Differentiation of neuron-like cells from mouse parthenogenetic embryonic stem cells

doi:10.3969/j.issn.1673-5374.2013.04.001

Abstract

Abstract:

Parthenogenetic embryonic stem cells have pluripotent differentiation potentials, akin to fertilized embryo-derived embryonic stem cells. The aim of this study was to compare the neuronal differentiation potential of parthenogenetic and fertilized embryo-derived embryonic stem cells. Before differentiation, karyotype analysis was performed, with normal karyotypes detected in both parthenogenetic and fertilized embryo-derived embryonic stem cells. Sex chromosomes were identified as XX. Immunocytochemistry and quantitative real-time PCR detected high expression of the pluripotent gene, Oct4, at both the mRNA and protein levels, indicating pluripotent differentiation potential of the two embryonic stem cell subtypes. Embryonic stem cells were induced with retinoic acid to form embryoid bodies, and then dispersed into single cells. Single cells were differentiated in N2 differentiation medium for 9 days. Immunocytochemistry showed parthenogenetic and fertilized embryo-derived embryonic stem cells both express the neuronal cell markers nestin, βIII-tubulin and myelin basic protein. Quantitative real-time PCR found expression of neurogenesis related genes (Sox-1, Nestin, GABA, Pax6, Zic5 and Pitx1) in both types of embryonic stem cells, and Oct4 expression was significantly decreased. Nestin and Pax6 expression in parthenogenetic embryonic stem cells was significantly higher than that in fertilized embryo-derived embryonic stem cells. Thus, our experimental findings indicate that parthenogenetic embryonic stem cells have stronger neuronal differentiation potential than fertilized embryo-derived embryonic stem cells.

Key words: neural regeneration, stem cells, parthenogenesis, parthenogenetic embryonic stem cells, embryonic stem cells, neuronal cells, karyotypes, Oct4, differentiation, embryoid body, mice, grants-supported paper, photographs-containing paper, neuroregeneration

Xingrong Yan, Yanhong Yang, Wei Liu, Wenxin Geng, Huichong Du, Jihong Cui, Xin Xie, Jinlian Hua, Shumin Yu, Liwen Li, Fulin Chen. Differentiation of neuron-like cells from mouse parthenogenetic embryonic stem cells[J]. Neural Regeneration Research, 2013, 8(4): 293-300.

References

[1] Alvarez CV, Garcia-Lavandeira M, Garcia-Rendueles ME, et al. Defining stem cell types: understanding the therapeutic potential of ESCs, ASCs, and iPS cells. J Mol Endocrinol. 2012;49(2):R89-111.

[2] Sills ES, Takeuchi T, Tanaka N, et al. Identification and isolation of embryonic stem cells in reproductive endocrinology: theoretical protocols for conservation of human embryos derived from in vitro fertilization. Theor Biol Med Model. 2005;2(25):1-8.

[3] Lin G, OuYang Q, Zhou X, et al. A highly homozygous and parthenogenetic human embryonic stem cell line derived from a one-pronuclear oocyte following in vitro fertilization procedure. Cell Res. 2007;17(12):999-1007.

[4] Okamoto S, Takahashi M. Induction of Retinal Pigment Epithelial Cells from Monkey Invest. Invest Ophthalmol Vis Sci. 2011;52(12):8785-8790.

[5] Jiang J, Ding G, Lin J, et al. Different developmental potential of pluripotent stem cells generated by different reprogramming strategies. J Mol Cell Biol. 2011;3(3): 197-199.

[6] Yang X, Smith SL, Tian XC, et al. Nuclear reprogramming of cloned embryos and its implications for therapeutic cloning. Nat Genet. 2007;39(3):295-302.

[7] Rideout WM 3rd, Hochedlinger K, Kyba M, et al. Correction of a genetic defect by nuclear transplantation and combined cell and gene therapy. Cell. 2002;109(1): 17-27.

[8] Robertson JA. Embryo stem cell research: ten years of controversy. J Law Med Ethics. 2010;38(2):191-203.

[9] Takahashi K, Yamanaka S. Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell. 2006;126(4):663-676.

[10] Zou J, Mali P, Huang X, et al. Site-specific gene correction of a point mutation in human iPS cells derived from an adult patient with sickle cell disease. Blood. 2011;118(17): 4599-4608.

[11] Alipio Z, Liao W, Roemer EJ, et al. Reversal of hyperglycemia in diabetic mouse models using induced-pluripotent stem (iPS)-derived pancreatic β-like cells. Proc Natl Acad Sci U S A. 2010;107(30): 13426-13431.

[12] Mou CL, Chen LY. Battle for pluripotency: Derivation of induced pluripotent stem cells. Recent Pat Regen Med. 2011;1:123-130.

[13] Zeitouni S, Krause U, Clough BH, et al. Human mesenchymal stem cell-derived matrices for enhanced osteoregeneration. Sci Transl Med. 2012;4(132): 132-155.

[14] Van Keymeulen A, Blanpain C. Tracing epithelial stem cells during development, homeostasis, and repair. J Cell Biol. 2012;197(5):575-584.

[15] Rode I, Boehm T. Regenerative capacity of adult cortical thymic epithelial cells. Proc Natl Acad Sci U S A. 2012; 109(9):3463-3468.

[16] Kaufman MH, Robertson EJ, Handyside AH, et al. Establishment of pluripotential cell lines from haploid mouse embryos. J Embryol Exp Morphol. 1983;73: 249-261.

[17] Kim K, Lerou P, Yabuuchi A, et al. Histocompatible embryonic stem cells by parthenogenesis. Science. 2007;315(5811):482-486.

[18] Revazova ES, Turovets NA, Kochetkova OD, et al. Patient-specific stem cell lines derived from human parthenogenetic blastocysts. Cloning Stem Cells. 2007; 9:432-449.

[19] Bibel M, Richter J, Schrenk K, et al. Differentiation of mouse embryonic stem cells into a defined neuronal lineage. Nat Neurosci. 2004;7(9):1003-1009.

[20] Stavridis MP, Smith AG. Neural differentiation of mouse embryonic stem cells. Biochem Soc Trans. 2003;31(Pt 1): 45-49.

[21] Yu SM, Yan XR, Chen DM, et al. Isolation and characterization of parthenogenetic embryonic stem (pES) cells containing genetic background of the Kunming mouse strain. Asian-Aust J Anim Sci. 2011;24(1):37-44.

[22] Yan X, Yu S, Lei A, et al. The four reprogramming factors and embryonic development in mice. Cell Reprogram. 2010;12(5):565-570.

[23] Gong SP, Kim H, Lee EJ, et al. Change in gene expression of mouse embryonic stem cells derived from parthenogenetic activation. Hum Reprod. 2009;24(4): 805-814.

[24] Jäderstad J, Jäderstad LM, Li J, et al. Communication via gap junctions underlies early functional and beneficial interactions between grafted neural stem cells and the host. Proc Natl Acad Sci U S A. 2010;107(11):5184-5189.

[25] Cao Q, He Q, Wang Y, et al. Transplantation of ciliary neurotrophic factor-expressing adult oligodendrocyte precursor cells promotes remyelination and functional recovery after spinal cord injury. J Neurosci. 2010;30(8): 2989-3001.

[26] Zhou J, Tian GP, Wang J, et al. In vitro differentiation of adipose-derived stem cells and bone marrow-derived stromal stem cells into neuronal-like cells. Neural Regen Res. 2011;6(19):1467-1472.

[27] Pevny LH, Sockanathan S, Placzek M, et al. A role for SOX1 in neural determination. Development. 1998;125: 1967-1978.

[28] Park D, Xiang AP, Mao FF, et al. Nestin is required for the proper self-renewal of neural stem cells. Stem Cells. 2101;28(12):2162-2171.

[29] Jia H, Tao H, Feng R, et al. Pax6 regulates the epidermal growth factor-responsive neural stem cells of the subventricular zone. Neuroreport. 2011;22(9):448-452.

[30] Corti S, Nizzardo M, Nardini M, et al. Embryonic stem cell-derived neural stem cells improve spinal muscular atrophy phenotype in mice. Brain. 2010;133(Pt 2): 465-481.

[31] Elkouris M, Balaskas N, Poulou M, et al. Sox1 maintains the undifferentiated state of cortical neural progenitor cells via the suppression of Prox1-mediated cell cycle exit and neurogenesis. Stem Cells. 2011;29(1):89-98.

[32] Jin Z, Liu L, Bian W, et al. Different transcription factors regulate nestin gene expression during P19 cell neural differentiation and central nervous system development. J Biol Chem. 2009;284(12):8160-8173.

[33] Suzuki S, Namiki J, Shibata S, et al. The neural stem/progenitor cell marker nestin is expressed in proliferative endothelial cells, but not in mature vasculature. J Histochem Cytochem. 2010;58(8):721-730.

[34] Zhang X, Huang CT, Chen J, et al. Pax6 is a human neuroectoderm cell fate determinant. Cell Stem Cell. 2010;7(1):90-100.

[35] Gao J, Wang J, Wang Y, et al. Regulation of Pax6 by CTCF during induction of mouse ES cell differentiation. PLoS One. 2011;6(6):e20954.

[36] Sansom SN, Griffiths DS, Faedo A, et al. The level of the transcription factor pax6 is essential for controlling the balance between neural stem cell self-renewal and neurogenesis. PLoS Genet. 2009;5(6):e1000511.

[37] Tan KR, Yvon C, Turiault M, et al. GABA neurons of the VTA drive conditioned place aversion. Neuron. 2012; 73(6):1173-1183.

[38] Inoue T, Hatayama M, Tohmonda T, et al. Mouse Zic5 deficiency results in neural tube defects and hypoplasia of cephalic neural crest derivatives. Dev Biol. 2004;270(1): 146-162.

[39] National Research Council (US) Committee for the Update of the Guide for the Care and Use of Laboratory Animals. Guide for the Care and Use of Laboratory Animals. 8th edition. Washington (DC): National Academies Press (US). 2011.

[40] Luo C, Zuñiga J, Edison E, et al. Superovulation strategies for 6 commonly used mouse strains. J Am Assoc Lab Anim Sci. 2011;50(4):471-478.

[41] González S, Ibáñez E, Santaló J. Establishment of mouse embryonic stem cells from isolated blastomeres and whole embryos using three derivation methods. J Assist Reprod Genet. 2010;27(12):671-682.

[42] Nagy A, Gertsenstein M, Vintersten K, et al. Karyotyping Mouse Cells. CSH Protoc. 2008;3(5):1-3.

[43] Cook MS, Munger SC, Nadeau JH, et al. Regulation of male germ cell cycle arrest and differentiation by DND1 is modulated by genetic background. Development. 2011; 138(1):23-32.