Primary epithelial cell cultures derived from canine prostate: Isolation, culture, and characterization

Abstract

Epithelial-cell-enriched primary cultures were established from canine prostate. Minced tissue was dissociated with 750 units/ml of collagenase in F12K tissue culture medium containing 1% fetal bovine serum. This treatment resulted in the gradual removal of stromal elements from the base of the epithelial cells. After 30 minutes of digestion, aggregates of epithelial cells free of stroma were dislodged from the minced pieces of prostate. These aggregates were washed and plated at high density in F12K plus 10% fetal bovine serum. After 12-16 hours in vitro the unattached cellular aggregates were removed from the culture dishes, washed, and reinoculated into new culture vessels containing fresh medium. After 48 hours in vitro, the aggregates had attached to the culture vessels and spread out to yield discrete patches of epithelial cells. By 120 hours in vitro the patches of cells had grown and coalesced to form a confluent monolayer of epithelial cells. Ultrastructural examination of these cultures indicated that adjacent cells were joined by desmosomes and tight junctions and had tonofilaments and microvilli, giving the cells an epithelial appearance. The cells contained rough endoplasmic reticulum, Golgi apparatus, and secretory granules similar to those of the epithelial cells in the intact organ. In addition, intracellular "blebs" containing acid phosphatase were observed in the monolayers and were found to increase in size and number with time in vitro. Differentiated function of the cultures was demonstrated by the presence of ornithine decarboxylase and acid phosphatase and the ability of the cultures to metabolize testosterone to primarily 5 alpha-reduced metabolites.

Stromal and epithelial cells of the canine prostate express parathyroid hormone-related protein, but not the PTH/PTHrP receptor

“…Their proliferative rate decreased, and some cells were shed from the monolayers and found in the culture medium. These features were consistent with the differentiation of nonsecretory cells to a secretory phenotype, as previously described [31][32][33]. Canine prostatic stromal cells were elongated with a typical morphology of mesenchymal cells, grew as isolated cells, and at confluence formed interweaving bundles (Fig.…”

Endothelin-1 production and agonist activities in cultured prostate-derived cells: Implications for regulation of endothelin bioactivity and bioavailability in prostatic hyperplasia

“…It has been suggested that ET-1 may promote sperm transport either in the male reproductive tract or in the uterine cavity by stimulating myometrial contractions [16]. In addition to the regulating ET-1 secretion, prostate secretory epithelial cells are polarized and possess distinct apical and basolateral surfaces formed by tight junctions between the cells [33][34][35][36]. Cultured endothelial cells secrete ET-1 through the basolateral surface [37] providing a mechanism for the paracrine action of ET-1 on vascular smooth muscle cells.…”

“…Cultured PE cell monolayers are organized in a manner reminiscent of columnar epithelial cells in vivo; the cells are polarized and possess distinct apical and basolateral surfaces formed by tight junctions between the cells [33][34][35][36]. Production of ET-1 by PE cells, and not by PS cells, therefore identifies another possible component of the endothelin axis, since the po- Figure 6A.…”

Requirements for attachment and subsequent growth of canine prostatic epithelial cells in culture

“…The protein composition of nECM elaborated by prostatic cells will be the subject of a latter report but it appears from the present study that commercial collagen type I, fibronectin, and laminin, used separately, do not have the capacity to enchance the attachment of prostatic cells. It has been reported that canine prostatic epithelial cells can be efficiently cultured in the presence of FBS in uncoated dishes [35] or in dishes coated with collagen type I in the presence of FBS [36]. It also appears that when canine prostatic epithelial cells are cultured on a ECM prepared from normal canine prostates, in steroid-free but serum-supplemented medium, the amount of secretory protein, a marker of their differentiation, is increased as compared with cells attached to plastic [37].…”