Human male germ cell development in vivo and in vitro
Final Report Abstract
The aim of this project was to define mechanisms involved in human male germ cell development and to apply novel culture systems to explore human male germ cell differentiation in vitro. Human embryonic stem cells (hESCs) and testicular germ cells were used with the ultimate goal to establish an in vitro model for human spermatogenesis. Because of lacking experimental models, cellular and molecular mechanisms of early germ cell development in the human testis are poorly understood. Novel approaches, such as the three-dimensional soft-agar-culture-system (SACS) or organ culture approaches were used to study male germ cells differentiation in vitro. Our results focussing on the establishment of a robust in vitro system to differentiate male germ cells, showed a positive effect of gonadotropins on colony formation and overall cell survival. However, the differentiation efficiency of the used cell culture system remains low and further studies are needed to establish a robust system for germ cell differentiation. Another aspect of the study addressed the early male germ cell development by characterization of testicular biopsies from prepubertal boys suffering cryoptorchidism. In human cryptorchid testis, the number of germ and Sertoli cells and testicular volume at 9 months were significantly larger than at 3 years. The intraabdominal testes showed the largest germ cell depletion at 3 years. At both ages, testicular volume correlated to the number of germ and Sertoli cells. None of the hormones measured during the first 6 months of life (LH, FSH, testosterone, and inhibin B) could predict the number of germ or Sertoli cells at either 9 or 36 months of age, nor could hormone levels predict whether spontaneous descent would occur or not. In the last part of the study, the ability of human ES cell to differentiate towards male germ cells, to obtain new insights into early stages of germ cell development which might be related to developmental failures, was investigated. The results obtained so far, show the importance of additive factors as BMPs, but also of the influence of the cell line itself to direct the future differentiation pathways by their genetic signature already in the undifferentiated stage. Combined with cultures of male germ cells, these experiments will lead to a better knowledge of cellular and molecular functions and might help to establish strategies for in vitro culture of human male germ cells for fertility preservation in prepubertal boys suffering side-effects of oncological treatments.
Publications
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”3D-culture systems to define early germ cell differentiation in vitro.” International Journal of Andrology (2010), 33:81
J.B. Stukenborg, L. Landreh, M. Hou, O. Söder
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”Ontogenesis of Ap-2γ expression in rat testes.” Sexual Development (2011) 5:188-196
M. Hou, J.B. Stukenborg, M. Nurmio, M. Andersson, J. Toppari, O. Söder and K. Jahnukainen
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Boys with undescended testes: Endocrine, volumetric and morphometric studies on testicular function before and after orchidopexy at 9 months or 3 years of age.” Journal of Clinical Endocrinology and Metabolism (2012) 97:4588-95
C. Kollin, J.B. Stukenborg, M. Nurmio, E. Sundqvist, T. Gustafsson, O. Söder, J. Toppari, A. Nordenskjöld and E.M. Ritzén
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Differentiation of stem cells towards the germ cell lineage upon deprivation of exogenous FGF2? – A general approach to study spontaneous differentiation of hESCs in vitro.” Systems Biology in Reproductive Medicine (2012) 58:330-8
K.R. Kjartansdóttir, A. Gabrielsen, A. Reda, O. Söder, R. Bergström-Tengzelius, C.Y. Andersen, O. Hovatta, J.B. Stukenborg, and J. Fedder
