Miroslav Mayer

Decrease in circulating DNA, IL-10 and BAFF levels in newly-diagnosed SLE patients after corticosteroid and chloroquine treatment

Arsenal of pattern-recognition receptors alongside antibody production machinery make B cells vulnerable to autoimmune response if an autoantigen elicits both pathways in a self-sustained fashion. Systemic lupus erythematosus is an autoimmune disease characterized by autoantibodies to DNA, RNA and related structures. Murine studies demonstrated autoreactive B cell activation upon TLR9 stimulation with DNA-containing immune complexes. This activation could be abolished with chloroquine, a drug used in SLE treatment that also blocks TLR9 signaling. We investigated whether chloroquine modulates TLR9 expression, circulating DNA levels and B cell-related cytokines in newly discovered, untreated SLE patients. TLR9 was measured in peripheral blood B cells by flow cytometry, serum DNA by real-time PCR, and IL-10 and BAFF by ELISA before treatment, after 3weeks on corticosteroids, and 3months after introduction of chloroquine. We found that circulating DNA is higher in SLE patients than in controls in every time-point and decreases significantly after chloroquine treatment. Untreated patients had higher serum IL-10 than controls or patients on corticosteroids. Also, corticosteroids decreased and chloroquine completely abolished CpG-mediated CD86 upregulation on B cells and IL-10 secretion in PBMC culture. Providing the TLR9 pathway activation demonstrates its importance in pathogenesis of human SLE, this data supports continuation of chloroquine in SLE treatment protocol. In addition, observed modulation of cytokine and DNA levels after immunomodulatory treatment prompts for inclusion of untreated patients in studies of human immune disorders.

Redefining a diagnosis: from meningeal plasma cell granuloma to rheumatoid meningitis. Report of a patient follow-up

Sir, In one of the previous issues of your Journal, we presented a patient with non-infectious meningitis, histologically characterized as plasma cell granuloma [1]. During the further months of follow-up, the patient developed mild normocytic anemia and a moderately elevated level of C reactive protein (CRP, 43.3 mg/l, normal values\5.0 mg/l) with unequivocally elevated levels of the rheumatoid factor (RF, 171.7 IU/ml, normal values \15.0 IU/ml) and antibodies against cyclic citrullinated peptides (anti-CCP, 405.3 IU/ml, normal values\7.0 IU/ ml). Although she was initially without signs and symptoms of arthritis, during the following weeks after obtaining these laboratory results the patient developed gradually worsening pain, swelling and morning stiffness of several joints, including both radiocarpal joints, the right elbow, right knee, as well as bilateral talocrural joints (Fig. 1). The clinical presentation consistent with polyarthritis in addition to elevated CRP levels and pronouncedly increased levels of RF and anti-CCP were sufficient for the diagnosis of rheumatoid arthritis. The patient was started on a regimen of a gradually tapered daily dose of methylprednisolone in addition to weekly methotrexate. Treatment led to stable remission of arthritis and further regression of the previously noted neurological deficit. The clinical course of the patient’s disease was followed by the development of our understanding of the clinical context in which the meningeal lesions took place. Although we previously concluded that the plasma cell granulomatous meningitis in our patient was an isolated finding and a primary disease of the meninges, the further disease course warranted a redefinition of the patient’s diagnosis, making rheumatoid arthritis the primary disease and (rheumatoid) meningitis its central nervous system manifestation. Rheumatoid meningitis (RM) is an extremely rare manifestation of rheumatoid arthritis (RA). The largest case series ofRM [2] involved 19 patients,most ofwhichwere diagnosed with RM post mortem based on autopsy findings (17 of 19 patients). That would probably not be the case nowadays, given the availability and development of sophisticated imaging methods, primarily magnetic resonance imaging. Patients from the case series [2] developed RM late in course of RA (after amedian of 14 years following diagnosis of RA). This makes the occurrence of RM as the initial disease manifestation evenmore unusual, although it was described in the literature in the recent years [3]. Both the leptoand the pachymeninges may be affected in patients with rheumatoid meningitis [2]. It is interesting tomention that the neurological manifestations observed in our patient (predominantly including seizures and paresis) are considered to be consistent with leptomeningitis, while symptoms such as headache and cranial neuropathies are ascribed to pachymeningitis [4]. Despite the unquestionable utility of imaging methods and I. Padjen (&) M. Mayer Division of Clinical Immunology and Rheumatology, Department of Internal Medicine, University Hospital Centre Zagreb, Kispaticeva 12, Zagreb, Croatia e-mail: ivan_padjen@yahoo.ca

FRI0284 Altered patterns of histone acetylation point to mechanisms of transcriptional dysregulation in patients with systemic lupus erythematosus

Background Systemic lupus erythematosus (SLE) is a chronic autoimmune disease designated by a heterogeneous course and systemic nature. It arises as a result of complex pathways, as well as the interaction of genetic and environmental factors, leading to the altered reactivity of the immune system that culminates in autoantibody formation. Epidemiological studies have shown an important role of the genetic component in the emergence of SLE and genome-wide association studies have identified more than 50 SLE-associated risk loci, pointing to a complex genetic background. Objectives The aim of the study was to further elucidate the genetic mechanisms influencing the development of SLE. Methods We performed chromatin immunoprecipitation experiments to ascertain the levels of histone acetylation in peripheral blood mononuclear cells collected from 5 recent onset and treatment naive SLE patients compared to 5 age and gender matched controls. Results The analysis revealed 16 379 significantly enriched genomic regions in control patients compared to 39 204 significantly enriched genomic regions in SLE patients. Among the SLE specific regions several pathways were significantly enriched including the adaptive immune system pathway, cytokine signalling in immune system, disease of immune system and inflammation mediated by chemokine and cytokine pathway. Conclusions The collective data point to a significant alteration of histone acetylation patterns in SLE patients possibly mediated by the DNA specific autoantibodies. The results of the study offer additional insight into the genetics of SLE pointing to putative mechanisms of transcriptional dysregulation. Disclosure of Interest None declared

FRI0282 Cardiovascular damage in deceased patients with systemic lupus erythematosus

Background Cardiovascular comorbidities are a major contributor of damage in patients with SLE. They are driven by classical, as well as SLE-related risk factors, i.e. disease activity and immunosuppressive treatment. Objectives We aimed to analyze cardiovascular damage (CVD) in a group of 90 deceased SLE patients regularly followed-up in a routine academic setting at our Department, and to identify features associated with accrual of CVD. Methods We retrospectively observed 90 SLE patients (68 females) deceased within the 2002–2011 period. All patients were ≥18 years of age and Croatian residents at the time of death, fulfilling ≥4 classification criteria of the American College of Rheumatology (ACR). We identified patients with CVD, including the following components of the Systemic Lupus International Collaborating Clinics (SLICC)/ACR damage index: cardiovascular damage as defined by the index (cardiac damage), peripheral vascular damage, cerebrovascular accident, pulmonary infarction, bowel infarction and avascular necrosis. An extensive set of variables was compared between patients with and without CVD: demographics, ACR criteria at diagnosis and death, damage (according to the SLICC/ACR index) and its components one year following diagnosis and at the time of death, disease activity at diagnosis (according to the European Consensus Lupus Activity Measurements index, ECLAM), as well as features of the metabolic syndrome, smoking and immunosuppressive treatment. Frequencies were compared using the χ2 and Fishers exact test, and continuous variables using the t-test and Mann-Whitney U test. Variables associated with CVD in the univariate analysis were included in a multivariate logistic regression model. Results We identified 63/90 patients with CVD, including 46/63 (73%) with cardiac damage, 19/63 (30%) with peripheral vascular damage, 21/63 (33%) with cerebrovascular accident, 4/63 (6%) with bowel infarction, 14/63 (22%) with avascular necrosis and a single patient with pulmonary infarction (Figure 1). Patients with CVD had a higher disease duration at time of death compared to patients without CVD (12±8 vs. 7±6 years), as well as higher cumulative proportions of hematologic disorder (60/63 vs. 15/27), lymphopenia (48/63 vs. 10/27), pulmonary damage (19/63 vs. 1/27), fractures (25/63 vs. 2/27), higher overall damage (6.0±3.0 vs. 2.4±2.0) and a higher proportion of secondary antiphospholipid syndrome (14/63 vs. 1/27) (p<0.05). Conversely, patients with CVD had a lower proportion of discoid lupus at diagnosis (7/49 vs. 9/24) and a lower proportion of skin damage one year following diagnosis (2/63 vs. 5/27) (p<0.05). Parameters associated with cardiovascular damage in the multivariate model were cumulative fulfillment of lymphopenia as a classification criterion (odds ratio, OR 4.7 (95% confidence interval, CI 1.3–17.0)) and accrual of pulmonary damage (OR 13.1 (95% CI 2.2–76.3)). Figure 1. Distribution of cardiovascular damage in the analyzed group (numbers represent frequencies of each subtype of cardiovascular damage) Conclusions More than two thirds of deceased patients accrued CVD over the disease course. Lymphopenia and pulmonary damage may be associated with CVD in deceased SLE patients. References Vila LM et al. Arthritis Rheum 2006;55:799–806. Becker-Merok A and Nossent JC. Lupus 2009;18:508–15. Disclosure of Interest None declared

AB0460 Causes of Early and Late Death of Patients with Systemic Lupus Erythematosus over A 10-Year Period

Background Causes of death (CODs) of patients with systemic lupus erythematosus (SLE) comprise active SLE and comorbidities that develop as a result of SLE and immunosuppressive therapy: infections, cardiovascular disease and malignant tumors. Active disease and infections are typical causes of early death, while cardiovascular disease typically causes late death. However, COD frequencies depend on the source population and data ascertainment method. Objectives To identify and compare causes of early and late death of SLE patients deceased during a ten-year period. Methods We retrospectively identified SLE patients followed-up by our Department, deceased between 2002 and 2011, and included patients with ≥4 classification criteria of the American College of Rheumatology (ACR), ≥18 years of age and Croatian residency at the time of death. Death and causes of death were retrospectively identified using patient medical records, as well as death certificates and autopsy reports, when available. We also matched data on all SLE patients that visited our Department from 2002 to 2011 with the National Mortality Database. We classified CODs into five categories: active SLE, cardiovascular disease, infection, malignant tumor and other. More than one COD category was possible in a single patient. We compared the frequencies of each COD category between patients deceased within and after 5 years following diagnosis (early vs. late death). Frequencies were compared using the χ2 and Fishers exact test, and continuous variables using the t-test and Mann-Whitney U test. The study was approved by the local ethics committee. Results We identified 90 deceased patients (68 females, 22 males; 21 in the early death group (EDG), 69 in the late death group (LDG)). EDG patients were older than LDG patients at diagnosis (mean age±SD: 56±15 vs. 46±17 years; p=0.005), but there was no difference between the age at death (mean age±SD: 58±15 years for all patients). Patients were followed-up for a median of 10 years (IQR: 5–15 years). LDG patients fulfilled a higher number of ACR criteria compared to EDG patients (median, IQR: 6, 5–7 vs. 5, 4–6; p=0.018). No difference between COD category frequencies was detected between EDG and LDG. Nevertheless, infections and active SLE were leading causes of early death (9/21 and 8/21, respectively), while cardiovascular disease was the most frequent cause of late death (30/69), followed by infection and active SLE (21/69 and 18/69, respectively)(Table). SLE was mentioned in the death-related medical records of only 41/90 patients. Cause of death Patients deceased % Early death Late death (N=90) (N=21) (N=69) Active disease (SLE) 26 29 8 18 Infection 30 33 9 21  a) Sepsis 20 22 6 14  b) Pneumonia 18 20 5 13  c) Urinary tract infection 8 9 3 5 Cardiovascular disease 36 40 6 30  a) Myocardial infarction + ischemic heart disease 12 (7+5) 13 1 11  b) Myocardial infarction 5 6 1 4  c) Stroke 8 9 0 8  d) Pulmonary embolism 3 3 1 2 Malignant tumor 15 17 1 14 Other 10 11 4 6 Unknown 8 9 1 7 Conclusions Infections and active SLE are leading causes of early death, while cardiovascular disease is the most frequent cause of late death in SLE. Lack of recording of SLE in death-related medical records requires matching of clinical data with a complementary source, such as a population-based mortality database, to identify deceased SLE patients. References Nossent J et al. Lupus 2007;16:309–17. Disclosure of Interest None declared

AB0508 Frequency of the American College of Rheumatology (ACR) Classification Criteria in A Group of Patients with Systemic Lupus Erythematosus (SLE) Deceased during A 6-Year Period

Background Although the number of fulfilled ACR criteria is not a feature of disease severity in patients with systemic lupus erythematosus (SLE), renal and neurologic involvement are associated with a severe disease course (1,2). Objectives Comparison of the number of fulfilled ACR criteria between SLE patients (a) deceased early and late during the disease course and (b) identified and not identified as deceased in the period from 2006 to 2011. Methods Data were retrieved from our hospital-based registry of SLE patients. Patients with a clinical diagnosis of SLE and at least one visit to the Centre in the 2006-2011 period or death in the same period were analyzed. Deceased patients were identified using the registry and death certificates. The number of fulfilled ACR criteria was counted for each patient. The χ2 test and Fisher exact test, as well as the Student t-test were used to evaluate differences between categorical and continuous variables, respectively. The study was approved by the hospital ethics committee. Results The total number of SLE patients with at least one visit to the Centre between 2006 and 2011 identified from the registry was 702. The frequency of ACR criteria was available for 693/702 patients. We identified 48 deceased patients (31 females and 17 males): 28 using the registry and 20 using death certificates - the latter were lost to follow-up before 2006. Data on the year of diagnosis and fulfillment of ACR criteria were not available for 4 deceased patients. The length of follow-up in the observed group of deceased patients was 12.09±7.38 years. Only 9/44 patients died within the first 5 years following diagnosis (early death). The rest of observed patients (35/44) died later during the disease course. The number of fulfilled criteria in the early and late death group was 4.11±1.05 and 4.71±1.56, respectively (difference not statistically significant). No difference was observed between the fulfillment of each of the criteria, except for the malar rash (19/35 in the late death group vs. 1/9 in the early death group, p=0.027). The frequency of renal disorder and discoid rash was significantly higher in the deceased patient group compared to patients not identified as deceased (21/44 vs. 167/665, χ2=9.702, p=0.002 and 20/44 vs. 128/665, χ2=15.610, p<0.001). No difference in frequencies of other criteria was observed between these two groups. Conclusions Renal disorder and discoid rash are more frequent among deceased SLE patients compared to patients not identified as deceased in the observed group. References Mak A, Cheung MW, Chiew HJ, Liu Y, Ho RC. Global trend of survival and damage of systemic lupus erythematosus: meta-analysis and meta-regression of observational studies from the 1950s to 2000s. Semin Arthritis Rheum. 2012;41:830-9. Doria A, Iaccarino L, Ghirardello A, Zampieri S, Arienti S, Sarzi-Puttini P, Atzeni F, Piccoli A, Todesco S. Long-term prognosis and causes of death in systemic lupus erythematosus. Am J Med. 2006;119:700-6. Disclosure of Interest None declared DOI 10.1136/annrheumdis-2014-eular.3838