Epididymal functions and their hormonal regulation

The epididymis is a complex organ which maintains a specific intraluminal environment thought to be important for effecting sperm maturation in proximal regions and sperm storage in distal regions of the duct. The composition of the internal milieu is achieved both by transport between blood and lumen (and vice versa) and by synthesis and secretion into the lumen. Several low-molecular weight organic molecules achieve high concentration in the epididymal lumen, but their functions in the events of sperm maturation and storage still remain unclear. Metabolic processes occurring within epididymal tissue and the absorptive and secretory activity of the epididymal epithelium are regulated by androgens. The synthesis of some, but not all, secretory proteins is also androgen-dependent. In addition to androgens, other hormones and local testicular factors may influence epididymal function. There is now increasing evidence that epididymal-specific and androgen-dependent secretory proteins play a fundamental role in modifying the surface characteristics of sperm in preparation for the events of fertilization.     

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.     

Androgens are necessary for the establishment of secretory protein expression in the guinea pig seminal vesicle epithelium.

The guinea pig seminal vesicle epithelium (GPSVE) synthesizes and secretes milligram quantities of four related secretory proteins in an androgen-dependent manner. To investigate the role of androgens in the establishment of secretory protein synthesis during the development of the GPSVE, animals were castrated at Day 5, approximately 10 days before secretory protein accumulation begins in intact animals. Castration did not eliminate secretory protein mRNA from the SVE, but it did indefinitely postpone the developmentally programmed increase in secretory protein mRNA. Injection of neonatally castrated guinea pigs with either estradiol or dexamethasone did not alter levels of secretory protein mRNAs. However, treatment of castrated neonates with either testosterone propionate or dihydrotestosterone (DHT) led to specific increases in secretory protein mRNAs within 4 days. Although neonatally castrated animals accumulated and translated significant amounts of secretory protein mRNA, the newly synthesized secretory proteins failed to accumulate until exogenous androgens were provided. This observation suggests that androgens regulate both the accumulation of secretory protein mRNA and the accumulation of secretory proteins in the GPSVE.     

Identification of glutathione S-transferase Yb1 mRNA as the androgen-repressed mRNA by cDNA cloning and sequence analysis

Androgens, while stimulating the growth of the rat ventral prostate, can also repress the levels of a limited number of mRNAs. The cDNA for one of the androgen-repressed mRNAs has been identified by nucleotide sequence analysis as coding for the glutathione S-transferase Yb1 subunit. The prostate cDNA is 1071 nucleotides long, and only 2 or 4 bases of this sequence do not match the two published sequences of the cDNA for the Yb1 subunit of rat liver glutathione S-transferase. The amino acids in the protein encoded by the prostate cDNA matched completely with that for one of the liver cDNAs and differ with the other cDNA only in two of 218 amino acids. The identification of the androgen-repressed mRNA as a glutathione S-transferase subunit may indicate that some of the cellular actions of the enzyme may be important in the control of androgen-dependent growth of the prostate. Since Yb forms of the transferases have been colocalized with uridylic acid-rich small nuclear RNAs at interchromatinic regions of the cell nucleus, autoregulation of prostate growth by androgens may be carried out through the modulation of RNA production or processing in this target organ.     

Studies on the mode of oestrogenic inhibition of hepatic synthesis of alpha2u-globulin and its corresponding messenger ribonucleic acid in rat liver.

1. The possible mechanism of the oestrogenic inhibition of the androgen-dependent synthesis of alpha2u-globulin in rat liver was explored by a correlative study of the amounts of alpha2u-globulin, its corresponding mRNA and circulating testosterone in oestrogen-treated male rats. 2. Daily treatments of mature male rats with oestradiol-17beta (10 microgram/100g body wt.) decreased and ultimately stopped the hepatic synthesis of alpha2u-globulin as determined by both hepatic and urinary concentrations of the protein. The oestrogen-mediated decrease in the hepatic synthesis of alpha2u-globulin was correlated with a decrease in the mRNA for this protein. 3. Withdrawal of oestrogen resulted in the recovery of alpha2u-globulin synthesis and an increase in mRNA for alpha2u-globulin. 4. At higher doses of oestradiol-17beta (50 microgram/100g body wt.), synthesis of alpha2u-globulin was totally suppressed. In addition, this treatment resulted in an extended period of androgen-insensitivity during which treatment with androgens induced synthesis of neither alpha2u-globulin nor its corresponding mtrna. 5. it is concluded that the oestrogenic inhibition of alpha2u-globulin synthesis is mediated by an oestrogen-dependent decrease in the hepatic content of translatable mRNA for alpha2u-globulin.     

The control of the form and function of the ribosomes in androgen-dependent tissues by testosterone

1. The ribosome content of the rat ventral prostate gland is controlled by the concentrations of circulating androgens and the polyribosomal complement of the total population of ribosomes is acutely dependent on androgenic stimulation. After the administration of testosterone to castrated rats in vivo, there is a pronounced increase in the amounts of heavy (150-240S) polyribosomes. 2. These results are consistent with a pronounced increase in the mRNA and rRNA content of the prostate gland after the administration of testosterone in vivo. 3. From studies conducted both in vitro, the heavy prostate polyribosomes formed after androgenic stimulation are particularly active in protein synthesis. 4. The androgen-stimulated increase in the formation of prostate polyribosomes has a mandatory requirement for sustained RNA and protein synthesis. 5. Since the androgen-mediated increase in prostate polyribosomes may also be suppressed by the concomitant administration of certain anti-androgenic steroids in vivo, the response in polyribosome formation is probably initiated by the binding of a metabolite of testosterone, 5alpha-dihydrotestosterone, in the prostate gland. 6. The relevance of these findings to the pronounced increase in protein synthesis in androgen-dependent tissues after hormonal stimulation is discussed.     

Calcitriol induces transcription of the placental transforming growth factor β gene in prostate cancer cells via an androgen-independent mechanism

Calcitriol (1α,25-dihydroxycholecalciferol) suppresses the growth of prostate cancer cells. Growth suppression of hormone-sensitive LNCaP prostate cancer cells by calcitriol is believed to depend on androgens, but the mechanisms of the interactions between the calcitriol-and androgen-dependent signaling pathways is unclear. A previous search for calcitriol-responsive genes in LNCaP cells with cDNA microarrays has shown that calcitriol regulates the expression of the gene for the placental transforming growth factor β (PTGF-β), which suppresses prostate cancer cell proliferation. A study was made of whether expression of the PTGF-β gene is regulated by 5α-dihydrotestosterone and whether induction of this gene by calcitriol is androgen-dependent. Quantitative PCR showed that 5α-dihydrotestosterone increases the level of the PTGF-β mRNA. Neither 5α-dihydrotestosterone nor the antiandrogen Casodex affected the calcitriol-induced level of the PTGF-β mRNA. It was assumed that calcitriol stimulates production of PTGF-β independently of 5α-dihydrotestosterone and that its effect on prostate cancer cell growth is partly mediated by an androgen-independent mechanism.     

Microarray analysis of androgen-regulated gene expression in testis: the use of the androgen-binding protein (ABP)-transgenic mouse as a model

Background  

Spermatogenesis is an androgen-dependent process, yet the molecular mechanisms of androgens' actions in testis are poorly understood. Transgenic mice overexpressing rat androgen-binding protein (ABP) in their testes have reduced levels of intratesticular androgens and, as a result, show a progressive impairment of spermatogenesis. We used this model to characterize changes in global gene expression in testis in response to reduced bioavailability of androgens.     

Elevated yolk androgen levels and the expression of multiple sexually selected male characters

Maternal hormones in bird eggs modulate not only offspring development, but recently it has also been shown that these effects can persist into adult life. A number of long-lasting effects concern traits of which the expression or development is modulated by androgens. This suggests that the nature of yolk hormone-mediated maternal effects may be organizational. Maternal androgens may therefore play an important role in sexual selection, since the expression of sexually selected male characters is often androgen-dependent.We experimentally manipulated the yolk androgen concentrations of European Starling (Sturnus vulgaris) eggs. Subsequently we followed 49 unrelated males from hatching until year of first reproduction. We investigated the expression of multiple sexually selected male characters (song, beak color and throat feather characteristics), taking into account whether a trait is androgen-dependent.Elevated levels of yolk androgens affected the length of the embryonic period, but did not modify the expression of either androgen-dependent or androgen-independent sexually selected male characters including song phenotype at adulthood. Thus the most important function of yolk androgens in starlings and possibly other bird species may relate to the early developmental period. The outcome of our study together with the results of our meta-analysis indicates that the effects of yolk androgens on sexually selected male characters may be comparatively small. Our results suggest that this may relate to the numerous other environmental and/or genetic factors influencing the expression of sexually selected male characters.     

Molecular cloning of the cDNA for androgen-dependent sperm-coating glycoproteins secreted by the rat epididymis

cDNA clones coding for two closely related androgen-dependent sperm-coating glycoproteins secreted by the rat epididymis were selected by screening an epididymal cDNA library constructed in lambda gt 11 with affinity-purified antibody directed against the glycoproteins. The largest clone of 956 nucleotides provided coding information for a protein of 246 amino acids of which the first 19 residues comprise a putative signal peptide sequence which when cleaved would produce a mature protein of 227 residues and a molecular mass of 26 kDa. Confirmation of the identity of the clone was provided by a match between the amino acid sequence predicted from the cDNA sequence and the actual amino acid sequence determined for a tryptic peptide fragment of one of the pure glycoproteins. It is probable that the primary amino acid sequence of the two glycoproteins is identical. Northern blot and slot-blot analysis revealed that the mRNA for the glycoproteins is approximately 1250 nucleotides long and that the concentration of the mRNA in the epididymis is androgen-dependent. The glycoproteins and their mRNAs were unique to the epididymis as determined by Western and Northern blots, respectively, since signals were absent from skin, brain, liver, kidney, heart, skeletal muscle and testis. Cross-reacting proteins of slightly smaller apparent molecular mass were detected in extracts of mouse and guinea-pig epididymis, but not rabbit or bull epididymis. Comparison with existing protein data bases revealed that the epididymal glycoproteins display significant sequence homology with yeast carboxypeptidase Y.     

CCAAT/enhancer binding protein-mediated role of thyroid hormone in the developmental expression of the kidney androgen-regulated protein gene in proximal convoluted tubules

The kidney androgen-regulated protein (KAP) gene is exclusively expressed in proximal tubules of mouse kidney and in the uterus of pregnant females before they give birth. It displays an exquisite and differential regulation of expression by steroid and thyroid hormones (THs) in different proximal tubule segments. Whereas the pars recta (PR cells) responds to thyroid and sexual hormones, the pars convoluta (PCT cells) represents a truly androgen-dependent compartment because expression occurs only in the presence of androgens and functional androgen receptors. Nevertheless, different hypothyroidism models have indicated that TH might also contribute to the androgenic response in PCT cells. In the present study, we aimed to determine the molecular mechanisms that ultimately control KAP expression in these cells. Using several genetically deficient mouse models and different pharmacologic and hormonal treatments, we determined that thyroid and GH modulate CCAAT/enhancer binding protein alpha and beta levels that, in turn, control KAP expression in PCT cells in a developmentally dependent manner. We demonstrated that these factors bind to sites in the proximal KAP promoter, thereby collaborating with androgens for full KAP expression. Finally, we propose that TH and GH, acting through CCAAT/enhancer binding protein, may constitute a general regulatory mechanism of androgen-dependent genes in mouse kidney.     

Effects of castration on the epididymis of a non-mammalian vertebrate: evolution of morphology, protein synthesis, and of specific mRNA levels

The lizard epididymis is an androgen-dependent organ undergoing large variations in its structure and in protein synthesis ability during its annual cycle. It produces a major androgen-dependent protein, the L protein. This work reports the effects of castration performed at 3 prominent points of the sexual cycle: stage 1 (epithelium reorganization) stage 3 (onset of secretory activity) stage 6 (full secretory activity). Evolution of various parameters (organ weight, histology, amount of soluble proteins, rate of soluble protein synthesis and of specific protein synthesis: L protein, and mRNA levels) was considered over a period ranging from 7 days to 15, 30 and 60 days. Deprivation of the testis was followed by an organ involution which was more or less accentuated or more or less rapid according to the stage of the operation but some peculiarities need to be emphasized. At first, the evolution of the organ was not stopped but it proceeded: at stage 1, there was cell division and a correlated increase in total protein synthesis (without L protein synthesis), at stage 3 total protein synthesis and L protein mRNA levels increased (synthesis of L protein proceeded up to 30 to 60 days), at stage 6 the involution was accelerated. These effects concerning stage 1 and particularly stage 3 appeared as a kind of a paradoxical induction. Secondly, the epithelium underwent phases of destruction and regeneration which were obviously not controlled by the testis.     

Neuroimmunoregulation of androgens in the adrenal gland and the skin

Human adrenals produce large quantities of androgens, especially DHEA which is the most abundant circulating hormone in the human body. Adrenal androgens are regulated by several factors, including pituitary ACTH and an intricate intraadrenal network involving immune cells, cytokines and neuroendocrine factors. The skin is a major target of androgens and androgen receptors are expressed in the epidermis, dermis, sebaceous glands and hair. In addition, the skin has the capacity to metabolize androgens into more powerful compounds. Similar to the adrenal gland, there is an intradermal neuro-immune network involving the local expression of cytokines and neuropeptides. Dysregulation of androgens in the adrenals and/or the skin is associated with acne, hirsutism and androgenic alopecia. Therefore, understanding the mechanisms of these intricate networks is of both basic and clinical relevance and may help to develop new strategies for the treatment of androgen-dependent skin disorders.