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OBJECTIVE: To develop a Childhood Lupus Improvement Index (CHILI) as a tool to measure response to therapy in childhood-onset systemic lupus erythematosus (cSLE), with a focus on clinically relevant improvement (CRIc SLE ). METHODS: Pediatric nephrology and rheumatology subspecialists (n = 213) experienced in cSLE management were invited to define CRIc SLE and rate a total of 433 unique patient profiles for the presence/absence of CRIc SLE . Patient profiles included the following cSLE core response variables (CRVs): global assessment of patient well-being (patient-global), physician assessment of cSLE activity (MD-global), disease activity index score (here, we used the Systemic Lupus Erythematosus Disease Activity Index), urine protein-to-creatinine ratio, and Child Health Questionnaire physical summary score. Percentage and absolute changes in these cSLE- CRVs (baseline versus follow-up) were considered in order to develop candidate algorithms and validate their performance (sensitivity, specificity, area under the receiver operating characteristic curve [AUC] ; range 0-1). RESULTS: During an international consensus conference, unanimous agreement on a definition of CRIc SLE was achieved ; cSLE experts (n = 13) concurred (100%) that the preferred CHILI algorithm considers absolute changes in the cSLE- CRVs. After transformation to a range of 0-100, a CHILI score of ≥54 had outstanding accuracy for identifying CRIc SLE (AUC 0.93, sensitivity 81.1%, and specificity 84.2%). CHILI scores also reflect minor, moderate, and major improvement for values exceeding 15, 68, and 92, respectively (all AUC ≥0.92, sensitivity ≥93.1%, and specificity ≥73.4%). CONCLUSION: The CHILI is a new, seemingly highly accurate index for measuring CRI in cSLE over time. This index is useful to categorize the degree of response to therapy in children and adolescents with cSLE.

Objective To study growth and puberty in a multinational longitudinal prospective cohort of children with juvenile dermatomyositis (DM). Methods Children from 31 countries who were ages DM in active phase were studied, and analyses of height, weight, and pubertal development were conducted in those who had follow-up visits during a 2-year period and for whom anthropometric data was available. Results A total of 196 of 275 children (71%) were included. We found a significant reduction in parent-adjusted height Z score over time in female patients (P < 0.0001) and male patients (P = 0.001), but with catch-up growth at the final study visit. Median body mass index Z score peaked at 6 months (P < 0.0001) and was still significantly above baseline at the final study visit, which was at a median of 26 months after baseline (P = 0.007), with no difference between sexes. Female patients with a disease duration >= 12 months after onset had significantly lower parent-adjusted height Z score (P = 0.002) and no 2-year catch-up growth. At the final study visit, growth failure was seen in 20 of 97 female patients (21%) and in 11 of 73 male patients (15%). Height deflection ( increment height Z score less than -0.25/year) was observed in 29 of 116 female patients (25%) and 25 of 80 male patients (31.3%). Delayed puberty was seen in 20 of 55 female patients (36.4%) and in 11 of 31 male patients (35.5%). Children in early pubertal stage at baseline had the highest risk of growth failure. Conclusion Juvenile DM in the active phase and/or its treatment has a significant impact on growth and puberty in affected children. Children with recent onset of puberty or previous growth failure have the highest risk of delayed pubertal development and further growth retardation.

Timeline for case report entitled â Unusual localization and presentation of osteoid osteoma mimicking juvenile spondyloarthritis: a case report.â (PDF 53 kb)

Introduction: in recent years a number of studies showed a connection between intestinal dysbiosis and various immunological diseases, hence it is not coincidence gastrointestinal abnormalities can often be found in patients with rheumatic diseases and musculoskeletal symptoms are not rare in variety of digestive system disorders. Nevertheless, when both musculoskeletal and gastrointestinal symptoms presents simultaneously at the beginning of the disease, it might be difficult to distinguish the primary disorder. Objectives:to present a patient with simultaneous presentation of characteristic gastrointestinal and musculoskeletal symptoms. Methods: case report. Informed consent to publish has been obtained from both the patient and parent. Results: a thirteen year old boy with medical history of infantile colic and a brother with HLA B27 positive juvenile spondyloarthritis was admitted to our department due to persistent inflammatory lumbosacral pain, Achiles enthesitis, and abdominal discomfort. During the six months period prior to the admission he had six episodes of fever, cough, abdominal pain and aphthae lasting for few days. Acute surgical disease was excluded on several occasions. Chest radiograph, throat and nose cultures showed no signs of infection. During the last few episodes he developed swelling of the right talocrural joint which resolved after short course of NSAID. Later, severe pain in sacroiliac (SI) joints with morning stiffness emerged, while his stools became loose and frothy. Abdominal ultrasound showed ileocecal mesenteric lymphadenitis and dilated intestinal loops filled with dense liquid content. Upon the first examination by paediatric rheumatologist he had palpatory periumbilical pain as well as tenderness of SI joints and Achiles entheses. His height and weight were below the 5th centile while estimated bone age was 12. ANA, ANCA and RF were negative. There was no sign of uveitis on ophthalmologic examination. HLA typing was positive for B27. Radiographic and magnetic resonance imaging of SI joints and spine showed no signs of inflammation. Immunoglobulin A anti-tissue transglutaminase antibody (IgA TTG) was negative while serum IgA levels were normal. No pathological agents were isolated from stool. Several faecal occult blood tests were negative and faecal calprotectin levels were normal. Endoscopy of upper and lower gastrointestinal tract revealed hiatal hernia, oesophagitis, gastritis and duodenitis. Histologic evaluation of several duodenal biopsy specimens displayed typical features of celiac disease. A month after initiation of gluten free diet and NSAID’s the improvement of all symptoms was noticed. Conclusion:although they differ in HLA predispositions, both JIA and CD are associated with 4q27 locus, which encodes IL-2 and IL-12 cytokines important for activation and regulation of immune system, and both are linked with a functional single nucleotide polymorphism in the PTPN22 gene (1858C>T). Intriguingly, lymphocyte cytotoxicity in the intestinal mucosa is abnormally increased in both entities, suggesting some luminal, possibly a nutritional factor, may be involved. This agent is well established in CD, but number of cases, including presented, exhibited beneficial role of gluten free diet in arthritis patients as well. Our patient clearly met ILAR criteria for ErA and had typical histological features of CD, but without elevated anti-TTG levels. Since his musculoskeletal symptoms could also be explained as a part of extra-intestinal CD manifestations, the real distinction of underlying disorders was challenging. Therefore it is reasonable to conclude both CD and certain subtypes of JIA have multiple shared mechanisms making them hard to separate in given circumstances.

Background/Purpose: 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. To elucidate the possible role of epigenetic modifications in the regulation of those genes, DNA methylation analysis was performed. Methods: DNA was isolated from PBMCs of 19 patients diagnosed with jSpA according to ILAR classification criteria for enthesitis related arthritis (ErA) and seven matched healthy children. None of the jSpA participant had symptoms for more than six months and all were untreated. Methylated DNA Immunoprecipitation (MeDIP) was performed in promotor region of differentially expressed genes (TLR4, CXCR4, NLRP3, PTPN12) using the Magmedip kit. Enrichment in MeDIP fraction was determined by quantitative RT-PCR using the AriaMx. MeDIP results were expressed as fold enrichment of immunoprecipitated DNA for each site. Results: Statistical analysis revealed significant hypermethylation of promoter sites in NLRP3 gene (p=0, 0220). No significant alterations in methylation status were observed in promotor regions of other genes (Table 1). Conclusion: Our study indicates the hypermethylation of NLRP3 gene promotor is probably responsible for expression alterations in jSpA patients in the initial phase of the disease. The NLRP3 gene has a crucial role in assembly of NLRP3 inflammasome, innate immune sensor that regulates “danger” response upon various signals. While increased expression of this gene has been found in many autoinflammatory diseases, decreased expression has been associated with IBD in which the microbiota is believed to contribute to the intestinal inflammation. Therefore, it is not entirely surprising decreased expression has also been observed in jSpA, the disease that has clinical and genetic similarities with IBD and is often characterized by subclinical gut inflammation. The growing number of evidence shows any modification of gut microbiota can lead to dysbiosis with long- term consequences for the whole organism. Specifically, in jSpA this could result in increased influx of TLR4 ligands and increased expression of TLR4 gene, as well as in reduction of commensal bacteria with anti-inflammatory properties, namely bacteria Faecalibacterium prausnitzii that inhibits NF-kB signaling, leading to TNF-a abundancy characteristic for jSpA. Since inflammasomes have been shown to shape the microbiota, it is reasonable to assume these processes can at least partially be explained by reduced NLRP3 expression due to hypermethylation. Therefore, our findings could have important implications in understanding of the disease mechanisms and possible therapeutic options.

Introduction: The introduction of biologic agents has revolutionized the treatment of juvenile idiopathic arthritis (JIA) due to their efficacy, speed of onset and tolerability. A numerous clinical practice guidelines and consensus statements on the criteria for biologic therapy (BT) introduction have been developed, however, the consensus on cessation of biologic agents has not been harmonized. Objectives: Presentation of our experience on discontinuation of biologic therapy in JIA patients. Methods: We conducted a retrospective two centre analysis of patients with JIA diagnosis according to ILAR criteria, treated with BT from January 2008 to May 2016. Demographic information, duration of the treatment, number of biologic agents used and discontinuation rate were extracted using medical charts. Successful discontinuation was defined as cessation of the drug due to disease control according to Wallace criteria and musculoskeletal ultrasound inactive disease. Results: Total of 92 patients (87% female, 13% male) with different JIA subtypes, non-responders or intolerant to syntetic DMARDs, were treated with one or more biologicals. Median disease duration, from onset to the introduction of first BT, was 3.4 years (0.4-13 years). Patients were diagnosed with poli JIA in 68.5%, oligo JIA in 18.5%, ERA in 6.4%, systemic JIA in 3.3% and psoriatic JIA in 3.3%. In 88 patients first biologic drug was anti-TNF agent ( etanercept in 54 pts, adalimumab in 18 pts, infliximab in 16 pts) and 3 pts were initially treated with tocilizumab and one patient with anakinra. By the May 2016, 4 patients were lost from the follow up, 9 patients were transferred to adult rheumatology department while on BT, and 52 patient were still on BT. In 27 patients (29.3%) BT was successfully withdrawn, out of which 13 (14.1%) discontinued all medications. Majority of the patients were on the first line BT (20 pts ; 74%), and another 7 patients were on the second line BT at the time of BT cessation. In another 37 patients BT discontinuation lasted for a short period of time (2-6 months), after which BT was reintroduced due to relapse. Median duration of BT before successful discontinuation was 2.96 years (1-7.2 years). The probability of the successful withdrawal of BT was associated with a shorter disease duration at the beginning of the treatment. Conclusion: The development of optimal timeline and modality of discontinuation of BT after documentation of inactive disease is needed. In our retrospective study probability of successful withdrawal of BT was associated with shorter disease duration at the beginning of the treatment. These data correlate with current literature overview. Future studies will hopefully identify other predictors of the successful discontinuation of biologic therapy.

CILJ: klavikularna kortikalna hiperostoza (CCH) rijetka je bolest nepoznate etiologije i ishoda, klinički karakterizirana bolnošću i/ili oticanjem medijalnog dijela klavikule. Neki je smatraju oblikom kroničnog nebakterijskog/rekurentnog multifokalnog osteomijelitisa (CNO/CRMO), no zbog izostanka drugih sijela upale i rekurentnosti mogla bi se smatrati samostalnom bolesti, zbog čega je važno rasvijetliti mehanizme odgovorne za nastanak i napredovanje. METODE: ukupna RNA izolirana je iz pune krvi 18 neliječenih bolesnika s CCH i osmero zdrave djece odgovarajuće dobi i spola. Ispitivanje cjelokupnoga genskoga izražaja na sitnopolju provedeno je u pet bolesnika i četvero zdrave djece, zajedno s opsežnom bioinformatičkom analizom. Pomoću qRT-PCR metode u svih sudionika ispitan je izražaj pomno odabranih gena koji su pokazali razliku u izražaju na sitnopolju (T RPM2, TRPM3, TRPM7, STAT3, EIF5A, ERBB2, TLR4, NLRP3, CD24, MYST3). Na kraju, u jednog bolesnika stanice su centrifugirane i podvrgnute imunoflourescentnoj mikroskopiji. REZULTATI: ispitivanje genskoga izražaja na sitnopolju pokazalo je razlike u izražaju za 974 gena, dok je qRT-PCR analiza pokazala značajno pojačan izražaj TRPM3 i TRPM7 gena te niži izražaj za ERBB2. Imunofluorescentna mikroskopija pokazala je značajno pojačan signal TRPM3 proteina u bolesnika s CCH. ZAKLJUČAK: ispitivanjem genskog izražaja na sitnopolju i detaljnom bioinformatičkom analizom dobivenih rezultata pokazalo se kako je većina gena s razlikom u izražaju uključena u različite upalne procese, dok je qRT-PCR analiza potvrdila statistički značajne razlike u izražaju tri gena. Dva od njih, TRPM3 i TRPM7, spadaju u skupinu gena čiji su proteinski produkti kationski kanali koji se ponašaju kao multimodalni senzori za različite stimulanse iz okoliša. Jedan od njih je temperatura, koja u bolesnika s CCH može porasti zbog prekomjerne upotrebe sterokalvikulatnog zgloba te dovesti do promjena u provođenju Ca2+ iona kroz TRP kanale. Takve promjene u provođenju iona mogu imati velik utjecaj na razne stanične i sistemske procese, uključujući aktivacije i regulaciju inflamasoma, koji pak ima važnu ulogu u patogenezi CRMO-a. ERBB2, treći gen s značajnom razlikom u izražaju, spada u skupinu gena koji kodiraju receptore za čimbenike rasta prisutne na mnogim stanicama. Nedavno se pokazalo da aktivacija navedenih receptroa štiti stanicu za vrijeme upale, iz čega proizlazi da smanjena ekspresija ovoga gena može dovesti do oštećenja prilikom upale. Naposljetku, rezultati dobiveni na razini genskog izražaja potvrđeni su i na razini proteina, čime se zaokružuje hipoteza prema kojoj je CCH autoinflamatorna bolest nastala zbog okolišnih (prekomjerna upotreba sternoklavikularnog zgloba) i genskih čimbenika (prekomjerni izražaj TRP kanala koji dovodi do aktivacije inflamasoma uz smanjenu zaštitu stanica tijekom upale).

Prikaz bolesnice s sistemskim eritemskim lupusom i ponavljajućim angioedemom.

CILJ: od samih početaka medicine toplina je jedan od četiri glavna znaka upale. Usprkos tome, još uvijek ne postoji pouzdana, jednostavna i objektivna metoda za mjerenje topline u upaljenom zglobu, već se u praksi najčešće procjenjuje samo kliničkim pregledom, što ima vrlo malu opservacijsku pouzdanost. Idealna metoda treba biti pouzdana, jeftina i jednostavna za upotrebu, najbolje na mjestu gdje se bolesniku pruža skrb (engl. point-of-care), a navedene kriterije moglo bi zadovoljiti termografsko snimanje zglobova FLIR ONE dodatkom za telefone. METODE: osmero djece u dobi od 2, 5 – 14, 5 godina s različitim oblicima artrtisa i/ili artralgije sudjelovalo je u ovom pilot presječnom istraživanju. Svim sudionicima snimljena su oba koljena FLIR ONE termografskom kamerom za pametne telefone, bez odjeće, uz jednaku sobnu temperaturu (20°C). Nakon toga dječji reumatolog certificiran za muskuloskelenti ultrazvuk svima je učinio pregled oba koljena i bodovanje PD (engl. power doppler) signala (0-3 boda). Dobivene slike analizirane su pomoću FLIR ONE TOOLS kompjutorskog programa a automatski predefiniranim rasponom temperatura, a na svakoj slici označena su koljena pomoću manualnog alata za mjerenje. Očitane su vrijednosti prosječne, najniže i najviše temperature unutar označene elipse, a prikupljeni podaci analizirani su pomoću Graphpad Prism statističkog programa. REZULTATI: šest sudionika istraživanja imalo je PD signal u oba koljena veći od 0, 5 (0, 5 – 2, 5), što je definirano kao aktivna upala, dok su dva imala signal 0 u oba koljena, što je definirano kao izostanak upale. Prosječna vrijednost prosječne temperature koljena u sudionika s aktivnom upalom bila je 34, 51°C, a u sudionika bez upale 30, 02°C. Prosječna vrijednost najviše očitane temperature u prvoj skupini bila je 35, 81°C, a u drugoj 31, 5°C, dok je prosječna vrijednost razlike najviše i najniže temperature bila 4, 95°C u prvoj skupini, a 2, 87°C u drugoj. Statistička analiza pokazala je da su razlike u svim navedenim mjerenjima između skupine sudionika s aktivnom upalom i skupine bez aktivne upale koljena bile značajne (p

Juvenilni spondiloartritis (jSpA) je skupina multifaktorijalnih bolesti u kojima dolazi do poremećenog međudjelovanja imunološkog sustava i čimbenika okoliša u ljudi s predisponirajućim genotipom, što dovodi do upale i strukturnih oštećenja ciljnog tkiva. Prvi simptomi jSpA rijetko su povezani s kralježnicom, a češće se javlja nesimetrični oligoartritis zglobova donjih ekstremiteta, daktilitis i periferni entezitis. Postoje brojni kriteriji za klasifikaciju jSpA, no većina pedijatrijskih reumatologa danas koristi kriterije Međunarodne lige reumatoloških udruženja (ILAR) prema kojima je najveći broj bolesnika s artirtisom i entezitisom ili artritisom ili entezitisom uz još dvije ili više od karakteristika poput bolnosti sakroilijakalnih zglobova na dodir i/ili bolnosti kralježnice zbog upalnog procesa, HLA B27 genotipa, bolesti povezane s HLA B27 genotipom u jednog ili više rođaka u prvom ili drugom koljenu, uveitisa te muškog spola uz više od osam godina starosti, svrstan u podskupinu juvenilnog idiopatskog artritisa (JIA) koju nazivamo entezitisu pridruženi artritis (ErA). Sukladno tome, dijagnoza bolesti postavlja se uglavnom na temelju kliničke slike i anamnestičkih podataka ; od laboratorijskih pretraga potrebno je učiniti antinuklearna antitijela (ANA), reumatoidni fakotr (RF) te HLA tipizaciju kako bi se utvrdila prisutnost HLA B27, B7 i/ili DR4 genotipa. Obzirom da velik broj bolesnika ima i subkliničku upalu crijeva, preporučljivo je provjeriti i fekalni kalprotektin. U slučaju da postoje simptomi perifernog entezitisa, potreban je pregled muskuloskeletnim ultrazvukom s osnaženim doplerom (PDUS), a pri znakovima zahvaćenosti kralježnice radiografsko snimanje te magnetska rezonancija (MR) s kontrastnim sredstvom. Najveći broj djece s jSpA-om liječi se fizikalnom terapijom i nesteroidnim protuupalnim lijekovima (NSAIL), dok se refraktorni oblici periferne bolesti mogu liječiti sintetskim lijekovima koji utječu na tijek bolesti (DMARD), poput sulfasalazina. Kada bolesti zahvati i kralježnicu, potrebno je liječenje biološkim DMARD-ovima poput adalimumaba, infliksimaba i etanercepta. Usprkos mnogim istraživanjima koja su pružala dobar uvid u patogenezu bolesti, odgovor na liječenje i prognozu teško je predvidjeti.

Naša iskustva u tranziciji djece s reumatskim bolestima u adultnu skrb.

Introduction: Gene expression profiling of juvenile spondyloarthritis (jSpA) patients revealed aberrant expression of some genes, but the mechanism of this expression alterations remained unknown (1). Objectives: To discover epigenetic modifications with effect on regulation of 4 genes (TLR4, NLRP3, CXCR4, PTPN12) differentially expressed in jSpA patients. Methods: DNA methylation analysis was performed in promotor region of differentially expressed genes by methylated DNA Immunoprecipitation (meDIP). Based on literature search, 4 microRNAs (miR-150, miR- 146a, miR-181a, miR-223) included in regulation of differentially expressed genes were selected and their expression was analyzed using RT-PCR with predeveloped Taqman microRNA assays. Both analysis were performed in 7 newly diagnosed untreated jSpA patients and 7 matched healthy children. Results: Statistical analysis revealed no difference in methylation of promotor sites for examined genes, but PTPN12 showed trend towards higher methylation (p=0, 076), compared with control group. There was no statistical difference in expression of selected miRs between two groups. Conclusion: jSpA is a multifactorial disease in which a complex interplay occurs between the immune system and environmental factors on a predisposing genetic background. One of the most important mechanisms by which environment can influence processes inside of an organism is gene regulation via epigenetic modifications. Therefore, we investigated possible influence of DNA methylation and posttranscriptional modifications on the genes differentially expressed in jSpA patients. The results didn't indicate any statistically significant abundance of miRs with described role in expression of examined genes, nor hypermethylation of their promotor sites, but there was a clear trend of higher methylation of PTPN12 in jSpA patients, while lack of statistical significance could be attributed to small number of study participants. This observation can easily explain decrease of PTPN12 expression in treated and untreated jSpA patients. PTPN12 expressed in dendritic cells is identified as a key regulator of dendritic cell migration as well as T cell-dependent immunity and autoimmunity (2). It is also a negative regulator of inflammation and intestinal cell migration, as well as a positive regulator of osteoclast activity, all of which are processes important for the pathophysiology of jSpA. Results of our study are in concordance with previous epigenetic studies of this gene in human breast cancer patients, which showed PTPN12 can be silenced by methylation. Interestingly, more recent study pointed to the potential of 5-Azac, a DNA hypomethylating agent, in increasing PTPN12 expression, and highlighted the therapeutic potential of this mechanism in treatment of breast cancer (6). Therefore, it is tempting to speculate the results of our study could after confirmation in larger cohorts give rise for new treatment options in jSpA patients as well. REFERENCES: 1. Lamot L, et al. Aberrant expression of shared master-key genes contributes to the immunopathogenesis in patients with juvenile spondyloarthritis. PLoS One. 2014 Dec 15 ; 9(12):e115416. 2. Inmoo Rhee, et al. Control of dendritic Cell Migration, T Cell-Dependent Immunity, and Autoimmunity by Protein Tyrosine Phosphatase PTPN12 Expressed in Dendritic Cells. Mol Cell Biol. 2014 Mar ; 34(5):888-99. 3. Thummuri D, et al. Epigenetic regulation of protein tyrosine phosphatase PTPN12 in triple-negative breast cancer. Life Sci. 2015 Jun 1 ; 130:73- 80.

Introduction: Since the dawn of the medicine, heat (lat. calor) is one of the four main signs of inflammation. Although it is recognized more than 2, 000 years ago, there is still no reliable, simple and objective method to measure the warmth of inflamed joint, which is therefore most often assessed only by clinical examination, with very low validity and reliability. Objectives: To investigate the use of smartphone attached mobile thermography for the assessment of joint inflammation. Methods: Twenty participants with various forms of arthritis and/or arthralgia participated in this pilot cross-sectional observational study. Average age of the participants was 10 years (2, 5 - 17, 5). In all of the participants Power Doppler vascularization signal of both knee joints was graded from 0-3 by EULAR certificated instructor in MSUS. FLIR ONE smartphone connected device was used to obtain infrared thermal image of every patient’s knee joint prior to the MSUS examination, without clothes and with same environment conditions (room temperature was set to 20°C). Acquired images were analyzed using FLIR Tools software for thermal image analysis with automatically set range of temperatures. On every image, both knee joints were selected using manual ellipse measurement tool and the data on average, minimum and maximum temperature inside the circle was collected. Subsequent statistical analysis of selected data was performed with Graphpad Prism software. Results: In sixteen knee joints of nine patients a PD signal was above 0, 5 (0, 5-2, 5), which was defined as an active inflammation, while in twenty-four knee joints of thirteen patients PD signal was 0, which was defined as no inflammation. Average value of average temperature in knee joints with inflammation was 33, 72°C (31, 3°C - 35, 4°C) and in patients without inflammation 29, 81°C (23, 9°C - 31, 9°C), average value of maximal temperature was 34, 98°C (32, 7°C - 36, 9°C) and 31, 08°C (25°C - 33, 3°C), while average value of difference between maximal and minimal temperature was 4, 73°C (2, 6°C - 7, 7°C) and 2, 96°C (1, 2°C - 5, 7°C) respectively. Statistical analysis revealed significant difference in all of the measurements between knee joints with and without active inflammation (p

CILJ: ranije provedeno ispitivanje genskoga izražaja bolesnika s juvenilnim spondiloartritisom (jSpA) pokazalo je promjene u izražaju četiri gena (TLR4, NLRP3, CXCR4, PTPN12) čiji proteinski produkti sudjeluju u procesima važnima za razvoj bolesti. Kako bi se otkrili mogući epigenetski mehanizmi regulacije navedenih gena ispitana je metilacija njihovih promotora te izražaj mikroRNK za koje se provedenim istraživanjima utvrdilo da imaju ulogu u njihovom prepisivanju i obradi. METODE: DNA metilacija promotora ispitana je metodom metilirane DNA imunoprecipitacije (meDIP). Pretragom baza podataka odabrano je 4 mikroRNK (miR-150, miR-146a, miR-181a, miR-223) za koje se utvrdilo da bi mogle imati ulogu u regulaciji gena važnih za razvoj jSpA te je ispitan njihov izražaj pomoću RT-PCR metode s odgovarajućim Taqman mikroRNK probama. Sve analize provedene su u skupini od sedmero novootkrivenih, neliječenih bolesnika s jSpA te u skupini od sedmero zdrave djece odgovarajuće dobi i spola. REZULTATI: statistička analiza nije ukazala na značajnu razliku u metilaciji promotorskih regija ispitivanih gena, no otkriven je trend pojačane metilacije gena PTPN12 u grupi bolesnika (p=0, 076). Nije bilo statističke razlike u izražaju ispitivanih mikroRNA između bolesnika i skupine zdrave djece. ZAKLJUČAK: jSpA je kompleksna bolest u kojoj dolazi do poremećaja međudjelovanja imunološkog sustava i čimbenika okoliša uz predisponirajući genotip. Jedan od najvažnijih mehanizama kojima okoliš utječe na procese unutar organizma je regulacija gena epigenetskim modifikacijama. U ovom istraživanju stoga je ispitan mogući utjecaj DNA metilacije i postranskripcijske modifikacije preko izabranih mikroRNK na gene koji su u bolesnika s jSpA pokazali razlike u izražaju. Rezultati nisu ukazali na statistički značajne razlike u metilaciji gena ili izražaju mikroRNK, što bi moglo biti i zbog malog broja sudionika, no opažen je jasan trend pojačane metilacije gena PTPN12, čime se može objasniti smanjen izražaj toga gena u liječenih i neliječenih bolesnika s jSpA. Produkt navedenog gena prepoznat je kao glavni regulator migracije dendritičkih stanica i imunološkog odgovora ovisnog o T- limfocitima, negativni regulator upale i migracije stanica crijeva te pozitivni regulator aktivnosti osteoklasta, što su sve procesi važni za razvoj jSpA. Rezultati našeg istraživanja u skladu su s rezultatima prijašnjih koji su pokazali kako se PTPN12 može utišati metilacijom. Nadalje, nedavno provedeno istraživanje pokazalo je da primjena hipometilacijskog agenta 5-Azac može pojačati izražaj PTPN12 gena u bolesnika s karcinomom dojke, čime se ukazalo i na mogući terapeutski potencijal ovog mehanizma. Stoga je primamljivo zaključiti kako bi i naši rezultati nakon potvrde u većim skupinama bolesnika mogli poslužiti za razvoj novih metoda liječenja bolesnika s jSpA.

CILJ: klavikularna kortikalna hiperostoza (CCH) rijetka je bolest nepoznate etiologije i ishoda, klinički karakterizirana bolnošću i/ili oticanjem medijalnog dijela klavikule. Neki je smatraju oblikom kroničnog nebakterijskog/rekurentnog multifokalnog osteomijelitisa (CNO/CRMO), no zbog izostanka drugih sijela upale i rekurentnosti mogla bi se smatrati samostalnom bolesti, zbog čega je važno rasvijetliti mehanizme odgovorne za nastanak i napredovanje. METODE: ukupna RNA izolirana je iz pune krvi 18 neliječenih bolesnika s CCH i osmero zdrave djece odgovarajuće dobi i spola. Ispitivanje cjelokupnoga genskoga izražaja na sitnopolju provedeno je u pet bolesnika i četvero zdrave djece, zajedno s opsežnom bioinformatičkom analizom. Pomoću qRT-PCR metode u svih sudionika ispitan je izražaj pomno odabranih gena koji su pokazali razliku u izražaju na sitnopolju (T RPM2, TRPM3, TRPM7, STAT3, EIF5A, ERBB2, TLR4, NLRP3, CD24, MYST3). Na kraju, u jednog bolesnika stanice su centrifugirane i podvrgnute imunoflourescentnoj mikroskopiji. REZULTATI: ispitivanje genskoga izražaja na sitnopolju pokazalo je razlike u izražaju za 974 gena, dok je qRT-PCR analiza pokazala značajno pojačan izražaj TRPM3 i TRPM7 gena te niži izražaj za ERBB2. Imunofluorescentna mikroskopija pokazala je značajno pojačan signal TRPM3 proteina u bolesnika s CCH. ZAKLJUČAK: ispitivanjem genskog izražaja na sitnopolju i detaljnom bioinformatičkom analizom dobivenih rezultata pokazalo se kako je većina gena s razlikom u izražaju uključena u različite upalne procese, dok je qRT-PCR analiza potvrdila statistički značajne razlike u izražaju tri gena. Dva od njih, TRPM3 i TRPM7, spadaju u skupinu gena čiji su proteinski produkti kationski kanali koji se ponašaju kao multimodalni senzori za različite stimulanse iz okoliša. Jedan od njih je temperatura, koja u bolesnika s CCH može porasti zbog prekomjerne upotrebe sterokalvikulatnog zgloba te dovesti do promjena u provođenju Ca2+ iona kroz TRP kanale. Takve promjene u provođenju iona mogu imati velik utjecaj na razne stanične i sistemske procese, uključujući aktivacije i regulaciju inflamasoma, koji pak ima važnu ulogu u patogenezi CRMO-a. ERBB2, treći gen s značajnom razlikom u izražaju, spada u skupinu gena koji kodiraju receptore za čimbenike rasta prisutne na mnogim stanicama. Nedavno se pokazalo da aktivacija navedenih receptroa štiti stanicu za vrijeme upale, iz čega proizlazi da smanjena ekspresija ovoga gena može dovesti do oštećenja prilikom upale. Naposljetku, rezultati dobiveni na razini genskog izražaja potvrđeni su i na razini proteina, čime se zaokružuje hipoteza prema kojoj je CCH autoinflamatorna bolest nastala zbog okolišnih (prekomjerna upotreba sternoklavikularnog zgloba) i genskih čimbenika (prekomjerni izražaj TRP kanala koji dovodi do aktivacije inflamasoma uz smanjenu zaštitu stanica tijekom upale).

OBJECTIVE: 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. Therefore, it is of high importance to elucidate the exact mechanisms responsible for the development and progression of the symptoms [1]. METHODS: Total RNA was isolated from whole blood of 18 new- onset, untreated CCH patients and 8 healthy controls. DNA microarray gene expression was performed 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. In one patient, the blood cells were processed using a cytosine for a immunofluorescence microscopy with TRPM3 and TRPM7 antibody. 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. Immunofluorescence microscopy showed high signal of TRPM3 in blood cells of one patient. CONCLUSIONS: Microarray data analysis revealed 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]. Finally, the results of transcriptome analysis were confirmed on a protein level with a proof-of-concept experiment which indicated high presence of TRPM3 in blood cells of a patient. 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: 1. Borzutzky A, Stern S, Reiff A, et al. Pediatric chronic nonbacterial osteomyelitis. Pediatrics. 2012 Nov ; 130(5):e1190-1197. 2. Shimizu S, Takahashi N, Mori Y. TRPs as chemosensors (ROS, RNS, RCS, gasotransmitters). Handbook of experimental pharmacology. 2014 ; 223:767-794. 3. Latz E, Xiao TS, Stutz A. Activation and regulation of the inflammasomes. Nature reviews Immunology. 2013 Jun ; 13(6):397-411. 4. 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. 5. 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.

U radu se prikazuju iskustva liječenja juvenilnog idopatskog artritisa biološkom terapijom u dva cetnra za pedijatrijsku reumatologiju.

CILJ: ranije provedeno ispitivanje genskoga izražaja bolesnika s juvenilnim spondiloartritisom (jSpA) pokazalo je promjene u izražaju četiri gena (TLR4, NLRP3, CXCR4, PTPN12) čiji proteinski produkti sudjeluju u procesima važnima za razvoj bolesti. Kako bi se otkrili mogući epigenetski mehanizmi regulacije navedenih gena ispitana je metilacija njihovih promotora te izražaj mikroRNK za koje se provedenim istraživanjima utvrdilo da imaju ulogu u njihovom prepisivanju i obradi. METODE: DNA metilacija promotora ispitana je metodom metilirane DNA imunoprecipitacije (meDIP). Pretragom baza podataka odabrano je 4 mikroRNK (miR-150, miR-146a, miR-181a, miR-223) za koje se utvrdilo da bi mogle imati ulogu u regulaciji gena važnih za razvoj jSpA te je ispitan njihov izražaj pomoću RT-PCR metode s odgovarajućim Taqman mikroRNK probama. Sve analize provedene su u skupini od sedmero novootkrivenih, neliječenih bolesnika s jSpA te u skupini od sedmero zdrave djece odgovarajuće dobi i spola. REZULTATI: statistička analiza nije ukazala na značajnu razliku u metilaciji promotorskih regija ispitivanih gena, no otkriven je trend pojačane metilacije gena PTPN12 u grupi bolesnika (p=0, 076). Nije bilo statističke razlike u izražaju ispitivanih mikroRNA između bolesnika i skupine zdrave djece. ZAKLJUČAK: jSpA je kompleksna bolest u kojoj dolazi do poremećaja međudjelovanja imunološkog sustava i čimbenika okoliša uz predisponirajući genotip. Jedan od najvažnijih mehanizama kojima okoliš utječe na procese unutar organizma je regulacija gena epigenetskim modifikacijama. U ovom istraživanju stoga je ispitan mogući utjecaj DNA metilacije i postranskripcijske modifikacije preko izabranih mikroRNK na gene koji su u bolesnika s jSpA pokazali razlike u izražaju. Rezultati nisu ukazali na statistički značajne razlike u metilaciji gena ili izražaju mikroRNK, što bi moglo biti i zbog malog broja sudionika, no opažen je jasan trend pojačane metilacije gena PTPN12, čime se može objasniti smanjen izražaj toga gena u liječenih i neliječenih bolesnika s jSpA. Produkt navedenog gena prepoznat je kao glavni regulator migracije dendritičkih stanica i imunološkog odgovora ovisnog o T- limfocitima, negativni regulator upale i migracije stanica crijeva te pozitivni regulator aktivnosti osteoklasta, što su sve procesi važni za razvoj jSpA. Rezultati našeg istraživanja u skladu su s rezultatima prijašnjih koji su pokazali kako se PTPN12 može utišati metilacijom. Nadalje, nedavno provedeno istraživanje pokazalo je da primjena hipometilacijskog agenta 5-Azac može pojačati izražaj PTPN12 gena u bolesnika s karcinomom dojke, čime se ukazalo i na mogući terapeutski potencijal ovog mehanizma. Stoga je primamljivo zaključiti kako bi i naši rezultati nakon potvrde u većim skupinama bolesnika mogli poslužiti za razvoj novih metoda liječenja bolesnika s jSpA.

CILJ: od samih početaka medicine toplina je jedan od četiri glavna znaka upale. Usprkos tome, još uvijek ne postoji pouzdana, jednostavna i objektivna metoda za mjerenje topline u upaljenom zglobu, već se u praksi najčešće procjenjuje samo kliničkim pregledom, što ima vrlo malu opservacijsku pouzdanost. Idealna metoda treba biti pouzdana, jeftina i jednostavna za upotrebu, najbolje na mjestu gdje se bolesniku pruža skrb (engl. point-of-care), a navedene kriterije moglo bi zadovoljiti termografsko snimanje zglobova FLIR ONE dodatkom za telefone. METODE: osmero djece u dobi od 2, 5 – 14, 5 godina s različitim oblicima artrtisa i/ili artralgije sudjelovalo je u ovom pilot presječnom istraživanju. Svim sudionicima snimljena su oba koljena FLIR ONE termografskom kamerom za pametne telefone, bez odjeće, uz jednaku sobnu temperaturu (20°C). Nakon toga dječji reumatolog certificiran za muskuloskelenti ultrazvuk svima je učinio pregled oba koljena i bodovanje PD (engl. power doppler) signala (0-3 boda). Dobivene slike analizirane su pomoću FLIR ONE TOOLS kompjutorskog programa a automatski predefiniranim rasponom temperatura, a na svakoj slici označena su koljena pomoću manualnog alata za mjerenje. Očitane su vrijednosti prosječne, najniže i najviše temperature unutar označene elipse, a prikupljeni podaci analizirani su pomoću Graphpad Prism statističkog programa. REZULTATI: šest sudionika istraživanja imalo je PD signal u oba koljena veći od 0, 5 (0, 5 – 2, 5), što je definirano kao aktivna upala, dok su dva imala signal 0 u oba koljena, što je definirano kao izostanak upale. Prosječna vrijednost prosječne temperature koljena u sudionika s aktivnom upalom bila je 34, 51°C, a u sudionika bez upale 30, 02°C. Prosječna vrijednost najviše očitane temperature u prvoj skupini bila je 35, 81°C, a u drugoj 31, 5°C, dok je prosječna vrijednost razlike najviše i najniže temperature bila 4, 95°C u prvoj skupini, a 2, 87°C u drugoj. Statistička analiza pokazala je da su razlike u svim navedenim mjerenjima između skupine sudionika s aktivnom upalom i skupine bez aktivne upale koljena bile značajne (p

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.

Sindrom SAPHO (Sinovitis, Akne, Pustuloza, Hiperostoza i Osteitis) rijetka je autoinfl amatorna bolest koja se prezentira širokom paletom kliničkih simptoma, a najčešće muskuloskeletalnim poremećajima i kožnim promjenama. Kako se simptomi muskuloskeletnog sustava i kože rijetko pojavljuju istodobno, postavljanje dijagnoze i započinjanje adekvatnog liječenja nerijetko je odgođeno. Prikazujemo slučaj 15-godišnjeg dječaka upućenog u našu ustanovu zbog poliartralgija i polimijalgija te mišićne slabosti koje su rezultirale otežanom pokretljivošću. Bolesnik je pet mjeseci prije hospitalizacije započeo liječenje izotretinoinom što mu ga je propisao dermatolog zbog profundnih akni na licu i gornjem dijelu trupa. Ubrzo nakon uvođenja izotretinoina u terapiju javljaju se bolovi u leđima, mišićima obiju natkoljenica, potom i u desnoj ruci i ramenu koji kroz sljedeća dva mjeseca progrediraju do bolesnikove potpune nepokretnosti, usprkos uvođenju terapije nesteroidnim antireumaticima (NSAR), peoralnim glukokortikoidima (GK) te ukidanju izotretinoina. Iz opsežne obrade izdvajaju se povišeni upalni parametri (SE 65, CRP do 82, 2 mg/l), nalaz sakroileitisa na učinjenom MR-u kralježnice uz negativne imunološke nalaze (ANA, anti ds-DNA, ENA , ANCA) i miopatski EMNG. Na scintigrafi ji skeleta Tc99m nađeno je multifokalno nakupljanje radiofarmaka u kostima ramenog obruča, 180 XI. KONGRES HRVATSKOG PEDIJATRIJSKOG DRUŠTVA PAEDIATR CROAT. 2014 ; 58 (SUPPL 2):65-198 torakalne kralježnice te pubične simfi ze uz uredan nalaz MR mozga, EEG-a te urednu kardiološku obradu. Nakon provođenja terapije klindamicinom i azitromicinom uz NSAR i GK dolazi do promptnog poboljšanja u kliničkom statusu uz urednu pokretljivost i normalizaciju hoda bolesnika, oporavka mišićne snage i smirivanja kožnih promjena. Scintigrafski nalaz učinjen sedam mjeseci nakon terapije bio je uredan, upalni parametri mirni, EMNG urednog nalaza. Kod bolesnika je postavljena dijagnoza sindroma SAPHO uz miopatiju proksimalne muskulature kao nuspojave liječenja izotretinoinom. Iako je poznato da terapija peroralnim izotretioninom uzrokuje miopatske nuspojave, uglavnom je riječ o blagim mijalgijama i ukočenosti. Pojava upalne miopatije ili rabdomiolize opisana je u malom broju pacijenata. Sindrom SAPHO se u literaturi klasifi cira kao izrazito rijedak entitet, zasad nedostatno razjašnjene etiologije. Prema postojećim hipotezama moguće je da je riječ o direktnom djelovanju mikroorganizama na kost ili, prema novijim istraživanjima, o autoinfl amatornom odgovoru potaknutom bakterijskim ili virusnim uzročnicima, što povezuje ovu bolest sa seronegativnim spondiloartritisima. Zato je liječenje, koje uz antibiotik uključuje i NSAR, brzo i učinkovito. Kad u kliničkoj slici dominiraju dermatološki poremećaji, vrijeme do postavljanja dijagnoze može se produžiti, često i komplicirati nuspojavama terapije, kao što je to bio slučaj kod prikazanog bolesnika. Brzo prepoznavanje simptoma i ciljana obrada dovest će do pravodobnog postavljanja dijagnoze, početka liječenja i brzog smirivanja tegoba.