Fig. 5

The self-assembled OTP-BTFPTU supramolecular liposomes inhibits the proliferation and migration of HOS cells in vitro. (A) Schematic illustration of osteosarcoma targeting peptide (OTP)-BTFPTU supramolecular liposomes prepared through self-assembly. (B) The TEM images of OTP-BTFPTU liposomes. Scale bar, 100 nm. (C) FITC (green), DID (red) and DAPI (blue) immunofluorescence of HOS cells with or without OTP-BTFPTU (2 µM) liposomes treatment for 3 h. Scale bar, 20 μm. (D) Cell viability of HOS cells with OTP-PTU, OTP-TFPTU or OTP-BTFPTU liposomes treatment for 24 h using Cell Counting Kit-8 assay. (E) HOS cells were treated with different concentrations of OTP-BTFPTU liposomes for 24 h and the cell proliferation was characterized by EdU assay. Scale bar, 200 μm. (F) Flow cytometry to evaluate apoptosis of HOS cells with or without OTP-BTFPTU liposomes (2 µM) treatment for 12 h by Annexin V-FITC/PI apoptosis detection. (G) Quantification of apoptosis rate of HOS cells. (H) Cell migration abilities of HOS cells treated with different concentrations of OTP-BTFPTU liposomes for 24 h were measured by Transwell migration assays. Scale bar, 200 μm. (I) Quantification of the migration abilities of HOS cells. (J) The wound healing assay was used to evaluate the migration abilities of HOS cells treated with different concentrations of OTP-BTFPTU liposomes for 24 h. Scale bar, 50 μm. (K) Quantification of the wound closure rate. (L) The protein expression of N-Cadherin, Vimentin and c-Myc was measured by Western blot analysis in HOS cells treated with varying concentrations of OTP-BTFPTU liposomes for 24 h. (M) Quantification and normalization of the gray levels of N-Cadherin, Vimentin and c-Myc proteins to that of GAPDH in HOS cells using Image J. The data represent the mean ± SD of three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001 for a comparison with the control group or as indicated