Abstract
BackgroundOsteosarcoma (OS) is the most common aggressive bone tumor in children and teenagers. Doxorubicin (DOX) is a chemotherapeutic drug for OS. This study aims to reveal the effects and underneath mechanism of DOX treatment in OS progression.MethodsThe expression of circular_0000006 (circ_0000006), microRNA-646 (miR-646) and brain-derived neurotrophic factor (BDNF) was detected by quantitative real-time polymerase chain reaction (qRT-PCR). BDNF protein expression was determined by western blot. Cell proliferation was illustrated by cell counting kit-8 (CCK-8) and cell colony formation assays. Cell migration and invasion were revealed by transwell migration and wound-healing assays and transwell invasion assay, respectively. Cell apoptosis was demonstrated by flow cytometry analysis. The binding relationship of miR-646 and circ_0000006 or BDNF was predicted by circRNA interactome and targetscan online database, respectively, and verified by dual-luciferase reporter assay. The effects of circ_0000006 knockdown on tumor growth in vivo were manifested by in vivo tumor formation assay.ResultsCirc_0000006 expression and the mRNA and protein levels of BDNF were dramatically upregulated, and miR-646 expression was effectively downregulated in OS tissues or cells compared with control groups. Circ_0000006 expression and BDNF protein expression were lower, and miR-646 expression was higher in DOX treatment groups than in control groups in OS cells. Circ_0000006 knockdown repressed cell proliferation, migration and invasion, whereas promoted cell apoptosis under DOX treatment in OS cells; however, these effects were attenuated by miR-646 inhibitor. Additionally, circ_0000006 sponged miR-646 to bind to BDNF. Circ_0000006 silencing suppressed tumor growth in vivo.ConclusionCirc_0000006 knockdown promoted DOX-mediated effects on OS development by miR-646/BDNF pathway, which provided a theoretical basis in treating OS with DOX.
Highlights
Osteosarcoma (OS) is a prime bone tumor caused by mesenchymal cells in children and teenagers [1, 2]
Circ_0000006 expression was dramatically downregulated by DOX exposure in OS cells In order to illustrate the characteristic of circ_0000006 in DOX-mediated OS progression, its expression was firstly determined in OS tissues and cells
RNase R treatment assay manifested that circ_0000006 expression had no dramatic change after RNase R treatment in U2OS and SJSA1 cells, whereas GAPDH mRNA level was obviously downregulated after RNase R exposure (Fig. 1C), which suggested that circ_0000006 was a circular RNA
Summary
Osteosarcoma (OS) is a prime bone tumor caused by mesenchymal cells in children and teenagers [1, 2]. Amuti et al J Orthop Surg Res (2021) 16:645 is chosen as the first-line drug in treating OS [6]. DOX is inoperative and results in drug resistance at a low dose and is highly toxic at a high dose [7]. Exploring the mechanism of DOX treatment in OS progression is urgent for its clinical application. Osteosarcoma (OS) is the most common aggressive bone tumor in children and teenagers. Doxorubicin (DOX) is a chemotherapeutic drug for OS. This study aims to reveal the effects and underneath mechanism of DOX treatment in OS progression
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