Page 104 - 2019_03-Haematologica-web

K. Jambrovics et al.
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Figure 4. Tissue transglutaminase (TG2) up-regulates both expression of tumor necrosis factor alpha (TNF-α), interleukin 1 beta (IL-1b), and monocyte chemoat- tractant protein 1 (MCP-1) through nuclear factor kappa (κ)-light-chain-enhancer of the activated B-cell (NF-κB) pathway transcriptional activation and their secre- tion. Relative mRNA expression of (A1) TNF-α, (B1) IL-1b and (C1) MCP-1 in 1 mM ATRA or 1 mM ATRA plus 30 mM NC9-treated NB4-WT, TG2-C, TG2-KD, TG2-ha, and TG2-KO cells measured at the indicated days by real-time Q-PCR and normalized to cyclophilin-D mRNA expression. In the cell culture, supernatant-secreted (A2) TNF- α chemokine, (B2) IL-1β and (C2) MCP-1 inflammatory cytokines were quantified by ELISA and normalized to 100 mg protein content of cell lysate. Figures show secreted protein levels from 3 independent experiments measured in triplicate and graphs show the representation of the mean±Standard Deviation. (D) All-trans retinoic acid (ATRA) induced NF-κB transcriptional activity on NF-κB responsive luciferase reporter gene. Treated and harvested cells were analyzed for luciferase reporter activity as relative light units (RLU). Luciferase activity measurements were performed in triplicates (n=9). Statistical significance was determined via the two-way analysis of variance (ANOVA; Bonferroni post-hoc test; *P<0.05, **P<0.01, ***P<0.005 ****P<0.001).
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haematologica | 2019; 104(3)