3D culturing: from individual cells to blastemic tissue (Revisited the phenomenon of epithelial - mesenchymal plasticity)

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The phenomenon of epithelial - mesenchymal plasticity of multipotent mesenchymal stromal cells CMMSCsJ observed in 2D cultures has not been clarified yet. MMSC 3D culturing to model the patterns of epithelial cell layer formation (lamination] gives new opportunities. The review of literature and own findings enabled a suggestion that it is spontaneous formation of MMSC spheroids that induces generation of blastemic tissue in fetal and adult organs when damaged. The feasibility of reprogramming dense MMSC spheroids into neuroectoderm and primitive entoderm has been demonstrated.

About the authors

I N Saburina

B C Repin

I N Saburina

The RAMS Institute of General Pathology and Pathophysiology, Moscow

The RAMS Institute of General Pathology and Pathophysiology, Moscow

VS Repin

The RAMS Institute of General Pathology and Pathophysiology, Moscow

The RAMS Institute of General Pathology and Pathophysiology, Moscow


  1. Репин B.C., Сабурина И.Н. Обратимые эпителио-мезенхимальные трансформации клеток в эмбриогенезе и постнатальном обновлении тканей. Клеточная трансплантология и тканевая инженерия 200В; 3: 64-73.
  2. Zipori D. MSCs: harnessing cell plasticity to tissue and organ repair. Blood Cell Mol. Dis. 2004; 33: 211-15.
  3. Zipori D. Biology of Stem Cells and the molecular basis of the stem state. Ed. Bosch G. Thomas C.G. Springer Verlag, Munich, Heidelberg. 2009.
  4. Zipori D. The stem state: mesenchymal plasticity as a paradigm. Curr. Stem Cell Res. Ther. 2006; 1: 95-102.
  5. Prindull G., Zipori D. Environmental guidance of normal and tumor cell plasticity: epithelio-to-mesenchymal transitions as a paradigm. Blood 2004; 8: 8-12.
  6. Thiery J.P., Acloque H., Huang R.Y. et al. Epithelial-to- mesenchymal transitions in development and disease. Cell 2009; 139: 871-90.
  7. Ryan P., Foty R.A., Kohn J. et al. Tissue spreading on implantable substrates is a competitive outcome of cell-cell vs cell-substratum adhesivity. Proc. Natl. Acad. Sci. US. 2001; 98: 4323-27.
  8. Ninomiya H., Winklbauer R. Epithelial coating controls mesenchymal shape change through tissue-positioning effects and reduction of surface-minimizing tension. Nat. Cell Biol. 2008; 10: 61-71.
  9. Marga F., Neagu A., Kostzin I. et al. Developmental Biology and Tissue Engineering. Birth Defects Res. Part С 2007; 81: 320-8.
  10. Takezawa T. A strategy for the development of tissue engineering scaffolds that regulates cell behavior. Biomaterials 2003; 24: 2267-75.
  11. Hoffman R.M. To do culture in 2D or 3D? That is the question. Stem Cells 1993; 11: 105-11.
  12. Walsh K., Megyesi J., Hammond R. Human CNS tissue cultures: a historical review and examination of recent advances. Neurobiol. Dis. 2005; 18: 2-18.
  13. Raiteri M. Functional pharmacology in human brain. Pharmacol. Rev. 2006; 58: 162-93.
  14. Gibbons H.M., Dragunow M. Adult human brain cell culture for neuroscience research. Int. J. Biochem. Cell Biol. 2010; 42C6): 844-56.
  15. Sutherland R.M. Cell and environment interactions in tumor genesis: the multicell spheroid model. Science 1988; 240: 177-84.
  16. Chun M.H. Serum signaling factors and spheroids. Crit. Rev. Oncol. Hematol. 2000; 36: 89-98.
  17. Morales J., Alpaugh M.L. Gain in cellular organization of inflammatory breast cancer: a 3D in vitro model that mimics the in vivo metastasis. BMC Cancer 2009; 9: 462-9.
  18. Miller B.E. Michelson S. Tumor micro-ecology and competitive interactions. J. Theoret. Biol. 1987; 128: 233-46.
  19. Kuwashima Y., Yamada Т., Saio M. et al. Formation and growth of multicellular spheroids in medium containing low concentration of agarose. Cancer Letters 1993; 71: 31-36.
  20. Emerman J.Т., Pitelka D.R. Maintenance and induction of morphologic differentiation in dissociated mammary epithelium on floating collagen membrane. In Vitro 1977; 13: 316-28.
  21. Li M.L., Aggler J., Farson D.A. Influence of reconstituted basement membrane and its components on casein gene expression and secretion in mouse mammary epithelial cells. Proc. Natl. Acad. Sci US. 1987; 84: 136-40.
  22. Moscona A., Moscona H. The dissociation and aggregation of cells from organ rudiments of the early chick embryos. J. Anat. 1952; 86: 287-303.
  23. Moscona A.A. Tissues from dissociated cells. Sci. Amer. 1959; 200: 132-4.
  24. Steinberg M.S. Reconsttution of tissues by dissociated cells: some morphogenetic tissue movements and the sorting out of embryonic cells may have a common explanation. Science 1963; 141: 401-8.
  25. Steinberg M.S. Differential adhesion and morphogenesis: a modern view. Curr. Opin. Genet. Dev. 2007; 17: 281-5.
  26. Mironov V., Visconti R.R., Kasjanov V. et al. Organ printing: tissue spheroids as a building blocks. Biomaterials 2009; 30: 2164-74.
  27. Takezawa Т., Mori Y., Yonaha T. et al. Characterization of morphology and cellular metabolism during spheroid formation. Exp. Cell Res. 1993; 208: 430-41.
  28. Takahashi K., Mitsui M., Takeuchi K. et al. Preservation of the characteristics of the cultured human type 2 alveolar epithelial cells. Lung 2004; 182: 213-28.
  29. Lin R.Z., Chang H.Y. Recent advances in 3-D-multicellular spheroid culture for biomedical research. Biotechnology 2008; 3: 1172-84.
  30. Kelm J.M., Dijonov V., Ittner M. Design of custom shaped vascularized microtissues using spheroids as a minimal building units. Tissue Eng. 2006; 12: 2151-60.
  31. Torisawa Y., Takagi A., Nashimoto Y. A multicellular spheroid assay to realize spheroid formation, culture and viability assay on a chip. Biomaterials 2007; 28: 559-66.
  32. Long S.H., Smith J., Hyde С Differentiation of prostate epithelial cell cultures by matrigel/stromal cell glandular reconstruction. In Vitro Cell Dev. Anim. 2006; 42: 273-80.
  33. Castillon N., Hinnrasky J., Zahm J.M. et al. Polarized expression of cystic fibrosis transmembrane conductance regulator and associated epithelial proteins during the regeneration of human airway surface epithelium in 3-D- culture. Lab. Invest. 2002; 82: 989-98.
  34. Hoem D., Dalen H., Andren-Sandberg A. et al. Nonadhesive organ culture of human exocrine pancreatic cells with their stroma. Pancreas 2002; 26: 71-7.
  35. Fjellbirkeland L., Bjerkvig R., et al. Non-adhesive stationary organ culture of human bronchial mucosa. Am. J. Resp. Cell Mol. Biol. 1996; 15: 197-206.
  36. Gamarra F., Bergner A., Stauss E. et al. Rotation frequency of human bronchial and nasal epithelial spheroids as an indicator of mucociliary function. Respiration 2006; 73: 864-72.
  37. Tesei A., Zoli W., Arienti С et al. Isolation of stem /progenitor cells from normal lung tissue of adult humans. Cell Prolif. 2009; 42: 298-308.
  38. Gati I., Danielsson G., Betmark T. et al. Culturing of diagnostic muscle biopsies as spheroid-like structures: a pilot study of morphology and viability. Neurol Res. 2009 Aug 5. Epub ahead of print.
  39. Sarig R., Baruchi Z., Fuchs 0. et al. Regeneration and transdifferentiation potential of muscle-derived stem cell propagated as myospheres. Stem Cells 200B; 24: 17B9-7B.
  40. Nomura Т., Ashihara E., Tateishi K. et al. Skeletal myosphere-derived progenitor cell transplantation promotes neovascularization in sarcoglycan knock down cardiomyopathy. Biochem. Biophys. Res. Comms. 2007; 352: 668-74.
  41. Arsic N., Mammaeva D., Lamb N.J. et al. Muscle-derived stem cells isolated as non-adherent population give rise to cardiac, skeletal muscle and neural lineages. Exp. Cell Res. 2008; 314: 1266-80.
  42. Gati I., Danielsson 0., Betkark T. Spheroid-like structures in cultures of muscle biopsies. Neuromusc. Disorders 2007; 17: 876-82.
  43. Teunissen C.E. Whole brain spheroids cultures as a model to study the development of NO-synthase guanylate cyclase signal transduction. Brain Res. 2000; 125: 99-115.
  44. Andersen R.M., Johansen M., Blaabjerg M. et al. Neural tissue spheres: a microexplant culture method for propagation of precursor cells from the rat forbrain subventricular zone. J. Neurosci. Methods. 2007; 165: 55-63.
  45. Andersen R.K., Zimmer J., Wahlberg L.U. et al. Effect of LIF and long term propagation of precursor cells derived from rat forebrain subventricular zone and ventral mesencephalon. Exp. Neurol. 2008; 211: 301-10.
  46. Funderburgh M.L., Du Y., Funerburgh J. Primary keratocytes form spheroid bodies in vitro. Invest. Ophtalmol. Vis. Sci. 2005: 46.
  47. Ushida S., Yokoo S., Yanagi Y. et al. Sphere formation and expression of neural proteins by human corneal stromal cells in vitro. Invest. Ophtalmol. Vis. Sci. 2005; 46: 1620-5.
  48. Mimura Т., Amano S., Yokoo S. et al. Isolation and distribution of rabbit keratocyte precursors. Mol. Vision. 2008; 14: 197-208.
  49. Layer R.G., Willbold E. Regeneraion of the avian retina by retinospheroid technology. Progr. Retinal Eye Res. 1994; 13: 197-207.
  50. Викторов И.В., Александрова О.П., Алексеева Н.Ю. Роллерная органная культура сетчатки постнатальных крыс. Бюлл. Эксп. Биол. мед. 2006; 142: 486-9.
  51. Tait I.S., Flint N., Cambell F.C. et al. Generation of neomucosa in vivo by transplantation of dissociated rat postnatal small intestinal epithelial cells. Differentiation 1994; 56: 91-100.
  52. Evans G.S., Flint N., Somers A.S. et al. The development of a method for the preparation of rat intestinal epithelial cell primary culture. J. Cell Sci. 1992; 101: 219-31.
  53. Slorach E.M., Campbell F.C, Dorin J.R. A mouse model of intestinal stem cell function and regeneration. J. Cell Sci. 1999; 112: 3029-38.
  54. Cottrill СР., Archer C.W., Wolpert L. Cell sorting and hondrogenic aggregate formation in micromass culture. Dev. Biol. 1987; 122: 503-15.
  55. Neufeld D.A. Formation of prechondrogenic mesenchymal cell aggregates in the regenerating newt limb. Dev. Biol. 1982; 93: 36-42.
  56. Gonzalez P., Epstein D.L., Luna С et al. Characterization of free-floating spheres from human trabecular meshwork cell culture in vitro. Exp. Eye Res. 2006; 82: 959-67.
  57. Chen M.H., Chen Y.J., Liao C.C. et al. Formation of salivary acinar cell spheroids in vitro above a polyvinyl alcohol-coated surface. J. Biomed. Mater. Res. 2009; 90: 1068-72.
  58. Hisatomi Y., Okumura K., Nakamura K. et al. Flow cytometric isolation of endodermal progenitors from mouse salivary gland differentiate into hepatic and pancreatic lineages. Hepatology 2004; 39: 667-75.
  59. Matsumoto S., Okumura K., Ogata A. et al. Isolation of tissue progenitor cells from duct-ligated salivary gland of swine. Cloning Stem Cells 2007; 9: 176-90.
  60. Carlsson J., Nilsson K., Westermark B. et al. Formation and growth of multicellular spheroids of human origin. Int. J. Cancer. 1983; 31: 523-33.
  61. Inoue S., Takaoka K., Endo T. et al. In vitro confirmation of a newly established lung cancer lines using flow cytometry and multicellular tumor spheroids. Lung Cancer 1997; 17: 85-101.
  62. Kubota A., Nishida K., Nakashima K. et al. Conversion of mammalian Muller glial cells into a neuronal lineage by in vitro aggregate-culture. Biochem. Biophys. Res. Comms. 2006; 351: 514-20.
  63. Willbold E., Berger J., Reinicke M. et al. On the role of Muller glia cells in histogenesis: Only retinal spheroids, but not tectal, telencephalic and cerebellar spheroids evelop histiotypical patterns. J. Hirnforsch. 1997; 38: 383-8.
  64. Ревищин А.В., Полтавцева P.А., Марей M.B. и др. Характеристика клеточных кластеров возникающих в культуре диссоциированных клеток эмбрионального мозга человека. Бюлл. Эксп. Биол. мед. 2001; 132: 856-60.
  65. Weinlich М., Baumstark С, Usta Е. et al. Human duodenal spheroids for non-invasive intracellular pH measurement and quantification of regulation mechanisms under physiological conditions. In Vitro Cell Dev. 2002; 38: 7-13.
  66. Frith J.E., Thomson В., Genever P. Dynamic 3D-culture methods enhance MSC cell properties and increase therapeutic potential. Tissue Eng Part С Methods. 2009 Oct 7. Epub ahead of print.
  67. Sauerzweig S., Munsch Т., Lessmann V. et al. A population of serum deprivation-induced bone marrow MSCs express markers typical for embryonic and neural stem cells. Exp. Cell Res. 2009; 315(1): 50-66.
  68. Howson K.M., Aplin A.C., Gelati M. et al. The postnatal rat aorta contains pericyte progenitor cells that form spheroidal colonies in suspension culture. Am. J. Physiol. Cell Physiol. 2005; 289: 1396-407.
  69. Laib A.M., Bartol A., Alajati A. et al. Spheroid-based human endothelial cell microvessel formation in vivo. Nat. Protoc. 2009; 4: 1202-15.
  70. BrophyC.M., Luebke-Wheeler J.L., Amiot B.P. etal. Rat hepatocyte spheroids formed by rocked technique maintain differentiated hepatocytes gene expression and function. Hepatology 2009; 49: 578-86.
  71. Yuasa C, Tomita Y., Shono M. etal. Importance of cell aggregation for expression of liver functions and regeneration demonstrated with the primary cultured hepatocytes, J. Cell. Physiol. 1993; 156: 522-30.
  72. Galan-Rodriguez В., del-Marco A., Flores J.A. et al. Grafts of extra-adrenal chromaffin cells as aggregates show better survival rate and regenerative effects on parkinsonian rats than despersed cell grafts. Neurbiol. Dis. 2008; 29: 529-42.
  73. Garzoni L.R. Rossi M.I., de Barros A.P. et al. Dissecting coronary angiogenesis: 3D culture of cardiomyocytes with endothelial or mesenchymal cells. Exp. Cell Res. 2009; 315: 3406-18.
  74. Thiery J.P., Sleeman J.P. Complex networks orchestrate epithelio-mesenchymal transition. Nat. Rev. Mol. Cell Biol. 2006; 7: 131-41.
  75. Grobstein С Mechanisms of organogenetic tissue interaction. J. Natl. Cancer Inst. Monogr. 1967; 26: 279-99.
  76. Cunha G.R. Mesenchymal-epithelial interactions: past, present and future. Differentiation 2008; 76: 578-86.
  77. Havlikova В., Biro Т., Mescalchin A. et al. Folliculoid microsphere assay for exploring epithelio-mesenchymal interactions in the human hair follicle. J. Investig. Dermatol. 2009; 129: 972-83.
  78. Dean C, Ito M., Makarenkova N.P. et al. BMP-7 regulates branching morphogenesis of the lacrimal gland by promoting mesenchymal proliferation and condensation. Development 2004; 131: 4155-65.
  79. Yang J., Weinberg R.A. Epithelial-mesenchymal transition: at the crossroads of development and tumor metastasis. Dev. Cell. 2008; 14: 318-28.
  80. Hay E.D., Zuk A. Transformation between epithelium and mesenchyme: normal, pathological and experimentally induced. Am. J. Kidney Dis. 1995; 26: 678-90.
  81. Hay E.D. The mesenchymal cell, its role in the embryo, and the remarkable signaling mechanism, that create it. Dev. Dyn. 2005; 233: 706-20.
  82. Shook D., Keller R. Mechanisms, mechanics and function of epithelio-mesenchymal transition in early development. Mech. Dev. 2003; 120: 1351-83.
  83. Hader C, Marlier A., Cantley L. Mesenchymal-epithelial transition n epithelial response to injury: the role of FoxC2. Oncogene 2009; 131: 1-10.
  84. Hudson L.G., Newkirk K.M., Chandler H.L. et al. Cutaneous wound reepithelialization is compromised in mice lacking functional Slug [Snail2]. J. Dermatol. Sci. 2009; 56: 19-26.
  85. Cornier J.Т., zur Nieden N.I., Rancourt D.E. et al. Expansion of undifferentiated murine embryonic stem cells as aggregates in suspension culture bioreactors. Tissue Eng. 2006; 12: 3233-45.
  86. zur Nieden N.I., Cormier J.I., Rancourt D.E. et al. Embryonic stem cells remain highly pluripotent following long term expansion as aggregates in suspension bioreactors. J. Biotechnol. 2007; 129: 421-32.
  87. Oh S.K., Chen A.K., Mok Y. et al. Long-term microcarrier suspension cultures of hESCs. Stem Cell Res. 2009; 2: 219-30.
  88. Djordjevic В., Lange C.S. Cell-cell interactions in ESC spheroids maintained in suspension. Acta Oncol. 2006; 45: 373-5.
  89. Mohr J.C., Pablo J.J., Palecek S.P. 3D microwell culture of human ESC. Biomaterials 2006; 27: 6032-42.
  90. Yim E.K., Wen J., Leong K.W. Enhanced extracellular matrix production and differentiation of hESCs derivatives in biodegradable poly-caprolactone-co-ethyl ethylenephosphate scaffold. Acta Biomaterialia 2006; 2: 365-76.
  91. Ouyang A., Ng R., Yang S.T. et al. Long term culturing of undifferentiated ESCs in conditioned media and 2D fibrous matrices. Stem Cells 2007; 25: 447-54.
  92. Gerecht S., Burdick J.A., Ferreira R.S. et al. Hyaluronic acid hydrogel for controlled self-renewal and differentiation of human embryonic stem cells. Proc. Natl. Acad. Sci. US. 2007; 104: 11298-303.
  93. Paquet-Durand F., Tan S., Bicker G. Turning teratocarcinoma cells into neurons: rapid differentiation of NT-2 cells in floating spheres. Dev. Brain Res. 2003; 142: 161-7.
  94. Barberi Т., Willis L.M., Socci N.D. et al. Derivation of multipotent MSC precursors from hESCs. PloS Medicine 2005; 2: 161-9.
  95. Stojkovich P., Lako M., Stewart R. et al. An autogenic feeder cells system that efficintly supports growth of undifferentiated hESCs. Stem Cells 2005; 23: 306-14.
  96. Hwang N.S., Varghese S., Lee H.J. et al. In vitro commitment and functional regeneration using hESC-derived MSCs. Proc. Natl. Acad. Sci US. 2008; 105: 20641-6.
  97. Pozzobon M., Picolli M., Ditadi A. et al. Mesenchymal stem cells can be derived from CD133+ cells: implication for therapy. Stem Cell Dev. 2009; 18: 497-507.
  98. Hewitt K.J., Shamis Y., Carlson M.W. et al. Three dimensional epithelial tissues generated from human ESCs. Tissue Eng. 2009; 15: 3417-27.
  99. Behr R., Heneweer C, Viebahn С et al. Epithelial-mesenchymal transition in colonies of rhesus monkey ESCs: a model for processes involved in gastrulation. Sem Cells 2005; 23: 805-16.
  100. Alacorn V.B., Marikawa Y. Molecular study of mouse peri-implantation development using the in vitro culture of aggregated Inner Cell Mass. Mol. Reprod. Dev. 2004; 67: 83-90.
  101. Masaki H., Nishida Т., Kitajima S. et al. Developmenta pluripotency associated 4 (DPPA4) localized in active chromatin inhibits mESC differentiation into a primitive ectoderm lineage. J. Biol.Chem. 2007; 282: 33034-42.
  102. Liu Z.J., Zhuge Y., Velazquez O.C. Trafficking and Differentiation of Mesenchymal Stem Cells. Journal of Cellular Biochemistry 2009; 106: 984-91.
  103. Potapoval.A., Gaudette G.R., Brink P.R., Robinson R.В., Rosen M.R., Cohen I.S., Doronin S.V. Mesenchymal stem cells support migration, extracellular matrix invasion, proliferation, and survival of endothelial cells in vitro. Stem Cells 2007; 25: 1761-8.
  104. Сабурина И.Н., Горкун A.A., Кошелева H.B., Семенова М.Л., Пулин А.А., Репин B.C. Сопоставление поведения стромальных клеток пупочного канатика и мультипотентных стромальных клеток взрослого костного мозга в 2-D и 3-D культуре: моделирование стромальной регенерации. Вестник новых медицинских технологий 2009; XVI [41: 9-11.
  105. Yoo J.U., Barthel T.S., Nishimura К. et al. The chondrogenic potential of human bone-marrow-derived mesenchymal progenitor cells. The Journal of Bone and Joint Surgery 1998; 80-AC12): 1745-57.
  106. Wang W., Itaka K., Ohba S. et al. 3D spheroid culture system on micropatterned substrates for improved differentiation efficiency of multipotent mesenchymal stem cells. Biomaterials 2009; 30: 2705-15.
  107. Penick K.J., Solchaga L.A., Welter J.F. High-throughput aggregate culture system to assess the chondrogenic potential of mesenchymal stem cells. Biotechniques 2005; 39C5): 687-91.
  108. Welter J.F., Solchaga L.A., Penick K.J. Simplification of aggregate culture of human mesenchymal stem cells as a chondrogenic screening assay. Biotechniques 2007; 42: 732-7.
  109. Lin R-Z., ChouL-F., Chien C-C.M. et al. Dynamic analysis of hepatoma spheroid formation: roles of E-cadherin and el-integrin. Cell and tissue Research. 2006; 324C3]; 411-422.
  110. Curcio E., Salerno S., Barbieri G. et al. Mass transfer and metabolic reactions in hepatocyte spheroids cultured in rotating wall gas-permeable membrane system. Biomaterials 2007; 28(36): 5287-97.
  111. Griffith L.G., Swartz M.A. Capturing complex 3D tissue physiology in vitro. Molecular Cell Biology 2006; 7: 211-24.

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