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Circ_0007534 Silencing Inhibits the Proliferation, Migration and Invasion and Induces the Apoptosis of Glioma Cells Partly Through Down-Regulating PROX1 Via Elevating miR-22-3p Level

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Abstract

Glioma is a common malignant brain neoplasm. The role and mechanism of circular RNA 0,007,534 (circ_0007534) in glioma progression were investigated in this study. The expression of circ_0007534, microRNA-22-3p (miR-22-3p) and prospero homeobox protein 1 (PROX1) messenger RNA (mRNA) was determined by quantitative real-time polymerase chain reaction (qRT-PCR). The proliferation, migration and invasion abilities were analyzed by colony formation assay, transwell migration assay and transwell invasion assay. Cell apoptosis was assessed through measuring the activity of Caspase-3 using the Caspase-3 kit and the apoptosis rate using flow cytometry. Dual-luciferase reporter assay was used to confirm the target interaction between miR-22-3p and circ_0007534 or PROX1. The protein level of PROX1 was examined by Western blot assay. Animal studies were conducted to analyze the influence of circ_0007534 interference on xenograft tumor growth in vivo. Circ_0007534 was highly expressed in glioma tissues and cell lines relative to that in normal tissues and NHA cell line. Circ_0007534 knockdown suppressed the proliferation and motility while induced the apoptosis of glioma cells. Circ_0007534 negatively regulated miR-22-3p level through targeting it in glioma cells. Circ_0007534 interference-induced influences in glioma cells were partly overturned by the silencing of miR-22-3p. PROX1 was a target of miR-22-3p, and circ_0007534 interference-mediated effects in glioma cells were largely diminished by the overexpression of PROX1. Circ_0007534 interference restrained glioma development in vivo. Circ_0007534 aggravated glioma progression through elevating PROX1 expression via targeting miR-22-3p, which provided new targets for the diagnosis and treatment of glioma.

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Data Availability

The datasets used and analyzed during the current study are available from the corresponding author on reasonable request.

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Funding

The project was funded by Shenzhen Sanming Project (SZSM201806067).

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Authors and Affiliations

  1. Department of Neurosurgery, The Second Affiliated Hospital of Shenzhen University (People’s Hospital of Shenzhen Baoan District), No. 118, Longjing Second Road, Xin’an Street, Baoan District, Shenzhen, 518101, China

    Yong Zheng, Rongkang Mai, Liang Liu, Zifeng Zhu & Yiyao Cao

  2. Department of General Medicine, The Second Affiliated Hospital of Shenzhen University (People’s Hospital of Shenzhen Baoan District), Shenzhen, China

    Yan Wang

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  1. Yong Zheng
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Contributions

RM designed and supervised the study. YZ conducted the experiments and drafted the manuscript. LL conducted the experiments and supervised the study. ZZ collected and analyzed the data. YW contributed the methodology and analyzed the data. YC edited the manuscript. All authors read and approved the final manuscript.

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Correspondence to Yong Zheng.

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The authors declare that they have no conflicts of interest.

Ethical Approval

This study was approved by the Ethics Review Committee of the Second Affiliated Hospital of Shenzhen University and was carried out according to the guidelines of Declaration of Helsinki.

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Supplementary Information

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10571_2021_1150_MOESM1_ESM.tif

Supplementary file1 Supplementary Figure 1. Circ_0007534 silencing has almost no effect on the apoptosis of human astrocytes NHA. Flow cytometry was performed to analyze the apoptosis rate in NHA cells with the silencing of circ_0007534 or not (TIF 771 kb)

10571_2021_1150_MOESM2_ESM.tif

Supplementary file2 Supplementary Figure 2. The candidate miRNA targets of circ_0007534. (A and B) The expression of miR-1236, miR-593, miR-22-3p, miR-625, and miR-502-5p was determined by qRT-PCR in LN229 and U251 cells transfected with si-NC or si-circ_0007534. *P<0.05, **P<0.01, ***P<0.001 (TIF 258 kb)

10571_2021_1150_MOESM3_ESM.tif

Supplementary file3 Supplementary Figure 3. The candidate mRNA targets of miR-22-3p. (A and B) qRT-PCR was conducted to detect the levels of GRM5, IL17RD, PROX1, and CDX2 in LN229 and U251 cells transfected with anti-NC or anti-miR-22-3p. *P<0.05, **P<0.01, ***P<0.001 (TIF 290 kb)

10571_2021_1150_MOESM4_ESM.tif

Supplementary file4 Supplementary Figure 4. PROX1 knockdown triggers the apoptosis of glioma cells. (A) The apoptosis of glioma cells transfected with si-NC or si-PROX1 was analyzed via flow cytometry. (B) Western blot assay was conducted to measure the expression of proteins associated with cell proliferation (Cyclin D1), motility (MMP2 and MMP9), and apoptosis (Bax). ***P<0.001 (TIF 809 kb)

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Zheng, Y., Wang, Y., Mai, R. et al. Circ_0007534 Silencing Inhibits the Proliferation, Migration and Invasion and Induces the Apoptosis of Glioma Cells Partly Through Down-Regulating PROX1 Via Elevating miR-22-3p Level. Cell Mol Neurobiol 42, 2819–2832 (2022). https://doi.org/10.1007/s10571-021-01150-y

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