Analiza proupalnog (IL-18) i protuupalnog (IL-10) citokina u bolesnika s juvenilnim idiopatskim artritisom [ Analysis of pro-inflammatory (IL-18) and anti-inflammatory (IL-10) cytokne and regulatory T cells in patients with juvenile idiopathic arthritis (JIA)

Jelušić, Marija (2005) Analiza proupalnog (IL-18) i protuupalnog (IL-10) citokina u bolesnika s juvenilnim idiopatskim artritisom [ Analysis of pro-inflammatory (IL-18) and anti-inflammatory (IL-10) cytokne and regulatory T cells in patients with juvenile idiopathic arthritis (JIA). PhD thesis, Sveučilište u Zagrebu.

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Abstract

Introduction: Juvenile idiopathic arthritis (JIA) is multisystemic autoimmune inflammatory disease. Although various factors have been proposed as the key mediators, immunopahtogenesis of JIA is still not clear. The aim of my study was to elucidate the role of regulatory T cells (TR) as well as interleukin (IL)-18 and IL-10, as representatives of pro-inflammatory and anti-inflammatory cytokines, respectively. Patients and Methods: Fifty children with JIA (mean age 5.5 years (range: 1 – 18); 31 girls, 19 boys; 25 with oligoarticular JIA, 15 with polyarticular JIA and 10 with systemic JIA) and 23 healthy children (mean age 7 years (range: 2 – 16)) were enrolled. Blood samples from patients were obtained in active and inactive phase of disease. Synovial fluid samples were collected from 16 patients with oligoarticular JIA. The levels of TR (defined as CD4+CD25++ or CD4+CD25++CCR4+) were analysed by flow cytometry. Concentrations of IL-18 and IL-10 were determined in blood serum, synovial fluid, and supernatants of phorbol-myristate-acetate -stimulated and unstimulated peripheral blood mononuclear cells (PBMC) using commercial ELISA kits. Results: Serum levels of IL-18 in systemic JIA were significantly higher than in other forms of disease, both in active and inactive phase (P < 0.001), while the IL-18 levels in children with oligoarticular and polyarticular JIA were comparable to the healthy controls. Although the concentrations of IL-18 in systemic JIA decreased significantly during remission (P = 0.004), they remained several-fold higher than in the other groups (P < 0.001). JIA patients had higher concentrations of IL-10 in serum than healthy children, but the difference was significant only during the remission (P = 0.004). On the other hand, PBMC of JIA patients produced more IL-10 in both active and inactive phase of disease (P < 0.001). In addition, more IL-10 was detected in synovial fluid than in blood serum (P = 0.001). The proportion and number of CD4+CD25++ lymphocytes in children with JIA increased with activity of the disease (from 0.16% to 0.30% of total lymphocytes), approaching the levels found in healthy children (0.39%). No consistent pattern of CD4+CD25++ and CD4+CD25++CCR4+ lymphocyte counts was found regarding the subtype of JIA or phase of disease. Conclusion: These results underline the importance of IL-18 in the pathogenesis of systemic JIA. The levels of IL-18 correlated rather well with the other indicators of systemic inflammation and were relatively high even during clinical remission. It seems that the IL-18 was produced systemically while the inflamed joints were the main site of production of IL-10. High concentrations of pro-inflammatory factors (IL-18) together with lower levels of anti-inflammatory mediators (IL-10 and TR) in inactive phase of disease, indicates that even in clinical remission there is activity of the immune system and that this “silent” phase should not be neglected, as it is usually. These findings imply that new strategies of clinical management of JIA and related disorders should be focused not only on suppression of overt inflammation in active phase, but on control of immune process in remission as well.

Abstract in Croatian

Uvod: Juvenilni idiopatski artritis (JIA) višesustavna je autoimunosna bolest. Unatoč mnogim istraživanjima ključnih medijatora u JIA, imunopatogeneza još je nepoznata. Cilj rada bio je istražiti ulogu regulacijskih limfocita T (TR), kao i IL-18 i IL-10, predstavnika proupalnih i protuupalnih citokina. Ispitanici i metode: U istraživanje je bilo uključeno 50-ero djece s dijagnozom JIA (prosječne dobi 5,5 godina (1-18); 31 djevojčica i 19 dječaka; 25-ero s oligoartikularnim tipom, 15-ero s poliartikularnim tipom i desetero sa sistemskim tipom JIA) i 23-oje djece kontrolne skupine (prosječne dobi 7 godina (2–6)). U svih bolesnika uzeta je venska krv u aktivnoj fazi i u remisiji bolesti. U 16 bolesnika s oligoartikularnim tipom uzeta je i sinovijalna tekućina. Broj TR (definiranih kao CD4+CD25++ i CD4+CD25++CCR4+) analiziran je protočnom citometrijom. Koncentracije citokina IL-18 i IL-10 u uzorcima seruma, sinovijalne tekućine i nadtaloga kultura stimuliranih (s forbol-miristat-acetatom) i nestimuliranih limfocita izmjerene su pomoću komercijalnih ELISA kitova. Rezultati: Serumske razine IL-18 u bolesnika sa sistemskim tipom JIA bile su značajno povišene u odnosu prema drugim tipovima JIA, u aktivnoj fazi i u remisiji bolesti (P < 0,001), dok nije nađena značajna razlika između bolesnika s oligoartikularnim i poliartikularnim tipom JIA i kontrolne skupine. Iako se razina IL-18 u bolesnika sa sistemskim tipom JIA značajno smanjila u remisiji bolesti (P = 0,004), i dalje je ostala nekoliko puta viša u usporedbi s drugim tipovima JIA (P < 0,001). Serumske razine IL-10 u bolesnika s JIA bile su više nego u kontrolnoj skupini, međutim razlika je bila statistički značajna samo u remisiji bolesti (P = 0,004), dok su razine IL-10 u nadtalogu kultura stimuliranih i nestimuliranih limfocita bolesnika bile značajno više i u aktivnoj fazi i u remisiji bolesti u odnosu prema kontrolnoj skupini (P < 0,001). U sinovijalnoj tekućini razina IL-10 značajno je viša od one u serumu (P = 0,001). Broj i udio CD4+CD25++ limfocita u bolesnika s JIA povećavao se s aktivnošću bolesti (od 0,16% do 0,30% ukupnih limfocita), dostižući nalaz kontrolne skupine (0,39%). Nije nađen dosljedan uzorak broja i udjela CD4+CD25++ i CD4+CD25++CCR4+ limfocita s obzirom na tip ili fazu bolesti. Zaključak: Navedeni rezultati govore u prilog važnoj ulozi IL-18 u patogenezi sistemskog tipa JIA. Razina IL-18 povezana je s laboratorijskim nalazima sustavne upale, a ostaje povišenom i za vrijeme remisije bolesti, što upućuje na to da se IL-18 proizvodi sustavno, dok su limfociti u upalom zahvaćenom zglobu glavni izvor IL-10. Nalaz visokih razina proupalnih medijatora (IL-18) zajedno s niskim razinama protuupalnih medijatora (IL-10 i TR) u remisiji bolesti upućuje na to da za vrijeme «klinički mirne» faze postoji aktivnost imunološkog sustava, te da remisija ne smije biti zanemarena kao što je to danas. Stoga, nove smjernice u liječenju JIA i srodnih bolesti trebaju biti usmjerene ne samo na supresiju upale u aktivnoj fazi, nego i na kontrolu imunosnog sustava u remisiji bolesti.

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