Immortalization of a human prostate stromal cell line using a recombinant retroviral approach.

摘要

PURPOSE: We established an immortalized human prostate stromal cell line with retained markers of cell differentiation and alpha1-adrenergic receptor expression. MATERIALS AND METHODS: Primary human prostate stromal explants were infected with an amphotrophic retrovirus encoding the E6/E7 open reading frame of the human papillomavirus type 16. Immunohistochemistry was used to verify the expression of prostate stromal markers. alpha1-Adrenergic receptor expression was investigated using ribonuclease protection assays and radioligand binding. Cell proliferation was measured by the WST-1 assay and cell counting. RESULTS: Clonal isolates of individual prostate stromal cells were isolated and passed in selection media. E6 and E7 expression was verified using reverse transcriptase polymerase chain reaction in the selected cell line. The new prostate stromal cell line PS30 was established which maintains the expression of alpha-smooth muscle actin and expresses 22 fmol./mg. of protein of alpha 1-adrenergic receptors, approximately equal to native human prostate alpha 1-adrenergic receptor expression. However, at a subtype level alpha 1a-adrenergic receptor expression is down-regulated and not detectable by ribonuclease protection assays or radioligand binding, while alpha 1b and alpha 1d-adrenergic receptor expression is enhanced. From a physiological prospective PS30 cells do not form tumors in nude mice and stimulation with phenylephrine does not increase cell proliferation. CONCLUSIONS: We successfully established and characterized an in vitro human prostate stromal cell line. This cell line should facilitate studies designed to characterize the role of the adrenergic nervous system in the regulation of prostate growth.