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Updated: Apr 20, 2026
Colony Formation Assay Detecting the Proliferative Capacity of LncRNA-knockdown Osteosarcoma Cells
Published on: January 16, 2026
A novel long non-coding RNA, hypoxia-inducible factor-2α promoter upstream transcript, functions as an inhibitor of
Yongcheng Wang1, Jie Yao2, Haoye Meng1
1Institute of Orthopedics, Chinese PLA General Hospital, Beijing 100853, P.R. China.
Abstract:
Long non‑coding RNAs (lncRNAs) have recently been identified as novel modulators of malignant tumors. However, the function of lncRNAs in cancer stem cells (CSCs) remains to be elucidated. The present study aimed to investigate the regulating role of a novel lncRNA, hypoxia‑inducible factor‑2α (HIF‑2α) promoter upstream transcript (HIF2PUT), in osteosarcoma stem cells. The expression levels of HIF2PUT were assessed by quantitative polymerase chain reaction in 17 osteosarcoma tissue specimens, and the correlation between the expression of HIF2PUT and its host transcript‑HIF‑2α was determined. In functional experiments, HIF2PUT expression was knocked down by small interfering RNAs, or overexpressed by transfection with pcDNA‑HIF2PUT, in order to evaluate the effects of HIF2PUT on cell proliferation, migration, expression rate of osteosarcoma stem cell marker CD133, and stem sphere‑forming ability in MG63 cells. HIF2PUT expression levels were positively correlated with HIF‑2α in osteosarcoma tissues. Overexpression of HIF2PUT markedly inhibited cell proliferation and migration, decreased the percentage of CD133 expressing cells, and impaired the osteosarcoma stem sphere‑forming ability of the MG63 cells. Whereas, knockdown of HIF2PUT expression had the opposite effect. Furthermore, altering the expression of HIF2PUT resulted in a concomitant change to HIF‑2α mRNA expression. These results indicate that the lncRNA HIF2PUT may be a novel regulatory factor of osteosarcoma stem cells, which may exert its function partly by controlling HIF‑2α expression. Further studies regarding HIF2PUT may provide a novel therapeutic target of osteosarcoma in the future.
Insights
Long non-coding RNA HIF2PUT regulates osteosarcoma stem cells by affecting proliferation and migration. This novel lncRNA may target cancer stem cells (CSCs) by influencing hypoxia-inducible factor-2α (HIF-2α) expression.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- Long non-coding RNAs (lncRNAs) are emerging as critical regulators in various cancers.
- The specific roles of lncRNAs in cancer stem cells (CSCs), particularly in osteosarcoma, require further investigation.
Purpose of the Study:
- To explore the regulatory function of a novel lncRNA, hypoxia-inducible factor-2α (HIF-2α) promoter upstream transcript (HIF2PUT), in osteosarcoma stem cells.
- To determine the correlation between HIF2PUT expression and its host gene, HIF-2α, in osteosarcoma tissues.
Main Methods:
- Quantitative polymerase chain reaction (qPCR) to assess HIF2PUT expression in 17 osteosarcoma tissues.
- Functional assays including small interfering RNA (siRNA)-mediated knockdown and plasmid-based overexpression of HIF2PUT in MG63 cells.
- Evaluation of cell proliferation, migration, CD133 expression, and stem sphere-forming ability.
Main Results:
- HIF2PUT expression was positively correlated with HIF-2α in osteosarcoma tissues.
- Overexpression of HIF2PUT inhibited cell proliferation and migration, reduced CD133+ cell percentage, and impaired stem sphere formation.
- Knockdown of HIF2PUT demonstrated opposite effects, and changes in HIF2PUT expression correlated with HIF-2α mRNA levels.
Conclusions:
- The lncRNA HIF2PUT acts as a novel regulator of osteosarcoma stem cells.
- HIF2PUT may exert its function, at least partly, by modulating HIF-2α expression.
- HIF2PUT presents a potential therapeutic target for osteosarcoma treatment.
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