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In vivo differentiation potential of buffalo (Bubalus bubalis) embryonic stem cell

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Abstract

Embryonic stem cells (ESCs) derived from inner cell mass (ICM) of mammalian blastocyst are having indefinite proliferation and differentiation capability for any type of cell lineages. In the present study, ICMs of in vitro-derived buffalo blastocysts were cultured into two different culture systems using buffalo fetal fibroblast as somatic cell support and Matrigel as synthetic support to obtain pluripotent buffalo embryonic stem cell (buESC) colonies. Pluripotency of the ESCs were characterised through pluripotency markers whereas, their differentiation capability was assessed by teratoma assay using immuno-compromised mice. Cumulus ooccyte complexes from slaughter house-derived ovaries were subjected to in vitro maturation, in vitro fertilization and in vitro culture to generate blastocysts. Total 262 blastocysts were derived through IVEP with 11.83 % (31/262) hatching rate. To generate buESCs, 15 ICMs from hatched blastocysts were cultured on mitomycin-C-treated homologous fetal fibroblast feeder layer, whereas the leftover 16 ICMs were cultured on extra-cellular matrix (Matrigel). No significant differences were observed for primary ESCs colony formation between two culture systems. Primary colonies as well as passaged ESCs were characterised by alkaline phosphatase staining, karyotyping and expression of transcription-based stem cell markers, OCT-4 and cell surface antigens SSEA-4 and TRA-1-60. Batch of ESCs found positive for pluripotency markers and showing normal karyotype after fifteenth passage were inoculated into eight immuno-compromised mice through subcutaneous and intramuscular route. Subcutaneous route of inoculation was found to be better than intramuscular route. Developed teratomas were excised surgically and subjected to histological analysis. Histological findings revealed presence of all the three germinal layer derivatives in teratoma sections. Presence of germinal layer derivatives were further confirmed by reverse transcriptase–polymerase chain reaction for the presence of differentiation markers like nerve cell adhesion molecule, fetal liver kinase-1 and alpha-feto protein for ectoderm, mesoderm and endoderm, respectively.

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References

  • Anand T.; Kumar D.; Singh M. K.; Shah R. A.; Chauhan M. S.; Manik R. S.; Singla S. K.; Palta P. Buffalo (Bubalus bubalis) embryonic stem cell-like cells and preimplantation embryos exhibit comparable expression of pluripotency-related antigens. Reprod. Dom. Anim. 46(1): 50–58; 2011.

    Article  CAS  Google Scholar 

  • Brivanlou A. H.; Gage F. H.; Jaenisch R.; Jessell T.; Melton D.; Rossant J. Setting standards for human embryonic stem cells. Science 300: 913–916; 2003.

    Article  PubMed  CAS  Google Scholar 

  • Carpenter M. K.; Rosler E.; Rao M. S: Characterization and differentiation of human embryonic stem cells. Cloning Stem Cells 5: 79–88; 2003.

    Google Scholar 

  • Dattena M.; Chessa B.; Lacerenza D.; Accardo C.; Pilichi S.; Mara L.; Chessa F.; Vincenti L.; Cappai P. Isolation, culture and characterization of embryonic cell lines from vitrified sheep blastocysts. Mol. Reprod. Dev. 73(1): 31–39; 2006.

    Article  PubMed  CAS  Google Scholar 

  • Evans M. J.; Kaufman M. Establishment in culture of pluripotent cells from mouse embryo. Nature 292: 154–156; 1981.

    Article  PubMed  CAS  Google Scholar 

  • Gertow K.; Wolban S.; Rozell B. Organized development from human embryonic stem cells after injection into immunodeficient mice. Stem Cells Dev. 13: 421–435; 2004.

    Article  PubMed  Google Scholar 

  • Ginis I.; Luo Y.; Miura T.; Thies S.; Brandenberger R.; Gerecht-Nir S.; Amit M.; Hoke A.; Carpenter M. K.; Itskovitz-Eldor J.; Rao M. S. Differences between human and mouse embryonic stem cells. Dev Bio. 269: 360–380; 2004.

    Article  CAS  Google Scholar 

  • Graves K. H.; Moreadith R. W. Derivation and characterization of putative pluripotential embryonic stem cells from preimplantation rabbit embryos. Mol. Reprod. Dev. 36: 424–433; 1993.

    Article  PubMed  CAS  Google Scholar 

  • Grunz H.; Tacke L. Neural differentiation of Xenopus laevis ectoderm takes place after disaggregation and delayed reaggregation without inducer. Cell Differ. Dev. 28: 211–218; 1989.

    Article  PubMed  CAS  Google Scholar 

  • Handyside A. H.; Hooper M. L.; Kaufman M. H.; Wilmut I. Towards the isolation of embryonal stem cell lines from the sheep. Roux’s Arch. Dev. Biol. 196: 185; 1987.

    Article  Google Scholar 

  • Hannes H.; Soong P. L.; Wang S.; Dunn N. R. Teratoma formation by human embryonic stem cell, evaluation of essential parameter for future safety studies. Stem Cell Res. 2: 198–210; 2009.

    Article  Google Scholar 

  • Hatoya S.; Torii R.; Kondo Y.; Okuno T.; Kobayashi K. Isolation and characterization of embryonic stem cell like cells from canine blastocysts. Mol. Reprod. Dev. 73: 298–305; 2006.

    Article  PubMed  CAS  Google Scholar 

  • Henderson J. K.; Draper J. S.; Baillie H. S.; Fishel S.; Thomson J. A.; Moore H.; Andrews P. W. Preimplantation human embryos and embryonic stem cells show comparable expression of stage-specific embryonic antigens. Stem Cells 20: 329–337; 2002.

    Google Scholar 

  • Hiata H.; Kawamata S.; Murakami Y.; Inoue K.; Nagahashi A.; Tosaka M.; Miyamoto Y.; Iwasaki H.; Asahara T.; Sawa Y. Coexpression of platelet-derived growth factor receptor alpha and fetal liver kinase-1 enhances cardiogenic potential in embryonic stem cell differentiation in vitro. J. Biosci. Bioeng. 103(5): 412–419; 2007.

    Article  PubMed  CAS  Google Scholar 

  • Hoffman L. M.; Carpenter M. K. Characterization and culture of human embryonic stem cells. Nat. Biotechnol. 23: 699–708; 2005.

    Google Scholar 

  • Huang B.; Li T.; Wang X. L.; Xie T. S.; Lu Y. Q.; da Silva F. M.; Shi D. S. Generation and characterization of embryonic stem-like cell lines derived from in vitro fertilization buffalo (Bubalus bubalis) embryos. Reprod. Domest. Anim. 45: 1220–128; 2010.

    Google Scholar 

  • Huang B.; Xie T.; Shi S.; Li D. S.; Wang Z. Q.; Li M. M. Isolation and characterization of EG-like cells from Chinese swamp buffalo (Bubalus bubalis). Cell Biol. Int. 31(10): 1079–1088; 2007.

    Article  PubMed  CAS  Google Scholar 

  • Kania G.; Corbeil D.; Fuchs J.; Tarasov K. V.; Blyszczuk P.; Huttner W. B.; Boheler K. R.; Wobus A. M. Somatic stem cell marker prominin-1/CD133 is expressed in embryonic stem cell-derived progenitors. Stem Cells 23: 791–804; 2005.

    Article  PubMed  CAS  Google Scholar 

  • Kitiyanant Y.; Saikhun J.; Kitiyanant N.; Sritanaudomchai H.; Faisaikarm T.; Pavasuthipaisit K. Establishment of buffalo embryonic stem-like cell lines from different sources of derived blastocysts. Proceedings of the annual conference of the international embryo transfer society. Portland, Oregon, USA. Reprod. Fertility Dev. 16: 216; 2004.

    Article  Google Scholar 

  • Li X.; Allen W. R. Isolation and proliferation of inner cell mass of horse blastocyst. Reprod. Ab. Ser. 30: 54; 2003.

    Google Scholar 

  • Niwa H. How is pluripotency determined and maintained? Development. 134: 635–646; 2007.

    Article  PubMed  CAS  Google Scholar 

  • Notarianni E.; Galli C.; Moor R. M.; Evans M. J. Derivation of pluripotent, embryonic cell lines from the pig and sheep blastocysts. Reprod. Fertil. Suppl. 43: 255–260; 1991.

    CAS  Google Scholar 

  • Pesce M.; Gross M. K.; Scholer H. R. In line with our ancestors: Oct-4 and the mammalian germ. Bio essays. 20: 722–732; 1998.

    CAS  Google Scholar 

  • Piedrahita J. A.; Anderson G. B.; Bondurant R. H. The isolation of embryonic stem cells: comparative behavior of murine, porcine and ovine embryos. Theriogenology 34: 23–79; 1990.

    Google Scholar 

  • Prokhorova T. A.; Harkness L. M.; Frandsen U.; Ditzel N.; Burns J. S.; Schroeder H. D.; Kassem M. Teratoma formation by human embryonic stem cells is site dependent and enhanced by the presence of Matrigel. Stem Cells Dev. 17: 255–267; 2008.

    Article  Google Scholar 

  • Resnick J. L.; Bixler L. S.; Cheng L.; Donovan P. J. Long term proliferation of mouse primordial germ cells in culture. Nature 359: 550–551; 1992.

    Article  PubMed  CAS  Google Scholar 

  • Richards M.; Tan S.; Fong C. Y.; Biswas A.; Chan W. K.; Bongso A. Comparative evaluation of various human feeders for prolonged undifferentiated growth of human embryonic stem cells. Stem Cells 21: 546–556; 2003.

    Article  PubMed  CAS  Google Scholar 

  • Saito S.; Ugai H.; Sawai K.; Yamamoto Y.; Minamihashi A.; Kurosaka K.; Kobayashi Y.; Murata T.; Obata Y.; Yokoyama K. Isolation of embryonic stem like cells from equine blastocysts and their differentiation in vitro. FEBS Lett. 531: 389–396; 2002.

    Article  PubMed  CAS  Google Scholar 

  • Sharma M.; Kumar R.; Dubey P. K.; Verma O. P.; Nath A.; G. Saikumar.; G. Taru Sharma. Expression and quantification of Oct-4 gene in blastocyst and embryonic stem cells derived from in vitro produced buffalo embryos. In Vitro Cellular & Developmental Biology-Animal; doi:10.1007/s11626-012-9491-2; 2012.

  • Solter D.; Gearhart J. Puffing stem cells to work. Science 283: 1468–1470; 1999.

    Article  PubMed  CAS  Google Scholar 

  • Solter D.; Knowles B. B. Monoclonal antibody defining a stage-specific mouse embryonic antigen (SSEA-1). Proc. Nat. Acad. Sci. 75: 5565–5569; 1978.

    Article  PubMed  CAS  Google Scholar 

  • Sritanaudomchai H.; Pavasuthipaisit K.; Kitiyanant Y.; Kupradinun P.; Mitalipov S.; Kusamran T. Characterization and multilineage differentiation of embryonic stem cells derived from a buffalo parthenogenetic embryo. Mol. Reprod. Dev. 74(10): 1295; 2007.

    Article  PubMed  CAS  Google Scholar 

  • Strelchenko N.; Stice S. Bovine embryonic pluripotent cell lines derived from morula stage embryos. Theriogenology 41: 304; 1994.

    Article  Google Scholar 

  • Strojek R. M.; Reed M. A.; Hoover J. L.; Wagner T. E. A method for cultivating morphologically undifferentiated embryonic stem cells from porcine blastocysts. Theriogenology 33: 901; 1990.

    Article  PubMed  CAS  Google Scholar 

  • Talbot N. C.; Rexroad C. E.; Pursel V. G.; Powell A. M. Alkaline phosphatase staining of pig and sheep epiblast cells in culture. Mol. Reprod. Dev. 36: 139–147; 1993.

    Article  PubMed  CAS  Google Scholar 

  • Thomson J. A.; Itskovitz-Eldor J.; Shapiro S. S.; Waknitz M. A.; Swiergiel J. J.; Marshall V. S.; Jones J. M. Embryonic stem cell lines derived from human blastocysts. Science 282: 1145–1147; 1998.

    Article  PubMed  CAS  Google Scholar 

  • Verma V.; Gautam S. K.; Singh B.; Manik R. S.; Palta P.; Singla S. K.; Goswami S. L.; Chauhan M. S. Isolation and characterization of embryonic stem cell-like cells from in vitro-produced buffalo (Bubalus bubalis) embryos. Mol. Reprod. Dev. 74: 520–529; 2007.

    Google Scholar 

  • Wang L.; Duan E.; Sung L. Y.; Jeong B. S.; Yang X.; Tian X. C. Generation and characterization of pluripotent stem cells from cloned bovine embryos. Biol. Reprod. 73: 149–155; 2005.

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

The study was supported by research grant from the Department of Biotechnology (DBT), Ministry of Science and Technology, Government of India. Authors thank Director, IVRI, Izatnagar, (Bareilly, U.P., India) for providing necessary facilities to support this work.

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Authors and Affiliations

  1. Division of Physiology and Climatology, Indian Veterinary Research Institute, Izatnagar-243 122, Bareilly, UP, India

    Om Prakash Verma, Amar Nath, Manjinder Sharma, Pawan Kumar Dubey & G. Taru Sharma

  2. Division of Animal Genetics and Breeding, Indian Veterinary Research Institute, Izatnagar-243 122, Bareilly, UP, India

    Rajesh Kumar

  3. Division of Pathology, Indian Veterinary Research Institute, Izatnagar-243 122, Bareilly, UP, India

    G. Sai Kumar

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  1. Om Prakash Verma
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  2. Rajesh Kumar
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  3. Amar Nath
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  4. Manjinder Sharma
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  5. Pawan Kumar Dubey
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Correspondence to Om Prakash Verma.

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Editor: T. Okamoto

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Verma, O.P., Kumar, R., Nath, A. et al. In vivo differentiation potential of buffalo (Bubalus bubalis) embryonic stem cell. In Vitro Cell.Dev.Biol.-Animal 48, 349–358 (2012). https://doi.org/10.1007/s11626-012-9515-y

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  • DOI: https://doi.org/10.1007/s11626-012-9515-y

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