Journal of Assisted Reproduction and Genetics

, Volume 36, Issue 6, pp 1211–1223 | Cite as

Development of an efficient perfusion-based protocol for whole-organ decellularization of the ovine uterus as a human-sized model and in vivo application of the bioscaffolds

  • Seyedeh Sima Daryabari
  • Abdol-Mohammad KajbafzadehEmail author
  • Kiarad Fendereski
  • Fariba Ghorbani
  • Mehrshad Dehnavi
  • Minoo Rostami
  • Bahram Azizi Garajegayeh
  • Seyed Mohammad Tavangar
Reproductive Physiology and Disease



The main purpose of this investigation was to determine an efficient whole-organ decellularization protocol of a human-sized uterus and evaluate the in vivo properties of the bioscaffold.


Twenty-four ovine uteri were included in this investigation and were decellularized by three different protocols (n 6). We performed histopathological and immunohistochemical evaluations, 4,6-diamidino-2-phenylindole (DAPI) staining, DNA quantification, MTT assay, scanning electron microscopy, biomechanical studies, and CT angiography to characterize the scaffolds. The optimized protocol was determined, and patches were grafted into the uterine horns of eight female Wistar rats. The grafts were extracted after 10 days; the opposite horns were harvested to be evaluated as controls.


Protocol III (perfusion with 0.25% and 0.5% SDS solution and preservation in 10% formalin) was determined as the optimized method with efficient removal of the cellular components while preserving the extracellular matrix. Also, the bioscaffolds demonstrated native-like biomechanical, structural, and vascular properties. Histological and immunohistochemical evaluations of the harvested grafts confirmed the biocompatibility and recellularization potential of bioscaffolds. Also, the grafts demonstrated higher positive reaction for CD31 and Ki67 markers compared with the control samples which indicated eminent angiogenesis properties and proliferative capacity of the implanted tissues.


This investigation introduces an optimized protocol for whole-organ decellularization of the human-sized uterus with native-like characteristics and a prominent potential for regeneration and angiogenesis which could be employed in in vitro and in vivo studies. To the best of our knowledge, this is the first study to report biomechanical properties and angiographic evaluations of a large animal uterine scaffold.


Uterus Infertility Tissue engineering Regeneration Bioscaffold 



We would like to express our sincere gratitude to Dr. Torabi for providing the organs and Mr. Reza Esmaili and Mr. Nourbakhsh for their kind cooperation during this project.


This study was funded by Tehran University of Medical Sciences (grant number 96-03-30-36497).

Compliance with ethical standards

All the animal procedures were approved by The Animal Ethics Committee of the Tehran University of Medical Sciences, School of Medicine and Education Section of Basic Sciences and were performed in accordance with the Animal Welfare Act and the Guide for the Care and Use of Laboratory Animals.

Conflict of interest

The authors declare that they have no conflict of interest.


  1. 1.
    Milliez J. Uterine transplantation FIGO Committee for the ethical aspects of human reproduction and women’s health. Int J Gynaecol Obstet. 2009;106:270.CrossRefGoogle Scholar
  2. 2.
    Kisu I, Mihara M, Banno K, Umene K, Araki J, Hara H, et al. Risks for donors in uterus transplantation. Reprod Sci. 2013 Dec;20(12):1406–15.
  3. 3.
    Kisu I, Banno K, Mihara M, Suganuma N, Aoki D. Current status of uterus transplantation in primates and issues for clinical application. Fertil Steril. 2013;100(1):280–94.CrossRefGoogle Scholar
  4. 4.
    Brännström M, Johannesson L, Bokström H, Kvarnström N, Mölne J, Dahm-Kähler P, et al. Live birth after uterus transplantation. Lancet. 2015;385:607–16.Google Scholar
  5. 5.
    Mats Brännström. Uterus transplantation and beyond. J Mater Sci Mater Med (2017) 28:70DOI, 70.
  6. 6.
    Hellström M, Bandstein S, Brännström M. Uterine tissue engineering and the future of uterus transplantation. Ann Biomed Eng. 2017 Jul;45(7):1718–30 Epub 2016 Dec 19.Google Scholar
  7. 7.
    Park DW, Choi DS, Ryu HS, Kwon HC, Joo H, Min CK. A well-defined in vitro three-dimensional culture of human endometrium and its applicability to endometrial cancer invasion. Cancer Lett. 2003;195:185–92.CrossRefGoogle Scholar
  8. 8.
    Heidari Kani M, Chan EC, Young RC, Butler T, Smith R, Paul JW. 3D cell culturing and possibilities for myometrial tissue engineering. Ann Biomed Eng. 2017;45(7):1746–57.CrossRefGoogle Scholar
  9. 9.
    Santoso EG, Yoshida K, Hirota Y, Aizawa M, Yoshino O, Kishida A, et al. Application of detergents or high hydrostatic pressure as decellularization processes in uterine tissues and their subsequent effects on in vivo uterine regeneration in murine models. PLoS One. 2014;9(7):e103201.Google Scholar
  10. 10.
    Hiraoka T, Hirota Y, Saito-Fujita T, Matsuo M, Egashira M, Matsumoto L, et al. STAT3 accelerates uterine epithelial regeneration in a mouse model of decellularized uterine matrix transplantation. JCI insight. 2016;1(8).Google Scholar
  11. 11.
    Hellström M, El-Akouri RR, Sihlbom C, Olsson BM, Lengqvist J, Backdahl H, et al. Towards the development of a bioengineered uterus: comparison of different protocols for rat uterus decellularization. Acta Biomater. 2014;10(12):5034–42.CrossRefGoogle Scholar
  12. 12.
    Miyazaki K, Maruyama T. Partial regeneration and reconstruction of the rat uterus through recellularization of a decellularized uterine matrix. Biomaterials. 2014;35(31):8791–800.CrossRefGoogle Scholar
  13. 13.
    Hellström M, Moreno-Moya JM, Bandstein S, Bom E, Akouri RR, Miyazaki K, et al. Bioengineered uterine tissue supports pregnancy in a rat model. Fertil Steril. 2016;106(2):487–96 e1.Google Scholar
  14. 14.
    Campo H, Baptista PM, Lopez-Perez N, Faus A, Cervello I, Simon C. De- and recellularization of the pig uterus: a bioengineering pilot study. Biol Reprod. 2017;96(1):34–45.CrossRefGoogle Scholar
  15. 15.
    Barakat O, Abbasi S, Rodriguez G, Rios J, Wood RP, Ozaki C, et al. Use of decellularized porcine liver for engineering humanized liver organ. J Surg Res. 2012;173(1):e11–25.Google Scholar
  16. 16.
    Hashemi J, Pasalar P, Soleimani M, Khorramirouz R, Fendereski K, Enderami SE, et al. Application of a novel bioreactor for in vivo engineering of pancreas tissue. J Cell Physiol. 2018;233(5):3805–16.Google Scholar
  17. 17.
    Kajbafzadeh AM, Khorramirouz R, Kameli SM, Fendereski K, Daryabari SS, Tavangar SM, et al. Three-year efficacy and patency follow-up of decellularized human internal mammary artery as a novel vascular graft in animal models. J Thorac Cardiovasc Surg. 2019;157(4):1494–502.Google Scholar
  18. 18.
    Khorramirouz R, Kameli SM, Fendereski K, Daryabari SS, Kajbafzadeh AM. Evaluating the efficacy of tissue-engineered human amniotic membrane in the treatment of myocardial infarction. Regen Med. 2019;14(2):113–26.CrossRefGoogle Scholar
  19. 19.
    Kitahara H, Yagi H, Tajima K, Okamoto K, Yoshitake A, Aeba R, et al. Heterotopic transplantation of a decellularized and recellularized whole porcine heart. Interact Cardiovasc Thorac Surg. 2016;22(5):571–9.Google Scholar
  20. 20.
    Young RC, Goloman G. Allo- and xeno-reassembly of human and rat myometrium from cells and scaffolds. Tissue Eng A. 2013;19(19–20):2112–9.CrossRefGoogle Scholar
  21. 21.
    Ott HC, Matthiesen TS, Goh SK, Black LD, Kren SM, Netoff TI, et al. Perfusion-decellularized matrix: using nature’s platform to engineer a bioartificial heart. Nat Med. 2008;14(2):213–21.Google Scholar
  22. 22.
    Myers KM, Elad D. Biomechanics of the human uterus. Wiley Interdiscip Rev Syst Biol Med. 2017;9(5).Google Scholar
  23. 23.
    Bhrany AD, Lien CJ, Beckstead BL, Futran ND, Muni NH, Giachelli CM, et al. Crosslinking of an oesophagus acellular matrix tissue scaffold. J Tissue Eng Regen Med. 2008;2(6):365–72.Google Scholar
  24. 24.
    Baptista PM, Siddiqui MM, Lozier G, Rodriguez SR, Atala A, Soker S. The use of whole organ decellularization for the generation of a vascularized liver organoid. Hepatology (Baltimore, Md). 2011;53(2):604–17.CrossRefGoogle Scholar
  25. 25.
    Williams C, Liao J, Joyce EM, Wang B, Leach JB, Sacks MS, et al. Altered structural and mechanical properties in decellularized rabbit carotid arteries. Acta Biomater. 2009;5(4):993–1005.Google Scholar
  26. 26.
    Arenas-Herrera JE, Ko IK, Atala A, Yoo JJ. Decellularization for whole organ bioengineering. Biomed Mater (Bristol, England). 2013;8(1):014106.CrossRefGoogle Scholar
  27. 27.
    Cebotari S, Tudorache I, Jaekel T, Hilfiker A, Dorfman S, Ternes W, et al. Detergent decellularization of heart valves for tissue engineering: toxicological effects of residual detergents on human endothelial cells. Artif Organs. 2010;34(3):206–10.Google Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Seyedeh Sima Daryabari
    • 1
  • Abdol-Mohammad Kajbafzadeh
    • 1
    Email author
  • Kiarad Fendereski
    • 1
  • Fariba Ghorbani
    • 1
  • Mehrshad Dehnavi
    • 1
  • Minoo Rostami
    • 1
  • Bahram Azizi Garajegayeh
    • 2
  • Seyed Mohammad Tavangar
    • 3
  1. 1.Section of Tissue Engineering and Stem Cell Therapy, Pediatric Urology and Regenerative Medicine Research Center, Children’s Medical Center, Pediatric Center of ExcellenceTehran University of Medical SciencesTehranIran
  2. 2.Imaging Center, Children’s Medical CenterTehran University of Medical SciencesTehranIran
  3. 3.Department of Pathology, Shariati HospitalTehran University of Medical SciencesTehranIran

Personalised recommendations