IL-33/ST2 osovina u eksperimentalnom modelu akutnog zapaljenja

Abstract

Interleukin-33 (IL-33) je jedini poznat ligand za ST2 receptor. Akutno zapaljenje je rani odgovor tkiva na oštecenje. Iako je njegova osnovna uloga zaštitna, može dovesti do dodatnog oštecenja tkiva i razlicitih poremecaja u organizmu. Cilj ovog istraživanja je bio razjašnjavanje uloge IL-33/ST2 osovine u patogenezi akutnog zapaljenja. Korišceni su genetski neizmenjeni miševi (WT) i miševi kojima je uklonjen gen za ST2 receptor (ST2-/-), i bili su rasporedeni u grupe: 1. WT-C (kontrolna WT grupa), 2. KO-C (kontrolna ST2-/- grupa), 3. WT-I (eksperimentalna WT grupa kod koje je izazvano akutno zapaljenje) i 4. KO-I (eksperimentalna ST2-/- grupa kod koje je izazvano akutno zapaljenje). Akutno zapaljenje je izazvano davanjem intramuskularne injekcije terpentinskog ulja, a kontrolne životinje su primile injekciju fiziološkog rastvora. Nakon 12 sati, životinje su uvedene u opštu anesteziju i sakupljeni su uzorci tretiranih mišica, slezine i krvi. Uzorci mišica su korišceni za analizu genske ekspresije ST2 receptora i citokina (IL-33, TNF-alfa, IL-6, IL-12p35, TGF-beta), za odredivanje koncentracije makro- i mikroelemenata (Mg, K, Fe, Cu, Mn, Se), i antioksidativnih parametara (GSH, GPx i SOD), histopatološku analizu i odredivanje volumenske gustine zapaljenjskog infiltrata (Vgz), volumenske gustine nekroticnih mišicnih celija (Vgn), fraktalne dimenzije (D), ugaonog drugog momenta (ASM) i teksturalne korelacije (COR). Odreden je procenat eritrocita u slezini. Uzorci krvi su korišceni za odredivanje broja eritrocita, hematokrita, koncentracije hemoglobina, prosecne kolicine hemoglobina u eritrocitu, broja neutrofila i limfocita i odredivanje aktivnosti CK i AST. Histopatološkom analizom tretiranog tkiva je utvrdeno postojanje zapaljenjskog infiltrata i nekroticnih celija u obe zapaljenjske grupe. Vgz, CK i AST su bili veci u prisustvu IL-33/ST2 osovine u akutnom zapaljenju. Genska ekspresija IL-33 i ST2 receptora je bila povecana na mestu akutnog zapaljenja. Genska ekspresije TGF-beta na mestu akutnog zapaljenja bila je veca u odsustvu IL-33/ST2 osovine. Fe je bilo vece u WT-I u odnosu na WT-C i KO-I, a znacajne razlike izmedu KO-C i KO-I nije bilo. Prosecna kolicina hemoglobina u eritrocitu bila je znacajno manja u WT-I u odnosu na WT-C, a nije bilo razlike izmedu KO-C i KO-I. Mg je bio manji u WT-I u odnosu na WT-C i KO-I, dok nije bilo razlike izmedu KO-C i KO-I. K se smanjuje u zapaljenju, ali je bio manji u WT-I u odnosu na KO-C. GSH je bio veci u KO-I u odnosu na KO-C i WT-I. Na osnovu rezultata ovog istraživanja može se zakljuciti da IL-33/ST2 osovina doprinosi intenzivnijem zapaljenju i oštecenju tkiva u akutnom zapaljenju uticuci na koncentraciju magnezijuma i gvožda, sadržaj redukovanog glutationa i gensku ekspresiju TGF-beta na mestu akutnog zapaljenja. U akutnom zapaljenju se povecava genska ekspresija interleukina-33 i ST2 receptora.

Interleukin-33 (IL-33) is the only known ligand for ST2 receptor. Acute inflammation is an early response to the tissue damage. Although a primarily role of inflammation is protective, it may also cause additional tissue damage and different disorders in organism. The aim of this study was to examine the role of IL-33/ST2 axis in pathogenesis of acute inflammation. Wild type (WT) and ST2 knockout (ST2-/-) mice were used and divided into groups: WT-C (wild-type control group), KO-C (ST2-/- control group), WT-I (wild-type inflammatory group), and KO-I (ST2-/- inflammatory group). Acute inflammation was induced by intramuscular injection of turpentine oil, while control animals received saline. After 12 hours, animals were euthanized and samples of treated muscles, spleen and blood were collected. Muscle samples were used for determination of ST2 and cytokine gene expression (IL-33, TNF-alpha, IL-6, IL-12p35, TGF-beta), macro- and microelement concentration (Mg, K, Fe, Cu, Mn, Se), antioxidative parameters (GSH, GPx and SOD), histopathological analysis and determination of volume density of inflammatory infiltrate (Vdz) and necrotic fiber (Vgn), fractal dimension (D), angular second moment (ASM), and textural correlation (COR). Erythrocytes percentage in spleen was determined. Blood samples were used for determination of erythrocyte blood count, hematocrit, hemoglobin concentration, mean corpuscular hemoglobin, neutrophil and lymphocyte blood count, and creatine kinase (CK) and aspartate aminotransferase (AST) activity. Presence of inflammatory infiltrate in the treated tissue was confirmed by histopathological analysis. Vgz, CK and AST were higher in presence of IL-33/ST2 axis in acute inflammation. IL-33 and ST2 gene expression increased in acute inflammation. TGF-beta gene expression at site of acute inflammation was higher in absence of IL-33/ST2 axis. Fe was higher in WT-I than in WT-C and KO-I, while there was no significant difference between KO-C and KO-I. Mean corpuscular hemoglobin was lower in WT-I than in WT-C, while there was no significant difference between KO-C and KO-I. Mg was lower in WT-I than in WT-C and KO-I, while there was no difference between KO-C and KO-I. K decreased in inflammation, but it was lower in WT-I than in KO-C. GSH was higher in KO-I when compared to KO-C and WT-I. These results indicate that IL-33/ST2 axis contributes to development of more intensive inflammation and tissue damage in acute inflammation by affecting concentration of magnesium, iron concentration, reduced glutathione and TGF-beta gene expression at the site of acute inflammation. Gene expression of IL-33 and ST2 increases in acute inflammation.