Figure 6

Transglutaminase 2 (TG2) inhibits the basal and the monosodium urate (MSU) crystal-induced JAK2 activation. (A) RAW264.7 cells stably transfected with TG2 or empty vector (mock) were stimulated with or without MSU crystals (1 mg/ml) for the indicated time points. Whole-cell lysates were analyzed by western blot analysis using antibodies against JAK2, phospho-JAK2 and β-actin. The ‘Fold’ induction was measured by densitometry analysis and normalized to β-actin. (B) Wild-type and TG2 null MEF cells were treated for 1 h with or without MSU crystals (2.0 mg/ml). Whole-cell lysates were analyzed by western blot analysis using antibodies against JAK2, phospho-JAK2 and β-actin. β-actin was used as a loading control. The ‘Fold’ induction was measured by densitometry analysis and normalized to β-actin. (C-D) Wild-type and TG2 null MEF cells were treated for 36 h with or without MSU crystals (2.0 mg/ml) or in the presence of the JAK2 inhibitor (AG490). (C) IL-1β and (D) TGF-β1 in the supernatants were evaluated by ELISA. (E) Experiments described in C were repeated in the presence of 0.5 g/ml neutralizing anti-TGF-β antibody and its isotype control. The results are expressed as the means ± SEM from three independent experiments (n ≧ 3, * = P < 0.05 compared with control (100%)).