Molecular and Cellular Endocrinology 224 (2004) 41–53 Spontaneously immortalized epithelial cells from mouse caput epididymidis A. Britan a , J-J. Lareyre b , A-M. Lefranc ¸ois-Martinez c , M. Manin c , V. Schwaab d , V. Greiffeuille a , P. Vernet a , J.R. Drevet a,∗ a Laboratoires “Epididyme and Maturation des Gam` etes”, Universit´ e Blaise Pascal-Clermont II, 24 Avenue des Landais, 63177 Aubi` ere Cedex, France b INRA SCRIBE, Campus de Beaulieu, 35042 Rennes, France c Physiologie Compar´ ee et Endocrinologie Mol´ eculaire, Universit´ e Blaise Pascal-Clermont II, 24 Avenue des Landais, 63177 Aubi` ere Cedex, France d LMD, Pra de Serre, 63960 Veyre-Monton, France Received 22 December 2003; received in revised form 28 June 2004; accepted 29 June 2004 Abstract We report here on the characterization of tissue-culture cell lines derived from primary cultures of the mouse caput epididymidis epithelium. The cell lines were spontaneously immortalized without the use of transforming oncogenes. In defined conditions, our epididymal cells adopted various morphological features that resembles that of the in vivo epididymis epithelium such as a polarized organization and the presence of junctional structures at their apical/lateral membranes as revealed by electron microscopy analyses. Flow cytometry analysis revealed that we were dealing with homogenous cell populations that had reached a near-tetraploid state. RT-PCR assays were used in order to show that several genes that can be considered as markers of in vivo caput epididymidis epithelium activity were expressed in our cell lines confirming that these cells were indeed in a differentiated state close to their endogenous state. © 2004 Elsevier Ireland Ltd. All rights reserved. Keywords: Epididymis epithelium; Tissue-culture cells; RT-PCR assays 1. Introduction The epithelium lining the mammalian epididymis is a particularly interesting tissue because of its highly tissue-, segment-, and cell-restricted pattern of gene expression (for reviews see: Cornwall and Hann, 1995; Kirchhoff, 1999). These characteristics, together with the fact that most of the genes expressed within the epididymis are developmentally- and hormonally-regulated make it a suitable model for understanding the molecular mechanisms governing gene ex- pression in higher eukaryotes. Despite these interesting fea- tures, the analysis of the fine regulation of epididymal gene expression as well as studies concerning the secretory ac- ∗ Corresponding author. Tel.: +33 473 407413; fax: +33 4734 05245. E-mail address: joel.drevet@univ-bpclermont.fr (J.R. Drevet). tivities of the epididymis epithelium have been impaired by the lack of tools such as homologous cell free transcription systems and, above all, established cell cultures. In the past 20 years, several attempts have been carried out to develop cultures of epididymal epithelial cells in mammals but the task is particularly difficult because of the complex- ity and the high degree of differentiation of this epithelium. Indeed, this long and convoluted duct although apparently ho- mogenous from the caput to the cauda, is prominently lined by columnar epithelial cells (i.e., the principal cells) but also exhibits other cell types such as apical cells, clear cells, halo cells and basal cells (Abe et al., 1983). Furthermore, its dif- ferentiated state is regulated postnatally by the increasing levels of circulating androgens and of paracrine factors en- tering the epididymis lumen with the testicular fluid. Based on its anatomy and also on its regionalized pattern of gene 0303-7207/$ – see front matter © 2004 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.mce.2004.06.010