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. 2024;24(4):463-475.
doi: 10.2174/1568009623666230817102104.

mirna-383-5p Functions as an Anti-oncogene in Glioma through the Akt/mTOR Signaling Pathway by Targeting VEGFA

Yan Liu  1   2 , Zhen Wang  2 , Zhi Tang  1 , Yao Fu  3 , Lei Wang  1   4
Affiliations

mirna-383-5p Functions as an Anti-oncogene in Glioma through the Akt/mTOR Signaling Pathway by Targeting VEGFA

Yan Liu et al. Curr Cancer Drug Targets. 2024.

Abstract

Background: Previously, we have screened 59 differentially expressed miRNAs and 419 mRNAs in the glioblastoma samples that have been compared to the peritumoral tissues using bioinformatics analyses, which included miRNA-383-5p and vascular endothelial growth factor A (VEGFA). miRNA-383-5p and VEGFA/Akt/mTOR pathway play important regulatory roles in the malignant biological behavior of glioma.

Methods: Glioma cell lines, U87 and U251, were collected for in vitro experiments. miRNA-383-5p and VEGFA expression levels were detected with qRT-PCR and WB. The protein expressions of Akt, mTOR, and VEGFR in U87 and U251 were detected with WB. The effect of miRNA-383-5p on the VEGFA activity was verified by dual-luciferase reporter assay. CCK-8 was used to examine the U87 and U251 cells' inhibition. Flow cytometry and transwell assays were used to detect cell apoptosis and invasion, respectively.

Results: Our research data indicated overexpression of miRNA-383-5p to suppress malignant biological behavior, which was manifested as promoting the apoptosis of U87 and U251 cells and inhibiting invasion, proliferation, and metastasis. VEGFA is one of the downstream target genes of miRNA-383- 5p. miRNA-383-5p could inhibit the expression of VEGFA and Akt/mTOR signaling pathways. Overexpression of VEGFA can reverse the inhibitory effect of miRNA-383-5p and reactivate the Akt/mTOR signaling pathway.

Conclusion: Our results indicate that miRNA-383-5p functions as an anti-oncogene by inhibiting the VEGFA/Akt/mTOR signaling pathway in glioma cells. These data provide potential therapeutic targets for glioblastoma.

Keywords: Akt/mTOR pathway; Glioma; VEGFA; cell apoptosis.; cytometry; miRNA383-5p.

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Conflict of interest statement

The authors declare no conflict of interest, financial or otherwise.

Figures

Fig. (1)
Fig. (1)
(a) The expression of miRNA-383-5p was found to be negatively correlated with the grade of glioma. (b) The prognostic value of miRNA-383-5p in gliomas of various grades. (c) miRNA-383-5p transfection increased miRNA-383-5p levels in U87 and U251 cells compared to NC-transfected cells. (d) qRT-PCR analysis of VEGFA expression in U251 and U87 cells treated with miRNA-383-5p mimic. (e-h) Western blotting was used to detect protein levels of VEGFA and VEGFR2 in U251 cells after transfection. (f-j) Western blotting was used to detect protein levels of VEGFA and VEGFR2 in U251 cells after transfection. Each experiment was repeated at least three times. ∗P < 0.05, ∗∗P < 0.01.
Fig. (2)
Fig. (2)
(a-d) Colony formation assays suggest that miR-383-5p mimic results in decreased U251 cell proliferation. (e) CCK8 experiments suggest that miR-383-5p mimic transfection reduces the cell viability of U87 and U251 cells. Each experiment was repeated at least three times. ∗P < 0.05, ∗∗P < 0.01.
Fig. (3)
Fig. (3)
(a-d) Flow cytometry analysis of miR-383-5p mimic and NC-mimic added to U87 and U251 cells showed that miR-383-5p mimic led to increased apoptosis in U87 and U251 cells. Each experiment was repeated at least three times. ∗P < 0.05, ∗∗P < 0.01.
Fig. (4)
Fig. (4)
(a-d) Transwell migration and invasion assays with U87 and U251 cells transfected with miR-383-5p mimics or NC. Each experiment was repeated at least three times. ∗P < 0.05, ∗∗P < 0.01.
Fig. (5)
Fig. (5)
(a) The predicted binding sites for miR-383-5p in the 3′-UTR of MYT1L VEGFA and the mutations in the binding sites are demonstrated. (b) Relative luciferase activity was measured in HEK293A cells co-transfected with miRNA-383-5p mimic/miRNA-NC and luciferase reporter plasmid. (c) qRT-PCR analysis of VEGFA expression in U251 and U87 cells transfected with NC mimic, miR-383-5p, and miR-383-5p+pcDNA3.1-VEGFA. (d-j) Western blot assay showing VEGFA, VEGFR2, AKT, and mTOR protein levels in U251 and U87 cells transfected with NC mimic, miR-383-5p, and miR-383-5p+pcDNA3.1-VEGFA. Each experiment was repeated at least three times. ∗P < 0.05, ∗∗P < 0.01.
Fig. (6)
Fig. (6)
(a, b) Comparison of U87 and U251 cell colonies two weeks after transfection of NC mimic, miRNA-383-5p mimic, and miRNA-383-5p+pcDNA3.1-VEGFA. (c, d) The reintroduction of VEGFA significantly increased the number of colonies in U87 and U251 cells compared to the miRNA-383-5p mimic alone transfection group. (e) CCK8 experiments suggest that overexpression of VEGFA reversed the tumor cell inactivation that was induced by miR-383-5p mimic transfection in U87 and U251 cells. Each experiment was repeated at least three times. ∗P < 0.05, ∗∗P < 0.01.
Fig. (7)
Fig. (7)
(a, c) Flow cytometry results suggest that miR-383-5p mimic transfection in U87 cells led to increased tumor apoptosis, which was partially alleviated by concurrent overexpression of VEGFA. (b, d) As in U87 cells, miR-383-5p mimic transfection also resulted in increased tumor cell apoptosis in U251 and U87 cells.
Fig. (8)
Fig. (8)
VEGFA expression inhibited the inhibitory effect of miR-383-5p on glioma cells. (a-d) Transwell invasion assay was used to assess the number of invaded U87 and U251 cells after transfection of NC mimic, miRNA-383-5p mimic, and miRNA-383-5p+pcDNA3.1-VEGFA.
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References

    1. Bray F., Ferlay J., Soerjomataram I., Siegel R.L., Torre L.A., Jemal A. Global cancer statistics 2018: GLOBOCAN estimates of inci-dence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J. Clin. 2018;68(6):394–424. doi: 10.3322/caac.21492. - DOI - PubMed
    1. Tan A.C., Ashley D.M., López G.Y., Malinzak M., Friedman H.S., Khasraw M. Management of glioblastoma: State of the art and fu-ture directions. CA Cancer J. Clin. 2020;70(4):299–312. doi: 10.3322/caac.21613. - DOI - PubMed
    1. Wang L., Liu Y., Yu Z., Gong J., Deng Z., Ren N., Zhong Z., Cai H., Tang Z., Cheng H., Chen S., He Z. Mir-139-5p inhibits glioma cell proliferation and progression by targeting GABRA1. J. Transl. Med. 2021;19(1):213. doi: 10.1186/s12967-021-02880-9. - DOI - PMC - PubMed
    1. Yu Z., Liu Y., Li Y., Zhang J., Peng J., Gong J., Xia Y., Wang L. miRNA-338-3p inhibits glioma cell proliferation and progression by targeting MYT1L. Brain Res. Bull. 2022;179:1–12. doi: 10.1016/j.brainresbull.2021.11.016. - DOI - PubMed
    1. Gong J., Tang Z., Yu Z., Deng Z., Liu Y., Ren N., Wang L., He Z. miR-138-5p inhibits the growth and invasion of glioma cells by regulating WEE1. Anal. Cell. Pathol. 2022;2022:1–12. doi: 10.1155/2022/7809882. - DOI - PMC - PubMed

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