Isolation and culture of epithelial cells from rat ductuli efferentes and initial segment epididymidis

To improve the study of epithelial function in rat ductuli efferentes (efferent ductules) and initial segment epididymis, we developed a primary cell culture system with modification of the Klinefelter method (1992). The cultured efferent ductal epithelium was grown to confluence and the cells maintained many of the organelles characteristic of these cells in vivo, including dense-staining granules, indented nuclei and apical cilia. Ciliary beat was observed for up to 10 days in culture, Cultured initial segment epithelial cells were elongated and characterized by apical branched microvilli. Electron microscopy revealed intact cell junctions, and endocytotic apparatus and lysosomal granules. Ultrastructurally, the initial segment epithelium contained a well developed Golgi apparatus. For both epithelia, cell characteristics were also confirmed by indirect immunofluorescent staining for cytokeratins 8, 18. Endocytotic activity was detected by the uptake of cationic ferritin at the apical surface and within vesicles. Estrogen receptor and clusterin mRNAs were expressed in the cultured epithelia and no difference was found in their expressions when cultured with or without 10(-9)M 17-beta estradiol. Indirect immunofluorescent staining for clusterin further indicated that this protein was present in the cultures. In conclusion, these in vitro methods will be useful for the investigation of epithelial function in the head of the epididymis.