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Cells, tissues, organs2014; 198(5); 377-389; doi: 10.1159/000358231

Bone morphogenetic protein-12 induces tenogenic differentiation of mesenchymal stem cells derived from equine amniotic fluid.

Abstract: Tendon injuries are common in race horses, and mesenchymal stem cells (MSCs) isolated from adult and foetal tissue have been used for tendon regeneration. In the present study, we evaluated equine amniotic fluid (AF) as a source of MSCs and standardised methodology and markers for their in vitro tenogenic differentiation. Plastic-adherent colonies were isolated from 12 of 20 AF samples by day 6 after seeding and 70-80% cell confluency was reached by day 17. These cells expressed mesenchymal surface markers [cluster of differentiation (CD)73, CD90 and CD105] by reverse transcription (RT)-polymerase chain reaction (PCR) and immunocytochemistry, but did not express haematopoietic markers (CD34, CD45 and CD14). In flow cytometry, the expression of CD29, CD44, CD73 and CD90 was observed in 68.83 ± 1.27, 93.66 ± 1.80, 96.96 ± 0.44 and 93.7 ± 1.89% of AF-MSCs, respectively. Osteogenic, chondrogenic and adipogenic differentiation of MSCs was confirmed by von Kossa and Alizarin red S, Alcian blue and oil red O staining, respectively. Upon supplementation of MSC growth media with 50 ng/ml bone morphogenetic protein (BMP)-12, AF-MSCs differentiated to tenocytes within 14 days. The differentiated cells were more slender, elongated and spindle shaped with thinner and longer cytoplasmic processes and showed expression of tenomodulin and decorin by RT-PCR and immunocytochemistry. In flow cytometry, 96.7 ± 1.90 and 80.9 ± 6.4% of differentiated cells expressed tenomodulin and decorin in comparison to 1.6 and 3.1% in undifferentiated control cells, respectively. Our results suggest that AF is an easily accessible and effective source of MSCs. On BMP-12 supplementation, AF-MSCs can be differentiated to tenocytes, which could be exploited for regeneration of ruptured or damaged tendon in race horses.
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Publication Date: 2014-03-21 PubMed ID: 24662023DOI: 10.1159/000358231Google Scholar: Lookup
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  • Journal Article
  • Research Support
  • Non-U.S. Gov't

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.

The research paper examines the use of mesenchymal stem cells (MSCs) from horse amniotic fluid (AF) to regenerate injured tendons in race horses, especially as these cells are found to differentiate into tendon cells (tenocytes) when treated with bone morphogenetic protein (BMP)-12.

Source and Properties of MSCs

  • The researchers collected amniotic fluid from 20 horses and found that 60 % of the samples yielded plastic-adherent colonies (a characteristic of MSCs) within 6 days from initial seeding.
  • The sampled cells reached a confluency rate (indicative of cell growth and division) of 70-80% by day 17.
  • These samples expressed mesenchymal surface markers (CD73, CD90, and CD105 as confirmed by RT-PCR and immunocytochemistry) and did not express haematopoietic markers (CD34, CD45, and CD14), confirming the presence of MSCs over other stem cells.

Characterization and Differentiation Capacity of MSCs

  • The presence of MSC markers (CD29, CD44, CD73, and CD90) was assessed and found in a high proportion of cells via flow cytometry.
  • The researchers tested the stem cells for their ability to differentiate into osteocytes (bone cells), chondrocytes (cartilage cells), and adipocytes (fat cells) by using specific staining techniques. Results confirmed that the cells exhibited differentiation into all these types, underlining the stem cells’ multipotency.

Tenogenic Differentiation of MSCs

  • The scientists exposed MSCs to 50 ng/ml BMP-12, a protein that induces tenogenic differentiation. After 14 days, they observed a change in the morphology of the MSCs, which became more slender and elongated, with thinner and longer cytoplasmic processes – features typical of tenocytes.
  • Increased expression of tenomodulin and decorin – markers for tenocytes – was observed via RT-PCR and immunocytochemistry. The flow cytometry further supported this, with a high proportion of cells showing these markers compared to the control group.

Conclusion

  • The research validated that amniotic fluid can be an effective and easily accessible source of MSCs for horses.
  • Upon exposure to BMP-12, these cells can differentiate into tenocytes – a process that could be utilized for the regeneration of ruptured or damaged tendons in racehorses, thus potentially providing an innovative method for equine injury treatment.

Cite This Article

APA
Gulati BR, Kumar R, Mohanty N, Kumar P, Somasundaram RK, Yadav PS. (2014). Bone morphogenetic protein-12 induces tenogenic differentiation of mesenchymal stem cells derived from equine amniotic fluid. Cells Tissues Organs, 198(5), 377-389. https://doi.org/10.1159/000358231

Publication

Cells, tissues, organs
ISSN: 1422-6421
NlmUniqueID: 100883360
Country: Switzerland
Language: English
Volume: 198
Issue: 5
Pages: 377-389

Researcher Affiliations

Gulati, Baldev R
  • National Research Centre on Equines, Hisar, India.
Kumar, Rajesh
    Mohanty, Niharika
      Kumar, Pawan
        Somasundaram, Rajesh K
          Yadav, Prem S

            MeSH Terms

            • Amniotic Fluid / cytology
            • Animals
            • Bone Morphogenetic Proteins / pharmacology
            • Cell Culture Techniques
            • Cell Differentiation / physiology
            • Cells, Cultured
            • Female
            • Horses
            • Mesenchymal Stem Cells / cytology

            Citations

            This article has been cited 12 times.
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