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THU0240 Defective regulation by ATP-GATED ionotropic P2X7 receptor drives T follicular helper cell expansion in systemic lupus erythematosus
  1. R Gualtierotti1,
  2. CE Faliti2,3,
  3. M Gerosa1,4,
  4. F Grassi3,5,
  5. PL Meroni1,4,6
  1. 1Lupus Clinic, Division of Rheumatology, ASST G. Pini
  2. 2Istituto Nazionale Genetica Molecolare “Romeo ed Enrica Invernizzi”, Milan, Italy
  3. 3Institute for Research in Biomedicine, Università della Svizzera Italiana, Bellinzona, Switzerland
  4. 4DISCCO, Department of Clinical Science and Community Health
  5. 5Department of Medical Biotechnology and Translational Medicine (BIOMETRA), University of Milan
  6. 6IRCCS Istituto Auxologico Italiano, Milan, Italy

Abstract

Background Systemic lupus erythematosus (SLE) is a chronic autoimmune disease of unknown aetiology. The deregulated activation of T follicular helper (Tfh) cells in secondary lymphoid organs may play a pivotal role in the activation of B cells and production of autoantibodies. Both murine and human Tfh cells were shown to be sensitive to extracellular ATP via purinergic P2X7 receptor. P2X7 is a non-selective ionic channel that in the presence of high concentrations of ATP or prolonged stimulation opens to a pore permeable to molecules up to 900 Da and causes cell death. Mice deleted for P2rx7 show a significantly worsened outcome of pristane-induced SLE. Expansion of circulating Tfh (cTfh) cells has been correlated with increased levels of autoantibodies and more severe clinical manifestations in SLE.

Objectives Our aim was to investigate the possible role of P2X7 receptor activity in driving cTfh expansion in a cohort of SLE patients. Patients with primary antiphospholipid syndrome (PAPS) and healthy donors (HD) served as normal controls.

Methods Forty-two adult patients with SLE (SLEDAI >4), 14 patients with primary antiphospholipid syndrome (PAPS) and 34 sex- and age-matched healthy donors (HD) were included. Circulating Tfh cells were isolated as a CCR7loPD1+ cells. In 32 patients with SLE, we investigated permeability of Tfh cells to Yo-Pro-1 staining over time at FACS upon stimulation with the P2X7 selective agonist BzATP and the presence of a correlation (Spearman's rho) between Tfh cells expansion and Yo-Pro uptake. We analysed in vitro differentiation of CXCR5+PD1+ Tfh cells and sensitivity of this pathway to BzATP in CD4 naïve cells isolated from 4 SLE and 4 healthy donors in the presence of a mixture of Activin A and IL-12, with or without BzATP.

Results As previously reported, SLE patients had a significant expansion of the CCR7loPD-1+ cTfh cells [SLE (n=42): 38.3±12.8% versus HD (n=34]): 21.6±4.5%, ****p<0.0001]. There was no significant difference in the representation of cTfh cells between PAPS patients ([n=14] 22.6±6.6%) and HD (p=0.6714). The analysis of BzATP induced Yo-Pro-1 permeability revealed a significantly increased resistance to P2X7-mediated cell death in SLE patients as compared to HD and PAPS patients (fold increase of Yo-Pro-1-positive cells at 450 sec, HD [n=28]: 5±2.3%; SLE [n=32]: 3±1.7%, ****p=0.003; PAPS [n=14]: 5±2.9%, **p=0.0042). Furthermore, Spearman's rho showed a significant correlation between the percentage of cTfh cells and the degree of Yo-Pro uptake (r -0.37). In vitro generation of Tfh cells from HD naïve cells revealed a significant response to inhibition by BzATP, which was not present when SLE naïve cells were used.

Conclusions The CCR7loPD-1+ cTfh cells are significantly expanded in SLE but not PAPS patients compared to HD. The degree of expansion correlates with diminished sensitivity to P2X7-mediated cell death. This defective regulation is present also within in vitro differentiation of Tfh from naïve cells isolated from SLE patients. Our data suggest a selective defect of P2X7-mediated control of Tfh cell generation and expansion in SLE.

Disclosure of Interest None declared

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