Phenotypical changes and proliferative potential of mesenchymal stem cells from humans Wharton’s jelly in the cultivation conditions

Vitaliy Tsymbaliuk, Elena Deriabina, Nadezhda Shuvalova, Olga Maslova, Ianina Pokholenko, Olena Toporova, Svitlana Shpileva, Vitaliy Kirik, Leonid Pichkur, Iuriy Kasianenko, Oleksandr Pichkur, Vitaliy Kordium

Abstract


Purpose. To develop a protocol for mesenchymal stem cells (MSCs) obtaining from wharton’s jelly of human umbilical cord, to study their phenotypic changes and proliferative potential in culture.

Materials and methods. MSCs from human umbilical cord were cultured for 6 passages. To study morphological features of the cells in a culture we used adapted staining with eosin and gematoksilіn, Romanovsky–Giemsa, and toluidine blue. For nuclear DNA coloring we used intercalators: Hoechst 33342, and DARІ Ethidiumbromid. MSCs surface markers (CD105, CD90, CD73, CD34) expression was studied by FACS-analysis.

Results. In the culture MSCs retain their morphological identity for 2 passages. Further a tendency to MSCs markers expression decreasing was observed, signs of culture degradation, spontaneous chondrogenic and adipogenic differentiation in late passages.

Conclusion. Cultured MSCs of human umbilical cord for 2 passages retain their morphological characteristics and proliferative potential, further they lose multipotent mesenchymal phenotype.


Keywords


mesenchymal stem cells; human umbilical cord; cultivation; proliferative potential

References


Augello A, Kurth T, De Bari C. Mesenchymal stem cells: a perspective from in vitro cultures to in vivo migration and niche. Eur Cell Mater. 2015;(20):121–133. [Publisher Full Text]

Bieback K, Brinkmann I. Mesenchymal stromal cells from human perinatal tissues: From biology to cell therapy. WJSC. 2010;2(4):81-92. CrossRef

Wang Z, Li Y, Ahmad A, Azmi A, Kong D, Banerjee S, Sarkar F. Targeting miRNAs involved in cancer stem cell and EMT regulation: An emerging concept in overcoming drug resistance. Drug Resistance Updates. 2010;13(4-5):109-118. CrossRef

Thiery J, Acloque H, Huang R, Nieto M. Epithelial-Mesenchymal Transitions in Development and Disease. Cell. 2009;139(5):871-890. CrossRef

Semenova VM, Lisianiy MI, Stayno LP, Belska LM, Yegorova DМ. [Proliferative and differentiated potential of mesenchymal stem cells from adipose tissue under cultivation conditions]. Ukrainian Neurosurgical Journal. 2014;(3):24–29. Russian. [Abstract/Full Text]

Selezneva T, Mishyn A, Barsukow V. Gistologiya. Polnyi Kurs Za Tri Dnya. Moscow: Eksmo; 2007. Russian.

Kalluri R. EMT: When epithelial cells decide to become mesenchymal-like cells. Journal of Clinical Investigation. 2009;119(6):1417-1419. CrossRef

Kalluri R, Zeisberg M. Fibroblasts in cancer. Nat Rev Cancer. 2006;6(5):392-401. CrossRef

Patel A, Park E, Kuzman M, Benetti F, Silva F, Allickson J. Multipotent Menstrual Blood Stromal Stem Cells: Isolation, Characterization, and Differentiation. cell transplant. 2008;17(3):303-311. CrossRef

Huang G, Gronthos S, Shi S. Mesenchymal Stem Cells Derived from Dental Tissues vs. Those from Other Sources: Their Biology and Role in Regenerative Medicine. Journal of Dental Research. 2009;88(9):792-806. CrossRef

Taghizadeh R, Cetrulo K, Cetrulo C. Wharton’s Jelly stem cells: Future clinical applications. Placenta. 2011;32:S311-S315. CrossRef

Maslova OO, Shuvalova NS, Sukhorada OM, Deryabina OG, Makarenko MV. Changes in morphofunctional characteristics of umbilical cord matrix mesenchimal cell вuring passaging. Problems of cryobiology. 2012;22(2):153–156. Available at: http://www.researchgate.net/publication/235248761_Changes_in_Morphofunctional_Characteristicsof_Umbilical_Cord_Matrix_Mesenchymal_Cells_During_Passaging. Accessed September 15, 2014.

Kozlovskaya LV, Nikolayev AYu. Uchebnoye posobiye po klinicheskim laboratornym metodam issledovaniya. Moscow: Meditsina, 1984. Russian.

Gimble J, Guilak F, Nuttall M, Sathishkumar S, Vidal M, Bunnell B. In vitro Differentiation Potential of Mesenchymal Stem Cells. Transfusion Medicine and Hemotherapy. 2008;35(3):228-238. CrossRef

Jennings B, Ridler P. Interaction of chromosomal stains with DNA. Biophys Struct Mechanism. 1983;10(1-2):71-79. CrossRef

Baraldi PG, Bovero A, Fruttarolo F, Preti D, Tabrizi MA, Pavani MG, Romagnoli R. DNA minor groove binders as potential antitumor and antimicrobial agents. Medicinal Research Reviews. 2004;24(4):475-528. CrossRef

Gray J, Darzynkiewicz Z. Techniques In Cell Cycle Analysis. Clifton, N.J.: Humana Press; 1987.

Kubista M, Aakerman B, Norden B. Characterization of interaction between DNA and 4',6-diamidino-2-phenylindole by optical spectroscopy. Biochemistry. 1987;26(14):4545-4553. CrossRef

Wexler S, Donaldson C, Denning-Kendall P, Rice C, Bradley B, Hows J. Adult bone marrow is a rich source of human mesenchymal 'stem' cells but umbilical cord and mobilized adult blood are not. British Journal of Haematology. 2003;121(2):368-374. CrossRef


GOST Style Citations


Augello A. Mesenchymal stem cells: a perspective from in vitro cultures to in vivo migration and niche / A. Augello, T.B. Kurth, C. De Bari // Eur. Cell Mater. — 2010. — V.20. — P.121–133.

Bieback K. Mesenchymal stromal cells from human perinatal tissues: From biology to cell therapy / K.Bieback, I.Brinkmann // World J. Stem Cells. — 2010. — V.2, N4. — P.81–92.

Targeting miRNAs involved in cancer stem cell and EMT regulation: An emerging concept in overcoming drug resistance / Z. Wang, Y. Li, A. Ahmad, A. Azmi, D. Kong, S. Banerjee, F. Sarkar // Drug Resistance Updates. — 2010. V.13, N4–5. — P.109–118.

Epithelial-mesenchymal transitions in development and disease / J-P. Thiery, H. Acloque, R.Y..J. Huang, M.A. Nieto // Cell. — 2009. — V.139, N5. — P.871–890.

Пролиферативный и дифференцировочный потенциал мезенхимальных стволовых клеток из жировой ткани в условиях культивирования / В.М.С еменова, Н.И. Лисяный, Л.П. Стайно, Л.Н. Бельськая, Д.М. Егорова // Укр. нейрохірург. журн. — 2014. — №3. — С.24–29.

Kalluri J. EMT: When epithelial cells decide to become mesenchymal-like cells / J. Kalluri // J. Clin. Invest. — 2009. — V.119, N6. — P.1417–1419.

Selezneva T. Gistologiya: polnyi kurs za tri dnya / T. Selezneva, A. Mishyn, V. Barsukow. — M.: Eksmo, 2007. — 354 p.

Kalluri R. Fibroblasts in cancer / R. Kalluri, M. Zeisberg // Nat. Rev. Cancer. — 2006. — V.6, N5. — P.392–401.

Multipotent menstrual blood stromal stem cells: isolation, characterization, and differentiation / A. Patel, E. Park, M. Kuzman, F. Benetti, F. Silva, J. Alickson // Cell Transplant. — 2008. — V.17, N3. — P.303–311.

Huang G.T.-J. Mesenchymal stem cells derived from dental tissues vs. those from other sources: their biology and role in regenerative medicine / G.T.-J. Huang, S. Gronthos, S. Shi // J.D.R. — 2009. — V.88, N9. — P.792–806.

Taghizadeh R.R. Wharton’s jelly stem cells: future clinical applications / R.R. Taghizadeh, K.J. Cetrulo, C.L. Cetrulo // Placenta. — 2011. — V.32, suppl.4. — P.311–315.

Changes in morphofunctional characteristics of umbilical cord matrix mesenchimal cell during passaging / O.O. Maslova, N.S. Shuvalova, O.M. Sukhorada, O.G. Deryabina, M.V. Makarenko // Probl. Cryobiol. — 2012. — V.22, N2. — P.153–156.

Козловская Л.В. Учебное пособие по клиническим лабораторным методам исследования / Л.В. Козловская, А.Ю. Николаев. — М.: Медицина, 1984. — 288 с.

In vitro differentiation potential of mesenchymal stem cells / J.M. Gimble, F. Guilak, M.E. Nuttall, S. Sathishkumar // Transfus. Med. Hemother. — 2008. — V.35. — P.228–238.

Jennings B.R. Interaction of chromosomal stains with DNA / B.R. Jennings, P.J. Ridler // Biophys. Struct. Mech. — 1983. — V.10. — P.71–79.

DNA minor grove binders as potential antitumor and antimicrobial agents / P.G. Baralddi, A. Bovero, F. Fruttarolo, D. Preti, M.A. Tbrizi, M.G. Pavani, R. Romagnoli // Med. Res. Rev. — 2004. — V.24. — P.475–528.

Gray J.W. Techniques in cell cycle analysis / J.W. Gray., Z. Darzynkiewicz. — N.J.: Humana Press, Clifton, 1986. — 407 p.

Kubista M. Characterization of interaction between DNA and 4’, 6-Dimidino-2-phenylindole by optical spectroscopy / M. Kubista, B. Akerman, B. Norden // Biochemistry. — 1987. — V.26. — P.4545–4553.

Adult bone marrow is a rich source of human mesenchymal “stem” cells but umbilical cord and mobilized adult blood are not / S. Wexler, C. Donaldson, P. Denning-Kendall, C. Rice, B. Bradley, J. Hows // Br. J. Haematol. — 2003. — V.121, N2. — P.368–374.





DOI: https://doi.org/10.25305/unj.45290

Copyright (c) 2015 Vitaliy Tsymbaliuk, Elena Deriabina, Nadezhda Shuvalova, Olga Maslova, Ianina Pokholenko, Olena Toporova, Svitlana Shpileva, Vitaliy Kirik, Leonid Pichkur, Iuriy Kasianenko, Oleksandr Pichkur, Vitaliy Kordium

Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 International License.