II. Characterization and development of the regional- and cellular-specific abnormalities in the epididymis of mice with beta-hexosaminidase A deficiency

Beta-hexosaminidase (Hex) is a lysosomal enzyme that exists as two isoenzymes: Hex A (subunit structure alphabeta) and Hex B (betabeta). Its presence in the testis and epididymis suggests important roles for Hex and its substrates in male fertility and reproductive functions. Disruption of the Hexa gene encoding the alpha-subunit of Hex has led to the generation of a mildly affected mouse model of human Tay-Sachs disease, allowing us the opportunity to analyze the effects of isolated Hex A deficiency on epithelial cellular morphology of the male reproductive tract. At 5 weeks and at 3, 5, and 12 months, the testes, efferent ducts and epididymides of Hex A-deficient (Hexa -/-) and wild-type (Hexa +/+) mice were perfuse fixed and analyzed by routine light and electron microscopy as well as with immunocytochemistry employing antibodies to lysosomal enzymes. In the testis, the seminiferous epithelium of Hexa -/- mice appeared comparable to that of wild-type mice in appearance and topographical arrangement of its cell types at all ages examined. Also, no differences were noted for the efferent ducts. In contrast, there were striking abnormalities in the epididymides of the mutant mice; however, the abnormalities were mainly restricted to the initial segment and intermediate zone. Principal cells of these regions at 5 weeks showed a dramatic increase in the number of lysosomes as compared with those from wild-type animals, and this progressed with increasing age. Furthermore, unlike the few small lysosomes present in wild-type mice, those of Hexa -/- mice were at times enlarged and often filled the supranuclear and basal regions of these cells. In the light microscope, large, dense cellular aggregates were noted at the base of the epithelium in the proximal initial segment that corresponded in the electron microscope to two different cell types, both of which increased in size with age. One aggregate was considered to belong to narrow cells on the basis of the presence of numerous cup-shaped vesicles characteristic of these cells; they appeared to be dislocated from the upper half of the epithelium. In the distal initial segment and intermediate zone, narrow cells were readily identified, but rather than being slender as in the control animals, they were greatly enlarged and filled with pale lysosomes in mutant mice. The second type of cellular aggregate noted in the proximal initial segment corresponded to halo cells. They contained numerous small and large lysosomes and small, Golgi-related, dense, core granules characteristic of halo cells. On the basis of the large size of these cells, they appeared to be actively internalizing substances from the intercellular space. In contrast, principal and clear cells of the caput, corpus, and cauda regions did not appear to show a significant increase in number or size of lysosomes as compared with those of wild-type animals. All structures identified as lysosomes in the various cell types were immunoreactive for cathepsin D. The present data thus reveal that isolated Hex A deficiency results in region- and cell-specific abnormalities in the epididymis but in no apparent abnormalities in the testis or efferent ducts. Specific roles for Hex A that cannot be compensated for by other isozymes of Hex appear to exist within lysosomes of epithelial cells predominantly of the initial segment and intermediate zone. Taken together, the results also suggest that the inability to degrade endocytosed substrates normally acted upon by Hex A in lysosomes of principal and narrow cells leads to their accumulation, eventual fusion, and increased size.     

Alterations in the testis and epididymis associated with loss of function of the cystatin-related epididymal spermatogenic (CRES) protein

Cystatin-related epididymal spermatogenic protein (CRES) or cystatin 8 (Cst8 gene) is a member of the cystatin superfamily of cysteine protease inhibitors. It differs from typical cystatins because it lacks consensus sites for cysteine protease inhibition and exhibits reproductive-specific expression. In the present study, we examined CRES expression within the testes, efferent ducts, and epididymides of normal mice by light microscope immunolocalization. Alterations to these tissues in male mice lacking the Cst8 gene (Cst8(-/-2)) were also characterized by histomorphometry and electron microscopy. In the normal testis, CRES was localized exclusively in mid and late elongating spermatids. In the efferent ducts, CRES was localized to the apical region of the epithelial cells suggestive of localization in the endosomes. In the initial segment of the epididymis, principal cells showed supranuclear and luminal reactions. In the cauda region, CRES was present exclusively as aggregates in the lumen and was detected in clear cells. Compared with wild-type mice (Cst8(+/+)), older (10-12 months) Cst8(-/-) mice had modest but statistically significant reductions in tubular, epithelial, and/or luminal profile areas in the testis and epididymis. By electron microscopy, some Cst8(-/-) tubules in the testis were normal in appearance, but others showed a vacuolated seminiferous epithelium, degenerating germ cells, and alterations to ectoplasmic specializations. In the epididymal lumen, abnormally shaped sperm heads and tails were noted along with immature germ cells. In addition, principal cells contained numerous large irregularly shaped lysosomes suggestive of disrupted lysosomal functions. In both the testis and epididymis, however, these abnormalities were not apparent in younger mice (4 months), only in the older (10-12 months) Cst8(-/-) mice. These findings suggest that the altered testicular and epididymal histology reflects a cumulative effect of the loss of CRES and support a role for CRES in maintaining the normal integrity and function of the testis and epididymis.     

Expression and regulation of aquaporins 1, 8, and 9 in the testis,efferent ducts,and epididymis of adult rats and during postnatal development

Aquaporins (AQPs) are membrane protein channels that allow the rapid passage of water through an epithelium containing tight junctions. In the present study, light microscope immunocytochemistry was utilized to localize several members of the AQP family in the testis, efferent ducts, and epididymis of normal adult animals during postnatal development and after various experimental procedures on adult animals. In the testis of normal adult animals, AQP-8 was expressed exclusively in Sertoli cells, while AQP-9 outlined Leydig cells. In the efferent ducts, AQP-1 was expressed on the microvilli, basolateral plasma membranes, and apical endosomes of the nonciliated cells and cilia of ciliated cells, while AQP-9 was present only on the microvilli of nonciliated cells. In the epididymis, AQP-9 was localized to the microvilli of the principal cells of all regions, with the most intense reaction being noted in the initial segment and cauda regions. The clear cells of the cauda region expressed only AQP-9. AQP-1 was not expressed in the testis or the epididymal epithelium, but it was expressed over the endothelial cells of the vascular channels of the efferent ducts and epididymis. After efferent duct ligation or orchidectomy, there was no change in the expression of AQP-1 or -9 over the microvilli or cilia of epithelial cells in the case of the efferent ducts, suggesting that testicular factors do not regulate their expression in this region. In contrast, AQP-9 expression in the principal cells of the initial segment, but not of other regions, and also in the clear cells of the cauda region was dramatically reduced after both treatments. As the expression was not restored to control levels by testosterone replacement, the data suggest that a luminal factor(s) derived from the testis regulates AQP-9 expression in the principal cells of the initial segment and in the clear cells of the cauda region. Postnatal studies revealed that the expression of AQP-1 and -9 in the different cell types of the efferent ducts and epididymis occurred between days 7 and 29, eliminating sperm and high androgen levels as possible regulating factors. Taken together, these data suggest cell specificity with respect to the expression of AQP-8 and -9 in the testis. In the efferent ducts and epididymis, specificity exists in cell, region, and tissue distribution with respect to the expression of AQP-1 and -9, and their expression does not appear to be regulated by androgens.     

Postnatal development and regulation of beta-hexosaminidase in epithelial cells of the rat epididymis

beta-Hexosaminidase (Hex) catalyzes the hydrolysis of terminal sugar residues from a number of substrates such as GM2 gangliosides, glycoproteins, glycolipids, and glycosaminoglycans. As an enzyme present in lysosomes of epithelial cells of the adult rat epididymis, it serves to degrade substances endocytosed from the epididymal lumen. In this way, it modifies and creates a luminal environment where sperm can undergo their maturational modifications. In this study, the postnatal developmental pattern of expression of Hex was examined in animals from days 7-56. In addition, the role of testicular factors on Hex expression in the different cell types and regions of the epididymis of adult rats was examined in orchidectomized and efferent duct-ligated rats. Both parameters were examined on Bouin-fixed epididymides in conjunction with light microscope immunocytochemistry. At postnatal day 7, the epithelium of the entire epididymis was unreactive for anti-Hex antibody. By day 21, narrow and clear cells of their respective regions became reactive, whereas basal cells became reactive only by day 29. Principal cells displayed only an occasional reactive lysosome at day 21, several by day 29, and numerous reactive lysosomes by day 39, comparable to the region-specific distribution noted for 90-day-old animals, and at an age when high androgen levels are attained. Thus, postnatal onset of Hex expression varies according to the different cell types of the epididymis, suggesting different regulatory factors. This finding was confirmed from studies employing adult orchidectomized and efferent duct-ligated adult rats. Indeed, in all experimental animals, Hex immunostaining in narrow, clear, and basal cells was intense and comparable to control animals. In contrast, there was a notable absence of lysosomal staining in principal cells at all time points after orchidectomy, which was restored, however, following testosterone replacement. No effect on Hex expression was observed in efferent duct-ligated animals. Taken together, the data suggest that Hex expression in lysosomes of principal cells is regulated by testosterone or one of its metabolites. However, the expression of Hex being independent of testicular factors in narrow, clear, and basal cells of adult animals, but occurring at different time points during postnatal development, suggests that different regulatory factors are responsible for onset of Hex expression in these cell types during development.     

一氧化氮合酶在食蟹猴睾丸及附睾中的表达及意义

目的:探索一氧化氮合酶(NOS)在食蟹猴睾丸及附睾中的表达及意义。方法:运用免疫组化染色法观察NOS在8只猴龄为8岁左右性成熟食蟹猴睾丸及附睾中的分布。结果:①神经元型NOS(nNOS)免疫反应阳性见于睾丸生精小管上皮内各级生精细胞、腔内精子、附睾输出小管上皮、血管内皮细胞。②诱导型NOS(iNOS)免疫反应阳性见于附睾输出小管上皮、腔内精子、管周类肌细胞、血管内皮细胞。③内皮型NOS(eNOS)免疫反应阳性见于睾丸间质细胞、附睾输出小管上皮、腔内精子、管周类肌细胞、血管内皮细胞。结论:NOS广泛表达于性成熟食蟹猴睾丸及附睾组织细胞中,推测其在参与精子发生、成熟及睾丸激素分泌等过程中起到重要作用。     

Cell specificity of aquaporins 0, 3, and 10 expressed in the testis, efferent ducts, and epididymis of adult rats

Aquaporins (AQPs) are transmembrane protein channels that allow the rapid passage of water across an epithelium at a low energy requirement, though some also transport glycerol, urea, and solutes of various sizes. At present, 11 members of the AQP family of proteins have been described in mammals, with several being localized to the testis (AQP-7 and AQP-8), efferent ducts (AQP-1 and AQP-9), and epididymis (AQP-1 and AQP-9) of adult rats. With the discovery of expression of multiple AQPs in different tissues, we undertook a systematic analysis of several other members of the AQP family on Bouin-fixed tissues of the male reproductive tract employing light microscope immunocytochemistry. In the testis, AQP-0 expression in the seminiferous epithelium was restricted to Sertoli cells and to Leydig cells of the interstitial space; no reaction was observed in the efferent ducts or epididymis. In Sertoli cells, a semicircular pattern of staining was noted, with only one fourth or one half of the Sertoli cells of a given tubule showing a reaction product. Furthermore, while Sertoli cells at stages VI-VIII of the cycle showed intense staining, those at stages IX-XIV were least reactive, with Sertoli cells at stages I-V showing intermediate levels of reaction product. The epithelial expression of AQP-10 was restricted to the microvilli of the nonciliated cells and the cilia of the ciliated cells of the efferent ducts; however, the endothelial cells of vascular channels of the efferent ducts and epididymis were also intensely reactive. AQP-3 expression was localized exclusively to the epididymis, where intense staining was noted exclusively over basal cells. Examination of orchidectomized rats revealed that AQP-3 expression was abolished over basal cells and that it was greatly diminished after efferent duct ligation. As the reaction was not fully restored in orchidectomized animals supplemented with high levels of testosterone, we suggest that AQP-3 expression in basal cells is regulated in part by testosterone, in addition to a luminal factor emanating from the testis. Together, the data indicate a cell- and tissue-specific expression for AQP-0, AQP-3, and AQP-10 in the testis, efferent ducts, and epididymis, as well as differential regulating factors for the expression of AQP-3 in basal cells.     

Epididymal growth and differentiation are altered in human cryptorchidism

Despite the knowledge and histological classification of testicular lesions, epididymal lesions associated with cryptorchidism are not well defined and only macroscopic alterations have been reported. We have evaluated the alterations in the growth of both the epithelium and muscular wall of efferent ducts and epididymis in human patients with cryptorchidism from infancy to adulthood. In addition, by cytokeratin immunostaining we have also evaluated the stage of differentiation of each segment along the human postnatal life in these patients. A decrease is shown in the size of efferent and epididymal ducts in cryptorchid children compared with normal, age-matched controls. The height of the epithelium, muscular wall, and lumen of the cryptorchid epididymis were reduced at every age studied. This decrease in all regions was seen even in the testicular quiescent period (1 to 4 years of age). In addition, the cryptorchid epididymis grows more slowly during the transition to the pubertal period. The smaller size of the cryptorchid epididymis in pubertal and adult men compared with that of normal men is due primarily to underdevelopment of the muscular wall and a reduction in epithelial height. The pattern of growth of cryptorchid efferent ducts and ductus epididymides parallels that in normal men, except that development of the lumen and muscular layer in the cauda epididymis region are delayed. Epithelial differentiation, monitored by cytokeratin expression, is minimal in efferent ducts and throughout the epididymis of the cryptorchid male, and this is already seen in children. In conclusion, our immunohistochemical and morphometric results show a reduced development of the human cryptorchid epididymis that is already evident in childhood. They indicate that cryptorchidism is a primary congenital illness of the testis and spermatic ducts, with evident lesions from the first years of life, and suggest that surgical descent would probably not be able to completely reverse these alterations.     

Morphologic changes in efferent ductules and epididymis in estrogen receptor-alpha knockout mice

Estrogen has been shown to have an important role in fluid reabsorption in efferent ductules of the testis. Our previous study of the estrogen receptor-alpha knockout mouse (ERKO) showed that the efferent ductules and rete testis were primary targets of estrogen receptor function. In the present study, a more comprehensive evaluation of the ERKO male reproductive tract was performed to determine the severity of effects in efferent ductules as well as the epididymis. The following observations were found in ERKO males: 1) blind-ending efferent ductules were more prevalent in ERKO than in wild type (WT) tissues; 2) glycogen-containing cells were observed at the rete testis-efferent ductule junction; 3) the tubular diameters of the efferent ductules and initial segment epididymides were dilated; 4) efferent ductules were dilated between 130 to 300% over wild type ductules; 5) efferent ductule epithelial height was reduced nearly 50%; 6) microvilli of nonciliated cells of efferent ductules were 64% shorter in length; 7) cilia were reduced in number; 8) initial segment epithelium was displaced into regions adjacent to the rete testis and in short segments of the common region of efferent ductule; 9) apical, narrow, and clear cells of the epididymis also were abnormal in some regions; 10) in the corpus and cauda regions, sperm granulomas were noted in one third of the ERKO males. In conclusion, the entire reproductive tract is affected in ERKO males. The cells showing the greatest effects were estrogen receptor-positive cells. It appears that in the ERKO mouse there are developmental anomalies that must be considered separately from adult dysfunctional changes in the male reproductive tract.     

Degeneration of the seminiferous epithelium with ageing is a cause of spermatoceles?

A spermatocele refers to the cystic accumulation of semen in the male reproductive tract. Although it is thought to be caused by narrowing of the lumen of the excurrent duct with resultant cystic dilatation of the duct, the pathogenesis of the narrowing remains unknown. In the present study, we histologically examined spontaneous spermatoceles in C3H/He mice to elucidate the pathogenesis of the lesions. Testes, efferent ducts, epididymides and vas deferens obtained from young and aged C3H/He mice were embedded in plastic for histological observation at the light microscopic level. It was found that spontaneous spermatoceles were localized in the rete testis and efferent ducts of aged mice, as seen in man. The dilated rete testis and efferent ducts contained many degenerated and aggregated germ cells derived from the exfoliated seminiferous epithelium in the aged testis. In particular, it was noted that the agglutinated germ cells obstructed the narrow lumen of the efferent ducts, resulting in the failure of transport of germ cells to the caput epididymis, and spermatoceles were consistently found in the region between the rete testis and the obstructed site in the efferent ducts. However, no inflammatory cell infiltration, traumatic injury or spermatic granulomas were found in the occluded region. These results suggest that agglutinated germ cells may occupy the narrow lumen of the efferent ducts, resulting in the formation of a spermatocele. It may be that a senile change to the seminiferous epithelium, which releases immature germ cells into the lumen of the seminiferous tubules, is the cause of this type of spermatocele.     

Targeted disruption of the mouse prosaposin gene affects the development of the prostate gland and other male reproductive organs

The prosaposin gene encodes a 65-70 kilodalton (kd) protein, which is secreted or targeted to lysosomes. In lysosomes, prosaposin is the precursor of 4 activator proteins, designated saposins A, B, C, and D, which promote by acidic hydrolases, the degradation of glycosphingolipids with short oligosaccharide chains. Mutations of the prosaposin gene have been linked to several lysosomal storage disorders. An animal model was recently developed by creating a null allele in embryonic stem cells through gene targeting in order to investigate the phenotypic diversity of prosaposin mutations, the involvement of this protein in lysosomal storage diseases, and to develop potential therapeutic approaches. Mutant homozygous mice die at 35-40 days of age and neurological disorders contribute to their early death. Secreted prosaposin is present in milk and in cerebrospinal and seminal fluids. In the nervous system, prosaposin exhibits a trophic activity. Examination of reproduc-tive organs in homozygous mutant males shows several abnormalities such as a decrease in testis size with reduced spermiogenesis, and an involution of the prostate, seminal vesicle, and epididymis, although levels of testosterone in blood remain normal. In the prostate of homozygous mutants, only basal cells appear to be present, whereas secretory cells are absent. The epithelia in efferent ducts is formed by ciliated cells, whereas heterozygotes exhibit a majority of nonciliated cells. Our data indicate that prosaposin is involved in the development and maintenance of male reproductive organs. In prostatic epithelium, targeted disruption of the prosaposin gene appears to inactivate the mitogen-activated protein kinase pathway and to interfere with differentiation of secretory cells.     

Regional variation in macrophage antigen expression by murine epididymal basal cells and their regulation by testicular factors

Factors controlling the appearance in the epididymis of basal cells and their expression of macrophage antigens were examined by ligating the efferent ducts to prevent the entry of spermatozoa in adult and juvenile mice. Fixation and antigen retrieval techniques were developed to preserve tissue morphology and expression of two macrophage antigens in paraffin-embedded epididymal tissue. A combination of periodate-lysine-phosphate fixation, low-temperature embedding and enzyme predigestion of sections permitted immunohistochemical detection of the mature macrophage antigen Mac-1, whereas the panmacrophage marker F4/80 required fixation in neutral-buffered formalin. Epididymal basal cells were immunostained for F4/80 and quantified with avidin-biotin-peroxidase. In the adult mouse, the total number of basal cells per millimeter length of tubule cross section perimeter and the percentage expressing the F4/80-antigen were significantly higher in the initial segment and caput region than in all other epididymal regions. In the initial segment, immunostained basal cells surrounded the tubule in a network, and some extended towards the lumen. Ligation of the efferent ducts to prevent inflow of testicular secretions significantly reduced the number of basal cells per cross section in the initial segment of the adult and juvenile; the percentage of basal cells expressing macrophage antigens in the initial segment and the caput epididymidis was also reduced. Since basal cells still appeared in the ligated postpubertal epididymis, it is concluded that testicular exocrine secretions entering the epididymal lumen around puberty are not the major influence on basal cell appearance in the murine epididymis, but they may modulate their expression of macrophage antigens.     

Thirsty business: cell, region, and membrane specificity of aquaporins in the testis, efferent ducts, and epididymis and factors regulating their expression

Water content within the male reproductive tract is stringently regulated in order to promote sperm differentiation and maturation. Aquaporins (AQP) are a family of integral membrane proteins allowing the transcellular transport of water, gases, urea, glycerol, and ions. Past studies from our lab have revealed the following. In the testis, Sertoli cells express AQP 8, whereas germ cells express AQP 7. In the efferent ducts (ED), AQP 1, 9, and 10 localize to microvilli of nonciliated cells, in addition to a basolateral staining for AQP 1, whereas AQP 1 and 10 localize to ciliated cells. AQP 7 and 11 are expressed in the ED epithelium of young but not adult rats, suggesting suppression of translation as rats age. In the adult epididymis, AQP 1 appears in endothelial cells of vascular channels and myoid cells, whereas AQP 3 delineates basal cells. In principal cells, AQP 9 and 11 appear on microvilli, whereas AQP 7 localizes to lateral then to basal plasma membranes in a region-specific manner; AQP 7 also associates with myoid cells. AQP 5 is expressed in corpus and cauda regions. Additionally, several AQPs are expressed by some but not all basal (AQP 7, 11), clear (AQP 7, 9), and halo (AQP 7, 11) cells. Regulation studies reveal a role for estrogen, androgens, and lumicrine factors. These findings indicate unique associations of AQPs with specific membrane domains in a cell type- and region-specific manner within the EDs and epididymis, as well as complex regulation patterns of expression.     

Stage and region-specific localization of lipocalin-type prostaglandin D synthase in the adult murine testis and epididymis

Lipocalin-type prostaglandin D synthase in semen has been associated with male fertility, although this relationship is not well defined. To gain insight into potential mechanisms, the objective of the present study was to immunocytochemically localize lipocalin-type prostaglandin D synthase within the testis, efferent ducts, and 4 segments of mouse epididymis. In the testis, immunoperoxidase staining was localized within the Sertoli cells only at stages VI-VIII of the spermatogenic cycle, which is just prior to spermiation. Intense staining was also evident throughout the interstitial tissue, including Leydig cells. The entire epithelium of the efferent ducts, including ciliated and nonciliated cells, was immunoreactive. A distinct pattern of immunostaining for lipocalin-type prostaglandin D synthase was observed in different regions of epididymis, suggesting a possible role in sperm maturation. Staining for lipocalin-type prostaglandin D synthase was strikingly absent in the initial segment. In caput epididymidis, staining was evident throughout the cell cytoplasm of principal cells with some cells more intensely stained than adjacent ones. In the corpus region, overall staining intensity decreased and appeared to be concentrated in the apical region of principal cells, but some cells were completely unreactive. Reaction product in the cauda region was heavily concentrated on microvilli and within the epididymal lumen. In all epididymal regions, expression of lipocalin-type prostaglandin D synthase was specific to epithelial principal cells; no immunoreactivity was apparent in other cell types. The specific localization of lipocalin-type prostaglandin D synthase within the testicular interstitial tissue, Sertoli cells, and principal cells of caput epididymidis strongly suggests that this protein plays an integral role in both the development and maturation of sperm.     

Prosaposin ablation inactivates the MAPK and Akt signaling pathways and interferes with the development of the prostate gland

The recent development of a prosaposin-/-mouse model has allowed the investigation of the role of prosaposin in the development of the male reproductive organs.A morphometric analysis of the male reproductive system of 37 days old mice revealed that prosaposin ablation produced a 30% reduction in size and weight of the testes,37% of the epididymis,75% of the seminal vesicles and 60% of the prostate glands.Light microscopy(LM) showed that smaller testis size from homozygous mutant mice was associated with reduced spermiogenesis.Both,dorsal and ventral lobules of the prostate glands were underdeveloped in the homozygous mutant.LM analysis also showed that prostatic alveoli were considerably smaller and lined by shorter epithelial cells in the homozygous mutant.Smaller tubular diameter and shorter undifferentiated epithelial cells were also observed in seminal vesicles and epididymis. In the efferent ducts of the homozygous mutant mice,the epithelium was composed exclusively of ciliated cells in contrast to the heterozygotes,which showed the presence of nonciliated cells.Radioimmunoassays demonstratedthat testosterone levels were normal or higher in mice with the inactivated prosaposin gene.Immunostaining of prostate sections with an anti-androgen receptor antibody showed that the epithelial cells lining the alveoli express androgen receptor in both the heterozygous and homozygous tissue.Similarly,sections immunostained with antibodies to the phosphorylated MAPKs and Akts strongly reacted with tall prostatic secretory cells in prostate from heterozygous mouse.On the other hand,the epithelial cells in the homozygous prostate remained unstained or weakly stained.These findings demonstrate that inactivation of the prosaposin gene affected the development of the prostate gland and some components of the MAP pathway.     

Localization of androgen and estrogen receptors in adult male mouse reproductive tract

There is considerable variation, both within and between species, in reports of nuclear steroid receptor localizations in the male reproductive tract. In this study, androgen receptor (AR) and estrogen receptors ERalpha and ERbeta were visualized by immunohistochemistry in adult male mice reproductive tracts, including testes, efferent ductules; initial segment, caput, corpus, and cauda epididymides; and vas deferens. Antibody specificity was demonstrated by Western blot and antibody competition. In testis, AR was expressed in Leydig cells, Sertoli cells, and most peritubular cells, but not in germ cells; Sertoli cells showed more intense staining in stages VI-VII; ERalpha was present in Leydig and some peritubular cells; ERbeta was in Leydig, some peritubular, all Sertoli and germ cells except in spermatids and meiotic spermatocytes. In efferent ductules, AR was strongly expressed in ciliated and nonciliated epithelial cells and in stromal cells; ERalpha was strongly expressed in ciliated and nonciliated epithelial cells; stromal cells were negative; and ERbeta was strongly expressed in ciliated and nonciliated epithelial cells and also in stromal cells. In epididymis, AR was strongly expressed in all epithelial cells (not in intraepithelial lymphocytes); ERalpha was strongly expressed in apical, narrow, and some basal cells of the initial segment, and in caput, principal cells of the caput, clear cells of the distal caput through cauda; stromal cells were negative in the initial segment, but more stromal cells were stained from caput to cauda; ERbeta was strongly expressed in most of epithelial cells of the epididymis, but stromal cells were inconsistently stained. In vas deferens, AR was weakly expressed or absent in principal cells but moderately stained in basal cells, smooth muscle cells of stroma were stained intensely, ERalpha was absent in epithelial cells but present in a subepithelial smooth muscle layer, and ERbeta was strongly expressed in all epithelial cells and most stromal cells. This study demonstrates that the reproductive tracts of male mice differ considerably from those of rats in expression of ARs and ERs and that caution is needed when extrapolating nuclear steroid receptor data across mammalian species.     

Role of prosaposin in the male reproductive system: effect of prosaposin inactivation on the testis, epididymis, prostate, and seminal vesicles

SGP-1/prosaposin can be secreted or targeted to the lysosomes where it is processed into smaller saposins (A, B, C, and D) required for the hydrolysis of glycosphingolipids. The deficiency of saposins B and C results in variant forms of metachromatic leukodystrophy and Gaucher's disease, respectively, which are characterized by lysosomal storage of undegraded glycosphingolipids. In the nervous system, prosaposin presents trophic activity. A mouse model was recently developed by creating a null allele in embryonic stem cells through gene targeting to investigate the phenotypic diversity of prosaposin mutations and the involvement of this protein in lysosomal storage diseases, and for the development of therapeutic approaches. Mice homozygous mutants die at the age of 35-40 days and neurological disorders contribute to the early demise of the mutant mice. The male reproductive organs in homozygous mutants show several abnormalities, such as a decrease in testis size with reduced spermiogenesis and an involution of the prostate, seminal vesicles, and epididymis. In these animals, the blood levels of testosterone remain normal. In the prostate of homozygous mutants, only the basal epithelial cells appear to be present, while the secretory cells are absent. These findings suggest that prosaposin may be involved in the development and maintenance of the male reproductive organs, as well as, in cellular differentiation.     

A morphological study of the efferent ducts of the human epididymis

Efferent ducts of the human epididymis were studied with a light microscope and a computerized three-dimensional image analyser by preparing complete serial sections. The efferent ducts were characterized by a columnar epithelium, which differed both from the cuboidal epithelium of the rete testis and from the tall columnar epithelium of the epididymal duct. Therefore, the junctions of the efferent ducts with the rete testis and epididymal duct could be identified morphologically. Five or six efferent ducts originated from the dilated extra-testicular rete testis, ran in an extremely tortuous manner and transformed into the epididymal ducts in an end-to-end pattern near the border of the epididymal head and corpus. Computerized image analysis confirmed light microscopical findings and demonstrated three-dimensional structures of the junctions of the efferent ducts with both the rete testis and the epididymal ducts.     

Differentiation and maturation of excurrent duct system of rat testes]

The present study was designed to investigate relations of gamma-Glutamyl transpeptidase (gamma-GTP) activities to morphological differentiation and maturation of rat testicular excurrent duct system including rete testis, efferent ductuli and epididymal ducts. Animals used were Wistar rats aged from 3 days to 12 weeks. Histochemical demonstration of gamma-GTP activities was carried out by the method of Rutenburg et al. (1969). Morphological differentiation of the epithelium and structure of the rete testis occurred between 2 and 3 weeks and completed by the age of 6 weeks, which was accompanied by induction and deletion of gamma-GTP activity. Morphological differentiation and maturation of the epithelium of the efferent ductuli and the epididymal ducts became evident by the age of 4 weeks and gamma-GTP was active in both epithelial cells throughout the period examined. Since the maturation of rete testis epithelium was found to proceed as observed as Sertoli cells, the origin and functions of the rete testis epithelium and Sertoli cells are considered to be identical. The simultaneous development of the efferent ductuli and the epididymal ducts suggests that both tissues originate from mesonephric ducts and have similar functions, although mature epithelial cells of both tissues are completely different.     

Attractin蛋白在大鼠睾丸和附睾中的免疫组织化学定位

目的 :探讨Attractin蛋白在成熟大鼠睾丸和附睾组织中的分布。　方法 :健康成年雄性SD大鼠 2 0只 ,灌流取睾丸和附睾组织固定 ,石蜡包埋和冰冻切片。用免疫组化法和间接免疫荧光方法检测Attractin蛋白在成熟大鼠睾丸和附睾组织中的表达。　结果 :睾丸组织间质细胞、睾丸精曲小管管周肌样细胞和各级生精细胞 (精原细胞、初级精母细胞和精子细胞 )、支持细胞呈阳性反应 ,主要表达于胞膜及胞质。睾丸间质细胞表达略强于生精细胞。附睾头、体、尾均未见表达。　结论 :Attractin蛋白在成熟大鼠睾丸组织间质细胞和生精细胞中有较强的表达 ,其生理功能尚有待进一步探讨。     

pH and bicarbonate in the ductuli efferentes testis of the rat

The pH and bicarbonate concentrations of luminal fluids in the efferent ducts of the rat were estimated from pH measurements of samples in vitro under conditions of controlled temperature and carbon dioxide tension. The pH of scrotal blood was estimated to be more acidic than systemic blood (mean pH=7.44) at either of the putative scrotal carbon dioxide tensions (5% and 7%, pH, respectively,=7.42 and 7.28). For PCO2 tensions of 5% and 7%, respectively, the data indicated that the pH in the efferent ducts was significantly higher (distal initial zone pH=7.55 or 7.41; coni vasculosi pH=7.66 or 7.51; p < 0.01) than in fluid entering (rete testis fluid, pH=7.34 or 7.20) or leaving the ducts (zone 1a of the epididymal duct 7.26 or 7.11). Bicarbonate concentrations were also significantly higher (p < 0.01) in the efferent ducts (35.4 +/- 4.7 mM, distal initial zone; 45.2 +/- 7.6 mM, coni vasculosi) than in fluids entering (22.9 +/- 3.6 mM) or leaving (20.4 +/- 4.9 mM) the ducts. Estimates of the reabsorption of bicarbonate and fluid indicated that 96% of the testicular output of bicarbonate was reabsorbed in the efferent ducts, but there was also some secretion of bicarbonate into the ducts. It is concluded that luminal pH and bicarbonate levels in the efferent ducts of the rat are high relative to those found in the epididymis where low pH and bicarbonate contributes to sperm quiescence during storage. Nevertheless, the high rate of bicarbonate reabsorption in the efferent ducts is a major contributor to the establishment of the low pH and bicarbonate milieu of the epididymis.