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Systemic lupus erythematosus (SLE) is characterized by an imbalance between proinflammatory and anti-inflammatory mediators. Single-nucleotide polymorphisms (SNPs) in genes coding IL10RA, IL10RB, and IL22RA could affect their expression or function and disrupt immune homeostasis. We aimed to analyze the associations of IL10RA, IL10RB, and IL22RA polymorphisms/haplotypes with patients’ susceptibility to and clinical manifestations of SLE. Our study included 103 SLE patients and 99 healthy controls. The genotypes of the selected polymorphisms within IL10RA (rs10892202, rs4252270, rs3135932, rs2228055, rs2229113, and rs9610), IL10RB (rs999788, rs2834167, and rs1058867), and IL22RA (rs3795299 and rs16829204) genes were determined by TaqMan® Assays. IL10RB rs1058867 G allele carriers were significantly more frequent among the controls than among the SLE patients (76.8% vs. 61.2%; p = 0.017, OR = 0.477, 95% CI: 0.258–0.879). The IL10RB CAA haplotype was more frequent among the SLE patients than in the control group (42.7% vs. 30.7%; p = 0.027). The IL22RA rs3795299 C allele and rs16829204 CC genotype were associated with Hashimoto thyroiditis in the SLE patients (n = 103; p = 0.002 and p = 0.026, respectively), and in all the included participants (n = 202, p < 0.000 and p = 0.007, respectively), and the IL22RA CC haplotype was more frequent in the SLE patients with Hashimoto thyroiditis (p = 0.047) and in the overall participants with Hashimoto thyroiditis (n = 32, p = 0.004). The IL10RA, IL10RB, and IL22RA polymorphisms/haplotypes could be associated with SLE susceptibility and various clinical manifestations, and the IL22RA CC haplotype could be associated with Hashimoto thyroiditis.

Although Epstein–Barr virus (EBV) reactivation has long been associated with the pathogenesis of systemic lupus erythematosus (SLE), many aspects of this relationship remain unclear. Our objective was to investigate the association between EBV reactivation and the achievement of SLE remission and lupus low disease activity state (LLDAS) over a six-month period. Clinical, laboratory, and virological tests (anti-EBV antibodies and EBV DNA) were performed among 51 patients with the active form of SLE on two occasions six months apart. SLE remission and LLDAS achievement were assessed at the end of the follow-up period. Active EBV infection was detected in 45% of active SLE patients at baseline, and 77% transitioned to latent EBV infection at six months (p < 0.001). Multivariate regression revealed a higher titer of anti-EA(D) IgM-Abs and the presence of anti-EA(D) IgM-Abs as independent predictors of remission and LLDAS in SLE patients with mucocutaneous manifestations (p = 0.042) and rash only (p = 0.023), respectively. Since a higher C3 level was an independent predictor of transition to latent EBV infection (p = 0.027), the estimated cut-off value that could identify active SLE patients who will transition to latent EBV infection after six months was ≥0.780 g/L with a sensitivity of 70.6% and a specificity of 75.0% (AUC = 0.756, p = 0.003). EBV reactivation is common in patients with active SLE, and most of them transition to latent EBV infection after six months. Achieving remission and LLDAS in SLE patients with mucocutaneous manifestations can be predicted by a higher titer, whereas in SLE patients who have only a rash, the presence of anti-EA (D) IgM-Abs was a predictor of remission and LLDAS.

Development of lymphoproliferative disorders (LPDs) is one of the well-known life-threatening complications in rheumatoid arthritis (RA) patients. However, there is a lack of definitive conclusions regarding the role of Epstein-Barr virus (EBV) activity in RA initiation and progression, especially in promoting LPDs. A systematic review and meta-analysis of studies that reported an EBV positive result in RA-LPD patients and controls were conducted. Studies published before 27 July 2021 were identified through PubMed, Web of Science, and SCOPUS. A total of 79 articles were included in the systematic review. The prevalence of EBV positive result among RA-LPD patients was 54% (OR = 1.54, 95% CI = 1.45–1.64). There was a statistically significant association between EBV presence and LPD susceptibility in RA patients in comparison with all controls (OR = 1.88, 95% CI = 1.29–2.73) and in comparison with LPD patients only (OR = 1.92, 95% CI = 1.15–3.19). This association was not shown in comparison with patients with autoimmune diseases other than RA who developed LPD (OR = 0.79, 95% CI = 0.30–2.09). This meta-analysis confirmed a high prevalence of EBV in the RA-LPD population. Furthermore, it provides evidence for the association between EBV presence and LPD susceptibility in RA patients, but not in those with other autoimmune diseases who developed LPD.

OBJECTIVE Cross-cultural validation of the Serbian version of the Modified Falls Efficacy Scale (MFES). METHODS This cross-sectional study involved 257 women aged 65 years and above who were referred for dual-energy x-ray absorptiometry examination at the Railway Healthcare Institute in Belgrade, Serbia, between January and April 2016. Data collection comprised of a sociodemographic questionnaire and Geriatric Depression Scale-Short Form (GDS-SF) questionnaire, and data related to fractures, level of physical activity, use of medications that can increase the risk of falls, and frequency of falls in the past 12 months. None of the study participants had been previously treated for osteoporosis. The internal consistency of the questionnaire items was assessed via Cronbach's alpha, whereas the interclass correlation coefficient (ICC) was used to calculate test-retest reliability based on the sample of 257 women. We also evaluated concurrent, convergent, and construct validity. RESULTS Cronbach's alpha for the total assay score was 0.98. Correlations among the items ranged from 0.84 to 0.93. While ICC for the scale as a whole was 0.99 (95% confidence interval 0.98-0.99), ICC pertaining to individual items ranged from 0.82 to 0.99. Concurrent validity analysis revealed a significant positive correlation between MFES scores and the reported level of physical activity (ρ = 0.34; P

Neutrophil extracellular traps (NETs) are the main source of autoantigens in systemic lupus erythematosus (SLE). The aim of this study was to evaluate the clinical importance of NETs-associated markers in SLE. We compared NETs-associated markers in SLE patients (n = 111) with healthy controls (n = 50). Moreover, in 35 patients with drug-naive SLE (n = 35), we investigated correlation between NETs-associated markers [DNase I concentration, myeloperoxidase (MPO) activity, anti-MPO antibodies, cell-free DNA (cfDNA), NETolytic activity] with serological parameters [anti-dsDNA antibodies, C3, C4 and B-cell activating factor (BAFF) levels] and disease activity measured by modified SLE Disease Activity Index (M-SLEDAI-2K). In comparison with healthy controls, SLE patients had higher cfDNA, MPO activity, anti-MPO antibodies (p

Background: The most reliable markers reflecting endothelial activation and injury in Systemic sclerosis (SSc), are intercellular adhesion molecule (ICAM1), vascular cell adhesion molecule (VCAM1), E selectin (Es) and P selectin (Ps) (1). Objectives: To assess concentrations of these vascular biomarkers in SSc patients (pts) in comparison to healthy controls (HC) and in relation to disease manifestations. Methods: Patients who fulfilled the 2013 aCR/EULAR SSc classification criteria and have never been treated with endothelin receptor antagonist, phosphodiesterase 5 inhibitors or prostanoids were eligibile. Exclusion criteria were overlap syndromes, other autoimmune and cardiovascular diseases, diabetes mellitus, thrombosis, pregnancy, active infection or neoplastic diseases. Our study included 53 SSc pts [34 limited (lcSSc) and 19 diffuse cutaneous SSc (dcSSc)] and 31 age- and sex matched HC. Serum concentration of ICAM1, VCAM1, Es and Ps were measured using a commercial ELISA kit (Quantikine; R&D Systems), expressed as ng/mL. Clinical evaluation of patients was obtained, including nailfold videocapillaroscopy (NVC). Disease activity was assessed by the revised EUSTAR activity index. Statistical analysis was done in R. Student’s t-test or aNOVA, otherwise Mann-Whitney U or Kruskal-Wallis tests were used. Pearson’s or Spearman’s correlation were done depends on nature of data. ICAM1 cut off value were assessed with ROC analysis. Association were performed with univariate logistic regression for NVC alterations and multivariate for anti-TopoI/Scl70 antibodies (aTSa). Results: Concentration of all markers were elevated in SSc pts compared to HC (ICAM1 p 0.05). Different disease subsets exhibited higher values of ICAM1 and VCAM1 respect to HC (lSSc p 3 years group (r 0.4, p 14 (p 0.05). Few megacapilaries were associated with ICAM1 (OR 1.2, 95% CI 1.06–1.34, p Conclusion: Our results support the role of vascular biomarkers in SSc pathogenesis. Besides anti-TopoI/Scl70 antibodies, ICAM-1 could be used as an additional marker of dSSc. Elevation of ICAM1 and Ps concentration might be associated with late NVC alterations. References [1] Mostmans Y, Cutolo M, Giddelo C, et al. The role of endothelial cells in the vasculopathy of systemic sclerosis: a systematic review. Autoimmun Rev2017;16:774–86. doi:10.1016/j.autrev.2017.05.024 Disclosure of interests: Jelena Colic: None declared, aleksandra antovic: None declared, Quan Tang: None declared, Maja Zlatanovic: None declared, Slavica Pavlov-Dolijanovic: None declared, Ivica Jeremic: None declared, aleksandra Kadic: None declared, Karin Gunnarsson: None declared, annika Nordin: None declared, Jelena Vojinovic: None declared, Mirjana Sefik Bukilica: None declared, Nemanja Damjanov Grant/research support from: abbVie, Pfizer and Roche, Consultant for: abbvie, Gedeon Richter, Merck, Novartis, Pfizer and Roche., Speakers bureau: abbvie, Gedeon Richter, Merck, Novartis, Pfizer and Roche.

Menopause is characterized by deep metabolic disturbances, including decreased insulin sensitivity, adiposity, and changes in lipid profiles. Estrogen replacement therapy can partially reverse these changes, and while it is safe in most healthy postmenopausal women, there are still existing concerns regarding an increased risk for breast and endometrial cancer as well as a risk for cardiovascular and thromboembolic disease. Therefore, certain natural compounds with positive metabolic effects may be considered as a possible alternative or adjunctive treatment in patients not willing to take estrogens or patients with contraindications for estrogens. The aim of this study was to investigate the influence of Sideritis scardica (mountain tea) extract on metabolic disturbances induced by ovariectomy in rats. The study included 24 rats divided into three groups: ovariectomized rats treated with 200 mg/kg S. scardica extract for 24 weeks (n = 8), ovariectomized non-treated (n = 8), and Sham-operated (n = 8) rats. Food intake, weight gain, body composition, fasting glucose levels, response to oral glucose challenge, liver glycogen content, catalase activity, thiol groups, and malondialdehyde concentrations as well as AMP-activated protein kinase activity in liver cells were studied. Ovariectomized rats treated with S. scardica extract had lower blood triglycerides, reduced fasting glucose levels, as well lower glucose peaks after oral glucose challenge, increased liver glycogen content, and significantly higher catalase activity and thiol group concentration than non-treated ovariectomized rats. The ability of S. scardica extract to attenuate metabolic disturbances associated with ovariectomy was associated with the activation of AMP-activated protein kinase in liver cells.

Old man’s beard (Usnea barbata) is one of the most investigated lichens which biological properties (such as antimicrobial, anti-inflammatory and cytotoxic activities) have been proven in several scien- tific studies. Stated properties of old man’s beard have been attributed mainly to its major secondary metabolite - a dibenzofuran derivative, usnic acid. The aim of the current study was to examine cyto- toxic activity of the quantified supercritical CO2 extract and pure usnic acid in parallel against L929 fibrosarcoma cells. Our results indicated the tested extract to be more toxic towards the investigated cell line compared to usnic acid per se, despite the high amount of this compound (81.41% (w/w)) revealed in the extract using HPLC. Such result may be connected to the unidentified compounds present in the supercritical CO2 extract that could be responsible for apoptosis and oxidative stress in the exerted cytotoxicity of the investigated extract, but not usnic acid. Our study supports further in- vestigations of supercritical CO2 extract of U. barbata as a prospective therapeutic agent with potential relevance in the treatment of sarcoma.

Incidence of gastrointestinal bleeding in most populations is about 1 per 1,000 inhabitants. More than 65% of all bleeding episodes are associated with drug use. The most often involved are non-steroidal antiinflammatory drugs and low doses of acetyl-salicylic acid. The mortality within the first month after the bleeding episode is about 10-12%, and has not significantly changed in the last decade. Therefore, bleeding prevention is of major importance. Appropriate selection of patients, proper drug choice, application of lowest efficient doses of potentially ulcerogenic drugs, and use of drugs that inhibit gastric acid secretion remain cornerstone preventive measures of gastrointestinal bleeding.

Background Adrenomedulin is a peptide firstly isolated from human phaeochromocytoma with vasodilatory effects. Secretion of adrenomedulin is modified by inflammatory cytokines and it may be important as a suppressor of lupus nephritis activity. Triggering receptor expressed on myeloid cells-1 (TREM) is a member of immunoglobulin superfamily clearly elevated in inflammatory conditions. Some subgroups of patients with systemic lupus erythematosus-SLE may have especially elevated levels of s-TREM. Objectives The aim of this study was to examine adrenomedulin and s-TREM levels in patients with SLE and healthy controls and to assess their relation with SLE disease activity index-SLEDAI Methods Serum samples from 50 SEL patients and 40 healthy blood donors were analysed. Serum levels of adrenomedulin, s-TREM, complement components C3 and C3, and anti-dsDNA antibodies were measured Results Patients with SLE had higher adrenomedulin (9.8±4.5 vs. 15.3±7.2 pg/mL, p Conclusions Increased adrenomedulin and s-TREM are found in lupus sera. Despite of correlation with SLEDAI score, s-TREM is not better then anti-dsDNA antibodies as a predictor of SLE disease activity. Disclosure of Interest None declared

BACKGROUND Despite successful primary percutaneous coronary intervention (PCI) after ST-segment elevation myocardial infarction (STEMI), some patients develop left ventricular systolic dysfunction (LVSD) and acute heart failure (HF). Identifying patients with an increased risk of developing LVSD by means of biomarkers may help select patients requiring more aggressive therapy. OBJECTIVES The aim of this study was to evaluate the relationship between the levels of oxidative stress markers and development of LVSD and acute HF early after STEMI. MATERIAL AND METHODS The study enrolled 148 patients with the first STEMI, who were treated by primary PCI < 12 h from the onset of symptoms. We assessed the impact of different biomarkers for developing LVSD and acute HF (Killip ≥ 2) including: markers of necrosis - peak creatine kinase (CK), markers of myocardial stretch - B-type natriuretic peptide (BNP), inflammatory markers - C-reactive protein (CRP), leucocyte and neutrophil count, as well as oxidative stress markers - total thiol groups, catalase, superoxide dismutase (SOD) and glutathione reductase (GR). RESULTS In multivariate analysis, thiol groups, peak CK, anterior wall infarction, and age were predictors of LVEF ≤ 40%. Out of 16 variables significantly associated with the Killip ≥ 2 in univariate logistic regression analysis, 5 appeared to be independently associated with acute HF in multivariate analysis: catalase, BNP, leucocytes, neutrophil count, and size of left atrium. CONCLUSIONS In this study, we have shown for the first time that thiol groups and catalase are independent predictors of STEMI complication - LVSD and acute HF, respectively. Beside routine used biomarkers of necrosis and myocardial stretch, thiol groups and catalase may provide additional information regarding the risk stratification.

Baseline sympathetic activity was found to be elevated in rheumatoid arthritis (RA) patients and it is related to increased cardiovascular risk in these patients. Although many studies have highlighted the association between RA and increased cardiac sympathetic activity, the underlying mechanistic links remain unclear. The aim of the present study was to understand how diseases-triggered changes in gene expression may result in maladaptive physiological changes. Our results suggest that the equilibrium between noradrenaline synthesis, release and reuptake was disrupted in the ventricles of arthritic rats. In the acute phase of the arthritic process, decreased gene expression of MAO-A might lead to accumulation of noradrenaline in myocardial interstitial space, whereas increased gene expression of NET protected cardiomyocytes from the deleterious effects of enhanced noradrenaline. During the chronic phase, reduced expression of beta(1)-adrenoceptor and decreased efficiency of noradrenaline reuptake contribute to progressive damage of the myocardium and limits heart efficiency.

We assessed the relationship between the serum levels of antibodies against double-stranded DNA (dsDNA), C1q, nucleosomes, histones, C3 and C4 complement components with one another, with organ involvement and overall disease activity in patients with systemic lupus erythematosus (SLE). One hundred seventy-five sera from 99 patients with SLE, 31 sera of patients with other connective tissue diseases, and 20 sera from healthy blood donors were tested. SLE disease activity was assessed by modified SLEDAI-2K (M-SLEDAI-2K), not including complement and anti-dsDNA descriptors. Anti-dsDNA antibodies were measured by indirect immunofluorescence on Crithidia luciliae (CLIFT), standard enzyme-linked immunosorbent assay (ELISA) and ELISA for high-avidity antibodies. The most significant risk factor for renal involvement were anti-C1q antibodies (OR = 3.88, p 0.05), high-avidity anti-dsDNA antibodies for polyserositis (OR = 7.99, p 0.01), anti-histone antibodies for joint involvement (OR = 2.75, p 0.05), and low C3 for cytopenia (OR = 11.96, p 0.001) and mucocutaneous lesions (OR = 3.32, p 0.01). Multiple linear regression analysis showed that disease activity in SLE could be predicted by the levels of antibodies against dsDNA determined by standard (p 0.05) and high-avidity (p 0.001) ELISA, and inversely associated with concentration of C3 (p 0.001). Using stepwise method, high-avidity anti-dsDNA antibodies were found to be in the closest association to M-SLEDAI-2K. Moreover, positive test for high-avidity anti-dsDNA antibodies appeared as an independent risk factor for moderately to severely active disease (M-SLEDAI-2K5) (OR = 5.5, p 0.01). The presence of high-avidity anti-dsDNA antibodies represented a risk for renal, joint, and most importantly for serosal involvement. Our results suggest that simple and reliable ELISA for high-avidity anti-dsDNA antibodies is the test of good clinical utility for the assessment of global SLE activity.

There is a worldwide ongoing investigation for novel natural constituents with cytotoxic and antioxidant properties. The aim of this study was to investigate chemical profile and stated biological activities of the supercritical CO2 extract (SCE) of old man's beard compared to the extracts obtained using the conventional techniques (Soxhlet extracts and macerate). The most abundant compound identified was usnic acid, which content was inversely proportional to the polarity of the solvent used and was the highest in the SCE, which was the sample revealing the highest cytotoxic activity in tested tumor cell lines (B16 mouse melanoma and C6 rat glioma), with lower IC50 values compared to pure usnic acid. Further investigations suggested both SCE and usnic acid to induce apoptosis and/ or autophagy in B16 and C6, indicating higher cytotoxicity of SCE to be related to the higher degree of ROS production. A good correlation of usnic acid content in the extracts and their antioxidant capacity was established, extricating SCE as the most active one. Presented results support further investigations of SCE of old man's beard as a prospective therapeutic agent with potential relevance in the treatment of cancer and/ or in oxidative stress-mediated conditions.

Contains fulltext : 172035.pdf (Publisher’s version ) (Open Access) Hyperuricemia is strongly linked to cardiovascular complications including atherosclerosis and thrombosis. In individuals with hyperuricemia, needle-shaped monosodium urate crystals (nsMSU) frequently form within joints or urine, giving rise to gouty arthritis or renal calculi, respectively. These nsMSU are potent instigators of neutrophil extracellular trap (NET) formation. Little is known on the mechanism(s) that prevent nsMSU formation within hyperuricemic blood, which would potentially cause detrimental consequences for the host. Here, we report that complement proteins and fetuins facilitate the continuous clearance by blood-borne phagocytes and resident macrophages of small urate microaggregates (UMA

Sideritis scardica Griseb. (ironwort, mountain tea), an endemic plant of the Balkan Peninsula, has been used in traditional medicine in the treatment of gastrointestinal complaints, inflammation, and rheumatic disorders. This study aimed to evaluate its gastroprotective and anti-inflammatory activities. Besides, continuously increasing interest in assessing the role of the plant active constituents preventing the risk of cancer was a reason to make a detailed examination of the investigated ethanol, diethyl ether, ethyl acetate, and N-butanol extracts regarding cytotoxicity. Oral administration of the investigated extracts caused a dose-dependent anti-inflammatory effect in a model of carrageenan-induced rat paw edema. Gastroprotective activity of the extracts was investigated using an ethanol-induced acute stress ulcer in rats. The cytotoxic activity of plant extracts was assessed on PBMC, B16, and HL-60 cells and compared to the cytotoxicity of phenolic compounds identified in extracts. Apoptotic and necrotic cell death were analyzed by double staining with fluoresceinisothiocyanate (FITC)-conjugated annexin V and PI. The developed HPLC method enabled qualitative fingerprint analysis of phenolic compounds in the investigated extracts. Compared to the effect of the positive control, the anti-inflammatory drug indomethacine (4 mg/kg), which produced a 50 % decrease in inflammation, diethyl ether and N-butanol extracts exhibited about the same effect in doses of 200 and 100 mg/kg (53.6 and 48.7 %; 48.4 and 49.9 %, respectively). All investigated extracts produced dose-dependent gastroprotective activity with the efficacy comparable to that of the reference drug ranitidine. The diethyl ether extract showed significant dose-dependent cytotoxicity on B16 cells and HL-60 cells, decreasing cell growth to 51.3 % and 77.5 % of control, respectively, when used at 100 µg/mL. It seems that phenolic compounds (apigenin, luteolin, and their corresponding glycosides) are responsible for the diethyl ether extract cytotoxic effect. It also appears that induction of oxidative stress might be involved in its cytotoxicity, since B16 and HL-60 cells increased their ROS production in response to treatment with diethyl ether extract. Neither of the tested extracts nor any phenolic compounds showed significant cytotoxic effect to human PBMC. These results demonstrated the potent anti-inflammatory and gastroprotective activities, as well as the promising cytotoxicity.

ObjectivesTo analyse the similarities and discrepancies between the official rheumatology specialty training programmes across Europe.MethodsA steering committee defined the main aspects of training to be assessed. In 2013, the rheumatology official training programmes were reviewed for each of the European League Against Rheumatism (EULAR) countries and two local physicians independently extracted data on the structure of training, included competencies and assessments performed. Analyses were descriptive.Results41 of the 45 EULAR countries currently provide specialist training in rheumatology; in the remaining four rheumatologists are trained abroad. 36 (88%) had a single national curriculum, one country had two national curricula and four had only local or university-specific curricula. The mean length of training programmes in rheumatology was 45 (SD 19) months, ranging between 3 and 72 months. General internal medicine training was mandatory in 40 (98%) countries, and was performed prior to and/or during the rheumatology training programme (mean length: 33 (19) months). 33 (80%) countries had a formal final examination.ConclusionsMost European countries provide training in rheumatology, but the length, structure, contents and assessments of these training programmes are quite heterogeneous. In order to promote excellence in standards of care and to support physicians’ mobility, a certain degree of harmonisation should be encouraged.

We present a 43-year-old woman with relapsing–remitting multiple sclerosis (MS) who developed lupus syndrome after 32 months of IFN-β-1a therapy. She presented with malaise, myalgia, arthralgia and fever. Laboratory tests showed high erythrocyte sedimentation rate, anaemia and lymphopenia. Antibodies to double stranded DNA (dsDNA) of IgG, IgM and IgA classes were detected on Critidia luciliae. Additionally, high levels of anti-nucleosomal antibodies, low levels of anti-histone and anti-Ro/SSA antibodies were also found. Diagnosis of drug-induced SLE was established. Treatment with IFN-β was discontinued and oral prednisone was started. Twelve weeks after cessation of IFN-β therapy, the patient’s symptoms completely resolved and autoantibodies disappeared. To the best of our knowledge, this is the first report of a patient with MS in whom treatment with IFN-β induced lupus syndrome and antibodies to dsDNA and nucleosome.

Objective: The aim of this paper is to report the first case of drug-induced eosinophilic myocarditis (EM) in a patient with hereditary periodic fever syndrome (PFS). Case: A 28-year-old man with hyper-IgD syndrome, one of the PFS, developed a sulfasalazine-induced systemic hypersensitivity reaction complicated by EM. Thirteen days after sulfasalazine introduction, which had been given for arthritis, the patient developed fever, facial/neck edema, rash and cardiogenic shock, and died within 8 h. The autopsy revealed hemophagocytosis, while acute heart failure caused by necrotizing EM was established as the cause of death. Conclusion: This was a case of drug-induced EM in a patient with PFS that had an atypical presentation, rapid evolution and poor outcome.

Healthy cells exhibit an asymmetric plasma membrane with phosphatidylserine (PS) located on the cytoplasmic leaflet of the plasma membrane bilayer. Annexin A5-FITC, a PS binding protein, is commonly used to evaluate apoptosis in flow cytometry. PS exposed by apoptotic cells serves as a major 'eat-me' signal for phagocytes. Although exposition of PS has been observed after alternative stimuli, no clearance of viable, PS exposing cells has been detected. Thus, besides PS exposure, membranes of viable and apoptotic cells might exhibit specific characteristics. Here, we show that Annexin A5 binds in a cooperative manner to different types of dead cells. Shrunken apoptotic cells thereby showed the highest Hill coefficient values. Contrarily, parafomaldehyde fixation of apoptotic cells completely abrogates the cooperativity effect seen with dead and dying cells. We tend to speculate that the cooperative binding of Annexin A5 to the membranes of apoptotic cells reflects higher fluidity of the exposed membranes facilitating PS clustering.

Sideritis scardica (mountain tea) is an endemic plant on the Balkan Peninsula traditionally used for treating different conditions, mainly of inflammatory nature. This study was aimed to examine the cytotoxic activity of different S. scardica extracts against the rat glioma C6 line and rat astrocytes in primary culture. The obtained data revealed that diethyl ether (extract 2) and ethyl acetate (extract 3) extracts of S. scardica exerted a cytotoxic effect on C6 rat glioma cells. Diethyl ether extract induced an increase in reactive oxygen species production, leading to apoptotic and autophagic cell death. Ethyl acetate extract induced G 2 M cell cycle arrest and autophagy. None of the tested extracts was cytotoxic to rat astrocytes in primary culture. Cytotoxic effects of S. scardica extracts were, at least in part, mediated by their flavonoid constituents apigenin and luteolin that, when applied alone, induced cell cycle arrest, apoptosis, and autophagy.

Background : Decreased activity of serum desoxyribonuclease I (DNase I) in systemic lupus erythematosus (SLE) has been reported, but its role as a biomarker in SLE is still unelucidated. Methods : Seventy-seven SLE patients (aged 39.6 ± 13.1 years) were studied for serum DNase I activity, levels of antinuclear (ANA), anti-dsDNA [high-avidity ELISA, conventional ELISA and indirect immunofluorescence (IIF)], anti-nucleosome, anti-histone antibodies, complement components C3 and C4. SLE disease activity was evaluated by disease activity index (SLEDAI-2K). Thirty-five patients were serologically and clinically followed for 3 – 12 months (mean 5.6 ± 2.8). Thirty-seven healthy blood donors were the control group. Results : DNase I activity in SLE patients was lower than in healthy controls (p < 0.01). DNase I activity was in positive correlation with SLEDAI-2K (p < 0.01), levels of ANA, anti-dsDNA, anti-nucleosome and anti-histone antibodies (p < 0.01) and in negative correlation with C3 concentration (p < 0.05). The highest correlation was found between DNase I activity and anti-dsDNA concentrations determined by high-avidity ELISA (r = 0.624), followed by IIF (r = 0.541) and conventional ELISA (r = 0.405). In the follow-up study, DNase I activity also correlated with SLEDAI-2K (p < 0.01). SLE patients with low DNase I activity more frequently had SLE-specific cutaneous lesions (p < 0.05). Conclusions : Monitoring of DNase I activity simultaneously with SLEDAI-2K might be a useful tool in the followup of SLE. An increase of DNase I activity characterized relapse in most SLE patients, although it did not reach the levels of healthy individuals. A decrease of DNase I activity in SLE flare-ups might be a functional biomarker of a subset of patients with specific dysfunction of apoptotic chromatin degradation.

We present a patient who developed carbamazepine (CBZ)-induced Drug Rash with Eosinophilia and Systemic Symptoms (DRESS) syndrome associated with high serum procalcitonin (PCT). The presentation (high fever, hepatosplenomegaly, leukocytosis), high PCT and CRP initially suggested sepsis, and he was treated with antibiotics, while CBZ was continued. The rash and hepatitis worsened. After withdrawal of CBZ, corticosteroid therapy was administered and the patient recovered with normalization of PCT. This case demonstrates that PCT may be increased in patients with DRESS. This is the first report of CBZ-induced DRESS associated with high PCT, and the second case of increased PCT in DRESS.

Background Osteoporosis is the most common metabolic bone disease. Postmenopausal loss of estrogen is a major risk factor. Objectives To assess if total ethanolic extract of Sideritis scardica could prevent osteoporosis in ovariectomized female rats. Methods Female Wistar rats were divided in three groups: I-sham operated (control), II-ovariectomized-OV and III-ovariectomized, treated orally with S.scardica extract (300 mg/kg) during 20 weeks-OVT. In 18th experimental week oral glucose tolerance test has been performed, as well measurement of bone mineral density-BMD. After killing, femurs, plasma and erytrocytes were isolated. The following parameters were measured: morfometric bone parameters, wet and dry bone mass, biomechanical testig (three point flexural test), Ca and P in urine and plasma, alkaline phosphatase in plasma, malondialdehyde, catalase activity, reduced gluthation (GSH), total SH groups, free fatty acids and fructozamines. Results There was no significant difference in Ca and P concentration in plasma between groups, but urinary Ca and P were significantly higher in OV rats (Ca 0.67±0.24 mM, P 3.29±0.77 mM) in comparison to control (Ca 0.434±0.21 mM, P 2.32±1.00 mM) and OVT group (Ca 0.52±0.19 mM, P 2.89±0.42 mM) (p 2 , for OV group 2726.68±93.4 N/m 2 and for OVT group 3257.2±86.5 N/m 2 (p Conclusions S. scardica extraxct prevents bone loss in ovariectomized rats, increases bone resistance to mechanical load and reduces cardiovascular metabolic risc factors associated with menopause. Disclosure of Interest None declared

Background and Objectives Besides phagocytosis as mechanism to combat pathogens, neutrophils are able to eject DNA decorated with microbicidal proteins (neutrophil extracellular traps-NETs), that can immobilize and kill microorganisms extracellularly. Procoagulative properties of NETs have already been shown, however, a role of NETs formation in atherosclerotic disease has not been described so far. The process of chromatin release is referred to as NETosis. The aim of this study was to assess if cholesterol crystals are able to induce NETosis in vivo and in vitro . Materials and Methods Polymorphonuclear cells-PMNs were isolated from blood obtained from healthy volunteers after informed consent. Isolated PMNs as well as whole blood were co/cultured with cholesterol crystals. After the treatment, cytospins were prepared and stained for DNA and neutrophil elastase. Extracelullar DNA was quantified by fluorometry. For in vivo experiments, cholesterol crystals were injected into murine air pouches. After 24h, air pouches were lavaged with PBS. After centrifugation cells were used for cytospins and supernatants for cytokines determination. Tophus-like aggregates were isolated and characterised by immune histochemistry and immunobloting. In addition, human autopsy specimens obtained from subject died of cardiac diseases were assessed for presence of NET markers. Results Cholesterol induced dose dependent NETosis in isolated PMNs as well as in whole blood samples. The maximal applied concentration of cholesterol crystals induced 236.5% higher DNA release when compared to control. Cholesterol crystal injected in air pouches induced tophus-like aggregates composed of aggregated NETs attached to the cholesterol crystals. These structures were highly positive for citrullinated histones, markers of NET formation. NET markers were also found in atherosclerotic blood vessels in zones surrounding necrotic core of atheromatous plaque, which is rich in cholesterol crystals Discussion This study showed for the first time ability of cholesterol crystals to induce NETosis in vitro , as well as in vivo . These data may explain prompt thrombus formation in atherosclerotic disease after rupture of fibrous cap of an atheromatous plaque containing huge amount of cholesterol crystals, which will be the subject of our future research.

Background and Objectives In autoimmune conditions, like rheumatoid arthritis (RA), the own immune system attacks the body and causes inflammation. One of the consequences of inflammation during autoimmune diseases is a changed glycosylation profile of IgG antibodies. RA is not only characterised by an altered N-glycan structure bound to the Fc portion of IgG molecules but also by circulating rheumatoid factors (RF) that recognizes the Fc portion of IgG as antigen. The aim of our work was to investigate the glycosylation of immune complexes circulating in the blood of patients with seropositive and seronegative RA. Material and Methods Sera from 70 patients with RA (RF positive n = 43 and RF negative n = 27) and 8 healthy donors (control) have been used in the analysis. (I) Random serum IgG-complexes were captured with F(ab’) 2 fragments of anti-human-IgG and analysed by ELISA for their interaction with anti-human-IgG, anti-human-IgM, AAL lectin (fucose-specific) and LCA lectin (manose-specific). (II) The IgG complexes eluted from the ELISA-plates were analysed by Western Blots for the presence of IgG, IgM and AAL as well as LCA binding. (III) IgG affinity purified with AAL-, SNA- and jacalin-agarose columns was tested by ELISA as target for IgM rheumatoid factor. Results (I) The accessibility of fucose residues for the AAL lectin of circulating IgG complexes was much higher in patients with RA when compared to healthy donors. This reactivity was independent of the presence of IgM-RF. The accessibility of mannose residues for the LCA lectin of circulating IgG complexes was much higher in patients with RA when compared to healthy donors. This reactivity was only to be observed in the presence of IgM-RF. (II) Western Blot confirmed an increased presence of IgM in IgG complexes from seropositive patients with RA. The fucosylation of IgG did not differ much between all groups. (III) No difference was detected in IgM-RF affinity towards positive and negative fractions of AAL- and SNA-reactive IgG. However, IgM-RF displayed a reduced binding to the minor IgG fraction isolated with jacalin-affinity chromatography. Conclusion In not-denatured IgG circulating in patients with RA the glycan shows a lower accessibility for the fucose-specific AAL lectin, independent of the presence of IgM-RF. In contrast, the increased binding of the LCA lectin is due to the presence of IgM complexed to IgG. IgM-RF did not discriminate between IgG reactive with AAL and SNA lectin. AAL – Aleuria aurantia lectin (α-1,6 core fucose) LCA – Lens culimaris agglutinin (α-mannose) SNA - Sambucus nigra agglutinin (α-2,6-sialic acid)

Background Many recent studies have suggested that smoking is an important risk factor for rheumatoid arthritis (RA), particulary IgM rheumatoid factor (IgM-RF) and antibodies to citrullinated peptide (ACPA) positive RA, as well as a predictor of severe disease. However, there is lack of data on relationship between history of smoking and response to tumor necrosis factor antagonists in patients with RA. Objectives To determine whether cigarette smoking influences the response to etanercept treatment in patients with RA. Methods A history of cigarette smoking was obtained from a questionnaire completed by each patient starting therapy with etanercept since 2008 (n=136). A core set of demographic and clinical variables was recorded at baseline and at 3 and 12 months. The extent of smoking was quantified in pack-years (py), with 1 py equivalent to 20 cigarettes per day for 1 year. The influence of cigarette smoking (current or past) on the response to therapy was evaluated by logistic regression, with never smokers as the referent group. Response to therapy was defined according to the European League Against Rheumatism improvement criteria, based on their 3 or 12-month Disease Activity Score (DAS28) and absolute change in DAS28 from baseline. Results A history of smoking was found in 68/136 (50%) patients. Of these, 50/136 (36.8%) were current smokers at the start of etanercept therapy. There was no significant difference in age, age of onset, sex and disease duration between nonsmokers, past smokers and current smokers. However, there was an increase in the frequency of patients with IgM-RF and ACPA who had smoked, although this did not achieve statistical significance. Compared with never smokers, current smokers were less likely to achieve a good response at 3 months following the start of etanercept therapy (27% versus 41%; p Conclusions RA patients with a history of smoking were more likely to show a poor response to etanercept. Response failure was associated with the intensity of smoking. References G. Westhoff, et al. Rheumatoid arthritis patients who smoke have a higher need for DMARDs and feel worse, but they do not have more joint damage than non-smokers of the same serological group. Rheumatology 2008; 47:849-854 Disclosure of Interest None Declared

Gout is characterized by an acute inflammatory reaction and the accumulation of neutrophils in response to monosodium urate (MSU) crystals. Inflammation resolves spontaneously within a few days, although MSU crystals can still be detected in the synovial fluid and affected tissues. Here we report that neutrophils recruited to sites of inflammation undergo oxidative burst and form neutrophil extracellular traps (NETs). Under high neutrophil densities, these NETs aggregate and degrade cytokines and chemokines via serine proteases. Tophi, the pathognomonic structures of chronic gout, share characteristics with aggregated NETs, and MSU crystals can induce NETosis and aggregation of NETs. In individuals with impaired NETosis, MSU crystals induce uncontrolled production of inflammatory mediators from neutrophils and persistent inflammation. Furthermore, in models of neutrophilic inflammation, NETosis-deficient mice develop exacerbated and chronic disease that can be reduced by adoptive transfer of aggregated NETs. These findings suggest that aggregated NETs promote the resolution of neutrophilic inflammation by degrading cytokines and chemokines and disrupting neutrophil recruitment and activation.