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Background: Thymic stromal lymphopoietin (TSLP), an epithelium-derived pro-inflammatory cytokine, activates distinct immune and non-immune cells. It has been shown to be a master regulator of type 2 immune responses. Limited information is available on TSLP in childhood asthma. The aim of the present study was to find out whether there is association between TSLP concentrations and asthma phenotypes or disease activity. Methods: A total of 207 children with asthma and 100 healthy children aged 1–13 years were enrolled. This study examined serum TSLP concentrations using ELISA Kit in asthma patients and controls, analyzed its correlation with asthma phenotypes and pulmonary function. We also examined TSLP concentrations in 23 patients during stable asthma and in acute asthma exacerbation. Results: The serum concentrations of TSLP were significantly elevated in asthma patients compared with healthy controls (p < 0.05), but there was no significant difference (p > 0.05) in TSLP concentrations between three different asthma phenotypes (allergic asthma, virus induced asthma and nonallergic asthma). There was no significant correlation between TSLP concentrations and FEV1pred% (r = 0.01, p > 0.05). In the acute asthma exacerbation TSLP concentrations were not significantly different than in stable phase of disease (p > 0.05). Conclusion: Children with asthma have higher serum TSLP concentrations when compared to healthy controls. TSLP does not seem to be a biomarker of disease exacerbation in children. Different asthma phenotypes have similar TSLP concentration profile in peripheral blood and TSLP does not seem to be useful biomarker in asthma phenotyping in children.

Although self-limiting viral infections are the principal cause of fever in the majority of cases, it is estimated that 5% of febrile children younger than three years of age would have a urinary tract infection (UTI). Despite the increased availability of laboratory testing and development of various clinical scores, there is still no reliable predictor of UTI in children. Recent advancements in the understanding of inflammatory process have highlighted the role of S100 proteins, with many studies suggesting their clinical value as a biomarker of inflammation. This research aims to investigate the possible role of serum calprotectin (sCAL), a heterodimer consisting of S100A8 and S100A9 proteins from S100 family, as a biomarker for bacterial urinary tract infection in children. Patients aged 0-36 months who came to Pediatric Emergency Department of Sestre milosrdnice University Hospital Center with fever ≥38°C, lasting

Juvenile idiopathic arthritis (JIA) is characterized by chronic joint inflammation lasting longer than six weeks, as opposed to the acute reaction in reactive arthritis (ReA) that develops in response to an infection, lasts shorter and usually ends with full resolution of symptoms, indicating different mechanisms responsible for those common forms of arthritis in children. A number of previous studies have indicated the critical role of adaptive immune system cells in the development of many immune- mediated diseases (IMD), including arthritis. Therefore, the objective of this study was to examine the differences in occurrence of various subsets of lymphoid cells in JIA and ReA patients: regulatory T (Treg) and regulatory B (Breg) cells as immunosuppressors, type 3 innate lymphoid cells (ILC3) associated with a wide range of inflammatory disorders by an increase in IL-17 producing T cells and Th17 cells that exhibit plasticity and can be shifted to produce IFN-g. Treg cells, Breg cells, ILC3 and Th17 derived Th1 cells were analyzed in whole blood of ten JIA and six ReA patients by flow cytometry, using directly conjugated monoclonal antibodies. The blood samples were collected during the first visit to Pediatric Rheumatology Clinic in Sestre milosrdnice University Hospital Center in Zagreb, Croatia, while the final diagnosis of JIA or ReA was made three months after. At each visit, juvenile arthritis disease activity score (JADAS- CRP) for each patient was calculated. The median ages of the JIA and ReA patients were 6.41 and 7.22, respectively. In patients with JIA, the CD3+CD45+CD25+CD4 +CD127-CD28- subpopulation of Treg cells was significantly abundant compared to ReA patients (P=0.04). No other significant differences in cell subpopulations among different patient groups were observed. Although Treg frequencies account for only ~5% of the total CD4+ T-cell population, they have a massive role in the immune response. Particularly, CD28- Treg cells are characterized by reduced suppression of effector T cells yielding a pro-inflammatory cytokine profile characteristic for JIA. Besides, they can be generated in vitro by stimulation of CD28+ Tregs with TNF-a, which is raised in JIA. Therefore, the increased occurrence of these cells in JIA patients found in our proof-of-concept study could partially explain the failure of the immunosuppressive mechanisms and the development of the chronic disease in JIA patients. This observation could have a broader clinical significance after confirmation in larger patient cohorts with multiple time points, considering Treg cells have already been shown as a novel therapeutic target in some IMDs

Juvenile fibromyalgia is a pain amplification syndrome characterized by diffuse idiopathic musculoskeletal pain, most frequently affecting children older than 10 years. Girls are more commonly affected than boys, with the ratio of 1: 4 – 8. American College of Rheumatology criteria for diagnosis include absence of other disorder that could explain the pain, symptoms lasting for ≥ 3 months, widespread pain index (WPI) ≥ 7 and symptom severity score (SSS) ≥ 5 or WPI 3 – 6 and SSS ≥ 9. We present a case of 15 years old girl with diffuse and persistent pain along with episodes of impaired consciousness. Those episodes occurred once a month for the last 3 years and were characterized by collapse and/or inability to move or speak for several minutes. They were preceded by paraesthesia of face, arms and feet, but have never resulted in injury. In addition, patient complained of pain in radiocarpal joints, hips, low back, groins and neck, along with extreme tiredness, poor sleeping and inability to concentrate. Extensive examination was undertaken. The neurological assessment included brain MRI and EEG, both of which were normal. The rheumatologic evaluation revealed multiple painful points with WPI 9 and SSS 8. The patient didn’t experience morning stiffness, joint swelling or fever. Extensive laboratory workup and MRI of sacroiliac joints along with a lumbar and thoracic portion of the spine excluded the inflammatory aetiology. The psychological assessment revealed poor stress coping strategies. The patient was diagnosed with fibromyalgia and conversion disorder manifested in the form of pseudoseizures. Physical therapy and psychotherapy were advised, along with melatonin for better sleep regulation, which led to a mild improvement of symptoms within a month. Fibromyalgia and diverse psychological distress are causally related. Accordingly, conversion symptoms are not uncommon. Therefore, a multidisciplinary team approach is necessary when caring for these patients, in addition to providing treatment not only for the affected individual but for the entire family as well.

Although self-limiting viral infections are the principal cause of fever in the majority of cases, it is estimated that 5% of febrile children younger than three years of age would have a urinary tract infection (UTI). Despite the increased availability of laboratory testing and development of various clinical scores, there is still no reliable predictor of UTI in children. Recent advancements in the understanding of inflammatory process have highlighted the role of S100 proteins, with many studies suggesting their clinical value as a biomarker of inflammation. This research aims to investigate the possible role of serum calprotectin (sCAL), a heterodimer consisting of S100A8 and S100A9 proteins from S100 family, as a biomarker for bacterial urinary tract infection in children. Patients aged 0-36 months who came to Pediatric Emergency Department of Sestre milosrdnice University Hospital Center with fever ≥38°C, lasting

Fever is the most common cause for which children and their caregivers seek medical attention. If accompanied by elevated blood inflammatory markers, infectious diseases are suspected. However, if microbiological findings are negative, and if there is no response to antimicrobial therapy, more careful and comprehensive evaluation is required. Amongst many disorders, one of the rare but often misdiagnosed cause is Castleman disease (CD). This is a heterogeneous group of lymphoproliferative disorders with similar histopathologic features divided into three types. Unicentric CD involves one or more enlarged lymph node(s) in a single region of a body while multicentric Castleman disease involves multiple regions of lymphadenopathy and is further subclassified in HHV-8 positive and HHV-8 negative/idiopathic type. We present a case of a 4.5-year-old boy who came to our pediatric emergency department with a history of intermittent fever for almost a month, cough, skin rash with spontaneous regression, fatigue, periodic leg pain, night sweats, and weight loss. The physical examination showed few mildly enlarged cervical lymph nodes, pale skin, antalgic gait and fever. Routine blood tests revealed elevated C- reactive protein level and erythrocyte sedimentation rate, anemia, and thrombocytosis. He was admitted to a Pediatric Rheumatology department for evaluation. Other laboratory tests showed elevated fibrinogen, ferritin and d-dimer levels, with high C3, IL-6 and TNF-alpha levels. Microbial cultures (blood cultures, nasopharyngeal and throat swab) were sterile, and serology for Cytomegalovirus, Epstein-Barr virus, Human parvovirus B19 and Bartonella henselae were negative, while serology for Mycoplasma pneumoniae (IgM and IgG) was positive. Azithromycin was prescribed, but fever and elevated inflammatory blood markers persisted and other procedures were performed to rule out rheumatic disease. Radiogram and bone scintigraphy were unremarkable, but an ultrasound revealed a retroperitoneal mass. MRI was performed, and retroperitoneal mass measuring 3x2.5 cm, arising from the pancreas and suspected to be a neuroendocrine tumor or enlarged lymph node, along with hepatosplenomegaly was described. FDG-positron emission tomography (PET)/CT showed a mass near the pancreas with moderately increased FDG uptake. Complete surgical resection of the mass was performed, and the histopathological evaluation revealed lymphoid hyperplasia consistent with the hyaline vascular type of Castleman disease. An immunohistochemical study excluded HHV-8 positivity. After the surgical procedure, the patient recovered completely. On follow up visits he was without fever or other symptoms suggestive of the disease, with normal laboratory findings. CD with systemic manifestations is rare in children emphasizing the need for thorough evaluation of unexplained prolonged symptoms.

Juvenile spondyloarthritis (jSpA) represents a spectrum of inflammatory arthritis with strong HLA-B27 association and involvement of enthesis and/or axial skeleton that appears in children and young adults. ILAR classification criteria for enthesitis related arthritis subtype of juvenile idiopathic arthritis, which is undifferentiated form of jSpA, includes arthritis, enthesitis, presence of sacroiliac joint tenderness, inflammatory lumbosacral pain, HLA-B27 positivity, positive family history and acute anterior uveitis. We present a case of 16 years old girl diagnosed with juvenile spondyloarthritis that first presented with recurrent monthly swelling of index finger at the age of 14. Symptoms progressed to low back pain in the morning which partly declined with activity. Additionally, her right knee was swollen and painful. She was examined on multiple occasions by pediatric orthopedic surgeon and in pediatric emergency department before seeing a pediatric rheumatologist. The first examination revealed sacroiliac joint tenderness with positive FABER test and abnormal modified Schober test. Family history was negative for rheumatic diseases and there were no signs of uveitis nor enthesitis, with ANA, RF and extensive laboratory workup being either negative or within reference range. However, HLA-B27 turned positive and MRI showed right sacroiliitis. NSAID was prescribed but symptoms nevertheless persisted. Thus, after the exclusion of TBC with quantiferon test, intraarticular corticosteroid injection was applied to the right knee and oral corticosteroid was introduced as Abstracts Arch Dis Child 2021 ; 106(Suppl 2):A1–A218 A187 copyright. on February 11, 2023 by guest. Protected by http://adc.bmj.com/ Arch Dis Child: first published as 10.1136/archdischild-2021- europaediatrics.448 on 11 October 2021. Downloaded from bridging therapy. Subsequently, according to ACR guidelines, a therapy with TNF-alpha inhibitor, adalimumab, was initiated. Within a month, a substantial reduction of the low back pain was noted, with a decrease in juvenile spondyloarthritis disease activity score (jSpADA). Low back pain often begins in childhood, with the prevalence in adolescence being similar to that in adulthood. Among many possible causes, inflammatory etiology should be thoroughly considered due to irreversible damage if not treated adequately. Therefore, children and adolescents complaining of a back pain, especially in the presence of other signs of arthritis, should be referred to pediatric rheumatologist for a further work-up and treatment.

The perplexing interactions between an organism and it’s gut commensal microbiota have both potentiating and detrimental effects on innate and adaptive immunity and consequentially the development of several immune-mediated diseases. Juvenile idiopathic (JIA) and reactive arthritis (ReA) are two adverse outcomes of such disorder in children. This study aimed to assess the differences in the presence of subtypes of Escherichia coli (E coli), one of the most abundant bacteria in the microbiota, in the stool of JIA and ReA patients. Stool samples of 14 patients with joint swelling were collected during their first visit to Pediatric Rheumatology Clinic in Sestre milosrdnice University Hospital Center in Zagreb, Croatia. Three months later, the diagnosis of JIA was made in seven patients, while the others were classified as ReA. The samples were analyzed by mass spectrometry on nanoLC- Synapt G2 Si instrument. To identify the most abundant E coli subtypes, specialized software named Protein Reader with implemented Dust algorithm have searched through the NCBI nr database, that contains the records of more than 400 E coli subtypes. The median age of patients was 7.14 and 7.11, respectively. Various E coli subtypes (P0301867.1-10, O104:H4, O103:H25, O111:H11, KTE and K) were three times more abundant in patients with JIA, while in children with ReA, the abundance of diarrheagenic E. coli (DEC) was detected. Many studies have speculated the influence of gut microbiota in the development of arthritis in children. Despite the technological advancements in the examination of microbiota composition, there are still many limitations imposed by patient selection, methodology and data analysis. Besides, without the proper definition of ‘healthy’ microbiome as a reference standard, it is challenging to distinguish alterations responsible for diseases development. In our proof-of-the-concept study microbiota was therefore compared in two groups of patients both presenting with arthritis. Since E coli is one of the paramount bacteria in gut microbiota with more than 600 recognized subtypes, it is reasonable to assume that described differences can have a potential impact on the gut environment, with the contribution to the development of the chronic disease in JIA patients or the resolution of symptoms in children with ReA. While this observation needs conformation in multiple time points and larger patient cohorts, it pinpoints gut microbiota as a potential new therapeutic target in the treatment of chronic inflammatory diseases.

Uvod: Entezitisu pridruženi artritis (ErA) posebni je oblik juvenilnog idiopatskog artritisa (JIA) koji predstavlja nediferencirani juvenilni spondiloartritis (jSpA). Uz artritis perifernih zglobova, jedna od najvažnijih značajki jSpA je upala enteza i/ili aksijalnog skeleta. I dok se u odraslih bolesnika sa SpA crijevo sve češće navodi kao polazište i/ ili ishodište subkliničke upale, učestalost i raširenost upalnog procesa u crijevima djece s ErA još uvijek nije poznata, uglavnom zbog izazova vezanih uz provođenje invazivnih pretraga neophodnih za otkrivanje upale crijeva u djece. Cilj našeg istraživanja stoga je bio procijeniti koncentraciju fekalnog kalprotektina (fCAL), surogatnog markera upale crijeva, u djece s različitim oblicima artritisa. Ispitanici i metode: U ovom presječnom istraživanju sudjelovalo je ukupno 71 dijete s postavljenom dijagnozom oligo i poliartikularnog oblika JIA (N=29), ErA (N=26), PsA (N=4) te neupalnim muskuloskeltnim stanjima (N=12). Svi bolesnici s ErA uključeni su neovisno o prisutnosti gastrointestinalnih simptoma, dok su ostali uključeni u slučaju izraženih simptoma. Uz standardne laboratorijske pretrage, svim sudionicima izmjeren je fCAL, a svim bolesnicima s ErA, kao i ostalima koji su se žalili na upalnu bol u leđima, učinjen je MR SIZ- ova. U svih bolesnika s ErA izmjerena je aktivnost bolesti pomoću jSpADA, a u ostalih oligo i poli JIA bolesnika pomoću JADAS-a. Rezultati: Medijan koncentracije fCAL bio je najviši u skupini bolesnika s ErA (33.2 mg/kg, p=0.043), s značajnom razlikom između bolesnika s aktivnom i inaktivnom bolesti (20.0 mg/kg vs 57.4 mg/kg, p=0.01), kao i između bolesnika s prisutnim znakovima upale te bolesnika s urednim nalazom MR-a (22.6 mg/kg vs 54.3 mg/ kg, p=0.048). Koncentracije fCAL nisu se značajno razlikovale između bolesnika koji su koristili i koji nisu koristili NSAIL-ove (23 mg/kg vs 20 mg/kg, p=0.18), mada je uočena slaba koleracija s trajanjem korištenja (r=0.25, p=0.03). Zaključak: Subklinička upala crijeva nije vezana samo uz odrasle bolesnike sa SpA, već i uz djecu s nediferenciranom oblikom jSpA, osobito ako je uz to prisutna aktivna bolest i/ili znakovi upale SIZ-ova na MR-u.

Bolni otok zgloba može biti simptom mnogih bolesti u djece, među kojima se ističu dvije vrlo različite imunološki posredovane bolesti: reaktivni artritis (ReA) i juvenilni idiopatski artritis (JIA). Za razliku od ReA-a koji traje kraće i nakon kojeg dolazi do potpunog nestanka simptoma, u bolesnika s JIA-om dolazi do razvoja kronične bolesti koja može dovesti do trajnog oštećenja zglobova. Brojna istraživanja dala su naslutiti da crijevna mikrobiota ima vrlo važnu ulogu u oblikovanju imunološkog odgovora, zbog čega bi razlog različitom odgovoru na poznati ili nepoznati patogen u bolesnika s ReA-om i JIA-om mogao ležati u različitom sastavu mikrobiote. Cilj ovog istraživanja stoga je bio ispitati razliku u zastupljenosti podvrsta Escherichiae coli (E coli), jedne od najzastupljenijih bakterija unutar crijevne mikrobiote, prilikom prve pojave simptoma juvenilnog idiopatskog i reaktivnog artritisa. Ispitanici i metode. Uzorci stolice 14 bolesnika prikupljeni su prilikom prvog posjeta Zavodu za reumatologiju i imunologiju Kliničkog bolničkog centra Sestre milosrdnice u Zagrebu, a konačna dijagnoza bolesti postavljena je 22 Reumatizam 2019 ; 66(Suppl 1):15–30 Sažeci / Abstracts Usmena priopćenja / Oral communications tri mjeseca kasnije: u sedmero bolesnika postavljena je dijagnoza JIA-a, a u drugih sedmero ReA-a. Svi uzorci su analizirani masenom spektrometrijom na nanoLC-Synapt G2 Si instrumentu u Institutu Ruđer Bošković. Kako bi se identificirale najučestalije vrste E coli korišten je posebni računalni program, Protein Reader s ugrađenim Dust algoritmom koji je pretraživao NCBI nr bazu podataka koja sadrži zapise o više od 400 vrsta E coli. Medijan dobi bolesnika oboljelih od JIA-a bio je 7, 14, a bolesnika oboljelih od ReA-a 7, 11. Rezultati. Pojedine podvrste E coli (P0301867.1- 10, O104:H4, O103:H25, O111:H11, KTE and K) bile su tri puta učestalije u bolesnika s JIA-om, dok je u bolesnika s ReAom uočeno dvostruko više podvrsta E coli koje uzrokuju infektivne proljeve (DEC). Zaključak. Rezultati ovog istraživanja ukazali su na razliku u podvrstama E coli u stolici bolesnika s ReA-om i JIA-om u samim počecima bolesti. Budući da je E coli jedna od ključnih bakterija u crijevnoj mikrobioti, s više od 600 do sada prepoznatih podvrsta, razumno je za pretpostaviti kako uočene promjene mogu utjecati na rav notežu crijevne mikrobiote i oblikovanje imunosnog odgovora te pridonijeti razvoju kronične bolesti ili potpune regresije simptoma.

Primarni Sjögrenov sindrom (pSS) je kronična autoimuna bolest koju karakterizira limfocitna infiltracija egzokrinih žlijezda što se očituje prvenstveno tzv. „sicca“ simptomima kseroftalmije i kserostomije te mogućom sistemnom zahvaćenosti sa serološkim nalazom pozitivnih antinuklearnih (ANA), anti-SSA i anti- SSB antitijela. U literaturi je objavljeno tek oko 250 slučajeva pSS u djece. Javlja se češće u djevojčica (83%) i tada uglavnom zahvaća parotidne žlijezde (53%) uz znakove sistemne bolesti (npr. febriliet, limfadenopatija) dok su sicca simptomi rjeđe izraženi (34%). Specifična protutijela mogu biti negativna u 30- 50% djece. Ekstraglandularna zahvaćenost bubrega u pSS u djece još je rjeđa (9, 9 %) a najčešće se radi o tubulointersticijskom nefritisu s mogućim razvojem renalne tubularne acidoze. Prikazujemo 12-godišnju djevojčicu s opterećenom obiteljskom anamnezom sistemnog eritematoznog lupusa kod majke koja je zaprimljena na obradu prethodno dijagnosticiranog sicca sindroma sa izraženim simptomima 2 godine prije postavljanja dijagnoze, prvenstveno se očitujući recidivirajućim epizodama parotitisa, a tek naknadno manjkom suza i suhoćom usta. Schimmerovim testom dokazana je hipolakrimacija, a u serološkom nalazu pozitivna antitijela (ANA > 1:1280, RF 762 U/mL, Anti-SS-A (Ro60) 122 U/mL, Anti-SS-A (Ro52) 176 U/ mL. Biopsijom žlijezde slinovnice usnice potvrđena je dijagnoza. Dodatnom obradom se verificira značajna perzistentna proteinurija u 24-satnom urinu (247 mg/24h) uz urednu globalnu bubrežnu funkciju te akutni intersticijski nefritis u nalazu biopsije bubrega. Provedena je pulsna kortikosteroidna terapija i započeta terapija hidroksiklorokinom na što dolazi do normalizacije nalaza proteinurije bez drugih znakova bubrežne bolesti. Dodatnom obradom utvrđena je i zahvaćenost štitnjače (T3 , 9 nmol/L, TSH 4, 070 mIU/L, Anti Tg 459 kiU/L) te prisustvo antileukocitih i antitrombocitnih protutijela. Zaključno, u dječjoj populaciji za sada ne postoje jedinstveni dijagnostički kriteriji za primarni Sjögrenov sindrom koji u početku često ostaje neprepoznat ne zadovoljavajući klasifikacijske AECG kriterije za odraslu populaciju, dok se ranim prepoznavanjem latentne zahvaćenosti bubrega i odgovarajućim imunosupresivnim liječenjem može smanjiti progresivno oštećenje bubrežne funkcije i druga sistemna oštećenja

Juvenilni idiopatski artritis (JIA) najčešća je kronična reumatska bolest dječje dobi. Međunarodna klasifikacija dijeli ga u 7 podvrsta, a poliartikularni oblik čini 15-25% i karakteriziran je težim tijekom i slabijim odgovorom na liječenje. Takvi su bolesnici izloženi većem riziku ranijeg razvoja trajnih oštećenja, nuspojava liječenja i lošijoj kvaliteti života. Velik su izazov za pedijatrijskog reumatologa, ali uz razvoj novih terapijskih mogućnosti ne i nepremostiv. Prikazujemo slučaj 15-godišnje pacijentice čija je dijagnoza poliartikularnog oblika JIA (RF i ANA negativni) postavljena u 3. godini života. U trenutku dijagnoze bolešću je bilo zahvaćeno više od 10 zglobova, a u dobi od 5 godina, unatoč liječenju pulsnim dozama glukokortikoida te metotreksatom uz održavajuće doze prednizona, dolazi do razvoja kontraktura. Od 2008. do 2014. godine pacijentica je podvrgnuta biološkim lijekovima (infliksimab, adalimumab, tocilizumab i etanercept) bez dugoročne učinkovitosti, uz stalnu potrebu za sistemnom, a često i intraartikularnom primjenom glukokortikoida koji dovode do razvoja teških nuspojava. Tijekom 2016. godine provedeno je liječenje rituksimabom i ciklofosfamidom prema protokolu. Unatoč svemu, bolest je trajno aktivna te se u lipnju 2018. godine u terapiju uvodi inhibitor JAK-STAT signalnog puta, tofacitinib. Postupno dolazi do smirivanja simptoma, poboljšanja kliničkog statusa, laboratorijskih i slikovnih nalaza te su ukinuti metotreksat i prednizon. Na kontrolnoj je obradi radiografski obostrano uočena Madelungova deformacija uz izrazito nizak rast što pobuđuje sumnju na Leri- Weill-ov sindrom (delecija u regiji SHOX) koji je sredinom 2019. godine potvrđen genetskom analizom, što djelomično objašnjava izrazito nizak rast i perzistentne bolove u ručnim zglobovima. Pacijentica je trenutno u stadiju remisije uz terapiju tofacitinibom, ibuprofenom i bisfosfonatima uz suplementaciju vitamina D. Zaključno, liječenje poliartikularnog oblika JIA refrakternog na lijekove predstavlja veliki izazov, posebice u bolesnika ovisnih o glukokortikoidima koji u dječjoj dobi izazivaju izraženije nuspojave i imaju negativan učinak na rast i razvoj. Jedna od mogućnosti liječenja uključuje inhibitore JAK-STAT signalnog puta koji su se u studijama, ali i pojedinačnim ili manjim serijama bolesnika pokazali učinkovitima.

INTRODUCTION: Painful joint swelling is a symptom of many childhood diseases, most notably reactive arthritis (ReA) and juvenile idiopathic arthritis (JIA). The growing body of evidence suggests that gut microbiota could orchestrate the immune response and development of those diseases. This study aimed to assess the differences in the presence of Escherichia coli (E coli) subtypes in the stool of JIA and ReA patients at the first occurrence of symptoms. METHODS: Stool samples of 14 patients with joint swelling were collected during their first visit to Pediatric Rheumatology Clinic in Sestre milosrdnice University Hospital Center in Zagreb, Croatia. The samples were analyzed by mass spectrometry on nanoLC-Synapt G2 Si instrument. To identify the most abundant E coli subtypes, specialized software named Protein Reader with implemented Dust algorithm searched through NCBI nr database. RESULTS: Various E coli subtypes (P0301867.1-10, O104:H4, O103:H25, O111:H11, KTE and K) were three times more abundant in patients with JIA, while increased abundance of diarrheagenic E. coli (DEC) was detected in children with ReA. CONCLUSION: This pilot study has shown differences in the subtypes of E. coli present in the stool of children with ReA and JIA in the early stage of the disease. Since E coli is one of the paramount bacteria in gut microbiota, it is reasonable to assume that the differences described in this study can have a potential impact on the gut environment, contributing to the development of the chronic disease in JIA patients, or the resolution of symptoms in children with ReA.

Comparison of various lymphoid cells in patients with juvenile idiopathic and reactive arthritis showed difference in Treg cells subpopulation

The variance of different subtypes of Escherichia coli in stool of children with juvenile idiopathic or reactive arthritis at the first occurance of symptom

Introduction: Rheumatic diseases are one of the most common chronic diseases in childhood. Although novel therapies are emerging, there is still a significant number of patients not reaching remission easily. That is why many parents/legal guardians turn to complementary and alternative medicine (CAM), usually not revealing it to their physicians. Even though valid tools for assessing CAM use exist, such as questionnaires, they are not routinely used probably due to their relative complexity. Objectives: To assess the prevalence and patterns of CAM use and variables associated with it in children with chronic rheumatic diseases in a single center pediatric rheumatology clinic. Methods: We conducted the cross-sectional study from April to May 2017 at the Department of Clinical Immunology and Rheumatology, Sestre Milosrdnice University Hospital Center, Zagreb, Croatia, on the consecutive sample of parents/legal guardians of the children diagnosed with chronic rheumatic diseases. The outcome was the self-reported frequency of CAM use. For the purposes of our study, we developed an anonymous questionnaire to answer our study questions and written informed consent was obtained from all the parents/legal guardians prior to enrollment in the study. Results: Out of the 100 children whose parents/legal guardians were interviewed, 67 (67%) had juvenile idiopathic arthritis (JIA) while other diagnosis included systemic lupus erythematosus, juvenile scleroderma, juvenile dermatomyositis, etc. 63 (63% ; CI 95% 54% −73%) of parents/legal guardians have used CAM in the treatment of their child. Out of those, 43/63 parents/legal guardians (68, 25%) did not inform any medical staff about it. The majority of them, 55/63 (87% ; CI 95% 77%−94%) used it concomitantly to their standard therapy and 40/63 (64% ; CI 95% 50%−75%) declared their intent to use it in the future as well. Only 6/63 (10% ; CI 95% 4%−20%) were sure they would no longer use it. Out of the listed CAM groups, the most commonly used were herbal remedies (30.49%) along with orthomolecular therapy (25%) such as vitamins, minerals, immune boosting products and probiotics. Statistical analyses of collected data showed that a significantly higher frequency of CAM use was associated with parents/legal guardians with lower monthly income (OR 9.66 ; p

Juvenile spondyloarthritis (jSpA) is a complex disease with both genetic and environmental factors contributing to the etiology. Recently obtained gene signatures in jSpA patients revealed TLR4 and CXCR4 gene had increased, while NLRP3 and PTPN12 had decreased expression, though the mechanism(s) responsible for those alterations remained unknown. Since it is well known that epigenetic mechanisms control gene expression and are influenced by external stimuli linking environment and gene function, the aim of the present study was to investigate the possible mechanistic role of DNA promoter region methylation and several non-coding microRNAs (miR- 150, miR-146a, miR-181a, miR-223) in jSpA patients related to the expression of genes with observed alterations (TLR4, CXCR4, NLRP3 and PTPN12). For that purpose, peripheral blood samples were obtained from 19 patients diagnosed with jSpA according to ILAR classification criteria for enthesitis related arthritis (ErA) and seven gender and age matched subjects without diagnosis of inflammatory disease. Methylated DNA Immunoprecipitation (MeDIP) analysis revealed significant hypermethylation of NLRP3 gene promotor while expression analysis of selected microRNAs showed no significant difference in fold change between two groups of participants. Our study therefore indicated epigenetic modifications are probably responsible for some of the expression alterations in jSpA patients in the initial phase of the disease. Since NLRP3 has a crucial role in inflammasome assembly and infllammasomes have been shown to shape microbiota, it is reasonable to assume that dysbiosis in jSpA patients can at least partially be explained by reduced NLRP3 expression due to hypermethylation, stressing for the first time the epigenetic contribution to jSpA pathophysiology. While it is still not clear if these epigenetic alterations are caused by genetic mutations in epigenetic factors or exposure to certain environmental factors that mediate the occurrence of aberrant epigenetic profiles, the possibility of reverting epigenetic modifications opens up whole new prospects for therapeutic treatment of these intricate disease.

Objectives: Juvenile idiopathic arthritis (JIA) is the most common chronic rheumatic disease in children, with heterogeneous clinical features. Although therapeutic options are wide and in majority of children symptoms improve with combination of nonsteroidal anti- inflammatory and disease modifying drugs, there are a number of patients who are not responding to conventional therapy and who do not meet criteria for systemic biologics, namely anti TNF alpha. Those patients are potential candidates for intraarticular therapy with biologics aind in this report we present the results of intra-articular infliximab treatment in a series of patients diagnosed with oligoarthritis subtype of JIA. Methods: Twenty patients (30 joints) were treated with intraarticular infliximab and monitored by power Doppler musculoskeletal ultrasound according to OMERACT and juvenile arthritis disease activity score (JADAS 10) before intraarticular application and during the follow up period of 18 months (0, 1, 12, 18 months). Results: The results showed statistically significant improvement in PD-MSUS measures and JADAS in both B mode and power Doppler mode scores (p

Introduction: in the past few years, various indicators of disease activity in juvenile idiopathic arthritis (JIA) patients have been investigated, allowing for the interpretation of different aspects of underlying inflammatory process (e.g. clinical, immunological, radiological etc.). Nevertheless, there is still evident lack of satisfactory support in diagnostic and prognostic evaluations in everyday clinical practice, focusing research activities on discovery of adequate biomarker that would support initial diagnosis and allow feasible disease monitoring. While growing number of evidence indicates phagocytic S100 proteins, like MRP8/14 complex (i.e. calprotectin), can be used to detect subclinical inflammatory activity in systemic JIA, their performance in other forms of JIA is still debatable, while in acute infectious diseases it is almost unknown. Objectives: compare serum calprotectin (sCAL), CRP and PCT levels in patients with various forms of non-systemic JIA and previously healthy children with acute febrile illness lasting

Background: Juvenile spondyloarthritis (jSpA) is a diverse group of related syndromes with shared symptoms and pathogenic mechanisms in which both extrinsic environmental factors and intrinsic genetic background perpetuate inflammatory response through immune system alterations. Recently obtained gene signatures in jSpA patients revealed TLR4 and CXCR4 gene had increased, while NLRP3 and PTPN12 had decreased expression.1 Although gene expression is regulated by various mechanisms, the increasing numbers of studies is showing the importance of epigenetic mechanisms in this fundamental biological process. Objectives: To investigate the possible mechanistic role of DNA promoter region methylation and several non-coding micro RNA (miR-150, miR-146a, miR-181a, miR-223) in jSpA patients regarding the expression of genes with previously observed alterations. Methods: The expression of specific microRNAs was analysed in 8 jSpA patients and 5 matched controls using RT-PCR with predeveloped microRNA assays. Methylated DNA Immunoprecipitation (MeDIP) was performed in 19 patients and 7 controls. Enrichment in MeDIP fraction was determined by qRT-PCR using the AriaMx. Results: The difference in fold enrichment of immunoprecipitated DNA was significant only for NLRP3 promotor site (p=0.0220). Expression analysis of selected miRs showed no significant difference in fold change between jSpA patients and healthy controls. Conclusions: Our study indicated epigenetic modifications are probably responsible for some of the expression alterations in jSpA patients in the initial phase of the disease. Since NLRP3 has a crucial role in inflammasome assembly and inflammasomes have been shown to shape microbiota, it is reasonable to assume dysbiosis in jSpA patients can at least partially be explained by reduced NLRP3 expression due to hypermethylation, stressing for the first time the epigenetic contribution to jSpA pathophysiology. While it is still not clear if these epigenetic alterations are caused by genetic mutations in epigenetic factors or exposure to certain environmental factors that mediate the occurrence of aberrant epigenetic profiles, the disocvery of DNA methylation-based signature of the NLRP3 gene could have important implications in addressing extrinsic and intrinsic contribution to jSpA pathophysiology, whereas the possibility of reverting epigenetic modifications opens whole new prospects for therapeutic treatment of this complex disease.

Uvod Klavikularna kortikalna hiperostoza (CCH) rijetka je bolest nepoznate etiologije i ishoda, klinički karakterizirana bolnošću i/ili oticanjem klavikule. Ispitivanje genskoga izražaja na sitnopolju u bolesnika s CCH pokazalo je razlike za 974 gena uključena u različite upalne procese, dok je qRT-PCR analiza potvrdila pojačan izraža TRPM3 i TRPM7 gena te niži izražaj ERBB2. K tome, imunofl uorescentna mikroskopija pokazala je značajno pojačan signal TRPM3 proteina u krvnim stanicama bolesnika. Kako bi se utvrdilo da li postoje varijante gena koje bi mogle uzrokovati bolest, potrebno je provesti i sekvencioniranje cjelokupnog egzoma (WES). Ispitanici i metode Genomska DNA izolirana je iz krvi tri bolesnika s CCH. Sekvencioniranje egzoma učinjeno je pomoću Nextra Rapid Capture Exome Kit na HiSeq 2000 sekvencioneru sljedeće generacije. Dobivene varijante su obilježene i fi ltrirane pomoću programa Variant Studio i Variant Interpreter. Rezultati WES analizom u bolesnika s CCH otkriveno je 428 zajedničkih identičnih varijanti, od kojih 30 nije povezano s genom, 121 je u ZNF717 genu i 277 je raspoređeno u 63 druga gena. Jedna heterozigotna varijanta u CTBP2, jedna u HYDIN i šest u ZNF717 genu klasifi cirane su kao vjerojatno patogene. Zaključak WES analiza pokazala je da se većina vjerojatno patogenih varijanti nalazi u ZNF717 genu. Navedeni gen kodira Kruppel-associated box (KRAB) zinc-fi nger protein koji spada u veliku grupu transkripcijskih regulatora te je stoga uključen u regulaciju krucijalnih fi zioloških i patoloških procesa. Mnogi aspekti navedenih proteina, poput uloge u upalnom procesu, nisu još poznati, no smatra se kako pojačavaju dostupnost DNK i transkripciju proupalnih gena. Ako još k tome dodamo i mogući utjecaj na opažene promjene u izražaju gena, svi navedeni procesi mogli bi doprinijeti razvoju CCH.

Clavicular cortical hyperostosis (CCH) is a rare inflammatory bone disorder of unknown aetiology represented by pain and swelling of the clavicle. Although similar to chronic nonbacetril/recurrent multifocal osteomyelitis (CNO/CRMO), due to lack of recurrence and additional inflammatory sites, it can be considered separate disease in the spectrum. In our previous study microarray gene expression profiling of CCH patients revealed 974 differentially expressed genes involved in various inflammatory processes. Subsequent qRT- PCR in additional number of patients confirmed significantly lower expression of ERBB2 gene and higher expression of TRPM3 and TPRM7 transient receptor potential (TRP) channel genes. Additionally, immunofluorescence microscopy showed high signal of TRPM3 in blood cells of one CCH patient. Based on described findings and other studies which showed TRP channels are involved in inflammasome activation while ERBB activation promotes protective cellular outcomes during inflammation, we hypothesized CCH could be autoinflammatory disease, but the mechanisms responsible for observed expression alterations remained unclear. Objectoive was to identify possible disease causing gene variants in CCH patients using whole exome sequencing (WES). Genomic DNA was extracted from peripheral whole blood samples of three CCH patients. Targeted exome sequencing was performed using the Nextera Rapid Capture Exome Kit (Illumina). The extracted variants were annotated and filtered using the Variant Studio software (Illumina) and Variant Interpreter (Illumina). WES analysis identified 428 shared identical variants among affected individuals. Thirty of these variants were not associated with a gene, 121 were in ZNF717 gene and 277 were distributed in 63 other genes. One heterozygous variant in CTBP2 gene, one in HYDIN gene and six in ZNF717 were classified as likely pathogenic. In this comprehensive study "bottom-up" approach was used in order to elucidate the molecular disease mechanisms of what we believe could be a new disease entity. Previously performed diligent transcriptome analysis and proof-of-concept experiment which confirmed the results on a protein level indicated CCH could be induced by sternoclavicular joint overuse, TRP channel overexpression, inflammasome activation and reduced protection during inflammation. Interestingly, WES analysis identified majority of likely pathogenic variants in ZNF717 gene. This gene encodes a Kruppel-associated box (KRAB) zinc-finger protein, which belongs to a large group of transcriptional regulators and is therefore involved in the regulation of crucial physiological and pathological processes. Many aspects of these proteins are still essentially unknown, as is their role in the inflammation, yet it is speculated they increase DNA accessibility and inflammatory gene transcription. Together with possible influence on observed expression patterns, all of these processes could contribute to CCH evolution.

Uvod Juvenilni spondiloartritis (jSpA) kompleksna je bolest koja se razvija pod utjecajem različitih genetskih i okolišnih čimbenika. Nedavno provedena analiza genskoga izražaja bolesnika s jSpA pokazala je pojačan izražaj TLR4 i CXCR4 te smanjen izražaj NLRP3 i PTPN12 gena, no mehanizmi odgovorni za opažene promjene ostali su nepoznati. Kako bi se otkrila moguća uloga epigenetskih modifi kacija na izražaj navedenih gena, ispitana je metilacija njihovih promotora. Ispitanici i metode Metilacija promotorskih regija ispitana je metodom metilirane DNK imunoprecipitacije (meDIP) u 19 novootkrivenih bolesnika u kojih je dijagnoza jSpA postavljena prema ILAR-ovim kriterijima za ErA-u te u sedmero zdrave djece. Rezultati Učinjena analiza pokazala je statistički značajnu hipermetilaciju promotorske regije NLRP3 gena (p=0.0220). Zaključak Produkt NLRP3 gena ima važnu ulogu u sastavljanju NLRP3 infl amasoma, senzora prirođene imunosti koji regulira „odgovor na opasnost” i sudjeluje u oblikovanju mikrobiote. Smanjen izražaj ovog gena dovodi se u vezu s poremećajem mikrobiote u upalnoj bolesti crijeva, što može dovesti do disbioze i dugotrajnih posljedica na cijeli organizam. Obzirom da navedena bolest ima mnogo sličnosti s jSpA, moguće je da hipermetilacijom uzrokovano smanjenje izražaja NLRP3 u bolesnika s jSpA u crijevima utječe na redukciju komenzalnih bakterija s protuupalnim svojstvima i posljedično stvaranje većih količina za jSpA karakterističnih citokina poput TNF-α. Tome zasigurno doprinosi i pojačan izražaj TLR4 i CXCR4 gena čiji su produkti odgovorni za prepoznavanje crijevnih mikroba te smanjen izražaj PTPN12 gena, produkt kojeg sudjeluje u regulaciji migracije stanica iz crijeva. Rezultati ovog opsežnog istraživanja pružaju nam bolji uvid u mehanizme odgovorne za razvoj jSpA, ali i moguće nove načine liječenja.

Introduction: Transition for patients with chronic diseases is defined as the purposeful, planned movement of adolescents and young adults from child-centered to adult oriented health care system.1 Rheumatic diseases of childhood extend to adulthood as active diseases in 30-70%, which is the reason for requiring rheumatologic care into adulthood. First coordinated transition of pediatric rheumatologic patients in Croatia was organized in 2012 as developmental type of transition, where patient is gradually prepared for the transition by a pediatrician rheumatologist throughout years preceding transfer. The moment of final transfer to adult care is organized in the pediatric rheumatology clinic, where patients and their parents are meeting adult rheumatologist in known environment (pediatric rheumatology ambulance). At the transfer, patient's history is being presented to the adult rheumatologist and afterwards patient is examined by both of the specialists. At the end of the examination, patient is given next checkup date at adult rheumatologist clinic, where the patient will continue with regular follow up visits. Objectives: Our objective is to verify successfulness of organized transition of pediatric rheumatologic patients to adult care. Methods: We have conducted a retrospective research of patients’ databases at Departments of Pediatric Rheumatology and correlated them with data from adult Clinical Immunology and Rheumatology Department in order to verify number of pediatric patients that have continued with regular follow up in adult care after coordinated transition. Results: During 5 year period (2012.-2017.) 78 patients have been transferred to adult rheumatologist. There were 50 JIA patients, 18 SLE patients, and 10 patients with other rheumatic diseases (8 patients with mixed connective tissue disease, 1 with Wegener's granulomatosis, 1 with fever of unknown origin). Median age at the time of transfer was 19, 6 years (14, 6-26, 6). Patients with SLE were older at the time of transfer than two other groups of patients (SLE 21, 29 ± Pediatric Rheumatology 2017, 15(Suppl 2):64 Page 71 of 191 2, 99 vs. JIA 18, 83 ± 1, 22 vs. other diagnosis 19, 42 ± 2, 8, p < 0, 01). Total of 63 patients (80, 8%) have continued with regular follow up at adult rheumatologist and 15 patients never showed up at adult rheumatologist clinic, probably due to remission of the disease or possible follow up by other chosen adult rheumatologist. Conclusion: Young adults with rheumatic diseases have complex medical and psychological needs and transition to adult care is a critical component of care. In order to maintain adequate level of follow up and prevent additional flares in turbulent adolescent period, continuity of rheumatologic care is essential. By organizing coordinated transition to adult rheumatologist, a most of the pediatric patients with rheumatic disease (80, 8%) are continuing with regular follow up in the adult care. Each transition should be an individually tailored process according to needs and psychological maturity of patients.

Low back pain (LBP) is very common complaint in adults which often begins in childhood. Despite the increasing frequency, it is estimated only 24% of children with LBP visit a doctor. Nevertheless, those patients are very often seen in pediatric rheumatology outpatient clinic. Although most of the cases of LBP after exclusion of trauma are caused by benign musculoskeletal disease, a history of sacroiliac (SI) joint tenderness and/or inflammatory lumbosacral (LS) pain is one of the ILAR classification criteria for Entesitis related arthritis (ErA), a form of juvenile idiopathic arthritis (JIA) that includes most of the patients with juvenile spondyloarthritis (jSpA). Recognition of jSpA particularly early in the course of the disease represents a unique set of challenges and therefore all the patients with inflammatory back pain (IBP) and arthritis or enthesitis should be suspected for jSpA with possible involvement of sacroiliac and other vertebral joints. Since magnetic resonance imaging (MRI) is the preferred method of assessment both for axial inflammation and other possible musculoskeletal, infectious and malignant causes of LBP in children, it might be advisable to use it in assessment of LBP in children early after recognition of symptoms characteristic for inflammation. Objectives Were to evaluate the usefulness of early MRI in discovering inflammation of spinal joints and other possible causes of LBP in children not fulfilling ILAR classification criteria for ErA at the time of investigation. Thirty five children referred to our pediatric rheumatology outpatient clinic due to LBP lasting for more than three months, who fulfilled ASAS criteria for IBP and had sacroilic joint tenderness and positive FABRE test on examination, participated in the study. Their average age was 14.2 years (6-18 years) and 11 (31.4%) of them were boys. Five of study participants (14.3%) had arthritis and 19 (54.3%) had enthesitis confirmed by ultrasound, but none of them fulfilled ILAR criteria for ErA at the time of examination. Twelve (out of 15) had B27 antigen and 13 (37.1%) had a history of SpA related disease in a first degree relative. One of the patients had a diagnosis of ulcerative colitis, and one of psoriasis. All of the participants had normal CBC and CRP values with negative antinuclear antibodies and rheuma factor. None had neurological symptoms. MRI of sacroiliac joints with lumbal and thoracical regions of the spine was performed in one week period after initial exam on 1.5 T machine and interpreted by experienced musculoskeletal radiologist. Sacroiliac joints were examined using short-tau inversion-recovery (STIR) sequence in coronal and axial planes, T1 weighted images (WI) in coronal plane, fat-suppressed T1-WI in axial plane before and after contrast admission. STIR sequence in coronal and sagittal planes and T1- WI in sagittal plane were used in imaging of thoracolumbar spine. Nineteen (54, 3%) of patients had various pathological findings detected by the MRI. Four (11, 4%) had signs of inflammation with one even having an active sacroiliitis according to ASAS criteria. Schmorl nodes were discovered in six patients and three of them fulfilled criteria for Scheuermann disease, including one with the signs of inflammation. Two patients had stress reaction in lumbosacral region. Five patients had incipient degeneration of intervertebral discs. Three patients had disc protrusion without and one with radial nerve compression. After three months of follow up, 19 patients (54.3%) satisfied ILAR criteria for ErA, one for psoriatic arthrits and one, who also had an ulcerative colitis, satisfied criteria for undifferentiated arthritis. Differential diagnosis of LBP in children is very wide and it is difficult to distinguish inflammatory and other causes upon the first encounter with pediatric rheumatologist on the grounds of history and examination alone. In our study, all of the patients with LBP had some characteristics of IBP and ErA, but definite diagnosis of jSpA was subsequently established in sixty percent ͞only͟. Interestingly, almost twenty percent of them already had inflammatory changes of LS and SI joints discovered by MRI, while many others had signs of additional causes of LBP. Therefore, MRI performed early after referral to pediatric rheumatologist in all children with suspected IBP can be very useful in elucidating the cause of LBP and differentiation of those who need only symptomatic relief, orthopedic consultation or further follow up in pediatric rheumatology clinic. High cost of this approach can be justified with the benefit of early therapeutic intervention in those with established axial inflammation and the avoidance of unnecessary treatment in those with other causes.

Background: Clavicular cortical hyperostosis (CCH) is a sterile in ammatory bone disorder of unknown etiology clinically characterized by pain and/or swelling of the clavicle. Objectives: Identi cation of speci c gene expression patterns in CCH patients. Methods: Total RNA was isolated from whole blood of 18 new-onset, untreated CCH patients and 8 healthy controls. DNA microarray gene expression was preformed in 5 CCH and 4 control patients along with bioinformatical analysis of retrieved data. Carefully selected di erentially expressed genes where analyzed by qRT-PCR in all participants of the study. Results: Microarray results and bioinformatical analysis revealed 974 di erentially expressed genes involved in various in ammatory processes, while qRT-PCR analysis showed signi cantly higher expression of TRPM3 and TRPM7, as well as lower expression of ERBB2. Conclusions: TRPM3 and TRPM7 are transient receptor potential (TRP) proteins that act as multimodal sensor cation channels for a wide variety of stimuli, one of which is environmental temperature that in the case of CCH could be elicited by overuse of sterno- clavicular joint (SCJ). Altered TRP channel function can have strong e ects on a variety of cellular and systemic processes, including the activation and the regulation of the in ammasomes. ERBB2 belongs to a family of genes that encodes for cell surface growth factor receptors. ErbB activation promotes protective cellular outcomes during in ammation, hence lower expression of this gene could cause damage due to in ammation. We hypothesize that CCH could be an autoin ammatory disease induced by SCJ overuse, TRP channel overexpression, in ammasome activation and reduced protection during in ammation.

ntroduction: Systemic lupus erythematosus (SLE) is an autoimmune rheumatic disease that results from the interaction of multiple enviromental, immunological and genetic factors, causing inflammation and eventually damage in multiple organs. Its prevalence ranges from approximately 40 cases per 100, 000 and 15– 20% of cases present in children under 16 years of age. It has been suggested that peadiatric lupus patients have more aggressive disease course and an increased rate of more unusual initial clinical presentations compared with their adult counterparts. Clinical feature may include urticarial vasculitis with prevalence of up to 2, 2%, while angioedema is very rare. Objectives: We present a 15 year old female patient with SLE, urticarial vasculitis and angioedema. Methods: The disease started with nonspecific symptoms of malaise and headache and the patient was even put on klonazepam because of irregular electroencephalogram (EEG). She also developed autoimmune thrombocytopaenia and prednisone therapy was included for several months. Few months prior to admittance to our Department because of recurrent urticarial rash and angioedema allergy tests were performed and all were negative. During an episode of urticarial rash and vascullitis, skin biopsy was performed and leukocytoclastic vasculitis was diagnosed. Direct immnofolurescency showed IgA, IgG, IgM, C3 and C1q depostis. With symptoms that included malaise, fever, poliarthralgias, headache that developed, laboratory tests and skin biopsy findings SLE was suspected. Results: Antinuclear antibodies (ANA) titer was positive up to 1: 1280 (anti Ro 3+, anti Ro- 52 3+ i anti La 1+) and with low concentration of C3 and C4 ACR- criteria were met for SLE. Patient was treated with "pulse“ doses of metilprednisolone followed by maintenance doses. Nevertheless, she developed angioedema of lips, tounge nad eyelids with obstructive breathing. She recived adrenaline, glucocorticoides and antihistamines with poor effect and only after infusion of fresh frozen plasma, symptoms decreased. At the time, levels of C1 esterase inhibitor (C1-INH) were normal, while C1-INH function and C1q levels are still expected Conclusion: Angioedema is an unusual occurence in SLE, especially in children and it's mechanisms are heterogenous . SLE can be associated with hereditary and acquired deficinecies or dysfunction of C1-INH which leads to hyperactivation of complement classic pathway, bradykinin synthesis and incrased vascular permeability. Anti C1-INH autoantibodies seem to be uncommon in acquired angioedema (AAE) in SLE. In our patient, angioedema is thought to be most likely of the acquired form secondary to SLE since in hereditary angioedema (HAE) there is usually a family history of angioedema and earlier onset of symptoms. Although angioedema is generally self- limited disease, it may be life threatening causing the airway obstruction with mortality rate 15-33%.

Background: Clavicular cortical hyperostosis (CCH) is a sterile inflammatory bone disorder of unknown etiology clinically characterized by pain and/or swelling of the clavicle. It has been regarded as a variant of chronic nonbacterial/recurrent multifocal osteomyelitis (CNO/CRMO) but due to lack of other inflammatory sites and recurrence it could also be regarded as a separate disease in the spectrum [1]. Objectives: Identification of specific gene expression patterns in CCH patients. Methods: Total RNA was isolated from whole blood of 18 new-onset, untreated CCH patients and 8 healthy controls. DNA microarray gene expression was preformed in 5 CCH and 4 control patients along with bioinformatical analysis of retrieved data. Carefully selected differentially expressed genes (TRPM2, TRPM3, TRPM7, CASP2, MEFV, STAT3, EIF5A, ERBB2, TLR4, NLRP3, CD24, MYST3) where analyzed by qRT-PCR in all participants of the study. Results: Microarray results and bioinformatical analysis revealed 974 differentially expressed genes, while qRT-PCR analysis showed significantly higher expression of TRPM3 and TRPM7, and lower expression of ERBB2. Conclusions: Microarray data analysis reveled that majority of differentially expressed genes in CCH patients are involved in various inflammatory processes, while qRT-PCR analysis confirmed statistically significant expression change of 3 genes. Among them, TRPM3 and TRPM7 are members of transient receptor potential (TRP) gene superfamily, which encodes proteins that act as multimodal sensor cation channels for a wide variety of stimuli, one of which is environmental temperature that in the case of CCH could be elicited by overuse of sterno- clavicular joint (SCJ) [2]. Upon stimulation, TRP channels transduce electrical and/or Ca2+ signals. Dysfunctions in Ca2+ signaling due to altered TRP channel function can have strong effects on a variety of cellular and systemic processes, including the activation and the regulation of the inflammasomes, which are reported to be involved in CRMO pathogenesis [3, 4]. ERBB2, third gene with significant expression change, belongs to a family of genes that encodes for widely expressed cell surface growth factor receptors. Recently it has been shown that ErbB activation promotes protective cellular outcomes during inflammation, hence lower expression of this gene could cause damage due to inflammation [5]. Based on the results of these and previous studies, we hypothesize that CCH could be an autoinflammatory disease induced by SCJ overuse, TRP channel overexpression, inflammasome activation and reduced protection during inflammation. References: Borzutzky A, Stern S, Reiff A, et al. Pediatric chronic nonbacterial osteomyelitis. Pediatrics. 2012 Nov ; 130(5):e1190-1197. Shimizu S, Takahashi N, Mori Y. TRPs as chemosensors (ROS, RNS, RCS, gasotransmitters). Handbook of experimental pharmacology. 2014 ; 223:767-794. Latz E, Xiao TS, Stutz A. Activation and regulation of the inflammasomes. Nature reviews Immunology. 2013 Jun ; 13(6):397-411. Scianaro R, Insalaco A, Bracci Laudiero L, et al. Deregulation of the IL- 1beta axis in chronic recurrent multifocal osteomyelitis. Pediatric rheumatology online journal. 2014 ; 12:30. Frey MR, Brent Polk D. ErbB receptors and their growth factor ligands in pediatric intestinal inflammation. Pediatric research. 2014 Jan ; 75(1-2):127-132.

Spondyloarthritis or spondyloarthropathy (SpA) is a multifactorial disease in which a disturbed interplay occurs between the immune system and environmental factors on a predisposing genetic background, which leads to inflammation and structural damage of target tissue. Many recent researches on development of SpA showed important role of innate and adaptive immunity as well as of prominent bone tissue remodeling which leads to osteoproliferation and ankylosis. It is believed that possible sites of inflammation in SpA are entheses, sinovium and gut. Current knowledge on inflammation and tissue destruction leads to conclusion that SpA is disease characterized by disorders on different levels. Disorder on the first level is disturbed pathogen recognition and immune response activation, on second level disturbed inflammatory cells migration and on third level disturbed immune response regulation. As follows, disease progress depends on range of disturbances: disease course can be short, as in reactive arthritis, or long-lasting with substantial structural damage, as in ankylosing spondylitis. Unfortunately, there are still no confident markers of disease progression, so at the mere beginning disease is often described as undifferentiated.

Microarray gene expression analysis is high-throughput method in which many different sized DNA molecules are attached to solid surface in designated spots. These molecules are used for the discovery of specific RNA molecules isolated from various biological samples of interest. Core principle of this method is hybridization of complementary nucleotides (A-T and G-C), which leads to creation of double stranded nucleic acids. Gene expression differences in two groups of samples are discovered and quantificated by comparison of signal intensity values in microarray spots. Systemic analysis of data gathered in microarray gene expression measurement is performed by various bioinformatic methods such as group analysis, annotation analysis as well as network and pathway analysis. Expression comparison of all genes in different cells of the same individual or same cells of different individuals provides an insight into the mechanism responsible for development of a certain condition or disease.

In addition to the long-established association of HLA-B27 antigen and spondyloarthritis, several studies have shown a similar association with HLA-B7 antigen. But since the whole MHC region carries less than half of the risk for the development of the disease, the main goal of many recently performed researches, which implemented various high-throughput methods, was to discover the influence of genes outside the MHC region on disease development. The results showed that genes closely linked to spondyloarthritis participate in antigen processing and coding of various cytokines. This can lead to the conclusion that diseases from the spondyloarthritis group are polygenic, affected by both autoinflammatory and autoimmune mechanisms.

Determination of efficacy of Intra-articular infliximab therapy in JIA patients by 3D/4D musculo-skeletal ultrasound.

Juvenilni idiopatski artritis (JIA) najčešća je kronična reumatska bolest u djece i jedan od glavnih uzroka kratkotrajnog ili dugotrajnog invaliditeta te smanjenja kvalitete života u djetinjstvu. Bez pravovremenog i adekvatnog liječenja bolest progredira i rezultira trajnim promjenama na zglobovima. Izbor terapije ovisi o tipu JIA, stupnju aktivnosti bolesti, prognostičkim čimbenicima te dokazanoj učinkovitosti i mogućim nuspojavama pojedinih lijekova. Cilj suvremenog liječenja JIA je postizanje potpune remisije bolesti, a ne samo ublažavanje simptoma i privremeno smirivanje upalnog procesa. Primjena bioloških lijekova dovela je do značajnog napretka u liječenju bolesnika s rezistentnim oblicima JIA. U radu donosimo hrvatske smjernice za primjenu biološke terapije u bolesnika s JIA.

Transplantacija krvotvornih matičnih stanica (TKMS) postala je učinkovita terapijska mogućnost za liječenje teških oblika autoimunih bolesti (AB). Krvotvorne matične stanice mogu se sakupiti od samog bolesnika (autologno), jednojajčanog blizanca (singeno) te HLA identičnog davatelja (alogeno). Alogenom transplantacijom mijenjaju se autoagresivne efektorne stanice imunološkog sustava bolesnika s neautoagresivnim stanicama davatelja, no ovaj oblik transplantacije manje je privlačan zbog moguće reakcije presatka protiv primatelja (GVHD, engl. Graft-versus-host disease). S druge strane, autologna transplantacija dovodi do “resetiranja” imunološkog sata i ne uzrokuje GVHD, no zahtijeva prethodno kondicioniranje. Kao alternativa ovim oblicima liječenja razvila se i transplantacija mezenhimskih matičnih stanica (MMS) koje imaju značajan imunosupresivni učinak, ne zahtijevaju prethodno kondicioniranje i ne dovode do GVHD. Ipak, s obzirom na još uvijek velik postotak smrtnih ishoda povezanih s liječenjem (TRM, engl. Treatment related mortality), transplantaciju matičnih stanica treba sačuvati za bolesnike s najtežim oblicima AB, kojima se ovakvim načinom liječenja pruža čak i mogućnost potpunog izlječenja.

Introduction Calprotectin is a neutrophil derived protein that binds calcium and belongs to the S100 family. It can be quantified in feces and has become established as a marker of gut inflammation, where increased levels are a direct result of neutrophil migration into the gut lumen across the inflamed mucosa. Since subclinical gut inflammation is present in the majority of adult and pediatric spondyloarthritis patients, fecal calprotectin (fcal) is emerging as possible noninvasive biomarker. There is strong evidence supporting the role of the VSL-3 probiotic in decreasing fcal, thus promoting and maintaining remission in patients with inflammatory bowel disease (IBD), but very little is known of its potential effects on disease activity in children with undifferentiated spondyloarthritis (ERA). Objectives To assess the effect of the VSL-3 probiotic on fcal levels, clinical symptoms and disease activity of children with undifferentiated spondyloarthritis (ERA). Methods Sixteen patients diagnosed with ERA, according to the ILAR criteria, were treated with VSL-3, in addition to standard therapy (NSAID's and DMARD's). All patients were negative for gastrointestinal (GI) symptoms, and/or poor growth. All four ankylosing spondylitis patients received biologics ; two were treated with adalimumab and further two with infliximab. In addition to general clinical data, patients completed the BASFI and BASDAI questioners, ESR and CRP were obtained, and fcal was measured by the Calprest ELISA method (Eurospital Spa, Italy). After VSL-3 was introduced, two follow up visits were scheduled ; the first after one month and the second after three months. At the three month visit a second fcal value was obtained, and BASFI and BASDAI were reevaluated. Results The baseline mean fcal level was 52, 3 mg/kg (normal < 50 mg/kg). Nine patients have completed all scheduled visits to date, and eight out of those nine had markedly decreased fcal levels (mean value 15, 6 mg/kg). The BASFI index was decreased in seven patients and the BASDAI index in eight patients. At the follow-up visit, none of the patients were found to have developed GI symptoms or other signs and symptoms suggestive of IBD. Conclusion VSL-3 is emerging as a possible potent adjuvant therapy in children and adults with IBD thus encouraging its use in patients with ERA. To our knowledge our study is the first in evaluating the effects of VSL-3, as adjuvant therapy, on disease activity in ERA patients. Preliminary data suggest that the use of VSL-3 can decrease fcal levels, and together with standard therapy can improve clinical symptoms and decrease disease activity in patients with ERA. A larger patient cohort is needed to confirm VSL-3 efficacy in ERA patients.

Introduction Single nucleotide polymorphisms of Toll like receptors modify cellular immune response and induce pro-inflammatory cytokine production and therefore could be associated with enthesitis related arthritis (ERA) and/or oligoarticular and polyarticular juvenile idiopathic arthritis (JIA). Objectives To determine whether polymorphisms of TLR2 and TLR4 influence susceptibility to ERA or JIA. Methods DNA was extracted from blood samples of 19 ERA patients, 10 patients with oligoarticular or polyarticular JIA and 40 healthy controls, all diagnosed according to ILAR criteria. Polymorphisms of the TLR2 (Arg753Gln) and TLR4 (Asp299Gly, Thr399Ile) were determined using real time and multiplex PCR. Results All JIA patients were carriers of wild type allele for all three polymorphisms. Regarding Arg753Gln polymorphism of TLR2, only one patient with ERA (5.56%) and 2 healthy controls (5%) were carriers of heterozygous allele. There were no homozygous mutants. All ERA patients had wild type allele for Asp299Gly polymorphism of TLR4. For Thr399Ile polymorphism of TLR4, 21.05% ERA patients were heterozygous (CT variant), and none of the ERA patients was homozygous (TT variant), (CC vs CT variant in ERA OR 3.7500, 95% CI 1.0498-13.3952, p 0.0419, CC vs TT variant OR 31.0000, 95% CI 1.7309-555.1867, p 0.0197). In group of healthy controls, TLR4 polymorphisms Asp299Gly and Thr399Ile were in linkage disequilibrium ; 2 controls were hetrozygous and 6 homozygous variant carriers for both polymorphisms, whereas linkage disequilibrium was not found among patient groups (CC vs CT in controls OR 16.0000, 95% CI 3.5905- 71.2994, p 0.0003, CC vs TT OR 5.3333, 95% CI 2.0099-14.1524, p 0.1944). Conclusion Polymorphisms of TLR2 and TLR4 are not associated with oligoarticular/polyarticular JIA. There was also no evidence that variants of TLR are major risk factors for ERA, however, lack of susceptibility should be confirmed on larger group of patients, since four out of 19 patients (21, 05%) were heterozygous. A study on larger cohort is currently ongoing.