THU0202 INTERFERON STIMULATED GENE 15 PROTECTS REGULATORY T CELL OF SYSTEMIC LUPUS ERYTHEMATOSUS PATIENTS FROM INTERFERON ALPHA-MEDIATED DEPLETION
Authors: Francesca Romana Spinelli, Ilenia Pacella, Silvia Piconese, Fulvia Ceccarelli, Francesca Miranda, Cristiano Alessandri, Vincenzo Barnaba, Guido Valesini, Fabrizio Conti
Journal: Annals of the Rheumatic Diseases (2019)

<h3>Background</h3> Data on T regulatory (T reg) cells number and functions in patients with Systemic Lupus Erythematosus (SLE) are conflicting (1). Type I interferon (IFN) decreases Treg proliferation and induces Treg apoptosis (2). IFN stimulated gene 15 (ISG15) is an IFN-induced protein with a negative effect on IFN pathway. <h3>Objectives</h3> Aim of the study was to evaluate the effect of IFN alpha and ISG15 on Treg number and on STAT1 phosphorilation in patients with SLE. <h3>Methods</h3> We recruited patients with SLE classified according to 1997 ACR criteria. We collected demographic and clinical data including disease duration, disease activity scored according to <i>Systemic Lupus Erythematosus Disease Activity Index</i> (SLEDAI) 2K. Peripheral blood mononuclear cells (PBMC) were isolated from whole blood samples. Intracellular phosphorilated STAT1 (pSTAT) levels were evaluated in Treg (CD4+FOXP3+CD127 low cells) and conventional T cell (Tconv, CD4+FOXP3- cells) by multiparametric flow cytometry, <i>ex vivo</i> or after stimulation with recombinant IFN alpha. PBMC were cultured for 48 hours with anti-CD3 or anti-CD3 and IFN alpha; the frequency of Treg was analysed by multiparametric flow cytometry. ISG15 mRNA expression was evaluated by RT-PCR on PBMC from SLE patients. Data were expressed as mean ± standard deviation or median (interquartile range(according) to the distribution; Mann-Whitney and Spearman tests were applied. P value &lt;0.05 were considered statistically significant. <h3>Results</h3> We enrolled 21 SLE patients [F:M 20:1; mean age 45.2 ± 12.9 years; mean disease duration 157.5± 103 months; median SLEDAI 2(4)] Overall, median baseline ISG15 mRNA expression was 0.06 (IQR 0.16); patients were divided into ISG15-high (n=9) and ISG15-low (n=12), according to the mean mRNA expression. <i>Ex vivo</i>, after short-term treatment with IFN alpha (15 and 25 minutes), we observed a significant increase in STAT1 phosphorilation compared to baseline both in Treg and Tconv (p&lt;0.00001), in ISG15-high and ISG15-low patients (Figure 1 A and B); however, high ISG15 levels protect both Treg and Tconv from STAT1 phosphorilation (Figure 1 C). After 48 of colture with IFN alpha, we observed a decrease in Treg and Tconv number, significantly greater in ISG15-low cells (Figure 1D). Treg frequency and ISG15 expression in Treg were higher in active vs inactive SLE patients [SLEDAI &gt; 4 vs SLEDAI &lt;4] (p&lt;0.05). <h3>Conclusion</h3> ISG15 protects T conventional cells from STAT1 phosphorilation and induces resistance to T regulatory cells depletion. The results of the study suggest that ISG15, a protein induced by IFN in the acute phase of the disease, exerts a negative feedback, allowing the recovery of T reg. <h3>References</h3> [1] Zhang SX, et al. J Immunol res. 2018 [2] Piconese S, et al. Cytokine Growth Factor rev. 2014 <h3>Disclosure of Interests</h3> None declared