MicroRNA-630 alleviates inflammatory reactions in rats with diabetic kidney disease by targeting toll-like receptor 4

Figure 3 Toll-like receptor 4 is the target gene of miR-630. A: Bioinformatic analysis showed the putative miR-630 target sites in the toll-like receptor 4 (TLR4) 3’-untranslated regions (UTR). The mutated nucleotides are underlined; B: The WT-TLR4 3’-UTR and the MUT-TLR4 3’-UTR reporters were cotransfected with miR-630 mimic or negative control into NRK-52Es. Forty-eight hours after transfection, the luciferase activities were measured; C: Quantitative reverse transcription polymerase chain reaction detection of miR-630-targeted downregulation of TLR4 mRNA expression levels; D: Enzyme-linked immunosorbent assay detection of tumor necrosis factor in miR-630-targeted downregulation of TLR4, α-smooth muscle actin (SMA), interleukin (IL)-1β, and IL-6 content; E: Western blotting was used to detect the protein expression of TLR4, α-SMA, collagen IV and E-cadherin downregulated by miR-630. Data are presented as mean ± SD, ^aP < 0.01 vs high glucose + mimic normal glucose, ^bP < 0.01 vs high glucose + miR-630 mimic + oe normal glucose. NG: Normal glucose, 5.6 mmol/L; HG: High glucose, 20 mmol/L; HM: High mannitol (5.6 mmol/L glucose + 14.4 mmol/L mannitol); TLR4: Toll-like receptor 4; α-SMA: α-smooth muscle actin; IL: Interleukin; TNF: Tumor necrosis factor; NC: Negative control.