Abstract
BackgroundOsteosarcoma (OS) is one of the most common malignant bone tumors in children and adolescents. Circular RNAs (circRNAs) are critical regulators involved in multiple physiological and pathological processes. However, the underlying regulatory mechanisms of circRNA in OS are still not fully understood.MethodsThe circRNA expression profiles were downloaded from the Gene Expression Omnibus (GEO) database and analyzed by GEO2R. Bioinformatics analysis was performed to predict the potential target miRNAs of hsa_circ_0069117 and its downstream mRNAs. The co-expression of hsa_circ_0069117/miR-875-3p/PF4V1 axis was further validated in OS tissue samples via quantitative real-time PCR (qRT-PCR). Luciferase reporter gene plasmids containing the sequence of PF4V1 and hsa_circ_0069117 were constructed to verify the putative sites of miR-875-3p. Gain/loss-of-function assays were performed to verify the effect of hsa_circ_0069117 on miR-875-3p/PF4V1 expression and related pathways via qRT-PCR and Western blot. Cell counting kit-8 (CCK-8) and wound-healing assays were performed to evaluate the effect of hsa_circ_0069117 on cell proliferation and migration of MG63 and U2OS, respectively.ResultsWe identified hsa_circ_0069117 as the most markedly dysregulated circRNA in OS cell lines. Bioinformatics analysis indicated that hsa_circ_0069117 might inhibit the expression of miR-875-3p, thereby promoting the expression of platelet factor 4 variant 1 (PF4V1). The expression of miR-875-3p was negatively correlated to hsa_circ_0069117 and PF4V1 in clinical samples. Luciferase reporter gene assays confirmed the binding sites of miR-875-3p on hsa_circ_0069117 and PF4V1. Gain/loss-of-function and rescue assays further indicated that hsa_circ_0069117 could significantly promote the expression of PF4V1 by sponging miR-875-3p, thereby inhibiting the proliferation and migration of OS cells by suppressing ERK1 and AKT.ConclusionOur study revealed that hsa_circ_0069117 is an anti-OS molecule that could substantially attenuate cell proliferation and migration of OS, which may provide a novel and reliable molecular target for the treatment of OS patients.
Highlights
Osteosarcoma (OS) is the most common primary malignant bone tumor accounting for 10% of solid tumors in children and adolescents [1, 2]
Cell culture Human osteosarcoma cell lines (OSCL) U2OS, 143B, MG63, HOS, and human mesenchymal stem cell line hMSCs were obtained from the American Type Culture Collection (ATCC)
A total of 58 upregulated and 126 downregulated miRNAs were detected in GSE70367 (Fig. 1B). Among these differentially expressed Circular RNA (circRNA), hsa_circ_0069117 was the most markedly dysregulated in OS cells lines
Summary
Osteosarcoma (OS) is the most common primary malignant bone tumor accounting for 10% of solid tumors in children and adolescents [1, 2]. Circular RNAs (circRNAs) were discovered decades ago [8, 9]. It is a type of ncRNA (non-coding RNA) that has been suggested to act as an essential biomolecule in regulating multiple progression of diseases [10, 11]. Circular RNAs have a unique feature to resist most RNases, enabling them to be ideal biomarker for detecting human diseases [9, 12]. It has been proven that circRNAs can bind miRNAs and proteins thereby regulating multiple physiological and pathological processes [13]. Circular RNAs (circRNAs) are critical regulators involved in multiple physiological and pathological processes. The underlying regulatory mechanisms of circRNA in OS are still not fully understood
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