Research Paper Volume 13, Issue 8 pp 11433—11454

Overexpression of long non-coding RNA AP001505.9 inhibits human hyaline chondrocyte dedifferentiation

Mechanism of AP001505.9-dependent inhibition of dedifferentiation. (A) Fluorescence in-situ hybridization for AP001505.9. U6 and 18S were used as internal reference probes in the nucleus and cytoplasm, respectively. (B) Competing endogenous RNA (ceRNA) analysis of AP001505.9; blue circles represent cartilage-related genes; red circles represent miRNAs; green circles represent lncRNAs. (C) Real-time qPCR for miRNAs expressed after dedifferentiation. (D) Real-time qPCR for miRNAs expressed after redifferentiation. (E) Real-time qPCR for miRNAs after AP001505.9 overexpression. (F) Real-time qPCR for miR-495. (G) Real-time qPCR for AP001505.9 after miR-495 overexpression. (H) Real-time qPCR for COL1A1, COL2A1, and SOX-9 after miR-495 overexpression. (I, J) Western blot for COL1, COL2, and SOX-9 after miR-495 overexpression. (K) Binding analysis of AP001505.9 and miR-495. (L) Double luciferase assay for AP001505.9 and miR-495. LNC represents chondrocytes transfected by AP001505.9 lentivirus. miR495 represents chondrocytes transfected by miR-495 mimic. NC represents chondrocytes transfected by negative control lentivirus or mimic. GAPDH was used as the internal reference. Data are represented as means ± standard deviation. *P P P

Figure 7. Mechanism of AP001505.9-dependent inhibition of dedifferentiation. (A) Fluorescence in-situ hybridization for AP001505.9. U6 and 18S were used as internal reference probes in the nucleus and cytoplasm, respectively. (B) Competing endogenous RNA (ceRNA) analysis of AP001505.9; blue circles represent cartilage-related genes; red circles represent miRNAs; green circles represent lncRNAs. (C) Real-time qPCR for miRNAs expressed after dedifferentiation. (D) Real-time qPCR for miRNAs expressed after redifferentiation. (E) Real-time qPCR for miRNAs after AP001505.9 overexpression. (F) Real-time qPCR for miR-495. (G) Real-time qPCR for AP001505.9 after miR-495 overexpression. (H) Real-time qPCR for COL1A1, COL2A1, and SOX-9 after miR-495 overexpression. (I, J) Western blot for COL1, COL2, and SOX-9 after miR-495 overexpression. (K) Binding analysis of AP001505.9 and miR-495. (L) Double luciferase assay for AP001505.9 and miR-495. LNC represents chondrocytes transfected by AP001505.9 lentivirus. miR495 represents chondrocytes transfected by miR-495 mimic. NC represents chondrocytes transfected by negative control lentivirus or mimic. GAPDH was used as the internal reference. Data are represented as means ± standard deviation. *P < 0.05, **P < 0.01, ***P < 0.001. NS, not significant. Scale bar, 100 μm.