Isolation and Characterization of the Progenitor Cells From the Blastema Tissue Formed at Experimentally-Created Rabbit Ear Hole

Document Type : Original Article


1Department of Stem Cells and Developmental Biology, Cell Science Research Centre, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran


Objective(s): Throughout evolution, mammalians have increasingly lost their ability to regenerate structures however rabbits are exceptional since they develop a blastema in their ear wound for regeneration purposes. Blastema consists of a group of undifferentiated cells capable of dividing and differentiating into the ear tissue. The objective of the present study is to isolate, culture expand, and characterize blastema progenitor cells in terms of their in vitro
differentiation capacity.
Materials and Methods:
Five New Zealand white male rabbits were used in the present study. Using a punching apparatus, a 4-mm hole was created in the animal ears. Following 4 days, the blastema ring which was created in the periphery of primary hole in the ears was removed and cultivated. The cells migrated from the blastema were expanded through 3 successive subcultures and characterized in terms of their potential differentiation, growth characteristics, and culture requirements.
The primary cultures tended to be morphologically heterogeneous having spindly-shaped fibroblast-like cells as well as flattened cells. Fibroblast-like cells survived and dominated the cultures. These cells tended to have the osteogenic, chondrogenic, and adipogenic differentiation potentials. They were highly colonogenic and maximum proliferation was achieved when the cells were plated at density of 100 cells/cm2 in a medium which contained 10% fetal bovine serum (FBS).
Taken together, blastema tissue-derived stem cells from rabbit ear are of mesenchymal stem cell-like population. Studies similar to this will assist scientist better understanding the nature of blastema tissue formed at rabbit ear to regenerate the wound.


Metcalfe AD, Willis H, Beare A, Ferguson MW. Characterizing re
generation in the vertebrate ear. J Anat 2006; 209:439-46.
2. Williams-Boyce PK, Daniel JC Jr. Comparison of ear tissue regeneration in mammals. J Anat 1986;149:55-63.
3. Joseph J, Dyson M. Tissue replacement in the rabbit’s ear. Br J Surg 1966; 53:372-380.
4. Goss RJ, Grimes LN. Epidermal downgrowths in regenerating rabbit ear holes. J Morphol 1975; 146:533-542.
5. Brockes JP, Kumar A. Plasticity and reprogramming of differentiated cells in amphibian regeneration. Nat Rev Mol Cell Biol 2002; 3:566–574.
6. Iten LE, Bryant SV. Forelimb regeneration from different levels of
amputation in the newt, otophthalmus viridescens: length, rate and stages. Roux’s Arch Dev Biol 1973; 173:263–282.
7. Goss RJ, Grimes N. Tissue interactions in the regeneration of rabbit ear holes. Am Zool 1972; 12:151-157.
8. Williams-Boyce PK, Daniel JC Jr. Regeneration of rabbit ear tissue. J Exp Zool 1980; 212:243-53.
9. Mahdavi-shahri N, Naseri F, Kheirabadi M, Babaie, Sadeghie- Shakib F, Azarnia M. The ultra structural study of blastema in pinna tissues of rabbits with trabnsmission electron microscope. J Biol Sci 2008; 8: 993-1000.
10. Mahmoudi Z, Moghaddam-Matin M, Saeinasab M, Nakhaei-Rad S, Mirahmadi M, Mahdavi-Shahri N,
et al
. Blastema cells derived from rabbit ear show stem cell characteristics. J Cell Mol Res 2011 3: 25-31.
11. Clark LD, Clark RK, Heber-Katz E. A new murine model for mammalian wound repair and regeneration. Clin Immunol Immunopathol 1998; 88:35-45.
12. Eslaminejad MB, Nikmahzar A, Taghiyar L, Dehghan MM, Kazemi Hossein, Farokhi A. Osteogenic, chondrogenic and adipogenic potential of canine marrow-derived mesenchymal stem cells. Yakhteh Med J 2007; 1:31-38.
13. Prochop DJ. Marrow stromal cells as stem cells for nonhematopoietic tissue. Science 1997; 276:71-74.
14. Dominici M, Le Blanc K, Mueller I, Slaper-Cortenbach I, Marini F, Krause D,
et al
. Minimal criteria for defining multipotent mesenchymal stromal cells. The international societry for cellular therapy position statement. Cytotherapy 2006; 4:315-317.
15. Friedenstein AJ, Gorskaja JF, Kulagina NN. Fibroblast precursors in normal and irradiated mouse hematopoietic organs. Exp Hematol 1976; 4:267-274.
16. Friedenstein AJ, Piatetzky-Shapiro II, Petrakova KV. Osteogenesis in transplants of bone marrow cells. J Embryol Exp Morphol 1966; 6:381-390.
17. Noth U, Osyczka AM, Tuli R, Hickok NJ, Danielson KG, Tuan RS. Multilineage mesenchymal differentiation of human trabecular bone-derived cells. J Ortho Res 2002; 20:1060-1069.
18. Wickham MQ, Erickson GR, Gimble JM, Vail TP, Guilak F. Multipotent stromal cells derived from infrapatellar fat pad of knee. Clin Ortho 2003; 196-212.
19. Jankowsk RJ, Deasy BM, Huard J. Muscle-derived stem cells. Gene therapy 2002; 9:642-647.
20. Zvaifler NJ, Marinova-Mutafchieva L, Adams G, Edwards CJ, Moss J, Burger JA. Mesenchyamal precursor cells in the blood of normal individuals. Artheritis Res 2000; 2:477-488.
21. You Q, Tong X, Guan Y, Zhang D, Huang M, Zhang Y,
et al
. biological characteristics of human third trimester amniotic fluid stem cells. J Int Med Res 2009; 37:105-112.
22. Erices A, Conget P, Miguell JJ. Mesenchymal progenitor cells in human umbilical cord blood. Br J Haematol 2000; 109:235-242.
23. Eslaminejad MB, Taghiyar L. Mesenchymal stem cell purification from the articular cartilage cell culture. Iran J Basic Med Sci 2008; 3:146-153.
24. Huang YC, Yang ZM, Chen XH, Tan MY, Wang J, Li XQ, Xie HQ, Deng L. Isolation of mesenchymal stem cells from human placental decidua basalis and resistance to hypoxia and serum deprivation. Stem Cell Rev 2009 5:247-255.