Isolation of bovine spermatogonial cells and co-culture with prepubertal sertoli cells in the presence of colony stimulating factor-1

Authors

1 Department of Clinical Sciences, Faculty of Veterinary Medicine, University of Tehran, Tehran, Iran

2 Department of Anatomy, Faculty of Medical Sciences, University of Tarbiat Modares, Tehran, Iran

3 Department of Animal Sciences, Faculty of Agriculture, University of Tabriz, Tabriz, Iran

Abstract

BACKGROUND: Spermatogonial stem cells (SSCs) are infrequent
self-renewing cells among the type A spermatogonia
within the seminiferous tubules and are the basis of spermatogenesis
in mammalian testis. An adequate number of SSCs is a
primary requirement for the study of their behavior, regulation, and
further biomanipulation. OBJECTIVES: In this paper, we studied
the development of the primary co-cultures of type A spermatogonia
and prepubertal bovine sertoli cells in the presence of Colony
Stimulating Factor 1 (CSF1), a potential contributor in the SSC
niche. METHODS: The effect of different concentrations of CSF1
(0, 10, 50 and 100 ng/mL) on the colonization activity of spermatogonial
cells was assessed 4, 7 and 11 days after the beginning of the
culture by counting the total number of colonies and measuring their
area in each group of the present experiment. Immunofluorescent
staining against OCT4 and vimentin led to the confirmation of the
nature of both the SSCs and sertoli cells. RESULTS: Results showed
that the total number of colonies from day 4 to 11 increased
significantly in all groups, independent of CSF1 concentration. In
addition, the total number and total area of colonies were higher (not
significant) in 10 and 50 ng/mL CSF1 treatments than the control
and 100 ng/mL CSF1 groups in all the three evaluations during the
experiment. However, this difference was only significant (p<0.05)
between the total area of colonies in the control and 10 ng/mLCSF1
groups at day 4 of co-culture. CONCLUSIONS:It was concluded that
CSF1 can be a suitable growth factor for improving SSCs colonization
in vitro, particularly during the first days of culture where
accompanying sertoli cells still have not proliferated sufficiently to
support the propagating spermatogonial cells.

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