Figure 2.

Effects of human Arg2 TSBs on Arg2, TNF-α, and IL-6 in human MDMs

(A) Schematic of ARG2 3′ UTR mRNA consisting of 785 base pairs (bp) and the predicted miRNA binding sites to the microRNA recognition elements. (B) Schematic illustrating complimentary base pairing of TSB-155-2 and miR-155 with a region in of the 3′ UTR of ARG2 mRNA. Pro-inflammatory cytokine secretion by MDMs transfected with TSBs (100 nM) and stimulated with LPS was analyzed for (C) TNF-α and (D) IL-6 by ELISA and graphed as a percentage relative to the negative control TSB (NC-TSB) stimulated with LPS (n = 8 donors, using experimental triplicates). (E) ARG2 was analyzed by qRT-PCR using TBP as the endogenous control (n = 4). (F) Arg2 protein was analyzed in TSB and LPS stimulated cells by western blot using glyceraldehyde 3-phosphate dehydrogenase (GAPDH) as the control. (G) Densitometry analysis was performed on the Arg2 western blots and normalized to GAPDH and graphed as relative expression to the NC-TSB with LPS (n = 5). Pro-inflammatory gene expression of (H) TNFA was analyzed by RT-PCR using TBP as the control (n = 5). Error bars are representative of the standard error of the mean (SEM). Statistical analysis was performed using multiple independent unpaired t tests for (C and D) and using a one-way ANOVA with Dunnett’s multiple comparison test for (E–H). ns, not statistically significant, ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001.