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Genes2025; 16(8); 936; doi: 10.3390/genes16080936

Comparative Transcriptome and MicroRNA Profiles of Equine Mesenchymal Stem Cells, Fibroblasts, and Their Extracellular Vesicles.

Abstract: Mesenchymal stem cells (MSCs) are a promising tool in regenerative medicine due to their ability to secrete paracrine factors that modulate tissue repair. Extracellular vesicles (EVs) released by MSCs contain bioactive molecules (e.g., mRNAs, miRNAs, proteins) and play a key role in intercellular communication. This study compared the transcriptomic profiles (mRNA and miRNA) of equine MSCs derived from adipose tissue (AT-MSCs), bone marrow (BM-MSCs), and ovarian fibroblasts (as a differentiated control). Additionally, miRNAs present in EVs secreted by these cells were characterized using next-generation sequencing. All cell types met ISCT criteria for MSCs, including CD90 expression, lack of MHC II, trilineage differentiation, and adherence. EVs were isolated using ultracentrifugation and validated with nanoparticle tracking analysis and flow cytometry (CD63, CD81). Differential expression analysis revealed distinct mRNA and miRNA profiles across cell types and their secreted EVs, correlating with tissue origin. BM-MSCs showed unique regulation of genes linked to early development and osteogenesis. EVs contained diverse RNA species, including miRNA, mRNA, lncRNA, rRNA, and others. In total, 227 and 256 mature miRNAs were detected in BM-MSCs and AT-MSCs, respectively, including two novel miRNAs per MSC type. Fibroblasts expressed 209 mature miRNAs, including one novel miRNA also found in MSCs. Compared to fibroblasts, 60 and 92 differentially expressed miRNAs were identified in AT-MSCs and BM-MSCs, respectively. The results indicate that MSC tissue origin influences both transcriptomic profiles and EV miRNA content, which may help to interpret their therapeutic potential. Identifying key mRNAs and miRNAs could aid in future optimizing of MSC-based therapies in horses.
Publication Date: 2025-08-05 PubMed ID: 40869984PubMed Central: PMC12386118DOI: 10.3390/genes16080936Google Scholar: Lookup
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  • Journal Article
  • Comparative Study

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

Overview

  • This research compares the gene expression (mRNA and miRNA) profiles of different types of equine mesenchymal stem cells (MSCs) and fibroblasts, as well as the microRNAs found in their secreted extracellular vesicles (EVs), to better understand their molecular characteristics and potential for regenerative medicine.

Introduction to Mesenchymal Stem Cells and Extracellular Vesicles

  • MSCs have regenerative abilities largely due to their secretion of paracrine factors—bioactive molecules that facilitate tissue repair and wound healing.
  • Extracellular vesicles (EVs) are small membrane-bound particles released by cells that carry various molecules such as mRNAs, miRNAs, proteins, and other RNAs, which function in intercellular communication.
  • The study focuses on equine MSCs derived from two tissue types: adipose tissue (AT-MSCs) and bone marrow (BM-MSCs), and compares them to ovarian fibroblasts which act as a differentiated control cell type.

Methods and Cell Characterization

  • All cell types used in the study were validated to meet the International Society for Cellular Therapy (ISCT) criteria for MSCs:
    • Positive for CD90 surface marker
    • Negative for Major Histocompatibility Complex class II (MHC II) expression
    • Ability to differentiate into three lineages (trilineage differentiation)
    • Adherence to plastic culture surfaces
  • Extracellular vesicles were isolated using ultracentrifugation methods to ensure purity.
  • EVs were verified through:
    • Nano-particle tracking analysis (to determine size and concentration)
    • Flow cytometry for EV surface markers CD63 and CD81

Transcriptomic and miRNA Profiling

  • Next-generation sequencing was used to analyze mRNA and miRNA expression profiles from the MSCs, fibroblasts, and their respective EVs.
  • Distinct expression profiles were observed between the MSC types and fibroblasts, correlating with their tissue origins:
    • BM-MSCs had unique gene expression, especially in genes related to early development and bone formation (osteogenesis).
    • EVs contained a diverse array of RNA types beyond miRNAs, including long non-coding RNAs (lncRNAs), ribosomal RNAs (rRNAs), and messenger RNAs (mRNAs).
  • In terms of miRNAs detected:
    • AT-MSCs had 256 mature miRNAs, including 2 previously unidentified novel miRNAs.
    • BM-MSCs had 227 mature miRNAs, also including 2 novel miRNAs.
    • Fibroblasts expressed 209 mature miRNAs, including 1 novel miRNA that was also shared with MSCs.
  • Comparison between cell types revealed significant differences:
    • 60 miRNAs were differentially expressed in AT-MSCs compared to fibroblasts.
    • 92 miRNAs were differentially expressed in BM-MSCs compared to fibroblasts.

Biological and Therapeutic Implications

  • The tissue source of MSCs significantly influences both the comprehensive transcriptomic signatures and the miRNA cargo of their extracellular vesicles.
  • These molecular differences may underlie the distinct therapeutic potentials observed in MSCs derived from different tissues.
  • Identification of key mRNAs and miRNAs that are unique or enriched in MSCs or their EVs could inform the development and optimization of MSC-based regenerative therapies in horses.
  • This detailed molecular profiling can help in selecting the most appropriate MSC source or in engineering EVs with desired therapeutic properties.

Conclusion

  • This study provides comprehensive transcriptomic and microRNA profiles of equine MSCs from adipose and bone marrow sources compared to fibroblasts, together with characterization of their EV miRNA content.
  • It highlights the importance of MSC tissue origin on gene and miRNA expression, offering molecular insights critical for enhancing MSC-based treatments in veterinary regenerative medicine.

Cite This Article

APA
Sawicki S, Bugno-Poniewierska M, Żurowski J, Szmatoła T, Semik-Gurgul E, Bochenek M, Karnas E, Gurgul A. (2025). Comparative Transcriptome and MicroRNA Profiles of Equine Mesenchymal Stem Cells, Fibroblasts, and Their Extracellular Vesicles. Genes (Basel), 16(8), 936. https://doi.org/10.3390/genes16080936

Publication

ISSN: 2073-4425
NlmUniqueID: 101551097
Country: Switzerland
Language: English
Volume: 16
Issue: 8
PII: 936

Researcher Affiliations

Sawicki, Sebastian
  • Department of Animal Reproduction, Anatomy and Genomics, University of Agriculture in Krakow, Mickiewicza 24/28, 30-059 Krakow, Poland.
Bugno-Poniewierska, Monika
  • Department of Animal Reproduction, Anatomy and Genomics, University of Agriculture in Krakow, Mickiewicza 24/28, 30-059 Krakow, Poland.
Żurowski, Jakub
  • Department of Basic Sciences, University of Agriculture in Krakow, Redzina 1C, 30-248 Krakow, Poland.
Szmatoła, Tomasz
  • Department of Basic Sciences, University of Agriculture in Krakow, Redzina 1C, 30-248 Krakow, Poland.
Semik-Gurgul, Ewelina
  • Department of Animal Molecular Biology, National Research Institute of Animal Production, Krakowska 1 St., 32-083 Balice, Poland.
Bochenek, Michał
  • Institute of Zoology and Biomedical Research, Jagiellonian University in Krakow, Gronostajowa 9, 30-387 Krakow, Poland.
Karnas, Elżbieta
  • Department of Cell Biology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Gronostajowa 7, 30-387 Krakow, Poland.
Gurgul, Artur
  • Department of Basic Sciences, University of Agriculture in Krakow, Redzina 1C, 30-248 Krakow, Poland.

MeSH Terms

  • Animals
  • Mesenchymal Stem Cells / metabolism
  • Mesenchymal Stem Cells / cytology
  • Horses / genetics
  • Extracellular Vesicles / metabolism
  • Extracellular Vesicles / genetics
  • MicroRNAs / genetics
  • Fibroblasts / metabolism
  • Transcriptome / genetics
  • Female
  • Adipose Tissue / cytology
  • Adipose Tissue / metabolism
  • Gene Expression Profiling
  • Cell Differentiation / genetics
  • RNA, Messenger / genetics

Conflict of Interest Statement

The authors declare no conflicts of interest.

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