CD45RO+ in the synovial fluid (SF) of chronic RA (cRA) patients (n=8) compared with peripheral blood (PB) and HV PB (n=6). The majority of Tim-3+ cells from SF co-express high levels of PD-1. Column graphs show percentage of positive gated cells as median (IQR). *P<0.05, **P<0.01, ***P<0.001.

Figure 3

SFMC were cultured for 48h from chronic RA (cRA) (n=9) with and without added neutralizing antibodies toward TIM3 and PD-1. Supernatants were collected and cytokines were measured by V-plex and ELISA for MCP-1. 0: untreated sample; I: anti-Tim-3 antibody (10 µg/ml); II: anti-PD-1 antibody (10 µg/ml); III: anti-Tim-3 (5 µg/ml) + anti-PD-1 (5 µg/ml) antibodies. Data are shown as median, bars are IQR. All significant results are shown. *P

Figure 4

Increased sTim-3 in synovial fluid (SF) and in plasma from RA patients (A, B). Plasma sTim-3 decreases in early RA (eRA, n=98) after 3 months of treatment (B). Chronic RA (cRA, n=17), healthy volunteers (HV, n=44). *P

Figure 5

Plasma concentrations of soluble (s) Tim-3 was examined in patients with early RA (eRA) at baseline and after 3 months (n=98). These were treated by a treat-to-target strategy for 2 years. Patients were divided into those who showed radiographic progression (+, n=55) or no progression (-, n=43) measured by their change in Total Sharp Score after 2 years of treatment TSS. Patients with progression had a greater reduction in sTim-3 from baseline to 3 months (ΔTim-3) compared with patients that did not progress in TSS. *P