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Objectives Changes of routine disease management associated with COVID-19 lockdown might have potentially affected the clinical course of juvenile idiopathic arthritis (JIA). Aim of our study was to assess the rate of disease flare before and during COVID-19 lockdown to investigate its impact on disease course in JIA children. Methods A single-center retrospective study was conducted, including patients presenting inactive JIA between September 1st , 2018 and March 9th , 2019 (group A) and between September 1st , 2019 and March 9th , 2020 (group B). For each patient, demographic and clinical data were collected. The rate of JIA flare from March 10th , 2019 to June 30th , 2019 for group A and from March 10th , 2020 to June 30th , 2020 for group B was compared. Results Group A included 126 patients and group B 124 patients. Statistical analysis did not show significant differences among the two cohorts with respect to age, sex, age of JIA onset, JIA subtype, co-occurrence of uveitis, ANA positivity and past or ongoing medications. The rate of disease flare during lockdown at time of first COVID-19 pandemic wave, was significantly higher in comparison to the previous year (16.9% vs 6.3%, p=0.009). Conclusion Our study showed that COVID-19 lockdown was associated with a higher rate of joint inflammation in JIA children. This finding has a considerable clinical implication, since restrictive measures may be necessary in order to contain pandemics. Our data highlight the need for rearrangement in the home and healthcare management of JIA children during lockdowns.

Background Abnormalities of the immune system are rarely reported in patients affected by RASopathies. Aim of the current study was to investigate the prevalence of immune system dysfunction in a cohort of patients affected by RASopathies. Study design A group of 69 patients was enrolled: 60 at the Federico II University, Naples, 7 at University Magna Graecia of Catanzaro, 2 at “Scuola Medica Salernitana”, Salerno. An age- and sex-matched control group was also enrolled. Autoimmune disorders were investigated according to international consensus criteria. Immune framework was also evaluated by immunoglobulin levels, CD3, CD4, CD8, CD19, CD56 lymphocyte subpopulations, autoantibodies levels and panel of inflammatory molecules, in both patients and controls. Results Frequent upper respiratory tract infections were recorded in 2 patients; pneumonia, psoriasis and alopecia in single patients. Low IgA levels were detected in 8/44 patients (18.18%), low CD8 T cells in 13/35 patients (37.14%). Anti-tg and anti-TPO antibodies were detected in 3/24 patients (12.5%), anti r-TSH in 2 cases (8.33%), all in euthyroidism. Serum IgA and CD8 levels were significantly lower in patients than in controls (p 0.00685; p 0.000656 respectively). All tested patients showed increased inflammatory molecules compared to controls. These findings may anticipate the detection of overt autoimmune disease. Conclusions Patients affected by RASopathies are at risk to develop autoimmune disorders. Routine screening for autoimmunity is recommended in patients with RASopathy.

Background and Aims COVID-19 is associated with liver injury and elevated interleukin-6 (IL-6). We hypothesized that IL-6 trans-signaling in liver sinusoidal endothelial cells (LSECs) leads to endotheliopathy (a proinflammatory and procoagulant state) and liver injury in COVID-19. Methods Coagulopathy, endotheliopathy, and alanine aminotransferase (ALT) were retrospectively analyzed in a subset (n = 68), followed by a larger cohort (n = 3,780) of patients with COVID-19. Liver histology from 43 patients with COVID-19 was analyzed for endotheliopathy and its relationship to liver injury. Primary human LSECs were used to establish the IL-6 trans-signaling mechanism. Results Factor VIII, fibrinogen, D-dimer, von Willebrand factor (vWF) activity/antigen (biomarkers of coagulopathy/endotheliopathy) were significantly elevated in patients with COVID-19 and liver injury (elevated ALT). IL-6 positively correlated with vWF antigen (p = 0.02), factor VIII activity (p = 0.02), and D-dimer (p, Graphical abstract

Aim Coronavirus disease (COVID‐19) is characterized by pneumonia with secondary damage to multiple organs including the liver. Liver injury (elevated alanine aminotransferase [ALT] and aspartate aminotransferase [AST]) often correlates with disease severity in COVID‐19 patients. The aim of this study is to identify pathological microthrombi in COVID‐19 patient livers by correlating their morphology with liver injury, and examine hyperfibrinogenemia and von Willebrand factor (vWF) as mechanisms of their formation. Methods Forty‐three post‐mortem liver biopsy samples from COVID‐19 patients were obtained from Papa Giovanni XXIII Hospital in Bergamo, Italy. Three morphological features of microthrombosis (sinusoidal erythrocyte aggregation [SEA], platelet microthrombi [PMT], and fibrous thrombi) were evaluated. Results We found liver sinusoidal microthrombosis in 23 COVID‐19 patients (53%) was associated with a higher serum ALT and AST level compared to those without (ALT: 10‐fold, p = 0.04; AST: 11‐fold, p = 0.08). Of 43 livers, PMT and SEA were observed in 14 (33%) and 19 (44%) cases, respectively. Fibrous thrombi were not observed. Platelet microthrombi were associated with increased ALT (p

Scurvy is a condition due to chronic ascorbic acid deficiency responsible for vascular fragility and impaired bone formation. The result is gingival bleeding, petechial lesions and articular and bone pain or limb swelling. Children may also present with irritability, limping or refusal to walk. Because of these symptoms it can be misdiagnosed with osteomyelitis, septic arthritis, bone and soft tissue tumour, leukaemia, bleeding disorders and other rheumatologic conditions. In highly developed countries it is rare, but should be suspected in children with underlying chronic condition characterized by selective eating as in developmental delay or autism spectrum disorders. The paper presents an emblematic case.

SARS2‐CoV‐2 breakout in Italy caused a huge number of severely ill patients with a serious increase in mortality. Although lungs seem to be the main target of the infection, very few information are available about liver involvement, possibly evocating a systemic disease. Post‐mortem wedge liver biopsies from 48 patients died from severe pulmonary COVID‐19 disease with respiratory failure were collected from two main hospitals in northern Italy. No patient had clinical symptoms of liver disease or signs of liver failure before and during hospitalization; for each of them liver function tests were available. All liver samples showed minimal inflammation features. Histological pictures compatible with vascular alterations were observed, characterized by increase in number of portal vein branches associated with lumen massive dilatation, partial or complete luminal thrombosis of portal and sinusoidal vessels, fibrosis of portal tract, focally markedly enlarged and fibrotic. SARS‐CoV‐2 was found in 15 of 22 samples tested by in situ hybridization method. Our preliminary results confirm the clinical impression that liver failure is not a main concern and this organ is not the target of significant inflammatory damage. Histopathological findings are highly suggestive for marked derangement of intrahepatic blood vessel network secondary to systemic changes induced by virus that could target not only lung parenchyma but also cardiovascular system, coagulation cascade and endothelial layer of blood vessels. It still remains unclear if the mentioned changes are directly related to virus infection or if SARS‐CoV‐2 triggers a series of reactions leading to striking vascular alterations.

Background Anti-mitochondrial autoantibodies (AMA) detected by indirect immunofluorescence (IIF) on rodent tissues are the diagnostic marker of primary biliary cholangitis (PBC). However, up to 15% of patients with PBC are AMA-negative by IIF. In the effort to close the serological gap and improve the diagnostic sensitivity of PBC testing, recently, novel autoantibodies specific for PBC, such as kelch-like 12 (KLHL12, KLp epitope) and hexokinase 1 (HK1) have been described. In this study, we evaluated the autoantibody profile in a large cohort of PBC patients and in patients with other liver disease, including anti-HK1 and anti-KLp autoantibodies. Methods Sera of 194 PBC patients (126 AMA-IIF-positive and 68 AMA-IIF-negative) and 138 disease controls were tested for a panel of PBC-specific antibodies (MIT3, sp100, gp210, HK1, KLp) using a new automated particle-based multi-analyte technology (PMAT) assay on the Aptiva instrument (Inova). Results Selecting a cutoff yielding a specificity of >95% for all the markers, the sensitivity for anti-MIT3, anti-sp100, anti-gp210, anti-HK1 and anti-KLp in the PBC AMA-IIF-negative cohort was 20.6%, 16.2%, 23.5%, 22.0%, 17.6 and 13.2%, respectively. Six out of the 68 (8.8%) AMA-IIF negative sera were positive for anti-HK1 or anti-KLp alone. Using these new markers in addition to anti-MIT3, anti-sp100 and anti-gp210, the overall sensitivity in this cohort of AMA-IIF-negative patients increased from 53% to 61.8%, reducing the serological gap in AMA-negative PBC patients. Conclusions PBC antibody profiling, made possible by the new Aptiva-PMAT technology, allows recognition of a higher number of AMA-negative PBC patients than conventional immunoassays and may represent a useful tool to evaluate the prognostic significance of autoantibody association in PBC patients.

BACKGROUND The aim of this study is to compare the performance of antitissue transglutaminase (atTG) chemiluminescence immunoassay (CLIA) with the standard enzyme-linked immunosorbent assay (ELISA) methods in monitoring celiac children after the start of gluten-free diet (GFD). METHODS Celiac children diagnosed between 2005 and 2016 at our centre were classified into 2 groups based on serum assay (ELISA vs CLIA) used for atTG monitoring, and were compared on percentage of decrease and time to normalization of atTG on GFD. RESULTS Among 260 included children, the rate of normalization of atTG levels at 30 months' follow-up was 86% and 70% in ELISA and CLIA group, respectively (P

Objective The objective of this study was to use real-world data to evaluate the effectiveness and safety of canakinumab in Italian patients with systemic JIA (sJIA). Methods A retrospective multicentre study of children with sJIA was performed. Clinical features, laboratory parameters and adverse events were collected at baseline, and 6 and 12 months after starting canakinumab. The primary outcome measure of effectiveness was clinically inactive disease (CID) off glucocorticoids (GCs) treatment at 6 months. Results A total of 80 children from 15 Italian centres were analysed. Of the 12 patients who started canakinumab in CID while receiving anakinra, all maintained CID. Of the 68 with active disease at baseline, 57.4% achieved CID off GCs at 6 months and 63.8% at 12 months. In univariate analysis, the variables significantly related to non-response were number of active joints (NAJs) ≥5, history of macrophage activation syndrome (MAS) and disease duration. Multivariate analysis confirmed the association between non-response and NAJs ≥5 [odds ratio (OR) 6.37 (95% CI: 1.69, 24.02), P = 0.006] and between non-response and history of MAS [OR 3.53 (95% CI: 1.06, 11.70), P = 0.039]. No serious adverse events were recorded in this series. There were two cases of MAS during canakinumab, leading to a rate of 2.9 episodes per 100 patient years. Conclusion We have confirmed, using real-world data, the efficacy of canakinumab in sJIA in a multicentric cohort. History of MAS and higher NAJ were associated with lower probability of achieving CID.

Familial Mediterranean fever (FMF) is characterized by self limited episodes of fever and polyserositis.1 MEFV gene en codes for a protein named Pyrin, which plays a pivotal role in the activation and secretion of IL-1.2 Daily colchicine is highly effective in preventing attacks in this disorder in a dose-related fashion.3 Many definitions of colchicine resistance are available in the literature. The European League Against Rheumatism (EULAR) guidelines defined resistance as one or more attacks per month in compliant patients who had been receiving the maxi mally tolerated dose for at least 6 months.4 A similar definition was confirmed by a recent consensus among experts.5 In the present national multicentric longitudinal study, we analyze the impact of colchicine treatment on disease activity and quality of life in real life in pediatric and adult patients with FMF.

Although, the association between celiac disease (CD) and selective immunoglobulin A deficiency (SIgAD) has been known for more than fifty years, the procedures for diagnosing and monitoring patients with both conditions are still far from definitive. When serological markers were introduced as pre-bioptic investigations, it was immediately clear that searching for specific IgA antibodies without checking total serum IgA could lead to a failure in diagnosing IgA-deficient CD patients, while specific IgG antibodies could be useful as additional tests, because they are frequently found in the serum of affected patients. Nonetheless, until recently the diagnosis of CD in IgA-deficient patients was based on the few, fragmentary and often contradictory data available in literature. The introduction of the European Society for Pediatric Gastroenterology, Hepatology and Nutrition (ESPGHAN) guidelines in 2012 provided the current criteria for diagnosing CD in IgA-deficient patients, although some issues remained open, such as the selection of patients who should undergo specific IgG antibody testing and the choice of the most reliable IgG-based test for both diagnosis and follow-up. A real-life study recently assessed the impact of the 2012 ESPGHAN guidelines in diagnosing and monitoring CD in SIgAD patients, highlighting several pitfalls that can lead to operational uncertainties and difficulties in patient management. In the present report, the evolution of diagnostic tools and criteria for CD in SIgAD patients has been critically assessed, both strengths and open issues have been highlighted, and future perspectives for improving the current diagnostic protocols have been suggested.

Mutations in the gene encoding tRNA nucleotidyltransferase 1 (TRNT1) are associated with heterogeneous phenotypes and multisystem involvement of variable severity and progression. Immunodeficiency and inflammation are recurrent-associated features. The use of cytokine inhibitors in suppressing the inflammatory phenotype has been recently reported, with a 3-year follow-up for patients treated with Etanercept. We report on two unrelated patients sharing the same clinical condition, who had been referred to our Pediatric Rheumatology Unit because of recurrent fever associated with cutaneous lesions and increased levels of inflammatory markers since their first months of life. Whole exome sequencing allowed to identify compound heterozygosity for functionally relevant variants in TRNT1 as the only molecular event shared by the two patients. Both patients have been treated with Etanercept during 11 years, documenting normalization of inflammatory indexes and resolution of recurrent fever and associated symptoms. This is the longest follow-up assessment of Etanercept treatment in patients with TRNT1 mutations. Our findings confirm efficacy and safety of the treatment. Key Points • Mutations in TRNT1 have been associated with phenotypic heterogeneity. • We report on two patients with early-onset autoinflammatory syndrome. • Whole exome sequencing led to reveal compound heterozygosity for two variants in TRNT1 in both patients. • The patients were successfully treated with Etanercept for more than 10 years, the longest follow-up described in literature.

COVID-19 breakout in Italy has caused a huge number of severely ill patients with a serious increase in mortality. Although lungs seem to be the main target of the infection very few information are available about liver involvement in COVID-19 infection, that could possibly evocate a systemic disease targeting a lot of organs. Since now there are no reports of large series of histological evaluation of liver morphology in this setting. Knowledge of histological liver findings connected to clinical data is crucial in management of this disease. Post-mortem wedge liver biopsies from 48 patients died for COVID-19 infection were available from two main hospitals located in northern Italy, Lombardy; all sample were obtained during autopsies. No patient has a significant clinical complain of liver disease or signs of liver failure before and during hospitalization; for each of them laboratory data focused on liver were available. All liver samples showed minimal inflammation features; on the other side, many histological pictures compatible with vascular alterations were observed, characterized by portal vein braches number increase associated with lumen massive dilatation, partial or complete recent luminal thrombosis of portal and sinusoidal vessels, fibrosis of portal tract, focally severely enlarged and fibrotic. Our preliminary results concerning histological liver involvement in COVID-19 infection confirm the clinical impression that liver failure is not a main concern and this organ is not the target of significant inflammatory damage; histopatological findings are highly suggestive for marked alteration of intrahepatic blood vessel network secondary to systemic alterations induced by virus that could target, besides lung parenchyma, cardiovascular system, coagulation cascade or endothelial layer of blood vessels.

BackgroundThe most severely COVID-19 patients need intensive care and show increased risk of thromboembolic events. Although some patients meet the diagnostic criteria for the Disseminated Intravascular Coagulation, the pathogenesis of the diffuse thrombotic status remains unclear.The aim of the present study is to evaluate the presence of antiphospholipid antibodies (aPL) in sera of deceased patients with autoptic proven thrombotic microangiopathy to evaluate if some patients may have developed Catastrophic Antiphospholipid Syndrome (CAPS).MethodsThirty-five patients were enrolled. The available medical history, comorbidities, therapies, laboratory and autopsy findings were collected post-mortem from clinical records.IgA, IgG and IgM anti cardiolipin (ACA) and anti β2 glycoprotein 1 (β2GP1) antibodies, IgG and IgM anti phosphatidylserine/prothrombin (PS/PT) antibodies were tested for all the patients.Results3/35 (8.6%) patients were slightly positive for aPL: one for ACA IgG and two for ACA IgM but values were low (< 3X the cut off). No patients tested positive for ACA IgA neither for β2GP1 isotypes. 3/35 (8.6%) patients were positive for PS/PT, one for IgG and two for IgM, but values were less than 2X the cut off. No patients showed simultaneous positivity for ACA and PS/ PT.ConclusionsIt is difficult to categorize the vascular events into a conventional disease: we did not find significant association with anti-phospholipid antibodies. It is most likely that several factors contribute to trigger the hypercoagulability status and the thromboembolism but, on the basis our results, CAPS is probably not involved into the pathogenesis of these phenomena.

A dense fine speckled pattern (DFS) caused by antibodies to the DFS70 kDa nuclear protein is a relatively common finding while testing for anti-nuclear antibodies (ANA) by indirect immunofluorescence (IIF) on HEp-2 cells. However, despite many efforts and numerous studies, the clinical significance of anti-DFS70 antibodies is still unknown as they can be found in patients with various disorders and even in healthy subjects. In this study we aimed at verifying whether these antibodies are associated with thrombotic events or with unexplained recurrent pregnancy loss (RPL). We studied 443 patients with venous or arterial thrombosis or RPL and 244 controls by IIF on HEp-2 cells and by a DFS70-specific chemiluminescent immunoassay (CIA). The DFS pattern was observed in IIF in 31/443 (7.0%) patients and in 6/244 (2.5%) controls (p = 0.01) while anti-DFS70 specific antibodies were detected by CIA in 11 (2.5%) patients and in one (0.4%) control (p = 0.06). Positive samples, either by IIF or by CIA, were then assayed by a second DFS70-specific line-immunoassay (LIA) method: 83.3% of the CIA positive samples were confirmed DFS70 positive versus only 29.7% of the IIF positive samples. These findings show that IIF overestimates anti-DFS70 antibody frequency and that results obtained by specific CIA and LIA assays do not indicate that venous or arterial thrombosis or RPL are linked to a higher prevalence of anti-DFS70 antibodies.

Background The serological screening for celiac disease (CD) is currently based on the detection of anti-transglutaminase (tTG) IgA antibodies, subsequently confirmed by positive endomysial antibodies (EMA). When an anti-tTG IgA positive/EMA IgA negative result occurs, it can be due either to the lower sensitivity of the EMA test or to the lower specificity of the anti-tTG test. This study aimed at verifying how variation in analytical specificity among different anti-tTG methods could account for this discrepancy. Methods A total of 130 consecutive anti-tTG IgA positive/EMA negative samples were collected from the local screening routine and tested using five anti-tTG IgA commercial assays: two chemiluminescence methods, one fluoroimmunoenzymatic method, one immunoenzymatic method and one multiplex flow immunoassay method. Results Twenty three/130 (17.7%) patients were diagnosed with CD. In the other 107 cases a diagnosis of CD was not confirmed. The overall agreement among the five anti-tTG methods ranged from 28.5% to 77.7%. CD condition was more likely linked to the positivity of more than one anti-tTG IgA assay (monopositive = 2.5%, positive with ≥ three methods = 29.5%; p = 0.0004), but it was not related to anti-tTG IgA antibody levels (either positive or borderline; p = 0.5). Conclusions Patients with positive anti-tTG/negative EMA have a low probability of being affected by CD. Given the high variability among methods to measure anti-tTG IgA antibodies, anti-tTG-positive/EMA-negative result must be considered with extreme caution. It is advisable that the laboratory report comments on any discordant results, suggesting to consider the data in the proper clinical context and to refer the patient to a CD reference center for prolonged follow up.

The commercial tests currently available as second-level tests to detect ANA sub-specificities are generally used independently from the ANA immunofluorescence (IIF) pattern. The aim of this study was to evaluate the efficacy of the use of a customizable pattern-oriented antigenic panel by immunoblot (IB) using the International Consensus on ANA Patterns (ICAP) classification scheme, in order to introduce a novel and updated autoimmune diagnostic flowchart. 710 sera referred for routine ANA testing were selected on the basis of the ANA pattern according to the ICAP nomenclature (nuclear speckled AC-2,4,5; nucleolar AC-8,9,10,29; cytoplasmic speckled AC-18,19,20) and on an IIF titer ≥1:320. They were then assayed by three experimental IB assays using a panel of selected antigens. ICAP-oriented IB detected 515 antibody reactivities vs. 457 of traditional anti-ENA in the nuclear speckled pattern group, 108 vs. 28 in the nucleolar pattern group, and 43 vs. 34 in the cytoplasmic speckled pattern. This pilot study may lead the way for a new approach introducing an ICAP pattern-oriented follow up testing as a valid alternative to the existing standard panels, thus enabling more patients with autoimmune rheumatic disease to be accurately diagnosed.

Background: COVID-19 breakout in Italy caused a huge number of severely ill patients with a serious increase in mortality. Although lungs seem to be the main target of the infection very few information are available about liver involvement, possibly evocating a systemic disease. Methods: Post-mortem wedge liver biopsies from 48 patients died for COVID-19 infection were collected from two main hospitals in the north of Italy: all samples were obtained during autopsies. No patient had clinical symptoms of liver disease or signs of liver failure before and during hospitalization; for each of them liver function tests were available. Findings: All liver samples showed minimal inflammation features. Histological pictures compatible with vascular alterations were observed, characterized by increase of number of portal vein branches associated with lumen massive dilatation, partial or complete luminal thrombosis of portal and sinusoidal vessels, fibrosis of portal tract, focally markedly enlarged and fibrotic. Interpretation: Our preliminary results confirm the clinical impression that liver failure is not a main concern and this organ is not the target of significant inflammatory damage. Histopathological findings are highly suggestive for marked derangement of intrahepatic blood vessel network secondary to systemic changes induced by virus that could target not only lung parenchyma but also cardiovascular system, coagulation cascade and endothelial layer of blood vessels. Funding Statement: This work has no financial support. Declaration of Interests: None of the authors declare a conflict of interest Ethics Approval Statement: This work is based exclusively on routine file autoptic material and is ethical approval exempt to use anonymous data. The study followed the Italian general rules used for research related to scientific purposes (official regulations n.72-26 /03/2012).

The Clinical Interview on the Sense of Grip on Chronic Disease has been administered to 68 mothers of children affected by Hereditary Angioedema (C1-Inh HAE), Type 1 Diabetes (T1D), Juvenile Rheumatoid Arthritis (JRA). The objectives are to detect general features of the experience of parenting children with chronic illness as well as the specificities of this experience related to the different conditions. Four Profiles of Sense of Grip were identified: Adempitive, Controlling, Reactive, Dynamic. The Sense of Grip Interview is an effective clinical tool for understanding the characteristics of the disease in daily life, which can help clinicians to encourage family adjustment to disease.

BackgroundCurrent guidelines recommend assessing and relieving pain in all children and in all instances; yet, in clinical practice, management is frequently suboptimal. We investigated the attitude of Italian family pediatricians towards the evaluation and treatment of different types of acute pain in children aged 7–12 years.MethodsThis is a cross-sectional study based on a 17-question survey accessible online from October 2017 to October 2018. Responders had to describe cases of children suffering from any type of acute pain among headache, sore throat, musculoskeletal/post-traumatic pain, and earache. Children’s characteristics, pain assessment modalities and therapeutic approaches were queried. The following tests were used: Z-proportion to evaluate the distribution of categorical data; chi-squared and Kruskall-Wallis to explore data heterogeneity across groups; Mann-Whitney for head-to-head comparisons.ResultsOverall, 929 pediatricians presented 6335 cases uniformly distributed across the types examined. Pain was more frequently of moderate intensity (42.2%,P P N = 4438), the most commonly used first-line non-opioids were ibuprofen (53.3%) and acetaminophen (44.4%). Importantly, a non-recommended dosage was prescribed in only 5.3% of acetaminophen-treated cases (overdosing). Among the misconceptions emerged, there were the following: i) ibuprofen and acetaminophen have different efficacy and safety profiles (when choosing the non-opioid, effectiveness weighted more for ibuprofen [79.7% vs 74.3%,P P ConclusionsSeveral gaps exist between the current practice of pain assessment and treatment and recommendations. Further efforts are needed to raise awareness and improve education on the possible exposure of the child to short- and long-term consequences in case of suboptimal pain management.

Introduction: The advent of biologic agents has revolutionized therapeutic approaches in systemic juvenile idiopatic arthritis (sJIA) as their introduction has been shown to modify disease course and improve overall outcomes, particularly when initiated early. Few studies have reported the drug retention rate (DRR) of biologic drugs in JIA, and none of them has specifically investigated the DRR of interleukin (IL)-1 inhibitors on sJIA. Objectives: The primary aim of the study was to examine the overall DRR of IL-1 blockers in sJIA patients. Secondary aims of our study were to: (i) explore the influence of biologic line of treatment, adverse events (AEs), type of anti-IL-1 agent and the concomitant use of conventional disease modifying anti-rheumatic drugs (cDMARDs) on DRR; (ii) find eventual predictive factors associated with events leading to drug discontinuation. The corticosteroid sparing effect and the impact of disease duration and treatment delay on survival constituted ancillary aims. Methods: sJIA patients – diagnosed according to the revised International League of Association for Rheumatology (ILAR) criteria – treated with anakinra (ANA) and canakinumab (CAN) were enrolled in 15 Italian tertiary referral centers. Demographic, clinical and therapeutic data collected from medical records were retrospectively collected and statistically analyzed. Results: Seventy seven patients were enrolled for a total of 86 treatment courses. The cumulative retention rate of the IL-1 inhibitors at 12-, 24-, 48-, and 60-months of follow-up was 79.9, 59.5, 53.5, and 53.5%, respectively, without any statistically significant differences between ANA and CAN (p = 0.056), and between patients treated in monotherapy compared to the subgroup co-administered with conventional immunosuppressors (p = 0.058). On the contrary, significant differences were found between biologic-naive patients and those previously treated with biologic drugs (p = 0.038) and when distinguishing according to AEs occurrence (p = 0.04). In regression analysis, patients pre-treated with other biologics (HR = 3.357 [CI: 1.341–8.406], p = 0.01) and those experiencing AEs (HR = 2.970 [CI: 1.186–7.435], p = 0.020) were associated with a higher hazard ratio of IL-1 inhibitors withdrawal. The mean treatment delay was significantly higher among patients discontinuing IL-1 inhibitors (p = 0.0002). Conclusion: Our findings suggest an excellent overall DRR for both ANA and CAN that might be further augmented by paying attention to AEs and employing these agents as first-line biologics in an early disease phase.

Key Points Question Are chilblain-like lesions of the toes associated with SARS-CoV-2 infection or is the association merely temporal? Findings This case series of 17 adolescents found that chilblain-like lesions of the toes emerged during the COVID-19 pandemic in otherwise healthy adolescents without signs of SARS-CoV-2 infection or other inflammatory, autoimmune, or thrombophilic phenomena. Meaning These results suggest that chilblain-like lesions are not associated with systemic or localized SARS-CoV-2 infection., Importance Chilblain-like lesions have been one of the most frequently described cutaneous manifestations during the COVID-19 pandemic. Their etiopathogenesis, including the role of SARS-CoV-2, remains elusive. Objective To examine the association of chilblain-like lesions with SARS-CoV-2 infection. Design, Setting, and Participants This prospective case series enrolled 17 adolescents who presented with chilblain-like lesions from April 1 to June 30, 2020, at a tertiary referral academic hospital in Italy. Main Outcomes and Measures Macroscopic (clinical and dermoscopic) and microscopic (histopathologic) analysis contributed to a thorough understanding of the lesions. Nasopharyngeal swab, serologic testing, and in situ hybridization of the skin biopsy specimens were performed to test for SARS-CoV-2 infection. Laboratory tests explored signs of systemic inflammation or thrombophilia. Structural changes in peripheral microcirculation were investigated by capillaroscopy. Results Of the 17 adolescents (9 [52.9%] male; median [interquartile range] age, 13.2 [12.5-14.3] years) enrolled during the first wave of the COVID-19 pandemic, 16 (94.1%) had bilaterally localized distal erythematous or cyanotic lesions. A triad of red dots (16 [100%]), white rosettes (11 [68.8%]), and white streaks (10 [62.5%]) characterized the dermoscopic picture. Histologic analysis revealed a remodeling of the dermal blood vessels with a lobular arrangement, wall thickening, and a mild perivascular lymphocytic infiltrate. SARS-CoV-2 infection was excluded by molecular and serologic testing. In situ hybridization did not highlight the viral genome in the lesions. Conclusions and Relevance This study delineated the clinical, histologic, and laboratory features of chilblain-like lesions that emerged during the COVID-19 pandemic, and its findings do not support their association with SARS-CoV-2 infection. The lesions occurred in otherwise healthy adolescents, had a long but benign course to self-resolution, and were characterized by a microvascular remodeling with perivascular lymphocytic infiltrate but no other signs of vasculitis. These results suggest that chilblain-like lesions do not imply a concomitant SARS-CoV-2 infection. Ongoing studies will help clarify the etiopathogenic mechanisms., This case series examines the association of chilblain-like lesions with SARS-CoV-2 infection.

Blau syndrome (BS) is a rare granulomatous auto-inflammatory disease, characterized by the classic clinical triad of joints, skin and ocular involvements. Ocular manifestation usually consists in a bilateral insidious chronic anterior uveitis with a potential evolution to panuveitis. We describe the case of two siblings, an 8-years old female and a 5-years old male, with a diagnosis of BS, evaluated by Anterior Segment-Optical Coherence Tomography (AS-OCT). In the female patient, slit-lamp examination revealed bilateral anterior granulomatous uveitis and inflammatory sequelae. AS-OCT revealed high intensity reflective layers in the anterior cornea, hyperreflective dots both in the aqueous humor and in the posterior corneal surface. In the male, no signs of inflammation were detected both on slit-lamp examination and AS-OCT scans. AS-OCT is a valuable, non-invasive tool that could improve the diagnosis of ocular involvement, better characterize and follow-up corneal alterations and anterior segment features in pediatric patients with BS.

Background: Juvenile idiopathic arthritis (JIA) is an autoimmune, chronic, inflammatory joint disease, affecting children and adolescents. Patients with JIA may have pain and fatigue in muscles. There are not studies evaluating the pressure pain thresholds (PPTs) of both masticatory muscles and temporomandibular joint (TMJ) in patients with JIA. Objective: This study aimed to investigate PPTs of masticatory muscles and TMJ in subjects with JIA. Methods: Fifty-one JIA patients and fifty-two healthy subjects were recruited. JIA group was assessed for with a standardised clinical examination for temporomandibular disorders. In all subjects, the PPT was evaluated in the following sites: anterior temporalis (AT) and masseter (MM) muscles, TMJ and thenar (TH) eminence. Comparisons between groups were assessed with unpaired t test and ANOVA (P

Purpose: The thymidine phosphorylase (TP) is a key enzyme involved in the metabolism of pyrimidines. Inhibition or downregulation of this enzyme causes accumulation of metabolites with consequences in DNA replication. TP regulates angiogenesis and chemotactic activity of endothelial cells. Different studies showed the presence of TP upregulation in human cancer but the correlation between TP expression and the microvascular density (MVD) in canine mammary tumors is unknown. The aim of this study was to investigate a possible correlation between the MVD and TP expression in tumor cells of canine mammary tumors of different degree of severity (G1–G3) by immunohistochemical analysis.Methods: Sixty-eight samples of spontaneous mammary neoplasia of 5–12 cm in diameter were collected from purebred and mixed-breed dogs (mean aged = 9.5 ± 7), not subject to chemotherapy treatments in veterinary clinics. Histopathological analysis and immunostaining were performed.Results: Carcinoma simple samples have been classified as 72.06% of tubule-papillary, 20.59% cysto-papillary, and 7.35% tubular carcinomas. Immunostainings revealed a marked cytoplasmic expression of TP in 30.88% of samples, mild in 32.35%, weaker in 22.07%, and negative in 14.70%. The correlation analysis and two-way ANOVA showed a linear correlation between MVD and TP with a coefficient of correlation (r) > 0.5 (p < 0.05) in G2 and G3. No correlation between variables was found in G1.Conclusions: These findings suggest that cytoplasmic TP overexpression is correlated with microvascular density in canine mammary tumors, in severe grade, and it can be a potential prognostic factor in breast cancer.

Background In 2008 the Paediatric Rheumatology European Society (PReS) promoted an International Project for the study of Autoinflammatory Diseases (AIDs) named Eurofever, whose main purpose is to create a web-based registry for the collection of information in AIDs patients. Objectives To implement the Registry with the new recently described AIDs and to increase the collection of longitudinal data. Methods The data were extracted from the Eurofever registry, which is hosted in the PRINTO website (http://www. printo.it). From February 2015 we started the longitudinal collection of follow-up data with particular focus on treatment, modification of the clinical picture, onset of complication/adverse events. We have enrolled patients included in the registry up to 28 September 2018. Results Up to date 4175 patients have been enrolled (3843 of them with complete demographic information, 1903 M e 1940 F) from 62 countries. For 3356 (87%) patients also clinical data from onset to diagnosis, collected during the first visit performed at referred pediatric or adult center, are available. For each disease the number of enrolled patients is: FMF 1086 pts (951 with complete clinical data); TRAPS 273 pts (256 complete); CAPS 298 pts (279 complete); MKD 205 pts (190 complete); Blau’s disease 49 pts (26 complete); PAPA 42 pts (41 complete); NLRP12 mediated periodic fever 13 pts (11 complete); DADA2 14 pts (9 complete); DIRA 3 pts (all complete); SAVI 3 pts (all complete); CANDLE 1 pt (complete) and Majeed 4 pts (all complete). Among multifactorial autoinflammatory diseases: PFAPA 676 pts (551 complete); CNO 581 pts (540 complete); Behcet 214 pts (186 complete), undefined periodic fever 368 pts (292 complete) and Schnitzler 13 pts (all complete). The median onset age is 4 years (range 1 month – 75 years), the median diagnosis age is 8 years (range 1 month – 78 years). Most of patients (3509, 91%) presented disease onset during pediatric age ( Conclusion The enrollment in Eurofever Registry is still ongoing. The analysis of data will improve our knowledge both on the natural history of the single disease and on the efficacy/safety of treatment commonly used in the clinical practice. Disclosure of Interests: Ilaria Gueli: None declared, Martina Finetti: None declared, Fabrizio De Benedetti Grant/research support from: Abbvie, SOBI, Novimmune, Roche, Novartis, Sanofi, Pfizer, Jordi Anton Grant/research support from: JA has received grant/research support, consulting fees and/or honoraria from AbbVie, Alexion, BMS, ChemoCentryx, Gebro, GSK, Novartis, Novimmune, Pfizer, Roche, Sanofi and Sobi, Consultant for: JA has received grant/research support, consulting fees and/or honoraria from AbbVie, Alexion, BMS, ChemoCentryx, Gebro, GSK, Novartis, Novimmune, Pfizer, Roche, Sanofi and Sobi, Maria Alessio: None declared, Joost Frenkel: None declared, Luca Cantarini: None declared, Romina Gallizzi: None declared, Judith Sanchez Manubens: None declared, Marco Cattalini: None declared, Efimia Papadopoulou-Alataki: None declared, Rolando Cimaz: None declared, Donato Rigante: None declared, Alma Nunzia Olivieri: None declared, Pavla Dolezalova: None declared, Alberto Martini Consultant for: I do not have any conflict of interest to declare since starting from 1 March 2016 I became the Scientific Director of the G. Gaslini Hospital; therefore, my role does not allow me to render private consultancies resulting in personal income. I perform consultancy activities on behalf of the Gaslini Institute for the companies listed below: AbbVie, Biogen, Boehringer Ingelheim, Bristol-Myers Squibb, EMD Serono, Janssen, Novartis, Pfizer, R-Pharm. The money received for these activities are directly transferred to the Gaslini Institute’s bank account. Before March 2016, I was the head of the Pediatric Rheumatology Department at the G. Gaslini Hospital, where the PRINTO Coordinating Centre is located. For the coordination activity of the PRINTO network, the Gaslini Hospital received contributions from the industries listed in this section. This money has been reinvested for the research activities of the hospital in fully independent manners besides any commitment with third parties., Nicolino Ruperto Grant/research support from: The Gaslini Hospital, where NR works as full-time public employee, has received contributions (> 10.000 USD each) from the following industries in the last 3 years: BMS, Eli-Lilly, GlaxoSmithKline, F Hoffmann-La Roche, Janssen, Novartis, Pfizer, Sobi. This funding has been reinvested for the research activities of the hospital in a fully independent manner, without any commitment with third parties., Consultant for: Received honoraria for consultancies or speaker bureaus (

Background: Dominantly gain-of-function mutations in the NLRP3 gene lead to Cryopyrin associated periodic syndromes (CAPS) characterized by constitutive activation of the inflammasome, increased secretion of interleukin (IL)-1beta and IL-18, and systemic inflammation. IL-1beta and active caspase-1 subunits are released in the serum together with the oligomeric particles of the adaptor ASC (apoptosis-associated Speck-like protein with a caspase-recruitment domain) after activation of the inflammosome complex and, as a consequence, patients with CAPS show an increased serum concentration of ASC+ particles. Objectives: Patients suffering from CAPS are characterized by clinical manifestation similar to other autoinflammatory diseases. This phenomenon together with the lack of specific laboratory tests makes difficult the diagnosis of CAPS. In this context the development of a test for the evaluation of serum ASC levels could provide novel biomarkers facilitating early disease diagnosis and able to monitor treatment responses. Methods: We analysed, with a novel ELISA assay, the levels of ASC particles in serum and plasma of normal subjects, CAPS patients and patients with autoimmune disorders (Multiple Sclerosis (MS), Type 1 Diabetes (T1D) and juvenile idiopathic arthritis), to confirm that ASC presence in the serum is not due to other chronic inflammatory processes characterizing autoimmunity. To evaluate the specificity of this biomarker in CAPS patients and not in individuals suffering from others inflammatory disorders, we also analysed sera from TNF receptor–associated periodic syndrome (TRAPS) patients. Results: ASC particles were higher in untreated CAPS patients with respect to healthy controls and patients suffering from MS and T1D. This tendency was also evident in patients with arthritis and TRAPS even if the difference was not statistically significant due to the small number of samples. In addition after pharmacological treatment there is a tendency to be confirmed through a reduction of ASC levels in CAPS patients. Conclusion: These data suggest that ELISA quantitation of ASC protein could represent a novel and additional strategy for the diagnosis and monitoring of CAPS. Disclosure of Interests:: Fortunata Carbone: None declared, Teresa Micillo: None declared, Luca Cantarini: None declared, Maria Alessio: None declared, Alma Nunzia Olivieri: None declared, Maria Francesca Gicchino: None declared, Antonella Insalaco: None declared, Maria Cristina Maggio: None declared, Orso Maria Lucherini: None declared, Roberto Scarpioni: None declared, Matteo Piga: None declared, Maria Maddalena Angioni: None declared, Laura Obici: None declared, Antonella Simpatico: None declared, Pietro Leccese: None declared, Rita Consolini: None declared, Raffaele Manna: None declared, Paolo Sfriso: None declared, Sara Bindoli: None declared, Paola Galozzi: None declared, Ida Orlando: None declared, Sabrina Chiesa: None declared, Marco Gattorno Grant/research support from: MG has received unrestricted grants from Sobi and Novartis, Giuseppe Matarese Grant/research support from: Matarese reports research grants from Merck-Serono, Biogen Idec, Novartis and IBSA.

Background and Objective: Only a few studies have reported long-term efficacy of interleukin (IL)-1 inhibition in systemic juvenile idiopathic arthritis (sJIA) and adult-onset Still disease (AOSD). Herein we report on the effectiveness of anakinra (ANA), expressed in terms of drug retention rate (DRR), and evaluate the predictive factors of drug survival in a cohort of patients with sJIA and AOSD. Patients and Methods: This is a multicenter study reviewing retrospectively the medical records from 61 patients with sJIA and 76 with AOSD, all treated with ANA in 25 Italian tertiary referral centers. Results: The cumulative retention rate of ANA at 12-, 24-, 48-, and 60-month of follow-up was 74.3%, 62.9%, 49.4%, and 49.4%, respectively, without any significant differences between sJIA and AOSD patients (p = 0.164), and between patients treated in monotherapy compared with the subgroup coadministered with conventional diseasemodifying antirheumatic drugs (cDMARDs) (p = 0.473). On the other hand, a significant difference in DRR was found between biologic-naïve patients and those previously treated with biotechnologic drugs (p = 0.009), which persisted even after adjustment for pathology (p = 0.013). In the regression analysis, patients experiencing adverse events (AEs) {hazards ratio (HR) = 3.029 [confidence interval (CI) 1.750–5.242], p < 0.0001} and those previously treated with other biologic agents [HR = 1.818 (CI 1.007–3.282), p = 0.047] were associated with a higher HR of ANA discontinuation. The median treatment delay was significantly higher among patients discontinuing ANA (p < 0.0001). Significant corticosteroid-sparing (p = 0.033) and cDMARD-sparing effects (p < 0.0001) were also recorded. Less than one-third of our cohort developed AEs, and 85% were deemed mild in nature, with 70% of them involving the skin. Conclusions: Our findings display an overall excellent DRR of ANA on the long run for both sJIA and AOSD, that may be further optimized by closely monitoring patient’s safety issues and employing this IL-1 inhibitor as a first-line biologic as early as possible. Moreover, ANA allowed a significant drug-sparing effect and showed an overall good safety profile.

Background In a generalist laboratory, the integration of the data obtained from hematology analyzers (HAs) with those from multiparametric flow cytometry (FMC) could increase the specificity and sensitivity of first level screening to identify the pathological samples. The aim of this study was to perform a preliminary evaluation of a new simple hybrid method (HM). The method was obtained by integration between HAs reagents into FCM, with a basic monoclonal antibodies panel for the leukocytes differential count. Methods Eighty-one peripheral blood samples, collected in K3EDTA tubes, were analyzed by XN-module, and CyFlow Space System, using both standard MoAbs and HM method analysis, and with the optical microscopy (OM). Within-run imprecision was carried out using normal samples, the carryover was evaluated, data comparison was performed with Passing-Bablok regression and Bland-Altman plots. Results The within-run imprecision of HM methods ranged between 1.4% for neutrophils (NE) and 10.1% for monocytes (MO) always equal or lower to the OM. The comparison between HM methods vs. OM shows Passing-Bablok regression slopes comprised between 0.83 for lymphocyte (LY) and 1.14 for MO, whilst the intercepts ranged between −0.18 for NE and 0.25 for LY. Bland-Altman relative bias was comprised between −12.43% for NE, and 19.77% for eosinophils. In all 11 pathological samples the agreement between the methods was 100%. Conclusions The new hybrid method generates a leukocytes differential count suitable for routine clinical use and it is also useful for identifying morphological abnormalities with a reduction in cost and improvement of screening for first level hematology workflow.

OBJECTIVES: Mevalonate kinase deficiency (MKD) is a rare metabolic disease characterized by recurrent inflammatory episodes. This study aimed to describe the genotype, phenotype and the response to treatment in an international cohort of MKD patients. METHODS: All MKD cases were extracted from the Eurofever registry (EAHC Project No. 2007332), an international, multicenter registry that retrospectively collects data on children and adults suffering from autoinflammatory diseases. RESULTS: One hundred and fourteen MKD patients were included in this study. The median age of onset was 0.5 years. Patients had on average 12 episodes per year. Most patients had gastrointestinal symptoms (n=112), mucocutaneous involvement (n=99), lymphadenopathy (n=102) or musculoskeletal symptoms (n=89). Neurological complaints included headache (n=43), but also cerebellar syndrome (n=2) and mental retardation (n=4). AA-amyloidosis was noted in five patients, almost twice as many as expected from previous cohorts. Macrophage activation syndrome occurred in one patient. Between attacks patients were generally well, but 10-20% patients suffered from constitutional symptoms, such as fatigue, between febrile attacks. Patients with p.V377I/p.I268T compound heterozygosity suffered significantly more often from AA-amyloidosis. Patients without a p.V377I mutation suffered more often from severe musculoskeletal involvement. Treatment with NSAIDs could relieve symptoms. Steroids given during attacks, anakinra and etanercept appeared to improve symptoms and could induce complete remission in MKD patients. CONCLUSION: This study described the clinical and genetic characteristics of 114 MKD patients, which is the largest cohort studied so far. The clinical manifestations confirm earlier reports. However, the prevalence of AA-amyloidosis was far higher than expected.

Objectives Anti-parietal cell antibodies (APCA) are a serologic marker of autoimmune gastritis. Their prevalence in healthy individuals is not well defined. Methods We evaluated APCA prevalence in 515 healthy blood-donors by rat/primate tissue indirect immunofluorescence (IIF), enzyme-linked immunosorbent assay (ELISA), and immunoblot. Results Fifty-three of 515 (10.3%) subjects were positive for APCA by at least one method: 18 only by ELISA, 10 by rodent tissue IIF, and one by primate tissue IIF; 18 were positive by ELISA and primate tissue IIF, and one by ELISA and rodent tissue IIF. Two were positive by both IIF methods, and three were triple positive. APCA positivity was confirmed by immunoblot in 100% of ELISA positive, in 95.8% of positive primate tissue IIF, and in 50% of positive rat tissue IIF. Conclusions A great discrepancy in APCA prevalence detected by different methods in this cohort was apparent. Thus, the results on APCA prevalence in healthy individuals are likely method-dependent.

Objectives: Chronic non-bacterial osteomyelitis (CNO) or chronic recurrent multifocal osteomyelitis (CRMO) is an autoinflammatory disorder characterized by sterile bone osteolytic lesions. The aim of this study was to evaluate the demographic data and clinical, instrumental and therapeutic features at baseline in a large series of CNO/CRMO patients enrolled in the Eurofever registry.Methods: A web-based registry collected retrospective data on patients affected by CRMO/CNO. Both paediatric and adult centres were involved.Results: Complete baseline information on 486 patients was available (176 male, 310 female). The mean age of onset was 9.9 years. Adult onset (>18 years of age) was observed in 31 (6.3%) patients. The mean time from disease onset to final diagnosis was 1 year (range 0-15). MRI was performed at baseline in 426 patients (88%), revealing a mean number of 4.1 lesions. More frequent manifestations not directly related to bone involvement were myalgia (12%), mucocutaneous manifestations (5% acne, 5% palmoplantar pustulosis, 4% psoriasis, 3% papulopustular lesions, 2% urticarial rash) and gastrointestinal symptoms (8%). A total of 361 patients have been treated with NSAIDs, 112 with glucocorticoids, 61 with bisphosphonates, 58 with MTX, 47 with SSZ, 26 with anti-TNF and 4 with anakinra, with a variable response.Conclusion: This is the largest reported case series of CNO patients, showing that the range of associated clinical manifestations is rather heterogeneous. The study confirms that the disease usually presents with an early teenage onset, but it may also occur in adults, even in the absence of mucocutaneous manifestations.

Pachydermoperiostosis (PDP), otherwise known as primary hypertrophic osteoarthropathy, is characterized by digital clubbing, pachydermia and subperiosteal new bone formation. Joint pain, polyarthritis, cutis verticis gyrata, seborrhea, and hyperhidrosis are frequently associated to this condition. We report a 17-year-old boy presented with pain and swelling of knees and ankles, and progressive thickening of skin face with seborrhea from about 4 years. At the admission he also showed digital clubbing of both hands and feet and palmoplantar hyperhidrosis. We hypothesized PDP and molecular analysis confirmed diagnosis showing a novel mutation in a homozygous state in the SLCO2A1 gene coding for prostaglandin transporter. He started therapy with hydroxychloroquine with a great improvement in joint pain and skin conditions. This is the first reported case of PDP who was successfully treated with hydroxychloroquine, with effects not only on arthralgia but also, surprisingly, on skin conditions.

Background Hereditary angioedema with C1-inhibitor deficiency (C1-INH-HAE) is characterized by recurrent edema of unpredictable frequency and severity. Stress, anxiety, and low mood are among the triggering factors most frequently reported. Impaired regulation and processing of emotions, also known as alexithymia, may influence outcomes. The aim of this study was to confirm the presence of alexithymia and stress in children with C1-INH-HAE, to determine whether they are also present in children affected by other chronic diseases, and to investigate their relationship with C1-INH-HAE severity. Data from children with C1-INH-HAE (n = 28) from four reference centers in Italy were compared with data from children with type 1 diabetes (T1D; n = 23) and rheumatoid arthritis (RA; n = 25). Alexithymia was assessed using the Alexithymia Questionnaire for Children scale; perceived stress was assessed using the Coddington Life Event Scale for Children (CLES-C). Results Mean age (standard deviation [SD]) in the C1-INH-HAE, T1D, and RA groups was 11.8 (3.3), 11.7 (2.9), and 11.1 (2.6) years, respectively. Mean C1-INH-HAE severity score was 5.9 (2.1), indicating moderate disease. Alexithymia scores were similar among disease groups and suggestive of difficulties in identifying and describing emotions; CLES-C scores tended to be worse in C1-INH-HAE children. C1-INH-HAE severity was found to correlate significantly and positively with alexithymia (p = 0.046), but not with perceived stress. Alexithymia correlated positively with perceived stress. Conclusions Alexithymia is common in children with chronic diseases. In C1-INH-HAE, it may result in increased perceived stress and act as a trigger of edema attacks. Comprehensive management of C1-INH-HAE children should consider psychological factors. Electronic supplementary material The online version of this article (10.1186/s13023-018-0871-x) contains supplementary material, which is available to authorized users.

Aim This study aimed to investigate whether it is useful both for diagnostic purposes and for cost savings to associate a solid-phase monotest called CTD screen to the immunofluorescence (IIF) test, to detect anti-nuclear and anti-cytoplasmic antibodies (ANA). Methods Fifteen Italian laboratories participated in the study enrolling at least 250-each consecutive unselected samples from patients routinely referred for ANA testing. They all used the conventional IIF method on HEp-2 cells associated to a solid phase CTD screen method. Seven centers used the EliA FEIA CTD screen method on the Phadia 250 instrument (ThermoFisher), five centers used the QUANTAFlash CTD Screen method (Inova Diagnostics) on the BioFlash CLIA instrument, while three centers used both methods, albeit in two different series of patients. Results A total of 5043 samples were tested. We focused attention to the 1674 discrepant results: 106 (2%) were ANA-IIF negative/CTD screen positive and 1568 (31%) were ANA-IIF positive/CTD screen negative. Specific antibodies were detected in 87/106 (82%) of the ANA-IIF negative/CTD screen positive samples, mainly Ro52, Ro60, dsDNA, PM/Scl and Jo1. Thirty four of these patients were diagnosed with an ANA-associated autoimmune rheumatic disease (AARD) (seven SLE, 15 Sjӧgren's syndrome, nine autoimmune inflammatory myositis, three systemic sclerosis). Among 1568 ANA-IIF positive/CTD screen negative discrepant patients, a specific antibody was found in 94 (6%) samples (28 patients with AARD, 13 with undifferentiated connective tissue disease, two with undifferentiated arthritis, and 51 without a rheumatic disease). Conclusions This study is the first that compares the IIF method vs. solid-phase assays in the detection of ANA, on a very large sample size from the routine work-up (real-life approach). Results show that the association of solid-phase CTD screen tests to the ANA-IIF test increases the sensitivity (from 89.2% to 97.4%) and the specificity (from 64.6% to 98.4%) of serological tests for ANA screening. Analysis of costs demonstrated that combination of the two tests is also cost-effective, reducing global costs for the immunoserological diagnosis of AARD by 22%.

Forbush Decrease (FD) effects on cloud composition and structure are under study and the results are still controversial. Time-scales are of paramount importance for supporting either a ‘microphysical hypothesis’, which hypothesizes a relation between FD and cloud microphysical parameters variability, or a different one. A most controversial question is related to the time delay between FD and cloud structure modification. The timescales of a radiative–dynamical mechanism, investigated through a simple model, are compatible with the observed variability with respect to cloud structure. Thus the delayed modification on cloud structure has to be put in relation with solar radiation variability, being coherent with the observed statistically significant Total Solar Irradiance (TSI), temperature and baric field variations, while not supporting the ‘microphysical hypothesis’.

Objectives 2012 European Society of Pediatric Gastroenterology, Hepatology and Nutrition guidelines allow to establish a celiac disease diagnosis without duodenal biopsy in symptomatic pediatric patients with antitissue transglutaminase (anti-tTG) titers >10 times the upper limit of normal. For some years now, new chemiluminescence immunoassays have been made available: it is important to establish the clinical performance of anti-tTG and to determine the cut-off best suited to predict Marsh ≥2 to avoid gastrointestinal endoscopy not only in children, but also in the adult population. Methods A total of 2565 patients performed duodenal biopsy from July 2012 to September 2016; we selected all the patients who had undergone QUANTA Flash anti-tTG immunoglobulin A (IgA) within -3 months of duodenal biopsy and before the start of gluten-free diet. A total of 827 patients fulfilled the criteria for selection. Results Using a cut-off of 20 chemiluminescent unit (CU; area under the curve: 0.995), sensitivity, specificity, positive, and negative predictive value were 98.2%, 98.4%, 97.9%, and 98.6%, respectively. For the correlation with Marsh ≥2, in the pediatric population, positive predictive values (PPV) were 92.1%, 99%, and 100% at 200 CU (10×), 560 CU (28×), and 1000 CU (50×), respectively. In the adult population PPV was 94.2%, 98.2%, and 100% at 200 CU (10×), 350 CU (15×), and 400 CU (20×). Conclusions Sensitivity, specificity, positive, and negative predictive value of QUANTA Flash h-tTG IgA were excellent. The cut-off providing an optimized PPV for histological lesions compatible for celiac disease (Marsh ≥2) for the QUANTA Flash h-tTG IgA is 350 CU (15×) in adult and 560 CU (28×) in children.

OBJECTIVE: To develop and validate a diagnostic score that assists in discriminating primary hemophagocytic lymphohistiocytosis (pHLH) from macrophage activation syndrome (MAS) related to systemic juvenile idiopathic arthritis. STUDY DESIGN: The clinical, laboratory, and histopathologic features of 362 patients with MAS and 258 patients with pHLH were collected in a multinational collaborative study. Eighty percent of the population was assessed to develop the score and the remaining 20% constituted the validation sample. Variables that entered the best fitted model of logistic regression were assigned a score, based on their statistical weight. The MAS/HLH (MH) score was made up with the individual scores of selected variables. The cutoff in the MH score that discriminated pHLH from MAS best was calculated by means of receiver operating characteristic curve analysis. Score performance was examined in both developmental and validation samples. RESULTS: Six variables composed the MH score: age at onset, neutrophil count, fibrinogen, splenomegaly, platelet count, and hemoglobin. The MH score ranged from 0 to 123, and its median value was 97 (1st-3rd quartile 75-123) and 12 (1st-3rd quartile 11-34) in pHLH and MAS, respectively. The probability of a diagnosis of pHLH ranged from 99% for a score of =123. A cutoff value of =60 revealed the best performance in discriminating pHLH from MAS. CONCLUSION: The MH score is a powerful tool that may aid practitioners to identify patients who are more likely to have pHLH and, thus, could be prioritized for functional and genetic testing.

Objectives To analyse the prevalence of CECR1 mutations in patients diagnosed with early onset livedo reticularis and/or haemorrhagic/ischaemic strokes in the context of inflammation or polyarteritis nodosa (PAN). Forty-eight patients from 43 families were included in the study. Methods Direct sequencing of CECR1 was performed by Sanger analysis. Adenosine deaminase 2 (ADA2) enzymatic activity was analysed in monocyte isolated from patients and healthy controls incubated with adenosine and with or without an ADA1 inhibitor. Results Biallelic homozygous or compound heterozygous CECR1 mutations were detected in 15/48 patients. A heterozygous disease-associated mutation (p.G47V) was observed in two affected brothers. The mean age of onset of the genetically positive patients was 24 months (6 months to 7 years). Ten patients displayed one or more cerebral strokes during their disease course. Low immunoglobulin levels were detected in six patients. Thalidomide and anti-TNF (tumour necrosis factor) blockers were the most effective drugs. Patients without CECR1 mutations had a later age at disease onset, a lower prevalence of neurological and skin manifestations; one of these patients displayed all the clinical features of adenosine deaminase 2deficiency (DADA2) and a defective enzymatic activity suggesting the presence of a missed mutation or a synthesis defect. Conclusions DADA2 accounts for paediatric patients diagnosed with PAN-like disease and strokes and might explain an unrecognised condition in patients followed by adult rheumatologist. Timely diagnosis and treatment with anti-TNF agents are crucial for the prevention of severe complications of the disease. Functional assay to measure ADA2 activity should complement genetic testing in patients with non-confirming genotypes.

Objective To describe the clinical, laboratory, and histopathologic features, current treatment, and outcome of patients with macrophage activation syndrome (MAS) complicating systemic juvenile idiopathic arthritis (JIA). Methods In this multinational, multicenter study, pediatric rheumatologists and hemato-oncologists entered patient data collected retrospectively into a web-based database. Results A total of 362 patients, 22% of whom had MAS at the onset of systemic JIA, were included in the study by 95 investigators from 33 countries. The most frequent clinical manifestations were fever (96%), hepatomegaly (70%), and splenomegaly (58%). Central nervous system dysfunction and hemorrhages were recorded in 35% and 20% of the patients, respectively. Platelet count and liver transaminase, ferritin, lactate dehydrogenase, triglyceride, and d-dimer levels were the sole laboratory biomarkers showing a percentage change of >50% between the pre-MAS visit and MAS onset. Evidence of macrophage hemophagocytosis was found in 60% of the patients who underwent bone marrow aspiration. MAS occurred most frequently in the setting of active underlying disease, in the absence of a specific trigger. Nearly all patients were given corticosteroids, and 61% received cyclosporine. Biologic medications and etoposide were given to 15% and 12% of the patients, respectively. Approximately one-third of the patients required admission to the intensive care unit (ICU), and the mortality rate was 8%. Conclusion This study provides information on the clinical spectrum and current management of systemic JIA–associated MAS through the analysis of a very large patient sample. MAS remains a serious condition, as a sizeable proportion of patients required admission to the ICU or died.

Background Autoimmune hepatitis (AIH) is a rare condition characterized by the presence of autoantibodies distinctive of type 1 AIH (AIH-1) and type 2 AIH (AIH-2). The aim of this study was to evaluate the autoantibody profile in a cohort of pediatric and adult AIH patients, using both indirect immunofluorescence (IIF) and a new multiplexed line-blot assay. Methods Sera from 63 pediatric and 53 adult AIH patients were tested for antinuclear (ANA), antismooth muscle (SMA), anti-liver kidney microsome 1 (anti-LKM1), anti-liver cytosol 1 (anti-LC1) autoantibodies using IIF methods; for anti-LKM1, anti-LC1, and soluble liver antigen/liver-pancreas (anti-SLA/LP) autoantibodies using the line-blot; for anti-F-actin autoantibodies using IIF both on VSM47 cell-line and on rat intestinal epithelial cells. Results AIH-1 was the most common type of AIH in the adult cohort (73.6%), while AIH-2 was the most common AIH in the pediatric cohort (61.9%). Both in adult and pediatric AIH-2 anti-LKM1 were the prevalent autoantibodies. In pediatric AIH-2 anti-LC1 autoantibodies were more frequent than in adult AIH-2 (59 vs. 28.6%), and in 35.9% of cases they were present alone. In 17 patients anti-LC1 autoantibodies were detected only with the line-blot assay. The levels of anti-LKM1 and of anti-LC1 were not different between adult and pediatric AIH, and the overall agreement between the results obtained with the two IIF methods for F-actin detection was 98.8% (CI 95%: 94.4-99.7%). Conclusions The line-blot assay showed a higher sensitivity than IIF for anti-LC1 detection. Anti-LKM1 and anti-LC1 autoantibody levels are not different in adults and children. An almost perfect agreement between the two IIF methods for anti-F-actin detection has been observed.

We evaluated whether perfusion brain abnormalities by single-photon emission computed tomography (SPECT) imaging improves diagnostic and prognostic assessment in Sydenham chorea. Twenty-three children with acute autoimmune chorea underwent technetium-99m hexamethylpropyleneamine oxime brain SPECT imaging. In 16 children, SPECT was repeated during the follow-up. A pattern of basal ganglia hyperperfusion was observed in 20 (87%) patients. In 4 of 10 patients with generalized chorea, perfusion was comparable in right and left striatum and right and left thalamus. In 13 patients with hemi-chorea and in 3 with generalized chorea, unilateral hyperperfusion was detected. Three patients with generalized chorea had normal perfusion. Tracer uptake of basal ganglia of the patients at the acute phase was higher than at the follow-up ( P < .001). SPECT seems a useful noninvasive tool in pediatric patients with Sydenham chorea to support the clinicians during the acute phase of disease and to monitor the course of autoimmune chorea.

To compare primary versus secondary intraocular lens (IOL) implantation after cataract removal in patients with juvenile idiopathic arthritis (JIA).Retrospective interventional study. Data were obtained for 40 children (40 eyes) with JIA-associated uveitis operated on for cataract before the age of 16 years between January 1998 and January 2005.Twenty-one patients underwent primary IOL implantation at a mean age of 13.1 ± 2.6 (9-16) years. Mean follow-up was 48.2 ± 5.4 (35-64) months. Nineteen patients underwent cataract removal at a mean age 12.3 ± 2.0 (9-16) years and secondary IOL implantation 13.6 ± 0.3 (11-16) months later. Mean follow-up was 47.2 ± 6.5 (32-64) months. BCVA difference between the two groups was significant at 24-month visit only (p = 0.001). Incidence of secondary glaucoma was significantly lower in the group that underwent secondary IOL implantation (p = 0.01).Secondary IOL implantation after cataract removal in children with JIA-related uveitis provided a significantly lower incidence of secondary glaucoma.

Summary Background: We evaluated the performance of Sysmex UF- 1000i for cell counting and differential cell count, as well as for assessment of bacteria load in cerebrospinal fluid (CSF), as a potential approach for the rapid screening of meningitis or bacterial encephalitis. Methods: We analyzed 77 consecutive CSF samples, 34 of which (44%) displayed leukocyte count >5 white blood cell (WBQ/^iL with optical microscopy. Results on the UF-1000i were compared with those obtained by microscopic analysis. Imprecision was evaluated by testing three CSF samples with leukocyte values between 3.5 and 28.8 WBC/nL in 10 repli- cates. Carry-over was evaluated with the Broughton formula on three CSF pools with leukocyte counts between 93.5 and 132.5 WBC/^L. Linearity was assessed according to CLSI document EP6-A. In the presence of bacteria, identification and antibiogram were performed with Vitex (Biomerieux), except for Neisserie meningitidis (ApiNH, Biomerieux). Sensitivity tests were performed with Vitex and disc diffusion. Results: Optimal correlation was found between UF-1000i and optical microscopy, displaying Pearson's correlation of 0.99 and mean bias of-3.5 WBC/^L (95% Cl, from -7.0 to 0.0 WBC/nL). Imprecision varied between 12 and 16%. Li- nearity was excellent, 4-278 WBC/nL. Carry-over was neg- ligible. ROC analysis yielded AUC of 0.99 for both WBC and bacterial counts. The agreement at threshold >4 WBC/nL was 0.91, with sensitivity and specificity of 1.00 and 0.84. At S19 bacteria^nL cut-off, accuracy was 0.98, sensitivity 1.00 and specificity 0.97. Conclusions: According to these results, CSF screening with UF-I000i seems a reliable approach in terms of instrument performance, turnaround time and overall laboratory effi- ciency.

Objective.Abatacept (ABA) has recently been proposed as second-line treatment in patients with juvenile idiopathic arthritis (JIA)–associated uveitis refractory to anti–tumor necrosis factor-α (anti-TNF) agents, but little is known about its efficacy as a first-line approach. The aim of the present study was to compare the safety and efficacy of ABA as a first-line biological agent (ABA-1) with that of ABA as a second-line treatment after 1 or more anti-TNF agents (ABA-2), in patients with severe JIA-related uveitis.Methods.In this multicenter study, we collected data on patients with severe JIA-related uveitis treated with ABA as a first-line or second-line biological agent. Changes in frequency of uveitis flares/year and ocular complications before and after ABA treatment, clinical remission, and side effects were recorded.Results.Thirty-five patients with a mean age of 10.8 years were treated with ABA for a mean period of 19.6 months. In 4 patients, ABA administration was discontinued, owing to inefficacy on arthritis in 3 cases and allergic reaction in 1. Thirty-one patients, 14 in the ABA-1 group and 17 in the ABA-2 group, completed the 12-month followup period; of these, 17 (54.8%) had clinical remission. The mean frequency of uveitis flares decreased from 4.1 to 1.2 in the ABA-1 group (p = 0.002) and from 3.7 to 1.2 in the ABA-2 group (p = 0.004). Preexisting ocular complications improved or remained stable in all but 5 patients, all in the ABA-2 group. No significant difference was found between the efficacy of the 2 treatment modalities. ABA confirmed its good safety profile.Conclusion.ABA, used as first-line biological treatment or after 1 or more anti-TNF agents, induces a comparable improvement in severe refractory JIA-related uveitis.

Objective To develop criteria for the classification of macrophage activation syndrome (MAS) in patients with systemic juvenile idiopathic arthritis (JIA). Methods A multistep process, based on a combination of expert consensus and analysis of real patient data, was conducted. A panel of 28 experts was first asked to classify 428 patient profiles as having or not having MAS, based on clinical and laboratory features at the time of disease onset. The 428 profiles comprised 161 patients with systemic JIA-associated MAS and 267 patients with a condition that could potentially be confused with MAS (active systemic JIA without evidence of MAS, or systemic infection). Next, the ability of candidate criteria to classify individual patients as having MAS or not having MAS was assessed by evaluating the agreement between the classification yielded using the criteria and the consensus classification of the experts. The final criteria were selected in a consensus conference. Results Experts achieved consensus on the classification of 391 of the 428 patient profiles (91.4%). A total of 982 candidate criteria were tested statistically. The 37 best-performing criteria and 8 criteria obtained from the literature were evaluated at the consensus conference. During the conference, 82% consensus among experts was reached on the final MAS classification criteria. In validation analyses, these criteria had a sensitivity of 0.73 and a specificity of 0.99. Agreement between the classification (MAS or not MAS) obtained using the criteria and the original diagnosis made by the treating physician was high (κ = 0.76). Conclusion We have developed a set of classification criteria for MAS complicating systemic JIA and provided preliminary evidence of its validity. Use of these criteria will potentially improve understanding of MAS in systemic JIA and enhance efforts to discover effective therapies, by ensuring appropriate patient enrollment in studies.