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Scientific reports2024; 14(1); 9012; doi: 10.1038/s41598-024-59471-z

De novo reconstruction of a functional in vivo-like equine endometrium using collagen-based tissue engineering.

Abstract: To better understand molecular aspects of equine endometrial function, there is a need for advanced in vitro culture systems that more closely imitate the intricate 3-dimensional (3D) in vivo endometrial structure than current techniques. However, development of a 3D in vitro model of this complex tissue is challenging. This study aimed to develop an in vitro 3D endometrial tissue (3D-ET) with an epithelial cell phenotype optimized by treatment with a Rho-associated protein kinase (ROCK) inhibitor. Equine endometrial epithelial (eECs) and mesenchymal stromal (eMSCs) cells were isolated separately, and eECs cultured in various concentrations of Rock inhibitor (0, 5, 10 µmol) in epithelial medium (EC-medium) containing 10% knock-out serum replacement (KSR). The optimal concentration of Rock inhibitor for enhancing eEC proliferation and viability was 10 µM. However, 10 µM Rock inhibitor in the 10% KSR EC-medium was able to maintain mucin1 (Muc1) gene expression for only a short period. In contrast, fetal bovine serum (FBS) was able to maintain Muc1 gene expression for longer culture durations. An in vitro 3D-ET was successfully constructed using a collagen-based scaffold to support the eECs and eMSCs. The 3D-ET closely mimicked in vivo endometrium by displaying gland-like eEC-derived structures positive for the endometrial gland marker, Fork headbox A2 (FOXA2), and by mimicking the 3D morphology of the stromal compartment. In addition, the 3D-ET expressed the secretory protein MUC1 on its glandular epithelial surface and responded to LPS challenge by upregulating the expression of the interleukin-6 (IL6) and prostaglandin F synthase (PGFS) genes (P < 0.01), along with an increase in their secretory products, IL-6 (P < 0.01) and prostaglandin F2alpha (PGF2α) (P < 0.001) respectively. In the future, this culture system can be used to study both normal physiology and pathological processes of the equine endometrium.
© 2024. The Author(s).
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Publication Date: 2024-04-19 PubMed ID: 38641671PubMed Central: PMC11031578DOI: 10.1038/s41598-024-59471-zGoogle Scholar: Lookup
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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.

This study focuses on the creation of a 3D in vitro model of an equine endometrium using tissue engineering techniques. This model can potentially assist in understanding the molecular aspects of equine endometrial function, and in studying both its normal physiology and potential pathologies.

Research Objectives and Methodology

  • The aim of the research was to develop a 3D model of equine endometrial tissue, which could closely imitate the in vivo endometrial structure more accurately than existing methods.
  • The creation of the model was achieved by using equine endometrial epithelial (eECs) and mesenchymal stromal cells (eMSCs), which were isolated and cultured separately using a Rho-associated protein kinase (ROCK) inhibitor. The inhibitor was used to optimize the epithelial cell phenotype.

Findings and Results

  • It was found that the concentration of ROCK inhibitor needed to enhance eEC proliferation and viability was 10 µM. However, preservation of mucin1 (Muc1) gene expression could be maintained only for a short while with this concentration of the inhibitor in the current epithelial medium.
  • Following this, fetal bovine serum (FBS) proved to be a more effective medium for maintaining Muc1 gene expression over longer culture durations.
  • Lastly, with a collagen-based scaffold to support the eECs and eMSCs, a 3D endometrial tissue was constructed in vitro. This tissue successfully mimicked the in vivo endometrium, presenting gland-like eEC-derived structures and the 3D morphology of the stromal compartment.

Potential Applications and Future Research

  • The constructed 3D endometrial tissue responded to challenges such as LPS to upregulate the expression of genes like interleukin-6 (IL6) and prostaglandin F synthase (PGFS), along with their secretory products, IL-6 and prostaglandin F2alpha (PGF2α) respectively. The successful recreation of these responses indicates the potential of this model to facilitate further research in the understanding of the equine endometrium’s normal physiology and potential pathological processes.

Cite This Article

APA
Santiviparat S, Swangchan-Uthai T, Stout TAE, Buranapraditkun S, Setthawong P, Taephatthanasagon T, Rodprasert W, Sawangmake C, Tharasanit T. (2024). De novo reconstruction of a functional in vivo-like equine endometrium using collagen-based tissue engineering. Sci Rep, 14(1), 9012. https://doi.org/10.1038/s41598-024-59471-z

Publication

Scientific reports
ISSN: 2045-2322
NlmUniqueID: 101563288
Country: England
Language: English
Volume: 14
Issue: 1
Pages: 9012
PII: 9012

Researcher Affiliations

Santiviparat, Sawita
  • Department of Obstetrics, Gynecology and Reproduction, Faculty of Veterinary Science, Chulalongkorn University, Bangkok, Thailand.
  • CU-Animal Fertility Research Unit, Chulalongkorn University, Bangkok, Thailand.
  • Veterinary Clinical Stem Cells and Bioengineering Research Unit, Chulalongkorn University, Bangkok, Thailand.
Swangchan-Uthai, Theerawat
  • Department of Obstetrics, Gynecology and Reproduction, Faculty of Veterinary Science, Chulalongkorn University, Bangkok, Thailand.
  • CU-Animal Fertility Research Unit, Chulalongkorn University, Bangkok, Thailand.
Stout, Tom A E
  • Department of Clinical Sciences, Utrecht University, Utrecht, The Netherlands.
Buranapraditkun, Supranee
  • Division of Allergy and Clinical Immunology, Department of Medicine, Faculty of Medicine, King Chulalongkorn Memorial Hospital, Chulalongkorn University, Thai Red Cross Society, Bangkok, 10330, Thailand.
  • Faculty of Medicine, Center of Excellence in Vaccine Research and Development (Chula Vaccine Research Center-Chula VRC), Chulalongkorn University, Bangkok, 10330, Thailand.
  • Thai Pediatric Gastroenterology, Hepatology and Immunology (TPGHAI) Research Unit, Faculty of Medicine, King Chulalongkorn Memorial Hospital, Chulalongkorn University, The Thai Red Cross Society, Bangkok, 10330, Thailand.
Setthawong, Piyathip
  • Department of Physiology, Faculty of Veterinary Medicine, Kasetsart University, Bangkok, Thailand.
Taephatthanasagon, Teeanutree
  • Veterinary Pharmacology and Stem Cell Research Laboratory, Faculty of Veterinary Science, Veterinary Stem Cell and Bioengineering Innovation Center (VSCBIC), Chulalongkorn University, Bangkok, Thailand.
  • Faculty of Veterinary Science, Veterinary Systems Pharmacology Center (VSPC), Chulalongkorn University, Bangkok, Thailand.
Rodprasert, Watchareewan
  • Veterinary Pharmacology and Stem Cell Research Laboratory, Faculty of Veterinary Science, Veterinary Stem Cell and Bioengineering Innovation Center (VSCBIC), Chulalongkorn University, Bangkok, Thailand.
  • Faculty of Veterinary Science, Veterinary Systems Pharmacology Center (VSPC), Chulalongkorn University, Bangkok, Thailand.
Sawangmake, Chenphop
  • Veterinary Pharmacology and Stem Cell Research Laboratory, Faculty of Veterinary Science, Veterinary Stem Cell and Bioengineering Innovation Center (VSCBIC), Chulalongkorn University, Bangkok, Thailand.
  • Faculty of Veterinary Science, Veterinary Systems Pharmacology Center (VSPC), Chulalongkorn University, Bangkok, Thailand.
  • Department of Pharmacology, Faculty of Veterinary Science, Chulalongkorn University, Bangkok, Thailand.
  • Faculty of Dentistry, Center of Excellence in Regenerative Dentistry, Chulalongkorn University, Bangkok, Thailand.
Tharasanit, Theerawat
  • Department of Obstetrics, Gynecology and Reproduction, Faculty of Veterinary Science, Chulalongkorn University, Bangkok, Thailand. Theerawat.t@chula.ac.th.
  • CU-Animal Fertility Research Unit, Chulalongkorn University, Bangkok, Thailand. Theerawat.t@chula.ac.th.
  • Veterinary Clinical Stem Cells and Bioengineering Research Unit, Chulalongkorn University, Bangkok, Thailand. Theerawat.t@chula.ac.th.

MeSH Terms

  • Female
  • Animals
  • Horses
  • Cells, Cultured
  • Tissue Engineering
  • rho-Associated Kinases / genetics
  • rho-Associated Kinases / metabolism
  • Endometrium / metabolism
  • Epithelial Cells / metabolism
  • Collagen / metabolism
  • Dinoprost / metabolism

Grant Funding

  • GCUGR1125662081D, No.1-81 / 90th Anniversary Ratchadaphiseksomphot Endowment fund
  • N41A660173 / the National Research Council of Thailand

Conflict of Interest Statement

The authors declare no competing interests.

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Citations

This article has been cited 2 times.
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