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Published on: February 6, 2018
DNM3OS Enhances the Apoptosis and Senescence of Spermatogonia Associated with Nonobstructive Azoospermia by Providing
Rui Hua1, Qingjun Chu1, Feiyan Guo1
1Center for Reproductive Medicine, Department of Obstetrics and Gynecology, Nanfang Hospital, Southern Medical University, Guangzhou, China.
Background:
Nonobstructive azoospermia (NOA) is a complex disease characterized by the spermatogenic dysfunction of testicular tissues. The roles played by long noncoding RNAs (lncRNAs) in NOA pathogenesis have not been extensively studied.
Methods:
Microarray assays were performed on samples of testicular biopsy tissue obtained from patients with NOA for the purpose of identifying differentially expressed lncRNAs and messenger RNA (mRNA) transcripts, and the results were verified by quantitative real-time polymerase chain reaction. Mouse-derived GC-1 spermatogonia (spg) cells undergoing treatment with Adriamycin (ADR) were used to investigate the biological functions of the selected lncRNAs in vitro. The target microRNAs (miRNAs) of lncRNAs and the target mRNAs of miRNAs were predicted by a bioinformatics analysis. Functional studies performed using the CCK-8 assay, EdU incorporation assay, apoptosis detection, and senescence-associated β-galactosidase (SA-β-Gal) staining were conducted using GC-1 spg cells.
Results:
Totals of 2,652 lncRNAs and 2,625 mRNAs were found to be differentially expressed in the testicular tissue of NOA patients when compared with patients in a control group. Dynamin 3 opposite strand (DNM3OS) was a provider of pe-miR-214-5p that positively regulates miR-214-5p expression in GC-1 spg cells. The E2 factor (E2F) family of transcription factor 2 (E2F2) was initially predicted and subsequently verified to be a downstream gene of miR-214-5p. E2F2 expression was upregulated after DNM3OS knockdown in ADR-treated GC-1 spg cells. Moreover, knockdown of either DNM3OS or miR-214-5p significantly alleviated ADR-induced decreases in cellular activity and proliferation, as well as increases in apoptosis and senescence of mouse spermatogonial GC-1 spg cells.
Conclusions:
DNM3OS was found to regulate the apoptosis and senescence of spermatogonia by providing miR-214-5p and decreasing E2F2 expression, suggesting it as a novel target for gene therapy of male infertility.
Insights
Long noncoding RNA DNM3OS regulates spermatogonia apoptosis and senescence via miR-214-5p and E2F2. This finding suggests DNM3OS as a potential gene therapy target for male infertility caused by nonobstructive azoospermia.
Area of Science:
- Reproductive Biology
- Molecular Genetics
- Cell Biology
Background:
- Nonobstructive azoospermia (NOA) involves testicular spermatogenic dysfunction.
- The role of long noncoding RNAs (lncRNAs) in NOA pathogenesis is not well understood.
Purpose of the Study:
- To identify differentially expressed lncRNAs and mRNAs in NOA testicular tissue.
- To investigate the functional role of identified lncRNAs in spermatogonial cells.
Main Methods:
- Microarray analysis of testicular biopsy samples from NOA patients and controls.
- Quantitative real-time PCR for verification.
- In vitro studies using Adriamycin-treated GC-1 spermatogonia cells.
- Bioinformatic prediction of miRNA-mRNA interactions.
- Functional assays including CCK-8, EdU, apoptosis, and senescence staining.
Main Results:
- Over 2,600 lncRNAs and mRNAs were differentially expressed in NOA patients.
- DNM3OS was identified as a regulator of miR-214-5p, which targets E2F2.
- Knockdown of DNM3OS or miR-214-5p ameliorated Adriamycin-induced damage in GC-1 cells, reducing apoptosis and senescence.
Conclusions:
- DNM3OS regulates spermatogonial apoptosis and senescence by modulating the miR-214-5p/E2F2 axis.
- DNM3OS represents a potential novel therapeutic target for male infertility associated with NOA.
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