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The Effect of Increased miR-16-1 Levels in Mouse Embryos on Epigenetic Modification, Target Gene Expression, and Developmental Processes

  • Reproductive Biology: Original Article
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Abstract

Changes in microRNA (miRNA) levels are present in numerous diseases. Although these changes are particularly noted in male infertility, little is known about the effects of increased miR-16-1 in sperm from infertile men. In this study, we assessed the effects of increased mir-16-1 expression on the developmental process, epigenetic changes, and target gene expressions. IVF embryos, 6 h after insemination, were divided into three groups: control, control negative (CN), and miR-16-1 harboring plasmid microinjection. The developmental rates of these embryos were recorded after 24, 48, 72, and 96 h of culture. The levels of histone H3 lysine 4 tri-methylation (H3K4me3) and histone H3 lysine 27 tri-methylation (H3K27me3) were assessed in the 2-cell and blastocyst stages by immunofluorescence staining. Expression profiles of the miR16-1, Bax, Bcl-2, Suz12, and Kmt2a genes were measured by quantitative real-time polymerase chain reaction (qRT-PCR). There was a significant decrease from the 8-cell stage to the blastocyst stage of embryo development in the miR-16-1 harboring plasmid microinjection group. We observed substantial reductions in the amounts of H3K4me3 and H3K27me3 in the 2-cell and the blastocyst stages in the miR-16-1 harboring plasmid microinjection group (P ≤ 0.05). The miR-16-1 level in the miRNA group was higher than the control group in the 2-cell and the blastocyst stages. There was a significant increase (P ≤ 0.05) in Bax and decreases in Bcl2, Suz12, and Kmt2a following the injection of the miR-16-1 harboring plasmid. These results suggest that a change in miR-16-1 expression can significantly affect embryo development, epigenetic changes, and target gene expressions.

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Acknowledgements

We deeply appreciate all those who assisted with the completion of this article. We would like to thank Ms. Dehghan for her assistance with the genetic analyses and Dr. Bakhtari for his comments that greatly improved the immunofluorescence staining.

Funding

This research was supported by the Cellular and Molecular Research Center of Shahid Beheshti University of Medical Sciences, and part of the cost of this research was provided by the School of Veterinary Medicine of Shiraz University. Also, part of this research was reinforced by the Department of Genetics of Shahid Beheshti University of Medical Sciences.

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Authors and Affiliations

  1. Department of Clinical Sciences, School of Veterinary Medicine, Shiraz University, Shiraz, Iran

    Maryam Kiani & Asghar Mogheiseh

  2. Cellular and Molecular Biology Research Center, Shahid Beheshti University of Medical Sciences, P.O Box 193954717, Tehran, Iran

    Mohammad Salehi, Samira Mohammadi-Yeganeh & Solmaz Shahidi

  3. Department of Biotechnology, School of Advanced Technologies in Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran

    Mohammad Salehi

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  1. Maryam Kiani
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Experimental design and data analysis were carried out by M. Salehi and the practical steps in the laboratory were carried out by M. Kiani and S. Shahidi. Samira Mohammadi Yeganeh was involved in the genetics department and A. Mogheiseh contributed in revisions and modifications for the final manuscript.

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Kiani, M., Salehi, M., Mogheiseh, A. et al. The Effect of Increased miR-16-1 Levels in Mouse Embryos on Epigenetic Modification, Target Gene Expression, and Developmental Processes. Reprod. Sci. 27, 2197–2210 (2020). https://doi.org/10.1007/s43032-020-00240-4

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