Document Type: Original Article

Authors

1 Department of Veterinary Medicine, Islamic Azad University, Eghlid Branch, Iran

2 PhD student, Faculty of Veterinary Medicine, Shahrekord University, Shahrekord, Iran

3 Research Institute of Animal Embryo Technology, Shahrekord University, Shahrekord, Iran

4 Reproductive Biotechnology Research Center, Avicenna Research Institute, ACECR, Tehran, Iran

5 Department of Clinical Sciences, Faculty of Veterinary Medicine, Shahrekord, University, Shahrekord, Iran

Abstract

Background: Background: Human-amnion membrane mesenchymal stem cells (hAMSCs) are the accessible cells that can be grown in in vitro condition to produce a great number of cells required for cell therapy in medicine. Several protocols have been proposed for isolation of hAMSCs, but the most of them are multi-step and expensive. The purpose of this article was to introduce a one-step, inexpensive protocol.
Methods: In this study human amnion membrane (hAM) was obtained from selected caesarean-sectioned births. The AM was sliced into small pieces and to isolate MSCs, it was digested only with one step instead of multi-step. Isolated cells were cultured in Dulbeco,s Modified Eagle,s Medium-Low Glucose (DMEM-LG) with 10% fetal bovine serum (FBS) without adding growth factors. After 80-90% confluency, the adherent cells were characterized by flow cytometry and multi- potentiality differentiation toward adipocyte-, and osteocyte -like cells.
Results: The results showed that hAMSCs isolated from hAM expressed CD105, CD90 and CD73 such as other MSCS, but did not express CD34 and CD45 hematopoietic markers. The osteogenic and adipogenic differentiation of the isolated cells were proven by Alizarin Red and Oil-Red-O straining, respectively.
Conclusions: The results showed that the stem cells derived from the AM belonged to the mesenchymal stem cells family. Furthermore, this method made it easier and cheaper to obtain this type of stem cells.

Keywords

Main Subjects

1. Barry F P, Murphy J M. (2004). Mesenchymal stem cells: Clinical applications and biological characterization Int. J. biochem. Cell Biol, 36(4): 568-584.

2. Chen F, Tuan R. (2008). Mesenchymal stem cells in arthric disease Arthric Res Ther, 10(5): 223.

3. Magana-Guerrero F, Dominguez-Lopez A, Martinez-Aboytes P, Buentello-Volante B, Yonathan-Garfas B. (2017). Human Amniotic Membrane Mesenchymal Stem Cells inhibit Neutrophil Extracellular Traps through TSG-6. SCLenTIFIC REPORTS, 7:12426. 10.1038/s41598-017-10962-2.

4. Zhang D, Jiang M, Miao D. (2011). Tranplanted humen amniotic membrane-derived mesenchymal stem cells ameliorate carbon tetrachloride-induced liver cirrhosis in mouse. PLoS One, 6(2): e16789.

5. Friedenstein A J, Piatetzky S, Petrakova K. (1966). Osteogenesis in transplants of bone marrow cells. J Embryol Exp Morphol, 16: 381-390.

6. Lu L-L, Liu Y-J, Yang S-G, Zhao Q-J, Gong W, al. e. (2006). Isolation and  characterization of human unbilical cord mesenchymal stem cells with hematopoiesis supprotive function and other potentials. Haematologica, 91(8): 1017-1026.

7. Da-Silva-Meirells L, Chagastelles P C, Nardi N B. (2006). Mesenchymal stem cells reside in virtually all postnatal organs and tissues. J Cell Sci, 119: 2204-2213.

8. dominici M, le Blanc K, Mueller L, Slaper-Cortenbach I, Marini F, Krause D, Deans R, Keating A, Prockop D, Horwitz E. (2006). Minimal criteria for defining multipotent mesenchymal stromal cells. The international Society for Cellular Therapy position statement. , Cytotherapy 8: 315-317.

9. Miao Z, Jin j, Chen L, Zhu J, Huang W, Zhao J. (2006). Isolation of mesenchymal stem cells from human placenta: comparison with human bone marrw mesenchymal stem cells. Cell Biol Int, 30(9): 981-987.

10. Scherjon S, Kleijburg-van der keur C, De-Groot-Swings G, Claas F, Fibbe W, Kanhai H. (2004). Isolation of mesenchymal stem cells of fetal and materna origin from human placenta. Stem Cells, 22(7): 1338-1345.

11. Cao Y, Sun Z, Liao L, Meng Y, Han Q, Zhao R. (2005). Human adipose tissue-derived stem cells differentiate into endothelial cells in vitro and improve postnatal neovascularization in vivo. Biochem Biophys Res Commun, 332(2): 370-379.

12. Diaz-Prado S, Muinos-Lopez A, Harmida-Gomez T, Rendal-Vazquez M, Fuentes-Boquete I, Francisco J. (2011). Isolation and  characterization of mesenchymal cells from human amniotic membrane Tissue Engineering Part-C: Methods., 17(1): 49-59.

13. Motedayyen H, Esmaeil N, Tajik N, Khadem F, S. G, Khani B, Rezaei A. (2017). Method and key points for isolation of human amniotic epithelial cells with high yield, viability and purity. BMC Res Notes, 10(552). 10.1186/s13104-017-2880-6

14. Razavi-Tousi M T, Amirizadeh N, Nasirinezhad F, Nikougoftar F, Ganjibkhsh M, Aboutaleb N. (2017). A rapid and cost-effective protocol for isolating mesnchymal stem cells from human amniotic membrane. GMJ, 6(3): 217-225.

15. Tsuji H, Miyoshi S, Ikegami Y, Hida N, Asada H, Togashi I. (2010). Xenografted human amniotic membrane-derived mesenchymal stem cells are immounologically tolerated and transdiffentiated into cardiomyocytes. Circul Res, 106(10): 1613-1623.

16. Kimotoshi K, Shunsuke O, Hidetaka H, Moto F, Ayano K, Ryosuke H, Takahiro Y, Reiz O, Kinichi Y, Hiroshi T, Naoya S. (2015). Human amniotic-derived mesenchymal stem cell transplantation ameliortes liver fibrosis in Rats. The Transplantation Society, 1(e16). 10.1097/TXD.0000000000000525

17. Alviano F, Fossati V, Marchionni C, Arpinati M, Bonsi L, Franchina M, al. e. (2007). Term amniotic membrane is a high throughput source for multiputent mesenchymal stem cells with the ability to diffenrentiate into endothelial  in vitro. DMC Dev Biol, 21(4): 11.

18. Kim S, Zhang H Z, Kim C E, An J M, Kim M H. (2012). Amniotic mesenchymal stem cells have rebust angiogenic properties and are effective in treating hindlimb ischaemia. Cardiovacular Research, 39(3): 525-534.

19. Solomon A, Wajngarten M, Alviano F, Anteby I, Elchalal U, peer J, al. e. (2005). Suppression of inflammatory and fibrotic responses in an in-vito model of allergic inflammation by the amniotic membrane stromal matrix. Clin Exp Allergy, 35(7): 941-948.

20. Dua H, Gomes J, King A, Maharajan V. (2004). The amniotic membrane in ophthalmolog. Surv Ophthalmol., 49(3): 51-77.

21. Chavez-Garcia C, al. e. (2016). Ophtalmic indications of amniotic membrane transplantation in Maxico: an eight years Amniotic Membrane Bank experience. Cell Tissue Bank, 17: 261-268.

22. Noth U, Steinert A, Tuan R. (2008). Technology insight: adult mesenchymal stem cells for osteoarthritis therapy. Nat Rev Rheumatol, 4(7): 371-380.

23. Ono M, al. e. (2015). Effects of human amnion-derived mesenchymal stromal cell transplantion in rats with radiation proctitis. Cytotherapy, 17: 1545-1559.

24. alviano F, Fossati V, Marchionni C, Arpinati M, Franchina M. (2007). Term amniotic membrane is a high throughput source for multipotent mesenchymal stem cells with the ability to differentiate into endothelial cells in-vitro. BMC Dev Biol, 21(7): 11.

25. Soncini M, Vertua E, Gibelli L, Zorzi F, Denegri M, Albertini A. (2007). Isolation and characterization of mesenchymal stem cells from human fetal membranes. J Tissue Eng Regen Med, 1(4): 296-305.

26. Toda A, Okabe M, Yoshida T, Nikaido T. (2007). The potential of amniotic membrane/amnion-derived cells for regeneration of  various tissues. J Pharmacol Sci, 105: 215-228.

27. Murphy S, Kidpoor A, Reid T, Atala A, Wallace E, Lim R. (2014). Isolation, cryopreservation and culture of human amniotic epithelail cells for clinical applications. J Vis Exp, org/10.3791/52085.

28. Mihu C M, Rus-Ciuca D, Soritau O, Susman S, Mihu D. (2009). Isolation and characterization of mesenchymal stem cells from the amniotic membrane. Morphology and Embryology, 50(1): 73-77.

29. Parolini O, al. e. (2008). Concise review: Isolation and characterization of cells from human term placenta outcome of the first international workshop on placenta derived stem cells. Stem Cells, 26: 300-311.

30. Kaboyashi M, Yakuwa T, Sasaki K, Sato K, Kikuchi A, Kamo I, Yokoyama Y, Sakuragawa N. (2008). Multilineage potential of side population cells from human amniotic mesenchymal layer. Cell Transplant, 17: 291.

31. Tamagawa T, Ishikawa H, Nakamura Y. (2008). Induced in-vitro differentiation of neurallike cells from human amnion-derived fibroblast-like cells. Hum Cell, 21: 38.