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Novel mRNA and vector-based covid-19 vaccinations have shown high efficacy in preventing symptomatic COVID-19 infections. Compared with the number of performed vaccinations, rates of severe side effects seem low. Rare prothrombotic coagulation disorders with suspected association to ChAdOx1 nCoV-19 (AstraZeneca) have been reported. These cases have gathered considerable media attention and caused a temporary pause of usage of the AstraZeneca vaccine in Europe and several other countries and are currently discussed as vaccine-induced immune thrombotic thrombocytopenia (VITT). However, hemorrhagic complications from ChAdOx1 nCoV-19 vaccination have also been reported but, so far, received less public attention despite considerable potential for life-threatening complications. Here we present a case of severe immune thrombocytopenia after ChAdOx1 covid-19 vaccination and its successful primary management.

Milk fat globule-epidermal growth factor 8 (MFG-E8) is an anti-inflammatory glycoprotein that mediates the clearance of apoptotic cells and is implicated in the pathogenesis of autoimmune and inflammatory diseases. Because MFG-E8 also controls bone metabolism, we investigated its role in rheumatoid arthritis (RA), focusing on inflammation and joint destruction. The regulation of MFG-E8 by inflammation was assessed in vitro using osteoblasts, in arthritic mice and in patients with RA. K/BxN serum transfer arthritis (STA) was applied to MFG-E8 knock-out mice to assess its role in the pathogenesis of arthritis. Stimulation of osteoblasts with lipopolysaccharide (LPS) and tumor necrosis factor (TNF)-α downregulated the expression of MFG-E8 by 30% to 35%. MFG-E8-deficient osteoblasts responded to LPS with a stronger production of pro-inflammatory cytokines. In vivo, MFG-E8 mRNA levels were 52% lower in the paws of collagen-induced arthritic (CIA) mice and 24% to 42% lower in the serum of arthritic mice using two different arthritis models (CIA and STA). Similarly, patients with RA (n = 93) had lower serum concentrations of MFG-E8 (-17%) compared with healthy controls (n = 140). In a subgroup of patients who had a moderate to high disease activity (n = 21), serum concentrations of MFG-E8 rose after complete or partial remission had been achieved (+67%). Finally, MFG-E8-deficient mice subjected to STA exhibited a stronger disease burden, an increased number of neutrophils in the joints, and a more extensive local and systemic bone loss. This was accompanied by an increased activation of osteoclasts and a suppression of osteoblast function in MFG-E8-deficient mice. Thus, MFG-E8 is a protective factor in the pathogenesis of RA and subsequent bone loss. Whether MFG-E8 qualifies as a novel biomarker or therapeutic target for the treatment of RA is worth addressing in further studies.

Milk fat globule-epidermal growth factor 8 (MFG-E8) is an anti-inflammatory glycoprotein that mediates the clearance of apoptotic cells and is implicated in the pathogenesis of autoimmune and inflammatory diseases. Because MFG-E8 also controls bone metabolism, we investigated its role in rheumatoid arthritis (RA), focusing on inflammation and joint destruction. The regulation of MFG-E8 by inflammation was assessed in vitro using osteoblasts, in arthritic mice and in patients with RA. K/BxN serum transfer arthritis (STA) was applied to MFG-E8 knock-out mice to assess its role in the pathogenesis of arthritis. Stimulation of osteoblasts with lipopolysaccharide (LPS) and tumor necrosis factor (TNF)-α downregulated the expression of MFG-E8 by 30% to 35%. MFG-E8-deficient osteoblasts responded to LPS with a stronger production of pro-inflammatory cytokines. In vivo, MFG-E8 mRNA levels were 52% lower in the paws of collagen-induced arthritic (CIA) mice and 24% to 42% lower in the serum of arthritic mice using two different arthritis models (CIA and STA). Similarly, patients with RA (n = 93) had lower serum concentrations of MFG-E8 (–17%) compared with healthy controls (n = 140). In a subgroup of patients who had a moderate to high disease activity (n = 21), serum concentrations of MFG-E8 rose after complete or partial remission had been achieved (+67%). Finally, MFG-E8-deficient mice subjected to STA exhibited a stronger disease burden, an increased number of neutrophils in the joints, and a more extensive local and systemic bone loss. This was accompanied by an increased activation of osteoclasts and a suppression of osteoblast function in MFG-E8-deficient mice. Thus, MFG-E8 is a protective factor in the pathogenesis of RA and subsequent bone loss. Whether MFG-E8 qualifies as a novel biomarker or therapeutic target for the treatment of RA is worth addressing in further studies.

The molecular mechanisms that lead to autoantibody production and autoimmune disease remain poorly defined. Based on a clinical observation in a patient with lupus erythematosus and autoimmune hepatitis we here provide the first evidence that (i) a particular cytogenetic aberration (deletion (13)(q14)) could be detected in a proportion of plasma cells and that (ii) the surface marker CD56 could be used to enrich for plasma cells with this cytogenetic aberration.

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 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

Background CRP levels are typically low in active SLE, but not in SLE patients with infections. However, IL-6, which directly leads to CRP increases, has been found increased in active SLE. Membrane-bound IL-6 receptor-α chains (CD126) are exclusively expressed on leukocytes, hepatocytes, and bowel epithelium. In contrast, gp130 (CD130), the second IL-6 receptor chain, is expressed on all cell types. IL-6 bound to soluble IL-6 receptors (sIL-6R), which are mainly derived by shedding CD126, can bind to CD130 on any cell and transmit signals, which is called trans-signalling. Objectives To analyze IL-6 and IL-6 receptor expression and function in SLE. Methods We prepared PBMC of 41 SLE patients and 71 healthy individuals (HC). CRP and disease activity (by ECLAM) were recorded. For determining the percentages of CD126 and CD130 positive cells, PBMC were directly stained with PE-labelled or control antibodies. For analyzing IL-6 induced Stat3 phosphorylation, PBMC were stimulated with rhIL-6 (250 ng/ml) for 15 min, fixed with formaldehyde (2%), permeabilized with methanol (80%), and stained with PE-labelled antibodies to phosphorylated Stat3 (pStat3) or control antibodies. For in vitro experiments on the influence on receptor expression, healthy PBMC were incubated for 24 hours with or without the addition of IL-6, IL-10, tumor necrosis factor (TNF), interferon-α (IFNα), or combinations of these cytokines. Stained cells were immediately analyzed on a Becton Dickinson FACSCalibur fluorocytometer, gating for lymphocytes. As a semiquantitative measure of pStat3 contents mean fluorescence intensity (mfi) was used. Serum IL-6 and sIL-6R were measured by ELISA. Results The serum IL-6 levels of SLE patients were significantly higher than those of HC (median (range) (3.6 (0.69-69.3) vs. 0.9 (0.12-10.5) pg/ml, p Conclusions In SLE, CD126 is reduced and sIL-6R increased, so that the increased IL-6 will more likely bind to cells only carrying CD130, but not CD126. Only in very high concentrations of IL-6, such as in infections or active serositis, will there be enough IL-6 signalling in cells carrying (reduced) CD126, and thus an increase in CRP. Combinations of IFNα (or TNF) with IL-6, all of which are increased in active SLE, explain this shift from classical IL-6 signalling via CD126 to trans-signalling via sIL-6R. Disclosure of Interest None declared

Background IL-6 signals via a receptor consisting of gp130 (CD130) and either membrane-bound IL-6 receptor α (CD126) present on leukocytes and hepatocytes or soluble (sIL-6Rα) on all other cells, the latter process being called trans-signalling. Objectives To investigate the IL-6/IL-6 receptor system in SLE and compare it to healthy individuals (HC) and RA patients. Methods We compared lymphocytes of 33 SLE, 16 RF and ACPA positive (RA+) and 12 seronegative (RA-) RA patients, and 50 HC. PBMC were stained with PE-labelled or control antibodies for CD126 and CD130. For determining Stat3 phosphorylation, PBMC were stimulated with IL-6 (250 ng/ml) for 10 min and 15 min, fixed with formaldehyde, permeabilized with methanol, and stained with PE-labelled anti-pStat3 or control antibodies. For in vitro experiments, HC PBMC were incubated for 24 hours in complete RPMI with or without the addition of IL-6, IL-10, TNF, interferon-α (IFNα) or combinations of these cytokines. CD126 staining was performed as above. Cells were analyzed on a Becton Dickinson FACSCalibur fluorocytometer, gating for lymphocytes. Increase in pStat3 mean fluorescence intensity (mfi) was used as an estimate of IL-6 signalling. Serum levels of IL-6 and soluble IL-6 receptor (sIL-6R) were measured by ELISA. Results As compared to IL-6 serum levels of HC (0.5 (0.5-12.8) median (range) pg/ml), serum IL-6 was increased in SLE (3.7 (0.97-69.3) pg/ml, p Conclusions IL-6 is increased in SLE, albeit less than in RA+. Cytokine combinations released in response to immune complexes induced a decrease in IL-6 receptors in vitro, which was similar to the in vivo situation. The concomitant increase in sIL-6R in SLE and RA+ sera points to receptor shedding. Reduction in surface receptors appeared to reduce IL-6 induced Stat3 phosphorylation. Thus, shedding may decrease leukocyte (and possibly hepatocyte) effects of IL-6, while increasing effects on other cells that rely on trans-signalling. Disclosure of Interest M. Skwarek: None declared, J. Fantana: None declared, B. Heschel: None declared, M. Aringer Consultant for: Roche, Speakers bureau: Roche DOI 10.1136/annrheumdis-2014-eular.4445

Background Despite common pathogenetic events in the effector phase, increasing evidence suggests that RA+ is different from RF and anti-CCP-negative RA-. In particular, genetic evidence and somewhat differential responses to therapy distinguish these subgroups. The IL-6 receptor, consisting of the IL-6 receptor α-chain (CD126) and of glycoprotein 130 (CD130), which is also part of other receptors, can be differentially regulated in inflammatory conditions, as can the homotrimeric TNF receptorsI (CD120a) and II (CD120b). Objectives To profile RA+ and RA- patients with regard to IL-6 and TNF receptors Methods Peripheral blood mononuclear cells (PBMC) of RA+, RA- patients, and healthy individuals (HC) were compared. Most (91%) of the RA+ patients were also positive for anti-CCP antibodies. PBMC were immediately prepared from peripheral venous blood. Serum levels of IL-6 and soluble IL-6R (sIL-6R) were measured by ELISA. For determining the percentages of CD126, CD130, CD120a, and CD120b, positive cells, PBMC were stained with phycoerythrin (PE)-labelled specific or control antibodies. Cells were analyzed on a Becton Dickinson FACSCalibur fluorocytometer, gating for lymphocytes. Results Disease activity (median (range) CDAI 15.5 (0.3-54.5) and 13.5 (4.4-33.4)) and disease duration (median 6 (0.04-66) vs. 3 (0.04-10) years) were not significantly different between RA+ and RA- patients, respectively. However, lymphocytes of RA+ and RA- patients differed in their lymphocyte expression of the IL-6 receptor and of CD120b+. The percentage of CD126+ lymphocytes in RA+ (48±13 (mean±SD) %) was decreased in comparison with RA- (61±12%, p=0.006) and HC (60±9%, p Conclusions Our findings show that sIL-6R was increased and membrane CD126 decreased, suggesting increased IL-6 and more shedding of IL-6 receptors in RA+ patients, but not RA- patients. Interestingly, while RA+ lymphocytes more commonly expressed membrane CD120b, CD120a was increased in the case of RA- lymphocytes. These differences between RA+ and RA- patients may have implications for the role of therapies targeting TNF and IL-6 (receptors), respectively. Disclosure of Interest None Declared

Background and Objectives Despite significant clinical overlap, genetic evidence and, in part, response to therapy distinguish seropositive (RA+), i.e. rheumatoid factor (RF) positive RA patients from patients with RF and anti-CCP-negative (RA-) disease. The aim of this study was to profile RA+ and RA- patients with regard to the differential expression of receptors for IL-6 and TNF, cytokines of established importance in RA. Materials and Methods PBMC of RA+ and RA- patients were compared to each other and to healthy individuals (HC). Most (93%) of the RA+ patients were also positive for anti-CCP antibodies. PBMC were immediately prepared from peripheral venous blood. For determining the percentage of IL-6Rα (CD126), gp130 (CD130), TNFR I (CD120a), and TNFR II (CD120b) positive cells, PBMC were stained with PE-labelled or control antibodies. Cells were analysed on a Becton Dickinson FACSCalibur fluorocytometer, gating for lymphocytes. Results Disease duration (median 7 (0.04–66) versus 3.5 (0.04–6) years) and disease activity (CDAI median 15.6 (5.3–54.5) versus 13.5 (4.4–28)) were comparable between RA+ and RA- patients. Lymphocytes of RA+ and RA- patients differed in their lymphocyte expression of CD126+ and CD120b+. The percentage of CD126+ lymphocytes in RA+ was decreased in comparison with RA- (mean ± SD, 49 ± 14 versus RA-58 ± 11, p = 0.05) and HC (59 ± 9%, p = 0.0007). The difference between RA- and HC was not significant. The percentage of CD130+ lymphocytes in RA+ (51 ± 11) was decreased when compared with HC (58 ± 11%, p = 0.007). While the mean values for CD130 were similar between RA+ and RA- (55 ± 14%), the latter values were not significantly different from HC. In contrast to the IL-6 receptor, CD120b+ lymphocytes were increased in RA+ patients (69 ± 12% versus RA- 58 ± 12%, p = 0.04, versus HC 57 ± 11, p = 0.0002). Again, the difference between RA- and HC was not significant. The percentages of CD120a+ lymphocytes were low in all groups. Nevertheless, mean percentages of CD120a+ lymphocytes from RA- (1.5 (0.53–2) % versus HC (median 1 (0.4–4)% p = 0.05) were somewhat higher than those of RA+ (median 1.1 (0.3–3)%, p = 0.9). Conclusions A direct comparison of (IL-6 mediated) downregulation of CD126 and (TNF mediated) upregulation of CD120b suggests that both are clearly more pronounced in RA+ than in RA-0. This was not explained by differences in disease activity.