Figure 7.

MCT4 was a directly target of miR‐92b‐5p in skeletal muscle. (A) Graphical representation of the conserved miR‐92b‐5p binding motifs within the 3′‐UTR of MCT4. Complementary sequences to the seed regions of miR‐92b‐5p within the 3′‐UTRs are conserved between human (Homo) and mouse (Mus) sequences. (B) C2C12 cells were treated with miR‐92b‐5p mimic or control (Ctrl) RNA and qPCR analysis was used to determine the mRNA level of MCT4 (n = 5). (C) Luciferase (luc) activity of the reporter constructs containing either wild‐type ormutated (MT) 3′‐UTR of murine MCT4 after treatment of C2C12 cells with miR‐92b‐5p mimic (M92M5) or Ctrl RNA (n = 5). (D) C2C12 cells treated with M92M5 (or miR‐92b‐5p inhibitor, M92I5) or Ctrl RNA under hypoxia, the protein level of MCT4 and β‐actin were determined by western blot and the quantitative result of western blot were shown (n = 3), intracellular lactate was detected by commercial kit (n = 3). (E) C2C12 cells treated with M92I5 and/or Mct4 siRNA under hypoxia, the protein level of MCT4 and β‐actin were determined by western blot, and the intracellular lactate was detected by commercial kit (n = 3). (F) C2C12 cells treated with M92M5 and/or AdMct4 under hypoxia, the protein level of MCT4 and β‐actin were determined by western blot, and the intracellular lactate was detected by commercial kit (n = 3). All results are expressed as means ± SD. *P < 0.05, **P < 0.01, ***P < 0.001, by unpaired Student's t test (B to F). *P < 0.05, **P < 0.01, ***P < 0.001, by a one‐way ANOVA with a Bonferroni correction test (G, H).