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Small RNA sequencing of small extracellular vesicles secreted by umbilical cord mesenchymal stem cells following replicative senescence

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Abstract

Background

Umbilical cord mesenchymal stem cells (UCMSC) are subsets of multipotent stem cells involved in immune modulation, tissue regeneration, and antimicrobial defense. Cellular senescence is associated with the onset of aging-related diseases and small extracellular vesicles (sEVs) are important mediators of senescence and aging.

Objective

However, little is known about the role and function of microRNAs (miRNAs) carried by UCMSC-derived sEVs. To analyze the expression profiles of miRNAs secreted by senescent UCMSC, small RNA sequencing of the miRNAs within the sEVs was performed in this study.

Methods

UCMSC cultures underwent serial passaging beyond passage number 20 to achieve replicative senescence, which was confirmed by various methods, including increased senescence-associated β-gal staining and cytokine secretion levels. sEVs derived from non-senescent and senescent UCMSC were isolated and characterized by nanoparticle tracking analysis, transmission electron microscopy, and immunoblot analysis.

Results

Small RNA sequencing of the miRNAs within the sEVs revealed senescence-associated differences in the miRNA composition, as shown by the upregulation of miR-122-5p and miR-146a-5p, and downregulation of miR-125b-5p and miR-29-3p. In addition, total RNA sequencing analysis showed that PENK, ITGA8, and TSIX were upregulated, whereas AKR1B10, UNC13D, and IL21R were downregulated by replicative senescence in UCMSC. In sEVs, upregulated genes were linked to downregulated miRNAs, and vice versa. In the gene-concept network analysis, five gynecologic terms were retrieved.

Conclusions

The study provides an insight into the cellular characteristics of UCMSC following replicative senescence and emphasizes the importance of monitoring passage numbers of UCMSC for further therapeutic use.

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Acknowledgements

This research was funded by MD-PhD/Medical Scientist Training Program through the Korea Health Industry Development Institute (KHIDI), funded by the Ministry of Health & Welfare, Republic of Korea, and the Basic Science Research Program of the National Research Foundation of Korea (NRF) by the Ministry of Science, ICT & Future Planning (NRF-2019R1A2C1005961). This research was also funded by the Basic Science Research Program through NRF by the Ministry of Education (NRF-2020R1I1A1A01052701).

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Author notes

  1. Chris Gunwoo Kim and Jae Kyung Lee contributed equally to this work.

Authors and Affiliations

  1. Department of Laboratory Medicine, College of Medicine, Korea University, Seoul, South Korea

    Chris Gunwoo Kim

  2. BK21 Graduate Program, Department of Biomedical Sciences, College of Medicine, Korea University Guro Hospital, Seoul, South Korea

    Jae Kyung Lee & Ok Sarah Shin

  3. Department of Obstetrics and Gynecology, College of Medicine, Korea University Guro Hospital, Seoul, South Korea

    Geum-Joon Cho

  4. Medical Science Research Center, College of Medicine, Korea University, Seoul, South Korea

    Jeong-An Gim

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  1. Chris Gunwoo Kim
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Correspondence to Ok Sarah Shin or Jeong-An Gim.

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Kim, C.G., Lee, J.K., Cho, GJ. et al. Small RNA sequencing of small extracellular vesicles secreted by umbilical cord mesenchymal stem cells following replicative senescence. Genes Genom 45, 347–358 (2023). https://doi.org/10.1007/s13258-022-01297-y

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