Defined medium for normal adult human prostatic stromal cells

Abstract

Stromal-epithelial interactions are pivotal in many aspects of prostatic biology. A defined culture system is critical for the investigation of factors that regulate the growth and differentiation of human prostatic stromal cells. We have identified conditions which promote stromal cell attachment and proliferation in serum-free medium. MCDB 201, originally developed for the clonal growth of chick embryo fibroblasts, proved to be a superior basal medium of those that we tested. Supplementation of MCDB 201 with basic fibroblast growth factor (FGF), insulin-like growth factor (IGF), and platelet-derived growth factor (PDGF) permitted attachment and exponential growth of cells throughout a 7-d period with an initial inoculum as low as 10(3) cells per well of a 96-well microtiter dish. Using these assay conditions, we subsequently verified that basic FGF and IGF, but not PDGF, were required for optimal growth. No activity was found for heparin, transferrin, or the androgen R1881. Epidermal growth factor (EGF) didn't stimulate growth when added to medium containing basic FGF and IGF, but was moderately stimulatory when added to basal medium alone. Cholera toxin inhibited growth. This simple and efficient culture medium provides a suitable assay system for more extensive studies of growth regulation and differentiation of human prostatic stromal cells, and will provide the basis for future development of a defined medium that supports clonal growth. Characterization of stromal-epithelial interactions will be facilitated by the use of this defined culture system for stromal cells in conjunction with the serum-free culture systems previously developed for human prostatic epithelial cells.

Basic FGF, EGF, and PDGF modify TGFβ-induction of smooth muscle cell phenotype in human prostatic stromal cells

“…Others, however, have presented evidence for mitogenic activity of EGF on prostatic stromal cells [25]. In a recent study, we noted that EGF did not stimulate growth of prostatic stromal cells if added to a serum-free medium containing basic FGF, but was moderately mitogenic if basic FGF was absent [26]. In this study, we observed that EGF and TGF␣ partially blocked the induction of ␣-smooth muscle actin by TGF␤, indicating that prostatic stromal cells are responsive to EGF and TGF␣.…”

Current models of human prostate contractility

“…Commonly, cell culture studies use explant or enzymatic digestion of human prostatic tissue and a variety of media formulations to generate prostatic stromal cells. 67,68,70,71,81,82 Often these media formulations contain additives to promote differentiation or proliferation. [81][82][83][84][85] Whatever the variations in media formulation, the resultant cultured prostatic stroma is generally a collection of fibroblasts, myofibroblasts and smooth muscle cells, somewhat reminiscent of the prostatic tissue of (mostly) ageing males.…”

“…67,68,70,71,81,82 Often these media formulations contain additives to promote differentiation or proliferation. [81][82][83][84][85] Whatever the variations in media formulation, the resultant cultured prostatic stroma is generally a collection of fibroblasts, myofibroblasts and smooth muscle cells, somewhat reminiscent of the prostatic tissue of (mostly) ageing males. 75 The relative proportion of these cell types in native human prostatic stroma is still debatable and the data linking BPH with an increase of individual tissue components is not clear; for example, the connective tissue mass in hyperplastic prostates has been shown to increase 86,87 or to remain static.…”

Elements regulating angiogenesis and correlative microvessel density in benign hyperplastic and malignant prostate tissue

“…MCDB 104 selects for prostate stromal cell growth and PrEGM for epithelial cell growth, yielding the two cell populations as described by Peehl. 25 Cultures were passaged 2 times before 24 h serum-free conditioned media (CM) was collected. Representative epithelial and stromal cultures were analyzed by immunohistochemistry (IHC) with antibodies against actin, pancytokeratin, pancytokeratin cocktail AE1/AE3 and vimentin to verify the purity of their respective cell populations (results not shown).…”

“…Additionally, it should be considered that the cultured epithelial and stromal cells are grown in different media. While this could potentially have a variable effect on cytokine production, these culture media and conditions as developed by Peehl 25 are felt to optimize the environment for each cell type.…”