Androgen regulation of androgen receptor content and distribution in the ventral and dorsolateral prostates of aging AXC rats

Total androgen receptor content of ventral or dorsolateral prostate of intact, aged (730–740 day old) rats is decreased 50% when compared to intact, young mature (150–170 day old) rats. Treatment with exogenous testosterone increased ventral and dorsolateral prostate androgen receptor content per cell in aged rats to values identical to those of prostates of young mature rats. The increase in prostate receptor content was not attributable to testosterone mediated cellular hypertrophy or hyperplasia. At 24 hr post-orchiectomy ventral prostate cytoplasmic androgen receptors are depleted of endogenous androgen, without any decrease in number of receptors per cell, and nuclear androgen receptors are undetectable. During 30 to 60 min after a single 200 μg testosterone injection, ventral prostate nuclear receptor content increased to the level of intact control rats without producing any reduction in total cytoplasmic androgen receptor content. Although dorsolateral prostate is devoid of cytoplasmic androgen receptor, the effects of orchiectomy and testosterone treatment upon nuclear androgen receptor are comparable to those seen in ventral prostate. These effects of orchiectomy and testosterone injection upon prostatic receptor content and distribution were identical in prostates of young and aged rats. Our studies show that receptor processing in prostates of young and aged rats does not involve a process by which nuclear receptor is derived by depletion of cytoplasmic receptor. Moreover, our studies of the effect of short-term (48 hr) exogenous testosterone treatment upon androgen receptor content in prostates of aged rats are the first demonstration that androgen receptor content may be enhanced independent of generalized androgen mediated anabolic effects in prostate.     

Intravesicular Fas localization in epithelial cells of castrated rat prostate glands

Androgenic steroids regulate the development and size of mammalian prostate epithelial cells. To evaluate the relationship between Fas-Fas ligand system and apoptosis in prostate epithelial cells of the castrated rats, we have examined immunocytochemical localization of Fas antigen in the castrated rat prostate glands at a series of different times. We used a rabbit polyclonal anti-Fas antibody with a streptavidin-biotin method and confocal laser scanning method or an immunogold method. Fas immunolocalization was examined in ventral lobes of prostate glands taken from intact or castrated adult male Wistar rats on day 1, 2, 3, 4 and 5 by light or electron microscopy. At a light microscopic level, the castrated prostate epithelial cells showed mostly Fas immunolocalization in their apical parts of cytoplasm on day 2 after the castration. In addition, their extent of the Fas expression was expanded throughout the cytoplasm in proportion to the androgen ablation periods, and later the Fas expression was detected at luminar or basolateral sides of the epithelial cells. Both immunogold labeling with ultrathin sections and immunoperoxidase technique with cryostat sections demonstrated that Fas was localized mainly in secretory granules of the castrated prostate epithelial cells and some parts of their cell membranes at later stages. Our immunocytochemical findings showed that Fas expression was time-dependently induced in most of the prostatic epithelial cells after castration of rats. The rate of Fas-expressing epithelial cells was too high and inconsistent with the previously reported rate of TUNEL-positive ones. The membrane-associated Fas may have little effect on the apoptosis in the present case, bacause a lot of soluble Fas was secreted from the prostatic epithelial cells. A further study is needed to clarify some significance of the secretory Fas in the prostatic epithelium after the rat castration.     

Rat ventral prostate as an independent source of inhibin-like material

Castration had no effect on prostatic inhibin-like activity as compared to normal controls. Consequent upon castration there is a significant reduction in the inhibin content of the ventral prostate on a per gland basis. However, when expressed as a function of protein content, there is no change. The data suggest that the very same epithelial cells which under the influence of androgen carry out the characteristic exocrine secretory activity are responsible for the synthesis and secretion of inhibin-like material.     

Castration induced changes in dog prostate gland associated with diminished activin and activin receptor expression

This study was conducted to evaluate the effect of androgen ablation on dog prostate gland structure and the proliferation capacity of the prostatic cells and their association with the expression of Activin A and Activin RIIA receptor. The effect of androgen on the prostate gland was compared in intact and castrated dogs after one and two weeks. Specific primary antibodies were used to immunolocalize activin-A, activin receptor type II A and the proliferation marker (PCNA). The results showed that the glandular acini of the prostate gland of intact dogs are lined by tall columnar secretory cells and less abundant flattened basal cells and surrounded by a thin fibromuscular tissue. The cytoplasm of the glandular cells exhibited an intense immunoreaction for activin A and activin RIIA receptor while basal cells expressed PCNA. Castration induced a remarkable atrophy of the prostatic acini associated with a progressive loss of secretory epithelial cells, which showed a dramatic decrease to complete disappearance of Activin A and Activin RIIA receptor immunoreactions. The remaining cells of the atrophied acini continue to express PCNA and the inter-acinar fibromuscular tissue showed a remarkable increase in its mass and are induced to express PCNA. These results indicated that androgen is required for the survival of epithelial cells and to maintain growth-quiescent fibromuscular cells, while basal cell proliferation is androgen independent. The changes in the Activin A and Activin RIIA receptor localization and their association with the dynamic pattern of prostate gland regression after castration suggested that Activin A and Activin RIIA receptor expression are androgen dependent.     

Effects of estradiol on prostate epithelial cells in the castrated rat.

There is evidence that estrogens can modulate the activity of prostate epithelial cells. To determine whether estradiol can have a direct influence on rat prostate, this study examined the effects of estradiol-17beta (E(2)) administered alone or in combination with dihydrotestosterone (DHT) to castrated rats for 3 weeks on prostate binding protein (PBP) C1 mRNA expression and androgen receptor (AR) localization. PBP C1 mRNA levels were measured by semi-quantitative in situ hybridization using a (35)S-labeled cDNA probe. In intact animals, strong hybridization signal could be observed in prostate sections after 12 hr of exposure to Kodak X-Omat films. In castrated rats, no PBP C1 mRNA could be detected even with longer exposure times, an effect that was prevented by administration of DHT. E(2) administered alone induced a detectable hybridization signal, and the concomitant administration of E(2) and DHT induced an increase in PBP C1 mRNA that significantly exceeded that obtained in animals that received only DHT. In prostate epithelial cells of intact animals, AR immunostaining was restricted to the nucleus. In castrated animals the alveoli were decreased in size and the epithelial cells were atrophied. AR staining was weak and was detected in both cytoplasm and nucleus. DHT administration completely obviated the effect of castration on epithelial cell histology and on AR immunostaining distribution and intensity. Interestingly, E(2) administration alone induced moderate hypertrophy of epithelial cells compared to the histological appearance of cells in untreated castrated rats. Moreover, in E(2)-treated animals the nuclear staining was much stronger than that detected in untreated castrated rats, whereas the cytoplasmic staining was not modified by the treatment. In animals that received both DHT and E(2), the staining was similar to that seen in DHT-treated rats. These results suggest that E(2) can influence the activity of rat prostate epithelial cells by mechanisms that remain to be fully clarified.     

Intracellular localization of Prostatic Binding Protein (PBP) in rat prostate by light and electron microscopic immunocytochemistry

Extra- and intracellular distribution of Prostatic Binding Protein (PBP) was studied in the different genital organs of the male rat by immunocytochemistry at the light and electron microscopic levels. PBP was extracted from cytosols of rat ventral prostate and used for immunization of rabbits. The specificity of the antiserum raised was tested by "western blotting" and immunoelectrophoresis. From the different fixatives tested for optimal structural and antigenic preservation of the ventral prostate a mixture containing 2.5% paraformaldehyde, 0.5% glutaraldehyde and 0.5% CaCl2 in cacodylate buffer, 0.05 M, pH 7.3 was selected. Using the immunofluorescence technique and the unlabeled antibody enzyme method PBP-immunoreactivity was detected at the light microscopic level in the luminal secretions of the ventral prostate. No reaction was observed with the seminal vesicle, the coagulating gland, the dorsal and lateral prostates, the epididymis and the testis. Intracellular secretory granules reacting with PBP antiserum were exclusively found in the secretory cells of the ventral prostate. Insufficiently fixed cells showed a diffuse generalized reaction of the cytoplasm indicating a leakage of the antigen from the secretory granules. Such artifacts were common in tissue sections processed with the preembedding-staining procedure. At the ultrastructural level therefore mostly the postembedding staining method was performed using both the unlabeled antibody enzyme method and the ferritin-labeled immunoglobulin technique in osmicated, Epon-embedded tissue. Labeling with either method was intense in the secretory granules and the condensing vacuoles, while the labeling density of the rough endoplasmic reticulum and the Golgi cisternae was in the background range. Castration experiments showed that secretory material displaying PBP immunoreactivity was retained within the acinar lumen of the gland for several days after castration, but was absent from most secretory cells already by four days after castration. Immunocytochemistry of PBP therefore is a very sensitive method for analysing the secretory activity and its androgen dependence of the prostate of the rat.     

Fibroblast growth factor receptor 2 tyrosine kinase is required for prostatic morphogenesis and the acquisition of strict androgen dependency for adult tissue homeostasis

The fibroblast growth factor (FGF) family consists of 22 members and regulates a broad spectrum of biological activities by activating diverse isotypes of FGF receptor tyrosine kinases (FGFRs). Among the FGFs, FGF7 and FGF10 have been implicated in the regulation of prostate development and prostate tissue homeostasis by signaling through the FGFR2 isoform. Using conditional gene ablation with the Cre-LoxP system in mice, we demonstrate a tissue-specific requirement for FGFR2 in urogenital epithelial cells--the precursors of prostatic epithelial cells--for prostatic branching morphogenesis and prostatic growth. Most Fgfr2 conditional null (Fgfr2(cn)) embryos developed only two dorsal prostatic (dp) and two lateral prostatic (lp) lobes. This contrasts to wild-type prostate, which has two anterior prostatic (ap), two dp, two lp and two ventral prostatic (vp) lobes. Unlike wild-type prostates, which are composed of well developed epithelial ductal networks, the Fgfr2(cn) prostates, despite retaining a compartmented tissue structure, exhibited a primitive epithelial architecture. Moreover, although Fgfr2(cn) prostates continued to produce secretory proteins in an androgen-dependent manner, they responded poorly to androgen with respect to tissue homeostasis. The results demonstrate that FGFR2 is important for prostate organogenesis and for the prostate to develop into a strictly androgen-dependent organ with respect to tissue homeostasis but not to the secretory function, implying that androgens may regulate tissue homeostasis and tissue function differently. Therefore, Fgfr2(cn) prostates provide a useful animal model for scrutinizing molecular mechanisms by which androgens regulate prostate growth, homeostasis and function, and may yield clues as to how advanced-tumor prostate cells escape strict androgen regulations.     

Intracellular localization of prostatic binding protein (PBP) in rat prostate by light and electron microscopic immunocytochemistry

Summary Extra- and intracellular distribution of Prostatic Binding Protein (PBP) was studied in the different genital organs of the male rat by immunocytochemistry at the light and electron microscopic levels. PBP was extracted from cytosols of rat ventral prostate and used for immunization of rabbits. The specificity of the antiserum raised was tested by western blotting and immunoelectrophoresis. From the different fixatives tested for optimal structural and antigenic preservation of the ventral prostate a mixture containing 2.5% paraformaldehyde, 0.5% glutaraldehyde and 0.5% CaCl₂ in cacodylate buffer, 0.05 M, pH 7.3 was selected. Using the immunofluorescence technique and the unlabeled antibody enzyme method PBP-immunoreactivity was detected at the light microscopic level in the luminal secretions of the ventral prostate. No reaction was observed with the seminal vesicle, the coagulating gland, the dorsal and lateral prostates, the epididymis and the testis. Intracellular secretory granules reacting with PBP antiserum were exclusively found in the secretory cells of the ventral prostate. Insufficiently fixed cells showed a diffuse generalized reaction of the cytoplasm indicating a leakage of the antigen from the secretory granules. Such artifacts were common in tissue sections processed with the preembedding-staining procedure. At the ultrastructural level therefore mostly the postembedding staining method was performed using both the unlabeled antibody enzyme method and the ferritin-labeled immunoglobulin technique in osmicated, Epon-embedded tissue. Labeling with either method was intense in the secretory granules and the condensing vacuoles, while the labeling density of the rough endoplasmic reticulum and the Golgi cisternae was in the background range. Castration experiments showed that secretory material displaying PBP immunoreactivity was retained within the acinar lumen of the gland for several days after castration, but was absent from most secretory cells already by four days after castration. Immunocytochemistry of PBP therefore is a very sensitive method for analysing the secretory activity and its androgen dependence of the prostate of the rat.Supported by a grant from the Deutsche Forschungsgemeinschaft (Au 48/7-6) and a grant from the Cystic Fibrosis Foundation (G 261 A)     

Androgen metabolism and actions in rat ventral prostate epithelial and stromal cell cultures

The rat ventral prostate requires androgens for normal development, growth, and function. To investigate the relationship between androgen metabolism and its effects in the prostate and to examine differences between the epithelial and stromal cells, we have established a system of primary cell cultures of immature rat ventral prostate cells. Cultures of both cell types after reaching confluency (6-7 days) actively metabolized 3H-labelled testosterone (T), 5 alpha-dihydrotestosterone (5 alpha-DHT), 5 alpha-androstane-3 alpha,17 beta-diol, and 5 alpha-androstane-3 beta,17 beta-diol. The epithelial cells actively reduced T to 5 alpha-DHT and formed significant amounts of 5 alpha-androstane-3,17-dione from T, 5 alpha-DHT, and 5 alpha-androstane-3 alpha,17 beta-diol. All substrates were converted to significant amounts of C19O3 metabolites. The stromal cells also metabolized all substrates, but very little 5 alpha-androstane-3,17-dione was formed. The metabolism studies indicate that both cell types have delta 4-5 alpha-reductase, 3 alpha- and 3 beta-hydroxysteroid oxidoreductase and hydroxylase activities. The epithelial cells have significant 17 beta-hydroxysteroid oxidoreductase activity. The epithelial cells cultures grown in the presence of T have higher acid phosphatase (AP) contents (demonstrated histochemically and by biochemical assay). Tartrate inhibition studies indicate that the epithelial cells grown in the presence of T are making secretory AP. Stromal cell AP is not influenced by T. The results indicate that the cultured cells maintain differentiated prostatic functions: ability to metabolize androgens and, in the case of the epithelial cells, synthesize secretory AP.     

Mesenchymal-epithelial interactions as factors influencing male accessory sex organ growth in the rat

Mesenchyme (UGM) and epithelium (UGE) isolated from the urogenital sinuses (UGS) of 17-day male and female rat embryos were separated by using a trypsinization procedure, grown on soft agar, transplanted into syngeneic pubertal male hosts as subcapsular renal grafts, and then collected after 29-30 days. Neither UGM nor UGE underwent prostatic morphogenesis when grown under these conditions. However, tissue recombinants composed of UGM + UGE grew and produced prostatic glands with acinar secretory material. Further, UGM + UGE recombinants were made by varying the proportions of mesenchymal and epithelial tissues. The size of the implants was a function of the absolute amount of mesenchyme; increasing the absolute amount of UGM produced larger specimens whereas varying the UGE had no effect. The UGM was also found to be essential for supporting the growth of small glandular elements derived from the ventral prostate of pubescent rats. Segments isolated from the terminal vesicles (TIPs) and from prostatic tissue adjacent to the urethra (PDCT) regressed when implanted alone under the kidney capsule. However, combination of the prostatic segments with UGM produced prostatic glands with relative wet weight and DNA content responses of the following order: UGM + TIP greater than UGM + PDCT = UGM + UGE. Two-dimensional gel electrophoretic protein patterns from UGM + PDCT and UGM + TIP specimens had differential expression of three protein regions unique to the ventral prostate Quantitative and qualitative responses of the TIP and PDCT segments to UGM inductive influences indicate that differences exist between the epithelia of the TIP and PDCT regions of the ventral lobes of the rat prostate.     

Does the lack of regression-associated mRNA expression render a rat ventral prostate epithelial cell line androgen independent?

A series of rapidly dividing epithelial (RDE) cell lines have been isolated from primary cultures of rat ventral prostate (RVP) epithelial cells. Unlike androgen-dependent secretory epithelial cells, the RDE cells in culture do not express the androgen-dependent secretory proteins, nor do they express the androgen-repressed cell death sequences (TRPM-2) found in the epithelial cells during prostatic regression. Screening of a cDNA clone library established from RDE cell mRNA has yielded a number of RDE cell-specific sequences. One of these, RDE-.25 is a 250-base mRNA. The sequence of RDE-.25 shows considerable homology with the rat growth hormone gene and two murine oncogene sequences. We believe that the absence of androgen-repressed cell death sequence expression confers androgen independence for survival and growth, while the expression of RDE-.25 may represent an autocrine growth stimulus which greatly increases the rate of cell division in these cells.     

Ductal heterogeneity of cytokeratins, gene expression, and cell death in the rat ventral prostate.

The rat ventral prostate is a complex gland composed of numerous ducts. The epithelial cells that line the lumen of the ducts are surrounded by stromal cells. The epithelial cells display a characteristic morphology that is dependent on their anatomical location within the ducts; the cells that line the lumen in the region of the ducts close to the urethra (the proximal region) are cuboidal, while those in the distal regions of the ducts are tall columnar cells. We have examined the regional expression of two genes that are expressed in the prostate: prostate steroid-binding protein (PSBP; a marker for androgen-dependent protein synthesis) and TRPM-2 (a marker for programmed cell death). We have demonstrated that the expression of PSBP, in the presence of androgens, and TRPM-2, in the absence of androgens, is restricted to the luminal epithelial cells in the distal regions of the prostatic ducts. Neither of the genes is expressed in the proximal regions of the ducts. In view of the probable effects of the epithelial-stromal interactions in the gland we have also characterized the cytokeratin composition of the epithelial cells lining the prostatic ducts. We have established that the basal epithelial cells of the prostate are primarily localized in the proximal region of the ducts. We propose that these cells may attenuate the influence of the stromal cells on the luminal epithelium and exert a negative influence on the cytodifferentiation of the secretory epithelial cells. The results also suggest that PSBP, which has been considered to be an androgen-dependent gene may, in fact, be a sequence that is constitutively expressed in the luminal cells that die in the absence of androgens. This has significant implications on the mechanism of androgen action in the rat ventral prostate.     

Proprotein convertases modulate budding and branching morphogenesis of rat ventral prostate

The onset of prostate morphogenesis is involved in the interaction between mesenchyme and epithelium. Proprotein convertases (PCs) activate a variety of growth and differentiation factors including mesenchymal and epithelial factors, such as insulin-like growth factor (IGF) and transforming growth factor-beta (TGF-beta), which induce ductal budding and branching. In this study, we provide evidence that PCs play a critical role in prostatic budding from the urogenital sinus (UGS) and ductal branching morphogenesis of the neonatal rat ventral prostate. PCs were expressed only in the epithelial cells of neonatal rat prostate. PC activity in the ventral prostate was modulated by endogenous androgen. PC inhibition suppressed prostatic budding and branching. Taken together, our data indicates that androgen-induced PCs initiate the development of the prostate.     

Inhibition of monoamine oxidase A promotes secretory differentiation in basal prostatic epithelial cells

Abstract Monoamine oxidase A (MAO-A) expression is associated with high-grade prostate cancer. Immunohistochemistry showed that MAO-A is also expressed in the basal epithelial cells of normal prostate glands. Using cultured primary prostatic epithelial cells as a model, we showed that MAO-A prevents basal epithelial cells from differentiating into secretory cells. Under differentiation-promoting conditions, clorgyline, an irreversible MAO-A inhibitor, induced secretory cell-like morphology and repressed expression of cytokeratin 14, a basal cell marker. More importantly, clorgyline induced mRNA and protein expression of androgen receptor (AR), a hallmark of secretory epithelial cells. In clorgyline-treated cells, androgen induced luciferase activity controlled by the promoter of prostate-specific antigen, an AR target gene, in a dose-dependent manner. This activity was blocked by the AR antagonist Casodex, showing that AR is functional. In turn, androgen decreased MAO-A expression in clorgyline-treated, secretory-like cells. Our results demonstrated that cultured basal epithelial cells have the potential to differentiate into secretory cells, and that inhibition of MAO-A is a key factor in promoting this process. Increased expression of MAO-A in high-grade prostate cancer may be an important contributor to its de-differentiated phenotype, raising the possibility that MAO-A inhibition may restore differentiation and reverse the aggressive behavior of high-grade cancer.     

The ultrastructure of the accessory sex organs of the male rat

Summary The fine structure of the nuclei of epithelial cells of the dorsal lobe of the rat prostate were studied 2, 3, 5, 7 and 21 days after castration. The nucleolus appears to undergo a progressive disorganisation with partial fragmentation and dispersion of its normal components.Changes in the nucleoplasm were primarily reflected by a condensation of chromatin, particularly along the nuclear membrane and adjacent to the nucleolus. Later, different types of intranuclear inclusions were observed.After 21 days, the nuclei were characterized by an irregular outline with large indentation. Within the nucleoplasm aggregates of coarse granular chromatin were found. No cell necrosis was observed, indicating that androgen deprivation results in a remodeling of the cell to a less active state with marked cellular alterations and cessation of secretion, but apparently with some of their basic functions still intact.Injections of testosterone completely reverse the castrated-induced alterations.The changes observed are assumed to be due to the withdrawal of the androgenic stimulus, with a direct influence on the secretory function of the cell. The findings support the view that the stimulating secretory effect of androgen is mediated via an intranuclear androgen receptor, probably located in the nucleolus-associated-chromatin. It is also proposed that the secretory function of the epithelial cells of the prostatic complex, initiated by androgens, may be regulated by an intranuclear secretory center.     

Farnesyl diphosphate synthase is abundantly expressed and regulated by androgen in rat prostatic epithelial cells

Farnesyl diphosphate synthase (FPPS) has been identified as an androgen-response gene in the rat ventral prostate using a highly sensitive PCR-based cDNA subtraction technique. FPPS is an essential enzyme that catalyzes the synthesis of farnesyl diphosphate (FPP), which is required for cholesterol biosynthesis as well as protein prenylation. We have characterized the expression of FPPS in the rat prostate in response to androgen manipulation. Northern blot analysis showed that castration induced a 10-fold down-regulation of FPPS mRNA within 24 h in the ventral prostate and androgen replacement up-regulated FPPS mRNA rapidly in the regressed ventral prostate of a castrated rat. The expression of FPPS was also regulated by androgen in the lateral and dorsal prostate, indicating that FPPS is important to androgen action in all three lobes of the prostate. Western blot analysis showed that FPPS protein level was also regulated by androgen in the prostate. Northern blot analysis of tissue specificity indicated that FPPS was most abundantly expressed in the ventral prostate of a mature rat and was responsive to androgen manipulation in the prostate and seminal vesicles, but not in other tissues. In situ hybridization study showed that FPPS mRNA was localized to the prostatic epithelium. Interestingly, the expression of FPPS was elevated in Dunning rat prostate tumor cell lines. The above findings suggest that FPPS has the potential to play an important role in androgen action and prostate cancer progression.     

The rat androgen receptor: primary structure, autoregulation of its messenger ribonucleic acid, and immunocytochemical localization of the receptor protein

A composite androgen receptor DNA sequence 4,181 base pairs in length was determined from three cDNA clones isolated from a rat epididymal bacteriophage lambda gt11 library. An open reading frame of 902 amino acids encodes a protein of 98,227 mol wt. Structural domains characteristic of the steroid receptor family include an amino-terminal region with five repeated amino acid motifs, a central DNA-binding domain homologous with other steroid receptors, and a carboxyl-terminal steroid-binding region. A receptor cDNA probe used in Northern blot analysis hybridized with a predominant 10-kilobase androgen receptor mRNA in male reproductive tissues of the rat. Autoregulation of androgen receptor mRNA was indicated in rat ventral prostate by an increase in the level of 10-kilobase mRNA after castration and suppression of receptor mRNA upon androgen restimulation. A 15 amino acid peptide with sequence derived from the deduced androgen receptor sequence was synthesized and used as immunogen in raising receptor antibodies in rabbits. Antisera reacted with high titer against the synthetic peptide by enzyme-linked immunosorbent assay and against the native [3H]dihydrotestosterone-labeled androgen receptor as evidenced by an increase in receptor sedimentation rate determined by sucrose gradient centrifugation. Immunocytochemical staining localized the androgen receptor to epithelial cell nuclei in rat ventral prostate.     

Glyceraldehyde-3-phosphate dehydrogenase expression during apoptosis and proliferation of rat ventral prostate.

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a multifunctional enzyme known to play a critical role in neuronal apoptosis. We undertook the current studies to determine whether GAPDH also plays a role in prostate epithelial cell apoptosis in response to androgen deprivation. To do so, we analyzed GAPDH staining by immunohistochemistry during castration-induced involution and androgen-induced regeneration of rat ventral prostate. We found that GAPDH was undetectable in secretory epithelial cells at baseline and that staining did not increase in the epithelium during the period of peak apoptosis from 1 to 3 days after castration. However, GAPDH levels did increase within nuclei of some basal epithelial cells 5 days after castration and within the cytoplasm of all secretory epithelial cells 7 days after castration. GAPDH was also abundant within the cytoplasm of secretory epithelial cells during the period of maximal cell proliferation from 2 to 3 days after androgen replacement and was clearly apparent within nuclei of some epithelial cells 4 days after androgen replacement. Our studies suggest that GAPDH plays multiple roles during prostate epithelial cell apoptosis and proliferation.