A quantitative assessment of the gametogenic and androgenic properties of testicular steroids in hypophysectomized rats

The ability of testicular steroids to maintain the quantitative aspects of spermatogenesis was compared with reference to their androgenic properties. Hypophysectomized rats were injected daily with 0.2 mg progesterone, 20 alpha-dihydroprogesterone, 3 beta-hydroxy-5 alpha-pregnan-20-one, testosterone or testosterone propionate for 30 days beginning 2 days after the operation. Testosterone propionate was the most potent steroid tested both in terms of its peripheral androgenic effects and its ability to prevent the post-operative decline in the weight of the testis and seminiferous tubules and the numbers of germ cells throughout their differentiation. The natural androgen, testosterone, exhibited weak gametogenic properties and only partly maintained the normal measures of spermatogenesis. Progesterone exhibited low intrinsic androgenic potency yet was significantly more effective than testosterone in maintaining spermatogenesis; it prevented the degeneration of spermatocytes during the later stages of meiotic prophase and the reduction divisions resulting in an increased yield of step 7 spermatids. Low androgenic and gametogenic properties were exhibited by 20 alpha-dihydroprogesterone and 3 beta-hydroxy-5 alpha-pregnan-20-one. These results may indicate that testosterone produced locally in the seminiferous tubules from progesterone is more effective in maintaining spermatogenesis than androgens entering from the circulation. Alternatively, progesterone may act more directly on the germ cells than previously envisaged.     

Loss and recovery of androgen receptor protein expression in the adult rat testis following androgen withdrawal by ethane dimethanesulfonate

Androgens are especially important for the maintenance of spermatogenesis in adulthood and the experimental withdrawal of testosterone (T) production by ethane dimenthanesulfonate (EDS) is a valuable tool for studying androgen-dependent events of spermatogenesis. The aim of the present study was to investigate the specific changes in immunoexpression of androgen receptor (AR) in the testis in relation to degeneration and regeneration of Leydig cell (LC) population and seminiferous epithelium. Immunohistochemistry for AR and 3beta-hydroxysteroid dehydrogenase (3beta-HSD) as well as TUNEL assay for apoptosis were performed on testicular sections of control and EDS-treated rats. Serum LH and T levels were measured by RIA. Our results revealed a total loss of AR immunoexpression from the nuclei of Sertoli (SCs), LCs and peritubular cells during the first week after EDS administration and that coincided with severe drop in T levels. Two weeks after EDS administration, the AR expression was recovered in these cells but normal stage-specificity in SCs was replaced by uniform intensity of AR immunostaining at all the stages of the spermatogenic cycle. The stage-specific pattern of androgen expression in SCs with a maximum at stages VII-VIII appeared 5 weeks after treatment. LC immunoreactivity for 3beta-HSD at different time points after EDS administration correlated with values of T concentration. The maximal germ cell apoptosis on day 7 was followed by total loss of elongated spermatids 2 weeks after EDS treatment. Regeneration of seminiferous epithelium 3 weeks after EDS administration and onwards occurred in tandem with the development of new LC population indicated by the appearance of 3beta-HSD-positive cells and gradual increase in T production. The specific changes in AR after EDS including their loss and recovery in Sertoli cells paralleled with degenerative and regenerative events in Leydig and germ cell populations, confirming close functional relationship between Sertoli, Leydig and germ cells.     

Cell-cell interactions in the control of spermatogenesis as studied using Leydig cell destruction and testosterone replacement

This review centers around studies which have used ethane dimethane sulphonate (EDS) selectively to destroy all of the Leydig cells in the adult rat testis. With additional manipulations such as testosterone replacement and/or experimental induction of severe seminiferous tubule damage in EDS-injected rats, the following questions have been addressed: 1) What are the roles and relative importance of testosterone and other non-androgenic Leydig cell products in normal spermatogenesis and testicular function in general? 2) What are the factors controlling Leydig cell proliferation and maturation? 3) Is it the Leydig cells or the seminiferous tubules (or both) which control the testicular vasculature? The findings emphasize that in the normal adult rat testis there is a complex interaction between the Leydig cells, the Sertoli (and/or peritubular) cells, the germ cells, and the vasculature, and that testosterone, but not other Leydig cell products, plays a central role in many of these interactions. The Leydig cells drive spermatogenesis via the secretion of testosterone which acts on the Sertoli and/or peritubular cells to create an environment which enables normal progression of germ cells through stage VII of the spermatogenic cycle. In addition, testosterone is involved in the control of the vasculature, and hence the formation of testicular interstitial fluid, presumably again via effects on the Sertoli and/or peritubular cells. When Leydig cells regenerate and mature after their destruction by EDS, it can be shown that both the rate and the location of regenerating Leydig cells is determined by an interplay between endocrine (LH and perhaps FSH) and paracrine factors; the latter emanate from the seminiferous tubules and are determined by the germ cell complement. Taken together with other data on the paracrine control of Leydig cell testosterone secretion by the seminiferous tubules, these findings demonstrate that the functions of all of the cell types in the testis are interwoven in a highly organized manner. This has considerable implications with regard to the concentration of research effort on in vitro studies of the testis, and is discussed together with the need for a multidisciplinary approach if the complex control of spermatogenesis is ever to be properly understood.     

Immunoexpression of androgen receptors and aromatase in testes of patient with Klinefelter's syndrome

Klinefelter's syndrome (47, XXY) is the most common chromosome aneuploidy in men and is usually characterized by underdeveloped testes and sterility. The aim of the present study was to detect cellular distribution of androgen receptors (AR) and aromatase in testes of patient with KS. The tissue sections were processed for morphological and immunohistochemical staining. Additionally, levels of FSH, LH, PRL, estradiol, and testosterone were measured in the plasma. Morphological analysis revealed a complete absence of spermatogenesis. No germ cells were present in seminiferous tubules. In some tubules, nests of apparently degenerating Sertoli cells were found. In the interstitium, Leydig cell hyperplasia was observed. Using immunohistochemistry, nuclear AR staining was detected in Sertoli cells and peritubular cells, whereas in Leydig cells the staining was exclusively cytoplasmic. The immunostaining of aromatase was detected in the cytoplasm of Sertoli cells and Leydig cells. Increased levels of gonadotropins and decreased level of testosterone concomitantly with the cytoplasmic localization of AR in Leydig cells might contribute to the impaired testicular function in patient with KS.     

Testicular histological examination of spermatogenetic activity in captive gorillas (Gorilla gorilla)

To clarify the reproductive state of male gorillas, we performed histological examinations on the testicles of 10 male gorillas (Gorilla gorilla). The testicular samples were obtained by autopsy, and ordinal histological preparations were made for light microscopy. The poor spermatogenesis of this species was characterized by the following findings: First, spermatogenesis was evident in only four samples. Meiosis progressed in two samples, but they lacked spermatogenesis. In the remaining four specimens, seminiferous tubules hyalinized without any sign of spermatogenesis. Second, seminiferous epithelia were thin even in the males in which spermatogenesis was observed. Third, degenerated seminiferous tubules were found in all specimens. Fourth, abnormally large syncytial cells were found in the tubules. Six stages in the epithelial cycle of the seminiferous tubules were identified. Testosterone staining made it clear that there were many Leydig cells with spherical or fusiform nuclei in an abundance of interstitial tissue. The relevance of the testicular architecture of gorillas to the mating system is discussed.     

Immunolocalization of androgen receptors in testicular cells of prepubertal and pubertal pigs

In the testis, androgen receptors are known to mediate autocrine and paracrine effects of androgens on Leydig cell function and spermatogenesis. The pig presents some unusual features with regard to the synthesis of testosterone and estrogens in the male gonads. In testes from prepubertal males, testosterone level was lower than in testes from adult boars, while estrogen secretion was relatively high and comparable to that of mature porcine gonad. Immunolocalization of androgen receptors and intensity of immunohistochemical staining was age-dependent. In testis sections from adult boars, androgen receptors were found in nuclei of all somatic cells such as Leydig cells, Sertoli cells, and peritubular-myoid cells, whereas in sections from immature pigs only in the Leydig cell cytoplasm showed positive immunoreaction for androgen receptors. In control tissue sections incubated with omission of the primary antibody, no positive staining was observed. Detection of the androgen receptors in testicular cells of the pig is important for understanding of their central role in mediating androgen action.     

Profiling of Androgen Response in Rainbow Trout Pubertal Testis: Relevance to Male Gonad Development and Spermatogenesis

The capacity of testicular somatic cells to promote and sustain germ cell differentiation is largely regulated by sexual steroids and notably androgens. In fish species the importance of androgens is emphasized by their ability to induce sex reversal of the developing fries and to trigger spermatogenesis. Here we studied the influence of androgens on testicular gene expression in trout testis using microarrays. Following treatment of immature males with physiological doses of testosterone or 11-ketotestosterone, 418 genes that exhibit changes in expression were identified. Interestingly, the activity of testosterone appeared stronger than that of 11-ketotestosterone. Expression profiles of responsive genes throughout testis development and in isolated germ cells confirmed androgens to mainly affect gene expression in somatic cells. Furthermore, specific clusters of genes that exhibit regulation coincidently with changes in the natural circulating levels of androgens during the reproductive cycle were highlighted, reinforcing the physiological significance of these data. Among somatic genes, a phylogenetic footprinting study identified putative androgen response elements within the proximal promoter regions of 42 potential direct androgen target genes. Finally, androgens were also found to alter the germ line towards meiotic expression profiles, supporting the hypothesis of a role for the somatic responsive genes in driving germ cell fate. This study significantly increases our understanding of molecular pathways regulated by androgens in vertebrates. The highly cyclic testicular development in trout together with functions associated with regulated genes reveal potential mechanisms for androgen actions in tubule formation, steroid production, germ cell development and sperm secretion.     

Expression of the c-Kit receptor in germ cells of the seminiferous epithelium in rats with hormonal imbalance

The aim of the study was to investigate the effects of pharmacologically induced hormonal imbalance in adult male rats treated with letrozole and rats exposed to soya isoflavones on the testicular morphology and c-Kit receptor (c-Kit-R) expression in germ cells. The study was conducted during all developmental periods: prenatal period, lactation, youth, and sexual maturity. Morphological and morphometrical analyses were performed on testicular section, and c-Kit-R was identified using immunohistochemistry. In addition, concentration of circulating steroids was measured in mature rats exposed to soya isoflavones. A significant reduction in testosterone level in rats exposed to soya isoflavones, and the sloughing of the premature germ cells into the lumen of the seminiferous tubules in the testes of both groups of rats were observed. Immunohistochemistry showed a decrease in c-Kit-R expression in germ cells of both experimental groups. Morphometric analysis indicated a decreased thickness of the layers occupied by c-Kit-R-positive spermatogonia, and a decreased diameter of the seminiferous tubules in the testes of both experimental groups of animals. In conclusion, the pharmacologically induced reduction of the estradiol level in adult rats and the diminished level of testosterone in rats exposed to soya isoflavones during the prenatal period, lactation and up to maturity caused similar morphological and functional changes associated with the decreased c-Kit-R expression in germ cells in the seminiferous epithelium. These findings demonstrate the importance of the estrogen/androgen balance for normal testicular morphology and spermatogenesis.     

Stage dependent and androgen inductive expression of orphan receptor TR4 in rat testis

In this study, we investigated the expression of TR4 in different stages of seminiferous tubules and the relationship between TR4 and androgen in rat testis. We found that TR4 was stage-dependently expressed in rat seminiferous tubules, T withdrawal induced by high doses of testosterone undecanoate and ethane dimethane sulfonate inhibit TR4 expression in rat testis, and testosterone induced TR4 expression in co-cultured primary germ/Sertoli cells. Furthermore, we demonstrated that androgen receptor could enhance TR4-mediated transactivation activity in testis cells in the presence of testosterone. Together, these data indicate that the expression of TR4 in rat testis is stage dependent and androgen inductive, and suggest the important role of orphan receptor TR4 in spermatogenesis.     

Testicular function in Klinefelter syndrome

Klinefelter syndrome (KS) is the most common genetic form of male hypogonadism, but the phenotype becomes evident only after puberty. During childhood, and even during early puberty, pituitary-gonadal function in 47,XXY subjects is relatively normal, but from midpuberty onwards, FSH and LH levels increase to hypergonadotropic levels, inhibin B decreases to undetectable levels, and testosterone after an initial increase levels off at a low or low-normal level. Hence, most adult KS males display a clear hypergonadotropism with a varying degree of androgen deficiency; subsequently, testosterone substitution therapy is widely used to prevent symptoms and sequels of androgen deficiency. Testicular biopsies of prepubertal KS boys have shown preservation of seminiferous tubules with reduced numbers of germ cells, but Sertoli and Leydig cells have appeared normal. The testes in the adult KS male are, however, characterized by extensive fibrosis and hyalinization of the seminiferous tubules, and hyperplasia of the interstitium, but the tubules may show residual foci of spermatogenesis. Introduction of testicular sperm extraction in combination with intracytoplasmic sperm injection techniques has allowed non-mosaic KS males to father children.     

Establishment of spermatogenesis in neonatal bovine testicular tissue following ectopic xenografting varies with donor age

Ectopic testicular xenografting can be used to investigate spermatogenesis and as an alternative means for generating transgenic spermatozoa in many species. Improving the efficiency of spermatogenesis in xenografted testicular tissue will aid in the application of using this approach. The present study was conducted to evaluate age-related differences in the establishment of spermatogenesis in grafted testicular tissue from bulls between 2 and 16 wk of life. Testicular tissue was ectopically xenografted under the skin on the backs of castrated nude mice and subsequently evaluated for growth, testosterone production, and establishment of spermatogenesis 24 wk after grafting. The greatest weight increases occurred in donor tissue from calves of the ages 2, 4, and 8 wk compared with the ages of 12 and 16 wk. Recipient mouse serum testosterone concentration was at normal physiological levels 24 wk after grafting and no significant differences were detected between recipients grafted with testicular tissue from bull calves of different ages. The development of germ cells to elongated spermatids were observed in seminiferous tubules of grafts from donor calves of the ages 4, 8, 12, and 16 wk but not observed in grafts from 2-wk donors, which contained round spermatids as the most advanced germ cell stage. Grafts from 8-wk donors contained a significantly higher (10-fold) average percentage of seminiferous tubules with elongated spermatids than all other donor ages. These data demonstrate differences in the ability of testicular tissue from donor animals of different ages to establish spermatogenesis following ectopic testicular xenografting.     

Localization of epidermal growth factor (EGF) and its receptor (EGFR) during postnatal testis development in the alpaca (Lama pacos)

The objective of the present studies was to determine the localization of epidermal growth factor (EGF) and the epidermal growth factor receptor (EGFR) in testicular tissue collected from male alpacas at 12 and 24 months of age. In the testes of 12-month-old alpacas, positive staining for EGF was not detected. EGFR was localized to Leydig cells within the 12-month-old alpaca testis, but staining was absent within seminiferous tubules. At 24 months of age, EGF was localized to Leydig cells, peritubular myoid cells, Sertoli cells and germ cells of the alpaca testis, with a preferential adluminal compartment staining within the seminiferous tubules. EGFR was also localized to the Leydig cells, peritubular myoid cells, Sertoli cells and germ cells within the 24-month-old alpaca testis, but staining within the tubules was primarily within the basal compartment. Results indicate distinct temporal and spatial regulation of EGF and EGFR in the alpaca testis and support a potential role for EGF and its related ligands in alpaca testis development and spermatogenesis.     

Intratesticular androgen levels, androgen receptor localization, and androgen receptor expression in adult rat Sertoli cells

In the rat, quantitatively normal spermatogenesis is maintained only when intratesticular testosterone (ITT) levels greatly exceed the peripheral T concentration. When ITT concentrations fall below a threshold, germ cells are lost at specific stages of the seminiferous cycle. Germ cells can be restored by high doses of T that binds to androgen receptors (AR) in Sertoli cells. However, the relationships between germ cell dynamics, AR-mediated molecular events, and ITT concentrations are not established. ITT levels may regulate germ cell life and death through an effect on AR localization and AR mRNA or protein levels within Sertoli cells at specific stages of the cycle. We determined AR localization and mRNA and protein expression in adult rat Sertoli cells in relation to reduced and then restored ITT concentrations in vivo. ITT levels were reduced by implanting rats with T- and estradiol (E)-filled capsules for 7-28 days and subsequently restored with large T-filled capsules. AR is normally localized within Sertoli cell nuclei at stages VII-VIII of the seminiferous epithelium. After T/E treatment, AR immunostaining in Sertoli cell nuclei became nondetectable by 14-28 days but was restored 6 h following T restoration. The loss of Sertoli cell nuclear AR localization correlated with increasing numbers of apoptotic germ cells. AR mRNA levels in isolated Sertoli cells did not change through 14 days of T/E treatment, increased significantly by Day 28, and remained elevated 24 h after T restoration. AR mRNA levels in microdissected tubules at stages II-IV, VI-VIII, and IX-XII did not decrease through 14 days of T/E treatment. In contrast, AR protein levels were reduced in seminiferous tubules by Day 14 and in testes at Day 28 post-T/E treatment but were restored within 24 h by T repletion. Therefore, the reduction of ITT concentration results in a time-dependent redistribution of AR and reduced AR protein but not AR mRNA levels in Sertoli cells. Repletion of T restored AR protein and it relocated to Sertoli cell nuclei. By an unknown mechanism, T regulates AR localization within Sertoli cells to determine germ cell life or death.     

Response of the seminiferous epithelium of the rat testis to withdrawal of androgen: evidence for direct effect upon intercellular spaces associated with Sertoli cell junctional complexes

The morphological response of the Sertoli cells to partial or complete withdrawal of testosterone was studied in adult rats following hypophysectomy or administration of ethane dimethanesulphonate (EDS), a toxicant known to destroy selectively the Leydig cells of the testis. To assess the role of germ cells in effecting changes to Sertoli cells following withdrawal of testosterone, germ cell-deficient rats with Sertoli-cell-only testes (SCO) were treated with EDS to remove the source of testosterone. At 6 days after hypophysectomy or 4,6 and 8 days after EDS treatment, stage VII and VIII seminiferous tubules showed degenerating germ cells and numerous basally-located vacuoles approximately 1–15 m in diameter. Ultrastructural analysis indicated that most of the vacuoles were multiple focal dilations of the intercellular space associated with Sertoli cell junctional complexes. In SCO rats, treatment with EDS resulted in a significant (P<0.05) increase in the formation of many vacuoles particularly in the base but also in the trunk of the Sertoli cells and again electron microscopic analysis showed multiple, localized expansions of the intercellular space associated with Sertoli cell junctional complexes. The appearance of intercellular spaces in SCO testes following androgen withdrawal cannot be attributed to shrinkage of degenerating germ cells since the seminiferous tubules did not contain germ cells. It is concluded that withdrawal of androgen induces early morphological alterations of the Sertoli cell junctional complexes in which the sites of membrane fusions representing tight junctions remain intact whereas the intercellular spaces exhibit major focal dilations. The results are discussed in relation to the fluid secretion by the seminiferous tubules which is regulated by the Sertoli cells.     

Impairment of spermatogenesis and enhancement of testicular germ cell apoptosis induced by exogenous all-trans-retinoic acid in adult lizard Podarcis sicula

In mammals, retinoic acid is involved in the regulation of testicular function by interaction with two families of nuclear receptors, retinoic acid receptor (RAR) and retinoid X receptor (RXR). Among RAR isoforms, the testicular cells of the lizard were found to express only RARalpha (3.7 kb) and RARbeta (3.4 kb) mRNAs, as reported here. In this study, the effects of exogenous all-trans-retinoic acid (atRA) on spermatogenesis of a non-mammalian seasonal reproducer were investigated. Daily intraperitoneal injections of atRA or atRA plus testosterone (atRA+T) were given for 2 weeks to adult males of the lizard Podarcis sicula. In animals treated with atRA, the seminiferous tubules were markedly reduced in cross-area. The seminiferous epithelium collapse was responsible for a sensible reduction in the number of germ cells and disruption in normal epithelial organization. In comparison, in atRA+T-treated lizards the loss of germinal cells was significantly less. The loss of germ cells observed in both experimental groups results from an induction of apoptotic process, as revealed by TUNEL analysis. Although low in number, apoptotic germ cells were also observed in the control groups (saline- and T-treated lizard), where the main germ cells undergoing apoptosis are primary spermatocytes (most frequently) and some spermatogonia.In conclusion, it is shown here that retinoic acid has deleterious effects on lizard spermatogenesis, causing a severe depletion of seminiferous epithelium, probably via induction of apoptotic processes. These effects are not completely inhibited by simultaneous administration of testosterone, although this hormone, once injected, is able to stimulate spermatogenesis and protect germinal cells from apoptotic cell death.     

The effect of prenatal treatment with busulfan on in vitro androgen production by testes from rats of various ages

Treatment of rats with busulfan in utero severely depletes the germ cell population of the seminiferous tubules. These studies have examined the in vitro capacity of testicular tissue and Leydig cells from such testes to secrete androgens. Leydig cells were identified by staining for 3 beta-hydroxy steroid dehydrogenase. Rats were studied at several ages to identify any developmental changes in the androgen-secreting capacity of control and treated gonads. At 30 days of age, no effect of treatment on serum androgen was found. At 60 and 90 days of age, treatment caused decreased androgen and increased LH content of the serum. At 12, 30, 60, and 90 days of age, the amount of androgen secreted per milligram of testicular tissue in response to LH was higher in busulfan-treated rats. Leydig cells from 60- and 90-day-old rats which had received busulfan were also hyperresponsive to LH. It was concluded that Leydig cells from testes essentially devoid of germ cells were hyperresponsive to LH. Serum androgen levels were decreased yet androgen production per Leydig cell was increased. A possible explanation of this apparent paradox is that busulfan treatment resulted in decreased numbers of Leydig cells in the gonads.     

Regulation of seminiferous tubular function by FSH and androgen.

Seminiferous tubules contain a cytoplasmic androgen receptor similar to the receptors in the epididymis and ventral prostate. The presence of a cytoplasmic receptor indicates that androgens maintain spermatogenesis by a direct action on certain types of cells within the seminiferous tubule. The Sertoli cell appears to be one of the cell types containing androgen receptors and the receptor might also be present in spermatogonia, primary spermatocytes, or peritubular cells. The Sertoli cell is stimulated by FSH to produce an androgen-binding protein which may serve to increase the accumulation of androgen in the seminiferous epithelium and make it available for binding by intracellular androgen receptors. This may be a way in which FSH enhances the action of androgen on spermatogenesis. Androgens act on the Sertoli cell to increase its response to FSH. This action of androgens on the Sertoli cell results in increased production of androgen-binding protein and may enhance the production of other substances which exert trophic effects on spermatogenesis.     

Testosterone and FSH have independent,synergistic and stage-dependent effects upon spermatogenesis in the rat testis

Summary Adult rats were hypophysectomized and treated with ethane dimethanesulphonate (EDS) selectively to eliminate the Leydig cells in the testis. By removing the source of endogenous gonadotrophins and androgens, the subsequent effects on the seminiferous epithelium were studied after 20 days of treatment with vehicle, or FSH (2x50 g/day) or a low dose of testosterone (0.6 mg testosterone esters every 3rd day) alone or in combination. Compared to vehicle-treated hypophysectomized rats with Leydig cells, testis weight in saline-treated hypophysectomized rats treated with EDS declined by 50%, spermatogenesis was disrupted severely and only 18% of the tubules contained spermatids, these being confined to stages I–VI of the spermatogenic cycle. Treatment with either FSH or testosterone esters alone significantly (P<0.01) increased testis weight compared to vehicle-treated hypophysectomized rats treated with EDS and 40% of tubules contained spermatids either at stages I–VI after FSH, or at all stages I–XIV after testosterone treatment. Treatment with FSH and testosterone esters together maintained testis weights approximately 20% above vehicle-treated hypophysectomized controls; over 70% of the seminiferous tubules contained spermatids and there was a marked stimulation of spermatogenesis at all stages of the spermatogenic cycle. The results suggest, that in the absence of the pituitary gland and the Leydig cells, FSH alone partially supports spermatogenesis up to the development of round spermatids whereas testosterone is capable of maintaining spermatid development at all 14 stages of the cycle. When FSH and testosterone were administered in combination, the effects upon spermatogenesis were far greater than the response expected if their individual effects were simply additive. It is therefore concluded that FSH may play a role in normal spermatogenesis and that this role is essentially that of augmenting the response of the testis to testosterone. The biochemical mechanisms via which this might occur are discussed and hypophysectomized rats treated with EDS used in the present studies should provide a useful approach for their identification.     

Androgen receptor in nuclei of rat testis.

Testis nuclei of hypophysectomized rats selectively accumulate labeled testosterone and 5alpha-dihydrotestosterone following the injection of tritiated testosterone in vivo. Testosterone and 5alpha-dihydrotestosterone are bound to macromolecules in nuclei and can be extracted with 0.5 M KCl. Accumulation of protein bound radioactive androgens in nuclei of isolated seminiferous tubules is similar to that of whole testis. The relative amounts of testosterone and dihydrotestosterone in purified nuclei were similar to the relative amounts bound to cytoplasmic receptors, suggesting that cytoplasmic androgen-receptor complexes may be transported into the nuclei. Binding of labeled androgen is saturable and inhibited by prior injection of unlabeled testosterone or cyproterone acetate. Nuclear binding sites are destroyed by the proteolytic enzyme pronase, but not by DNase. Like the cytoplasmic androgen-receptor complexes in rat testis, nuclear androgen-protein complexes are heat labile and dissociate slowly at 0 degrees C. androgens fail to accumulate in testis nuclei of the Stanley-Gumbreck androgen insensitive rat, a species lacking cytoplasmic androgen receptors in testis and other androgen target tissues.     

Restoration of spermatogenesis in infertile mice by Sertoli cell transplantation

The niche is considered to play an important role in stem cell biology. Sertoli cells are the only somatic cells in the seminiferous tubule that closely interact with germ cells to create a favorable environment for spermatogenesis. However, little is known about how Sertoli cells develop to form the male germ line niche. We report here that Sertoli cells recovered and dissociated from testes of donor male mice can be microinjected into recipient testes, form mature seminiferous tubule structures, and support spermatogenesis. Sertoli cells from perinatal donors had a dramatically greater capacity for generating seminiferous tubules than those from adult donors. Furthermore, transplantation of wild-type Sertoli cells into infertile Steel/Steel(dickie) testes created a permissive testicular microenvironment for generating spermatogenesis and spermatozoa. Thus, our results demonstrate that the male germ line stem cell niche can be transferred between animals. In addition, the technique provides a novel tool with which to analyze spermatogenesis and might provide a mechanism for correcting fertility in males suffering from supporting cell defects.