2.11. Animal model

Animal experiments were approved by the Animal Care and Use Committee of Shanghai Jiao Tong University Affiliated Sixth People's Hospital. Male BALB/c nude mice (4–6 weeks old) were kept in specific pathogen-free conditions. MG63 cells transduced with lentivirus expressing shTRIM7 or shNC, and SAOS2 cells transduced with lentivirus expressing TRIM7, BRMS1, TRIM7 plus BRMS1 or control vector, were injected via the tail vein into the nude mice (1 × 10⁶ cells/mouse) (n = 11 per group). At 21 days after injection, mice were sacrificed and the lung tissues were collected and stained in hematoxylin and eosin (H&E) as previously described [19]. Otherwise, mice were injected via the tail vein with MG63 or SAOS2 cells and intraperitoneal injection of 1 mg/kg ADR every 2 days or 0.2 mg/kg MTX every 7 days (n = 20 per group), and then, lung metastasis and survival time were measured. To establish the patient-derived xenograft (PDX) model, tumour tissues from osteosarcoma patients, divided into two groups according to the TRIM7 expression (high vs low according to IHC staining), were collected at the time of surgery at Shanghai Jiao Tong University Affiliated Sixth People's Hospital. These tissues (about 5 × 5 × 3 mm³) were subcutaneously transplanted into 6–8-week-old nude mice within 1 h of removal of tissues. ADR (1 mg/kg; every 2 days) and MTX (0.2 mg/kg; every 7 days) chemotherapy were initiated when tumour volumes reached around 700 mm³ (n = 5 per group). On day 21 days following the injection, mice were sacrificed and tumour size was measured.