Circular RNA circBLNK promotes osteosarcoma progression and inhibits ferroptosis in osteosarcoma cells - Abstract and study methods

Sep 24, 2026

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Circular RNA circBLNK is examined for its function in osteosarcoma tumorigenesis and progression. The study quantifies mRNA and protein levels using RT-qPCR and western blotting, determines circBLNK subcellular localization, and measures ferroptosis, proliferation, and apoptosis with MTT, CCK-8, clonogenic assays, and flow cytometry. circBLNK/miR-188-3p/GPX4 interactions are tested via luciferase reporter, RNA pull-down, and RNA immunoprecipitation, with stability confirmed by RNase R. Clinical association and mechanistic effects are evaluated through survival analysis and tissue expression statistics.

LanguageEnglish

CategoryHealthcare

FormatPDF

Pages11

Reading Time28 min

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Abstract

Study objective

Methods and assays

circBLNK–miR-188-3p–GPX4 axis

Introduction

Osteosarcoma background and challenges

Circular RNAs and ceRNA mechanisms

miR-188-3p and GPX4/ferroptosis context

ONCOLOGY REPORTS 50: 192, 2023 Circular RNA circBLNK promotes osteosarcoma progression and inhibits ferroptosis in osteosarcoma cells by sponging miR‑188‑3p and regulating GPX4 expression ZHONGJUN LI 1* , YI LUO2* , CHUNBO WANG 1* , DUNXIN HAN 1 and WEIPING SUN 1 1Department of Orthopaedics, Weihai Medical District of The 970th Hospital of The PLA Joint Logistic Support Force, Weihai, Shandong 264200; 2Department of Orthopaedics, The 970th Hospital of The PLA Joint Logistic Support Force, Yantai, Shandong 264002, P.R. China Received November 10, 2022; Accepted July 4, 2023 DOI: 10. 3892/or.2023.8629 Abstract. As a newly identified circular RNA (circRNA), the role of circBLNK in cancer progression has not been probed. The objective of the present study was to functionally dissect the role of circBLNK in osteosarcoma (OS) tumorigenesis and progression. With regards of the experimental procedure, the levels of mRNAs and proteins were assessed using reverse transcription‑quantitative PCR and western blot analysis, respectively. The subcellular location of circBLNK in OS cells was determined by cell cytosolic/nuclear fractionation assay. Cell ferroptosis ability was assessed through MTT assay. Cell proliferative abilities were assessed by clonogenic and Cell Counting Kit‑8 assays, and cell apoptosis was measured using flow cytometry. The relationships among circBLNK, miR‑188‑3p, and glutathione peroxidase 4 (GPX4) were vali‑ dated by luciferase reporter and RNA pull‑down assays, as well as RNA immunoprecipitation. The stability of circBLNK and linear BLNK was confirmed using RNase R treatment assay. The association between circBLNK expression and overall survival rate was assessed by Kaplan‑Meier plot. The correla‑ tion between the expression levels of circBLNK, miR‑188‑3p, and GPX4 in OS tissues was assessed by Pearson's χ2 test. The results revealed that CircBLNK and GPX4 were significantly upregulated in OS tissues, which predicted the poor prognosis. CircBLNK knockdown led to suppressed cell proliferation and enhanced cell apoptosis, an effect that could be reversed by the inhibition of miR‑188‑3p. In an in vivo circBLNK Correspondence to: Dr Dunxin Han or Dr Weiping Sun, Department of Orthopaedics, Weihai Medical District of The 970th Hospital of The PLA Joint Logistic Support Force, 8 Baoquan Road, Weihai, Shandong 264200, P.R. China E‑mail: [13573518855@163.com](13573518855@163.com) E‑mail: [swp411@163.com](swp411@163.com) *Contributed equally Key words: osteosarcoma, CircBLNK, microRNA‑188‑3p, glutathione peroxidase 4 deficiency model, tumor growth was observed to be markedly suppressed. Moreover, circBLNK deficiency elevated levels of intracellular free iron (Fe2+), malondialdehyde, lipid reactive oxygen species and mitochondrial superoxide, while dimin‑ ishing mitochondrial membrane potential in Erastin‑treated OS cells, which were eliminated by overexpressing GPX4 . Furthermore, mechanistic investigations revealed that circBLNK sponged miR‑188‑3p to regulate the expression of GPX4, thereby affecting OS progression . In conclusion, the present study delineated a new regulatory axis involving circBLNK/miR‑188‑3p/GPX4 in OS progression, adding to the growing evidence that circRNAs are critical gene regulators in cancer progression. Introduction As a form of primary malignant bone tumor, osteosarcoma (OS) is ranked first in the cause of cancer‑associated death in children and adolescents (1,2) . OS is derived from mesen‑ chymal cells, characterized by a high recurrence rate, early lung metastasis, and rapid infiltrating growth (3) . Surgical resection, adjuvant chemotherapy, and small molecule targeted therapy have been used for OS treatment (4,5); however, because of the high propensity of metastasis and relapse the prognosis remains poor (6). Thus, it would be beneficial to identify novel OS thera‑ pies by studying complex gene regulatory networks. Circular RNAs (circRNAs) do not code for protei

FAQs

What is the main objective of the present study on circBLNK in osteosarcoma?

To functionally dissect how circBLNK contributes to osteosarcoma tumorigenesis and progression.

How do the experiments assess circBLNK effects on cell growth and death?

Cell proliferation is measured by clonogenic and CCK-8 assays, while apoptosis is quantified by flow cytometry; ferroptosis ability is assessed by MTT assay.

Which molecular axis is proposed to explain circBLNK’s role in osteosarcoma progression?

circBLNK is reported to sponge miR-188-3p to regulate GPX4 expression, thereby affecting osteosarcoma progression.

Sep 24, 2026

Uploaded by Adam