Amplification of rabbit adipose-derived stem cells using explants culture method

doi:10.3969/j.issn.2095-4344.2014.01.015

Abstract

Abstract:

BACKGROUND: The rabbit adipose-derived stem cells are mostly isolated by type I collagenase digestion, but rarely by explants culture method.
OBJECTIVE: To isolate rabbit adipose-derived stem cells for adipogenic and osteogenic differentiation.
METHODS: The rabbit adipose-derived stem cells were isolated from rabbit adipose by explants culture method, and cultured in vitro followed by morphological observation. The grow curve and cell surface markers CD29, CD44, CD45 of passage 3 cells were analyzed respectively by 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide and ?ow cytometry; cells from the third passages were induced for adipogenic and osteogenic differentiation by different revulsants, and cells were examined by oil red O staining and alizarin red staining .
RESULTS AND CONCLUSION: The rabbit adipose-derived stem cells cultured in vitro exhibited a spindle-shaped appearance and could rapidly expand. Flow cytometry analysis revealed that the third passage of rabbit adipose-derived stem cells was positive for CD29, CD44, but negative for CD45. Rabbit adipose-derived stem cells were positive for oil red O staining at 14 days of adipogenic induction, and positive for alizarin red staining at 14 days of osteogenic induction. In conclusion, we could successfully isolate rabbit adipose-derived stem cells using explants culture method.

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

全文链接：

Key words: stem cells, phenotype, adipogenesis, adipose tissue, flow cytometry

CLC Number:

R94.2

Liu Qin, Wang Li-ping, Yu Jing, Chen Fang, Diao Bo, Zhang Yi . Amplification of rabbit adipose-derived stem cells using explants culture method[J]. Chinese Journal of Tissue Engineering Research, 2014, 18(1): 88-93.

Figures/Tables 3

References

[1] de Girolamo L, Arrigoni E, Stanco D, et al. Role of autologous rabbit adipose-derived stem cells in the early phases of the repairing process of critical bone defects. J Orthop Res. 2011; 29(1):100-108.

[2] Arrigoni E, de Girolamo L, Di Giancamillo A, et al. Adipose-derived stem cells and rabbit bone regeneration: histomorphometric, immunohistochemical and mechanical characterization. J Orthop Sci. 2013;18(2):331-339.

[3] Li H, Xu Y, Fu Q, et al. Effects of multiple agents on epithelial differentiation of rabbit adipose-derived stem cells in 3D culture. Tissue Eng Part A. 2012;18(17-18):1760-1770.

[4] Arrigoni E, de Girolamo L, Di Giancamillo A, et al. Adipose-derived stem cells and rabbit bone regeneration: histomorphometric, immunohistochemical and mechanical characterization. J Orthop Sci. 2013;18(2):331-339.

[5] Baptista LS, do Amaral RJ, Carias RB, et al. An alternative method for the isolation of mesenchymal stromal cells derived from lipoaspirate samples. Cytotherapy. 2009;11(6):706-715.

[6] 彭智,陈崎,贾振华,等.组织块培养法扩增人脂肪源性干细胞的生物学特征鉴定[J].中国组织工程研究, 2010, 14(36):6689-6694.

[7] Sunay O, Can G, Cakir Z, et al. Autologous rabbit adipose tissue-derived mesenchymal stromal cells for the treatment of bone injuries with distraction osteogenesis. Cytotherapy. 2013;15(6):690-702.

[8] Sgodda M, Aurich H, Kleist S, et al. Hepatocyte differentiation of mesenchymal stem cells from rat peritoneal adipose tissue in vitro and in vivo. Exp Cell Res. 2007;313(13):2875-2886.

[9] Toghraie F, Razmkhah M, Gholipour MA, et al. Scaffold-free adipose-derived stem cells (ASCs) improve experimentally induced osteoarthritis in rabbits. Arch Iran Med. 2012; 15(8): 495-499.

[10] 庞荣清,何洁,李福兵,等.一种简单的人脐带间充质干细胞分离培养方法[J].中华细胞与干细胞杂志:电子版,2011,1(2):162-167.

[11] Yang X, Qu L, Wang X, et al. Plasticity of epidermal adult stem cells derived from adult goat ear skin. Mol Reprod Dev. 2007;74(3):386-396.

[12] Spath L, Rotilio V, Alessandrini M, et al. Explant-derived human dental pulp stem cells enhance differentiation and proliferation potentials. J Cell Mol Med. 2010;14(6B):1635- 1644.

[13] Priya N, Sarcar S, Majumdar AS, et al. Explant culture: a simple, reproducible, efficient and economic technique for isolation of mesenchymal stromal cells from human adipose tissue and lipoaspirate. J Tissue Eng Regen Med. 2012; 1-9.

[14] Ishige I, Nagamura-Inoue T, Honda MJ, et al. Comparison of mesenchymal stem cells derived from arterial, venous, and Wharton's jelly explants of human umbilical cord. Int J Hematol. 2009;90(2):261-269.

[15] Miranville A, Heeschen C, Sengenès C, et al. Improvement of postnatal neovascularization by human adipose tissue-derived stem cells. Circulation. 2004;110(3):349-355.

[16] Yoshimura K, Shigeura T, Matsumoto D, et al. Characterization of freshly isolated and cultured cells derived from the fatty and luid portions of liposuction aspirates. J Cell Physiol. 2006;208(1): 64-76.

[17] Brzoska M, Geiger H, Gauer S, et al. Epithelial differentiation of human adipose tissue-derived adult stem cells. Biochem Biophys Res Commun. 2005;330(1):142-150.

[18] Sotiropoulou PA, Perez SA, Salagianni M, et al. Characterization of the optimal culture conditions for clinical scale production of human mesenchymal stem cells. Stem Cells. 2006;24(2):462-471.

[19] Ou Y, Yuan XD, Cai YN, et al. A novel ethanol-based method to induce differentiation of adipose-derived stromal cel ls into astrocytes. Neural Regen Res. 2011;6(10): 738-743.

[20] Baer PC, Griesche N, Luttmann W, et al. Human adipose-derived mesenchymal stem cells in vitro: evaluation of an optimal expansion medium preserving stemness. Cytotherapy. 2010;12(1):96-106.

[21] Campagnoli C, Roberts IA, Kumar S, et al. Identification of mesenchymal stem/progenitor cells in human firsttrim ester fetal blood, liver and bone marrow. Blood. 2001;98(8): 2396-2402.

[22] Cai YN, Yuan XD, Ou Y, et al. Apoptosis during β-mercaptoethanol-induced differentiation of adult adipose-derived stromal cel ls into neurons. Neural Regen Res. 2011;6(10):750-755.

[23] Gronthos S, Franklin DM, Leddy HA, et al. Surface protein characterization of human adipose tissue-derived stromal cells. J Cell Physiol. 2001;189(1):54-63.

[24] Gu F, Gao JH, Lu F. Culture of the single cell clones of human adipose-derived stem cells and identification of their surface antigen expressions. Nan Fang Yi Ke Da Xue Xue Bao. 2008; 28(6):1067-1069.

[25] Zuk PA, Zhu M, Ashjian P, et al. Human adipose tissue is a source of multipotent stem cells. Mol Biol Cell. 2002;13(12): 4279-4295.

[26] Dominici M, Le Blanc K, Mueller I, et al. Minimal criteria for de?ning multipotent mesenchymal stromal cells. The international Society for Cellular Therapy position statement. Cytotherapy. 2006;8(4):315-317.

[27] Varma MJ, Breuls RG, Schouten TE, et al. Phenotypical and functional characterization of freshly isolated adipose tissue-derived stem cells. Stem Cells Dev. 2007; 16(1): 91-104.

[28] Lindroos B, Suuronen R, Miettinen S. The potential of adipose stem cells in regenerative medicine. Stem Cell Rev. 2011;7(2):269-291.

[29] Baer PC, Geiger H. Adipose-derived mesenchymal stromal/stem cells: tissue localization, characterization, and heterogeneity. Stem Cells Int. 2012;2012:812693.

[30] Xiao J, Yang X, Jing W, et al. Adipogenic and osteogenic differentiation of Lin(-)CD271(+)Sca-1(+) adipose-derived stem cells. Mol Cell Biochem, 2013. [Epub ahead of print].

[31] Zuk P. Adipose-Derived Stem Cells in Tissue Regeneration: A Review. ISRN Stem Cells. 2013;2013:1-35.

[32] Yuan XD, Cai YN, Ou Y, et al. Adult adipose-derived stromal cells differentiate into neurons with normal electrophysiological functions. Neural Regen Res. 2011;6(34): 2681-2686.

[33] 王清富,陈庄洪,蔡贤华,等.兔脂肪干细胞的多向诱导分化[J].中国组织工程研究, 2012, 16(49):9162-9167.

[34] Cawthorn WP, Scheller EL, MacDougald OA. Adipose tissue stem cells: the great WAT hope.Trends Endocrinol Metab. 2012;23(6):270-277.

[35] Macotela Y, Emanuelli B, Mori MA, et al. Intrinsic differences in adipocyte precursor cells from different white fat depots. Diabetes. 2012;61(7):1691-1699.

[36] Suga H, Matsumoto D, Eto H, et al. Functional implications of CD34 expression in human adipose-derived stem/progenitor cells. Stem Cells Dev. 2009;18(8):1201-1209.

[37] Rowlands AS, George PA, Cooper-White JJ. Directing osteogenic and myogenic differentiation of MSCs: interplay of stiffness and adhesive ligand presentation.Am J Physiol Cell Physiol. 2008;295(4):C1037-1044.

[38] Buehrer BM, Cheatham B. Isolation and characterization of human adipose-derived stem cells for use in tissue engineering. Methods Mol Biol. 2013;1001:1-11.