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. 2019 Jan 21;25(1-2):44–54. doi: 10.1089/ten.tea.2017.0484

FIG. 2.

FIG. 2.

Inhibition of PHD2 and miR-210. (A, B) Dose/response curves of PHD2 inhibition by sshRNAs targeting human (A) and mouse (B) PHD2 mRNAs. Efficacies of unmodified and 2′-OMe-modified sshRNAs (SG302 vs. SG302m (human) and SG402 vs. SG404 (mouse)) are shown over the indicated concentration range. Total RNA was isolated 48 h after transfections in NHEK cells (human) and NIH-3T3 (mouse). PHD2 transcripts were quantified by RT-qPCR using the ΔΔCt method, normalizing to GAPDH. Quantification is expressed as fold-inhibition relative to cells that were not transfected with inhibitors. (C) Luciferase biosensor assay to measure inhibition of miR-210 by antimiRs. Derepression of rLuc signal shows specific activity of miR-210-targeting antimiR SG608 relative to NSC control antimiR. (D) Specific inhibition of PHD2 and miR-210 by sshRNA and antimiR in cell culture. qPCR analysis of PHD2 transcript levels and miR-210 levels (red) in keratinocytes (HaCaT) 48 h after transfection with PHD2 sshRNA, miR-210 antimiR, or a combination of both. Quantification is expressed as fold-change relative to cells that were not transfected with inhibitors. mRNA, messenger RNA; NSC, nonspecific control; RT-qPCR, real-time quantitative polymerase chain reaction.