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Investigation of the Role of Induced Overexpression of the Isolated Secreted Klotho on the A-172 Human Glioblastoma Cells

Abstract

Klotho gene, identified in 1997 as an anti-aging gene, can manufacture two protein products: transmembrane and secreted forms. The later research revealed the involvement of klotho in carcinogenesis. However, little is known about the action of different Klotho forms on antitumor effects is still. The purpose of this article is to evaluate the effect of isolated secreted Klotho overexpression on the growth features of human glioblastoma cell line A-172.

A-172 was transfected by a plasmid vector incorporating secreted Klotho sequence by the liposomal method. Overexpression assay was carried out quantitatively on both mRNA and protein using RT-qPCR and ELISA, correspondingly. It was shown that the relative expression of secreted Klotho in the experimental group was significantly higher than in the untransfected group by both methods (p < 0.001). At the same time, the growth curves and MTT proliferation assay demonstrated significantly decreased values under induced overexpression (p < 0.01). The increased amount of cells with activated caspases and annexin V (p < 0.001) corresponded with the expression of secreted Klotho. This mechanism, as suggested, maybe causative of the observed effects.

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Acknowledgements

Authors also express our gratitude to Gleb Derbyshev, Anna Shcheglova, Elisaveta Yakovleva for all kinds of technical assistance accompanying most of the experimental procedures related to this article.

Funding

This study was in part supported by the grant of The Russian Ministry of Health (AAAA-A18-118041890078–5) as a part of the project “Investigation of the role of the Klotho genes family in carcinogenesis.” The funding body had no role in the design of the study and collection, analysis, and interpretation of data, and in writing the manuscript.

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All authors contributed to the study’s conception and design. Material preparation, data collection, and analysis were performed by VM, AP, and MD. The first draft of the manuscript was written by VM, AP, and OG, and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.

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Correspondence to Alexander I. Ponomarev.

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Melekhin, V.V., Ponomarev, A.I., Desyatova, M.A. et al. Investigation of the Role of Induced Overexpression of the Isolated Secreted Klotho on the A-172 Human Glioblastoma Cells. J Mol Neurosci 72, 401–409 (2022). https://doi.org/10.1007/s12031-021-01960-1

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  • DOI: https://doi.org/10.1007/s12031-021-01960-1

Keywords

  • Secreted Klotho
  • Lipofection
  • Plasmid vector
  • Gene expression
  • Apoptosis