{"title":"Mechanism of miR-4465 targeting PTEN-mediated autophagy of astrocytes in epilepsy.","authors":"Jinhua Zhao, Jihong Tang, Xiaoyan Shi","doi":"10.5937/jomb0-52626","DOIUrl":null,"url":null,"abstract":"<p><strong>Background: </strong>To investigate the mechanism of miR-4465 targeting PTEN-mediated autophagy of astrocytes in epilepsy.</p><p><strong>Methods: </strong>Serum samples were collected from epileptic children and healthy children. Extract foreign bodies from serum samples and determine their quality. The exosomes were sequenced, and the abnormal expression of miRNA in patients' serum exosomes was analysed, and the expression of miR-4465 was verified by quantitative PCR. Bioinformatics predicts the size of miR-4465 and makes GO and KEGG analyses. HEK293 cells were cultured, and the relationship between miR-4465 and its target was detected using the double luciferase reporting method. Astrocytes were cultured, and quantitative PCR and WB were used to detect the expression of miR-4465 and PTEN after overexpression. In addition, CCK-8 and WB were used to detect the growth of miR-4465 and the changes of autophagy-related proteins ATG5 and Beclin1, respectively.</p><p><strong>Results: </strong>miR-4465 was markedly increased in exosomes. The bioinformatic analysis found the differentially expressed target genes of miR-4465 were mainly enriched in molecular binding, molecular function regulation, and other molecular functions and participated in cell adhesion, cell-extracellular matrix receptor interaction, and the Rap1 signalling pathway. PTEN has been predicted as a target gene of miR-4465; meanwhile, the results of the dual-luciferase reporter assay confirmed the interaction between miR-4465 and PTEN. Quantitative PCR, as well as WB results, suggested the level of PTEN was decreased in serum exosomes of patients with epilepsy, while increased miR-4465 expression inhibited expressions of PTEN. CCK-8, as well as WB results, suggested miR-4465 could suppress the growth of astrocytes and promote ATG5 as well as Beclin1 expression; finally, up-regulation of PTEN partially alleviated effects of miR-4465 on astrocytes growth as well as autophagy.</p><p><strong>Conclusions: </strong>In children with epilepsy, miR-4465 can target and regulate PTEN to promote autophagy in astrocytes.</p>","PeriodicalId":16175,"journal":{"name":"Journal of Medical Biochemistry","volume":"44 6","pages":"1347-1356"},"PeriodicalIF":1.5000,"publicationDate":"2025-09-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12497466/pdf/","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of Medical Biochemistry","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.5937/jomb0-52626","RegionNum":4,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q4","JCRName":"BIOCHEMISTRY & MOLECULAR BIOLOGY","Score":null,"Total":0}
引用次数: 0
Abstract
Background: To investigate the mechanism of miR-4465 targeting PTEN-mediated autophagy of astrocytes in epilepsy.
Methods: Serum samples were collected from epileptic children and healthy children. Extract foreign bodies from serum samples and determine their quality. The exosomes were sequenced, and the abnormal expression of miRNA in patients' serum exosomes was analysed, and the expression of miR-4465 was verified by quantitative PCR. Bioinformatics predicts the size of miR-4465 and makes GO and KEGG analyses. HEK293 cells were cultured, and the relationship between miR-4465 and its target was detected using the double luciferase reporting method. Astrocytes were cultured, and quantitative PCR and WB were used to detect the expression of miR-4465 and PTEN after overexpression. In addition, CCK-8 and WB were used to detect the growth of miR-4465 and the changes of autophagy-related proteins ATG5 and Beclin1, respectively.
Results: miR-4465 was markedly increased in exosomes. The bioinformatic analysis found the differentially expressed target genes of miR-4465 were mainly enriched in molecular binding, molecular function regulation, and other molecular functions and participated in cell adhesion, cell-extracellular matrix receptor interaction, and the Rap1 signalling pathway. PTEN has been predicted as a target gene of miR-4465; meanwhile, the results of the dual-luciferase reporter assay confirmed the interaction between miR-4465 and PTEN. Quantitative PCR, as well as WB results, suggested the level of PTEN was decreased in serum exosomes of patients with epilepsy, while increased miR-4465 expression inhibited expressions of PTEN. CCK-8, as well as WB results, suggested miR-4465 could suppress the growth of astrocytes and promote ATG5 as well as Beclin1 expression; finally, up-regulation of PTEN partially alleviated effects of miR-4465 on astrocytes growth as well as autophagy.
Conclusions: In children with epilepsy, miR-4465 can target and regulate PTEN to promote autophagy in astrocytes.
期刊介绍：
The JOURNAL OF MEDICAL BIOCHEMISTRY (J MED BIOCHEM) is the official journal of the Society of Medical Biochemists of Serbia with international peer-review. Papers are independently reviewed by at least two reviewers selected by the Editors as Blind Peer Reviews. The Journal of Medical Biochemistry is published quarterly.
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reference materials and methods,
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all related scientific disciplines where chemistry, biochemistry, molecular biology and immunochemistry deal with the study of normal and pathologic processes in human beings.
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