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| Status |
Public on Jan 31, 2023 |
| Title |
miR-197-3p promotes osteosarcoma stemness and chemo-resistance by inhibiting SPOPL |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by RT-PCR
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| Summary |
First-line treatment for osteosarcoma includes chemotherapy and surgery. However, the five-year survival rate of refractory osteosarcoma remains unsatisfactory. Osteosarcoma cancer stem cells, possessing stemness and chemoresistance, are one of the critical causes for poor response to chemotherapy. Elucidating regulatory signaling pathways of osteosarcoma cancer stem cells may provide a rationale for improving regimens against chemoresistant osteosarcoma. Methotrexate (MTX)-resistant osteosarcoma cells were established. microRNA expression profiles were used for detecting differentially expressed microRNA in resistant clones and the parental cells. microRNA target databases were employed to predict potential microRNA and mRNA interaction. Flow cytometry was performed to measure stem cell marker Prominin-1 (CD133) positive cells. Im-munofluorescence staining was applied to detect CD133 expression. miR-197-3p mimic or an-ti-miR-197-3p stably transfected cells were used to generate xenograft models. In the study, we found miR-197-3p was increased in MTX-resistant cell lines. Overexpression of miR-197-3p en-hanced the expression of cancer stem cell markers CD133, Octamer-binding protein 4 (OCT4), Transcription factor SOX-2 (SOX2), and Homeobox protein NANOG (NANOG), as well as chemoresistance-associated genes ATP-dependent translocase ABCB1 (ABCB1) and Broad substrate specificity ATP-binding cassette transporter ABCG2 (ABCG2), whereas miR-197-3p knockdown inhibited stemness and recovered sensitivity to MTX. We also classified the tumor suppressor Speckle-type POZ protein-like (SPOPL) as a target of miR-197-3p. The miR-197-3p mutation that cannot combine SPOPL promoter regions was unable to sustain stemness or chemoresistance. Collectively, we discovered miR-197-3p conferred osteosarcoma stemness and chemotherapy resistance by targeting SPOPL, prompting promising therapeutic candidates for refractory oste-osarcoma treatment.
miRNA qPCR assay
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| Overall design |
MicroRNA (miRNA) were enriched and purified by miRNeasy Micro Kit (catalog: 217084, QIAGEN, Hilden, Germany), followed by cDNA synthesis with miRCURY LNA RT Kit (catalog: 339340, QIAGEN). miRCURY LNA miRNA Custom PCR Panels (catalog: 339330, QIAGEN) were then used to investigate differentially expressed miRNA in the parental U2OS cells and MTX-resistant U2OS cells.
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| Contributor(s) |
He M |
| Citation missing |
Has this study been published? Please login to update or notify GEO. |
| Submission date |
Jan 27, 2023 |
| Last update date |
Jan 31, 2023 |
| Contact name |
Ming He |
| E-mail(s) |
topminghe56@outlook.com
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| Organization name |
Shengjing Hospital of China Medical University
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| Street address |
No. 36 Sanhao Street
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| City |
Shenyang |
| State/province |
Liaoning |
| ZIP/Postal code |
110004 |
| Country |
China |
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| Platforms (1) |
| GPL33063 |
Qiagen Custom miScript miRNA PCR Arrays |
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| Samples (6)
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| Relations |
| BioProject |
PRJNA928683 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE223857_data_sheets.xlsx |
38.5 Kb |
(ftp)(http) |
XLSX |
| Processed data included within Sample table |
| Processed data are available on Series record |
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