The hFOB1.19 (#CRL-11372) human osteoblast cell line and the DAN (#CRL-2130), MG63 (#CRL-1427), HOS (#CRL-1543), T1-73 (#CRL-7943), 143B (#CRL-8303), Saos2 (#HTB-85), and U2OS (#HTB-96) osteosarcoma cell lines were purchased from American Type Culture Collection (ATCC) (Manassas, VA, USA). All cell lines were cultured in Dulbecco's Modified Eagle's Medium (DMEM) (Corning, USA, #10-017-CM) containing 10% fetal bovine serum (FBS) (Sigma-Aldrich, China, #F2442) and 100 units/mL of penicillin-streptomycin (Sigma-Aldrich, #P4333). All osteosarcoma cell lines were grown in a 37 °C incubator, and the hFOB1.19 cells were cultured at 34 °C. For cell transfection with shRNAs, the lentiviral transduction particles of CtBP1 (Sigma-Aldrich, #TRCN0000285086) and HIPK2 (Sigma-Aldrich, # TRCN0000433047) and one control particle containing pLKO.1 empty vector were transfected into cells using FuGene 6 (Roche Diagnostics Corp., USA, #E2691) following the manufacturer's protocol. The transfected cells were allowed to recover for 12 h and then selected with 1 μg/mL puromycin (Sigma-Aldrich, #540411). Single puromycin-resistant cells were collected and used to confirm gene expression. Cells showing successful knockdown of CtBP1 and HIPK2 were used for subsequent experiments. For cell transfection with plasmids, the pCDNA3-HIPK2 and pCDNA3 empty vectors were individually transfected into cells with the Lipofectamine 3000 reagent (Thermo Fisher Scientific, China, #L3000015). Cells showing successful transfection of HIPK2 were used in subsequent experiments.

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