Hostname: page-component-77c89778f8-7drxs Total loading time: 0 Render date: 2024-07-17T13:58:20.075Z Has data issue: false hasContentIssue false

Separation and in vitro culture of goat fetal testicular cells and behaviour of male germ-line stem cells (mGSCs)

Published online by Cambridge University Press:  13 June 2008

Dong Wu-Zi
Affiliation:
Shaanxi Province Branch Center of National Stem Cell Engineering and Technology Center, Northwest Sci-tech University of Agriculture and Forestry, Yangling 712100, China
Hua Jin-Lian
Affiliation:
Shaanxi Province Branch Center of National Stem Cell Engineering and Technology Center, Northwest Sci-tech University of Agriculture and Forestry, Yangling 712100, China
Zhuang Shu-Zhen
Affiliation:
Shaanxi Province Branch Center of National Stem Cell Engineering and Technology Center, Northwest Sci-tech University of Agriculture and Forestry, Yangling 712100, China
Shen Wen-Zheng
Affiliation:
Shaanxi Province Branch Center of National Stem Cell Engineering and Technology Center, Northwest Sci-tech University of Agriculture and Forestry, Yangling 712100, China Yangling Vocational and Technical College, Yangling 712100, China
Dou Zhong-Ying*
Affiliation:
Shaanxi Province Branch Center of National Stem Cell Engineering and Technology Center, Northwest Sci-tech University of Agriculture and Forestry, Yangling 712100, China
*
*Corresponding author. Email: douzhongying@china.com

Abstract

Seminiferous tubule tissues from goat fetal testis were treated using four methods. One of the treatments, with 0.1% collagenase for 15 min and then with an enzyme mixture (containing 0.25% trypsin, 10 μg/ml DNase I, 0.04% EDTA) for 5 min showed ideal cell dispersal after centrifugation and washing. The in vitro culture of the cells separated with a different medium system showed that mulberry-shaped male germ-line stem cell (mGSC) clusters and a single layer of Sertoli cells appeared as the original generation in 120 h. The mGSC clusters developed in suspension and were distributed in different locations from the Sertoli cells in the plate. The cells in the marginal area of incompletely digested mGSCs clusters, and the single cells from them, were obviously homogenized after culture for 5 days in a fibre cell feeding layer from fetal mice (MEF feeding layer), whereas identically treated mGSCs clusters co-cultured with Sertoli cells in the same culture system did not displayed clear homogenization in marginal cells of the clusters. Also, the first generation of mGSC clusters co-cultured with Sertoli cells could form the same morphological properties with the original generation; cells in clusters were tight and mGSCs divided more slowly than those in the MEF feeding layer.

Type
Research Article
Copyright
Copyright © China Agricultural University and Cambridge University Press 2005

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

Allard, K, Hall, SJ and Boekelheide, K (1995) Stem cell kinetics in rat testis after irreversible injury induced by 2,5-hexanedione. Biology of Reproduction 53: 186192.CrossRefGoogle ScholarPubMed
Brook, PF, Radford, JA, Shalet, SM et al. , (2001) Isolation of germ cells from human testicular tissue for low temperature storage and autotransplantation. Fertility Sterility 75: 269274.CrossRefGoogle ScholarPubMed
Dirami, G, Ravindranath, N, Pursel, V et al. , (1999) Effects of stem cell factor and granulocyte macrophage-colony stimulating factor on survival of porcine type A spermatogonia cultured in KSOM. Biology of Reproduction 61: 225230.CrossRefGoogle ScholarPubMed
Hua, JL, Dou, ZY, Zhang, RP et al. , (2002) Isolation and culture of human embryonic stem like cells derived from primordial germ cells. Journal of Agricultural Biotechnology 10: 6466 (in Chinese with English abstract).Google Scholar
Izadyar, F, Creemers, LB, van Dissel-Emiliani, FMF et al. , (2000) Spermatogonial stem cell transplantation. Molecular and Cellular Endocrinology 169: 2126.CrossRefGoogle ScholarPubMed
Kubota, H, Avarbock, MR and Brinster, RL (2003) Spermatogonial stem cells share some, but not all, phenotypic and functional characteristics with other stem cells. Proceedings of the National Academy of Sciences USA 100 (11): 64876492.CrossRefGoogle Scholar
Lui, WY, Lee, MW and Cheng, CY (2003) The sertoli–germ cell adherens junction (AJ) dynamics in the testis are regulated by RhoB GTPase via the ROCK/LIMK signaling pathway. Biology of Reproduction 68: 21892206.CrossRefGoogle ScholarPubMed
Nagano, M, Shinohara, T, Avarbock, MR et al. , (2000) Retrovirus-mediated gene delivery into male germ line stem cells. FEBS Letters 475: 710.CrossRefGoogle ScholarPubMed
Nagano, M, McCarrey, JR and Brinster, RL (2001) Primate spermatogonial stem cells colonize mouse testes. Biology of Reproduction 64: 14091416.CrossRefGoogle ScholarPubMed
Ohta, H, Yomogida, K, Dohmae, K et al. , (2000) Regulation of proliferation and differentiation in spermatogonial stem cells: the role of c-kit and its ligand SCF. Development 127: 21252131.CrossRefGoogle ScholarPubMed
Orwig, KE, Ryu, BY, Avarbock, MR et al. , (2002a) Male germ-line stem cell potential is predicted by morphology of cells in neonatal rat testes. Proceedings of the National Academy of Sciences USA 99: 1170611711.CrossRefGoogle ScholarPubMed
Orwig, KE, Shinohara, T, Avarbock, MR et al. , (2002b) Functional analysis of stem cells in the adult rat testis. Biology of Reproduction 66: 944949.CrossRefGoogle ScholarPubMed
Shinohara, T, Avarbock, MR and Brinster, R (1999) β1- and α6-integrin are surface markers on mouse spermatogonial stem cells. Proceedings of the National Academy of Sciences USA 96: 55045509.CrossRefGoogle ScholarPubMed
Shinohara, T, Orwig, KE, Avarbock, MR et al. , (2000) Spermatogonial stem cell enrichment by multiparameter selection of mouse testis cells. Proceedings of the National Academy of Sciences USA 97 (15): 83468351.CrossRefGoogle ScholarPubMed
Xie, T and Spradling, AC (2000) A niche maintaining germ line stem cells in the Drosophila ovary. Science 290: 328330.CrossRefGoogle ScholarPubMed
Zhang, XM, Wen, XH, Lai, LX et al. , (2000a) Developmental process of germ cells in prenuberal mouse. Chinese Veterinary Science 20 (3): 273279 (in Chinese with English abstract).Google Scholar
Zhang, XM, Lai, LX, Li, DX et al. , (2000b) Separation and purification of spermatogonia in mouse. Chinese Veterinary Science 1 (3): 235239 (in Chinese with English abstract).Google Scholar