Male germ-line stem cell potential is predicted by morphology of cells in neonatal rat testes

Department of Animal Biology, School of Veterinary Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA.
Proceedings of the National Academy of Sciences (Impact Factor: 9.67). 10/2002; 99(18):11706-11. DOI: 10.1073/pnas.182412099
Source: PubMed


Gonocytes are a transient population of male germ-line stem cells that are derived from primordial germ cells in the embryo and give rise to spermatogonial stem cells, which establish and maintain spermatogenesis in the postnatal testis. In contrast to spermatogonial stem cells, gonocytes can be identified easily in neonatal rat testis cell suspensions based on their large size and distinct morphology. Furthermore, histological analysis of testes from neonatal transgenic rats demonstrated that gonocytes are the only cells that express a lacZ reporter transgene. Two gonocyte subpopulations, designated pseudopod and round, were identified and isolated from neonatal (0-4 days postpartum) rat testis cell suspensions. Male germ-line stem cells, identified by their ability to produce and maintain colonies of spermatogenesis upon transplantation into infertile recipient testes, were present almost exclusively in the pseudopod gonocyte subpopulation. In contrast, annexin V staining indicated that the majority of round gonocytes undergo apoptosis. These results indicate that a nearly pure population of male germ-line stem cells can be prospectively identified in neonatal rat testis cell suspensions by morphological criteria. Together, the pseudopod and round gonocyte populations will provide powerful tools for the study of cellular mechanisms that control cell fates and the establishment of spermatogenesis in the postnatal testis.

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    • "Gonocytes, preceding the formation of spermatogonia, represent the transient germ cell population with stem cell characteristics in neonatal testes. Like SSCs, gonocytes are able to colonize and produce donorderived spermatogenesis after transplantation into infertile recipient testes (Jiang & Short 1998, Orwig et al. 2002). In pigs and goats, gonocytes transform to spermatogonia at about 2 months after birth (Byskov & Hoyer 1994), while in sheep and cattle, this transition commences at w3 and 4 months after birth respectively (Curtis & Amann 1981, Pedrana et al. 2008, Borjigin et al. 2010, Sarma & Devi 2012). "
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    ABSTRACT: Spermatogenesis, an elaborate and male-specific process in adult testes by which a number of spermatozoa are produced constantly for male fertility, relies on spermatogonial stem cells (SSCs). As a sub-population of undifferentiated spermatogonia, SSCs are capable of both self-renewal (to maintain sufficient quantities) and differentiation into mature spermatozoa. SSCs are able to convert to pluripotent stem cells during in vitro culture, thus they could function as substitutes for human embryonic stem (ES) cells without ethical issues. In addition, this process does not require exogenous transcription factors necessary to produce induced-pluripotent stem (iPS) cells from somatic cells. Moreover, combining genetic engineering with germ cell transplantation would greatly facilitate the generation of transgenic animals. Since germ cell transplantation into infertile recipient testes was first established in 1994, in vivo and in vitro study and manipulation of SSCs in rodent testes have been progressing at a staggering rate. By contrast, their counterparts in domestic animals, despite the failure to reach a comparable level, still burgeoned and showed striking advances. This review will outline the recent progressions of characterization, isolation, in vitro propagation and transplantation of spermatogonia/SSCs from domestic animals, thereby shedding light on future exploration of these cells with high value, as well as contributing to the development of reproductive technology for large animals.
    Reproduction 12/2013; 147(3). DOI:10.1530/REP-13-0466 · 3.17 Impact Factor
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    • "They resume proliferation and migrate to the basement membrane of the seminiferous tubules after birth, presumably differentiating into undifferentiated spermatogonia and initiating spermatogenesis (Culty 2009). Gonocytes, when transplanted into infertile recipient testes, are able to initiate and maintain colonies of spermatogenesis (Jiang and Short 1998; Orwig et al. 2002). "
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    ABSTRACT: Gonocytes are important for the study of spermatogenesis. Identification and isolation of gonocytes has been reported in rodents but not in pigs due to a lack of molecular markers for gonocytes. The objective of this study was to identify THY1 expression in porcine testicular tissue and subsequently utilise THY1 as a marker to isolate and enrich porcine gonocytes from testes of newborn piglets. Immunohistochemical analysis showed that THY1 was expressed in gonocytes. Double-immunofluorescent analysis of THY1 and ZBTB16 indicated that THY1 and ZBTB16 were partially co-localised in gonocytes. Double-immunofluorescent analysis of both THY1 and GATA4 suggested that THY1(+) cells were not Sertoli cells. Magnetic-activated cell sorting of THY1(+) cells yielded a cell population with an enrichment of UCHL1(+) gonocytes 3.4-fold of that of the unsorted testicular cell population. Western blot and quantitative reverse transcription-polymerase chain reaction analyses confirmed that the selected THY1(+) fraction had a higher expression of UCHL1 than the unsorted cells. In conclusion, the study demonstrated that THY1 is a surface marker of gonocytes in testes of pre-pubertal boars and could be utilised to identify and isolate porcine gonocytes. The findings will also facilitate culture and manipulation of male germline stem cells.
    Reproduction Fertility and Development 05/2013; 26(4). DOI:10.1071/RD13075 · 2.40 Impact Factor
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    • "Compared with PGCs and SSCs, gonocytes are the least investigated germline progenitor cells [17]; therefore, obtaining new knowledge about gonocytes may also shed light on the germline stem cells as a whole. Although gonocytes can be identified in situ and in histological cross-sections by their distinctive topography within the seminiferous cords/tubules and unique morphological attributes [12] [18], specific biomarkers are required for their accurate quantification. The unique expression of biomarkers in/on gonocytes may also indicate specific cellular functions or uncover important biological information about them. "
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    ABSTRACT: Significant intrinsic fluorescence in tissues and in disassociated cells can interfere with fluorescence identification of target cells. The objectives of the present study were (1) to examine an intrinsic fluorescence we observed in both the piglet testis tissue and cells and (2) to test an effective method to block the autofluorescence. We observed that a number of granules within the testis interstitial cells were inherently fluorescent, detectable using epifluorescence microscopy, confocal laser scanning microscopy, and flow cytometry. The emission wavelength of the autofluorescent substance ranged from 425 to 700 nm, a range sufficiently broad that could potentially interfere with fluorescence techniques. When we treated the samples with Sudan Black B for different incubation times, the intrinsic fluorescence was completely masked after treatment for 10-15 min of the testis tissue sections or for 8 min of the testis cells, without compromising specific fluorescence labeling of gonocytes with lectin Dolichos biflorus agglutinin (DBA). We speculate that the lipofuscin or lipofuscin-like pigments within Leydig cell granules were mainly responsible for the observed intrinsic fluorescence in piglet testes. The method described in the present study can facilitate the identification and characterization of piglet gonocytes using fluorescence microscopy.
    08/2012; 2012(1):820120. DOI:10.1155/2012/820120
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