Background Increased interleukin-6 (IL-6) was found in SSc skin, and early studies suggest therapeutic efficacy of IL-6 receptor blockade in SSc. Similarly, an increased interferon (IFN) signature was found in SSc patients. Accordingly, both IL-6 and interferons may be upregulated in SSc, and both act via Stat (signal transducer and activator of transcription) molecules, namely Stat3, and Stat1, respectively.
Objectives To analyze the interleukin-6 (IL-6) receptor-Stat3 axis, as well as Stat1 and phosphorylated Stat1, in lymphocytes of patients with SSc.
Methods PBMC of 30 patients with SSc and 42 healthy individuals (HC) were prepared. CRP, ESR, modified Rodnan skin score (mRSS) and the EULAR activity score were recorded. For determining the percentages of CD126 and CD130 positive cells, PBMC were directly stained with PE-labelled or control antibodies. Ex vivo and after 15 minutes of incubation with or without recombinant human interferon-γ (IFNγ) or interleukin-6 (IL-6), PBMC were fixed with formaldehyde (2%), permeabilized with methanol (80%), and stained with PE-labelled control antibodies or antibodies to Stat1 or phosphorylated Stat1 (pStat1) or pStat3, respectively. Stained cells were immediately analyzed on a Becton Dickinson FACSCalibur fluorocytometer, gating for lymphocytes. As a semiquantitative measure of Stat and pStat contents, mean fluorescence intensity (mfi) was used. Data were expressed as mean±SD.
Results Both chains of the IL-6 receptor, CD126 and CD130, were expressed on decreased percentages of SSc lymphocytes (50.2±18.1% vs. 60.0±7.9%, p=0.0086, and 52.6±15.1% vs. 59.6±9.5%, p=0.0212, respectively). This translated into a diminished increase of pStat3 in SSc lymphocytes upon IL-6 stimulation (Δmfi 17.2±12.1 vs. 20.5±9.3, p=0.03). In contrast, the IFNγ-induced increase in pStat1was more pronounced in SSc lymphocytes (Δmfi 10.5±11.2 vs. 4.8±3.7, p=0.0035), despite their increased Stat1 contents (mfi 51.2±27.4 vs. 16.7±9.1, p<0.0001), thus suggesting activity of the IFN system in SSc. Somewhat unexpectedly, no differences were found between diffuse cutaneous (or anti-topoisomerase positive) and limited cutaneous (or anti-centromere-positive) SSc.
Conclusions In combination of increased Stat1 and Stat1 phosphorylation and decreased IL-6 receptors, our results suggest activity of both the IFN/Stat1 and the IL-6/Stat3 axis in SSc, with a shift towards phosphorylated Stat1.
Acknowledgement This work was supported by DFG grant AR-757/1–1.
Disclosure of Interest C. Kittel: None declared, M. Skwarek: None declared, B. Heschel: None declared, J. Fantana: None declared, M. Aringer Consultant for: Chugai, Pfizer, Roche, Sanofi, Speakers bureau: Chugai, Pfizer, Roche