Experimental cryptorchidism in the adult mouse: I. Qualitative and quantitative light microscopic morphology

Morphologic changes in the testes of adult mice after experimentally induced cryptorchidism were studied by light microscopy and stereology. Increasing duration of cryptorchidism resulted in a gradual decrease in the volume of seminiferous tubules per testis, and this was associated with germ cell degeneration. The volumes of Sertoli cell lipid droplets increased, and dilations of the intercellular space between the Sertoli cell junctions was observed in the cryptorchid testis. The luminal volume of the seminiferous tubule was reduced by 50% after 28 days of cryptorchidism. However, the volumes of intertubular tissue and Leydig cells in control and cryptorchid testes were not significantly different. Leydig cell number per testis increased, and the average volume of a Leydig cell decreased gradually with the progression of the cryptorchid state. The volume of the connective tissue cells in the intertubular area increased, but no significant volume change was observed in the volume of intertubular macrophages. After 28 days, the cryptorchid testis contained a significantly increased volume of blood vessels and a reduced volume of lymphatic space per testis. These observations clearly demonstrate that, although the mouse is a species closely related to the rat, the morphologic changes that occur in the Leydig cell population after induction of experimental cryptorchidism in this species is different.     

A mouse model of spinal and bulbar muscular atrophy

Spinal and bulbar muscular atrophy (SBMA) is an adult-onset motor neuron disease, caused by the expansion of a trinucleotide repeat (TNR) in exon 1 of the androgen receptor (AR) gene. This disorder is characterized by degeneration of motor and sensory neurons, proximal muscular atrophy, and endocrine abnormalities, such as gynecomastia and reduced fertility. We describe the development of a transgenic model of SBMA expressing a full-length human AR (hAR) cDNA carrying 65 (AR(65)) or 120 CAG repeats (AR(120)), with widespread expression driven by the cytomegalovirus promoter. Mice carrying the AR(120) transgene displayed behavioral and motor dysfunction, while mice carrying 65 CAG repeats showed a mild phenotype. Progressive muscle weakness and atrophy was observed in AR(120) mice and was associated with the loss of alpha-motor neurons in the spinal cord. There was no evidence of neurodegeneration in other brain structures. Motor dysfunction was observed in both male and female animals, showing that in SBMA the polyglutamine repeat expansion causes a dominant gain-of-function mutation in the AR. The male mice displayed a progressive reduction in sperm production consistent with testis defects reported in human patients. These mice represent the first model to reproduce the key features of SBMA, making them a useful resource for characterizing disease progression, and for testing therapeutic strategies for both polyglutamine and motor neuron diseases.     

Isolation and characterization of human and rabbit sperm tail fibrous sheath

Using mechanical and chemical dissection methods, fibrous sheath was isolated both from normal ejaculated human spermatozoa and from rabbit cauda epididymal spermatozoa. The same techniques did not produce a pure preparation of fibrous sheath from ejaculated rabbit spermatozoa, suggesting that further cross-linking and stabilization of sperm structures occurs in response to components of the seminal plasma. The isolation procedures were monitored by phase contrast microscopy and the purity of the fibrous sheath was verified by electron microscopy. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) of isolated human fibrous sheath revealed at least 14 protein bands of which the most intensely stained were of molecular weight 84, 72, 66.2, 57, 32 and 28.5 kDa. The rabbit fibrous sheath revealed at least 10 protein bands, of which the most intensely stained were 35.2, 32.7 and 28.5 kDa. The amino acid composition of the purified fibrous sheath from human and rabbit spermatozoa was similar, being high in aspartic acid and/or asparagine and glutamic acid and/or glutamine, serine, alanine, leucine, lysine and glycine, but low in histidine, tyrosine and isoleucine. This composition is similar to that reported for the rat and suggests that mammalian sperm tail fibrous sheaths are composed of similar types of proteins, although there are apparent differences in protein components between species.      

Experimental cryptorchidism in the adult mouse. III. Qualitative and quantitative electron microscopic morphology of Leydig cells.

The morphology of Leydig cells of control and 28-day-old cryptorchid mice was studied by electron microscopy and stereologic techniques. Leydig cell profiles of control mice were larger in section when compared to cryptorchid mice, but no differences were observed in the distribution of organelles in Leydig cells in the two groups. Quantitatively, the absolute volumes of smooth endoplasmic reticulum (SER), rough endoplasmic reticulum (RER), mitochondria, lysosomes, multivesicular bodies, peroxisomes, cytoplasmic matrix, nucleus, lipid droplets, membrane whorls, ribosomal aggregates, and annulate lamellae per Leydig cell were reduced significantly after 28 days of cryptorchidism. However, the absolute volumes of these organelles per testis were not significantly different between control and cryptorchid mice, due to the increase in Leydig cell number per testis in the cryptorchid testis, compared to the controls, except that the absolute volume of Golgi per Leydig cell was not significantly different between control and cryptorchid rats, but the absolute volume of Leydig cell Golgi was significantly lower in control rats. Based on these results, we conclude that, morphologically, a 28-day cryptorchid mouse Leydig cell clearly approximates a "half unit" of a control Leydig cell.     

Serum inhibin levels in normal men and men with testicular disorders

Serum concentrations of inhibin, FSH and LH were measured in 39 normal men and 127 men with testicular disorders resulting in infertility. The infertile men were divided into groups on the basis of their mean sperm count, FSH levels and karyotype. The mean (+/- S.D) serum concentrations of inhibin in the normal men was 554 +/- 156 U/l and did not differ significantly from those groups with oligospermia, azoospermia or Klinefelter's syndrome. Combined analyses of all groups did not reveal any significant correlation between serum concentrations of inhibin and FSH or with any other parameter measured. Serum concentrations of FSH and LH were positively correlated, and Leydig cell dysfunction, as evidenced by increased serum LH levels, low testosterone levels or a declining testosterone/LH ratio were found with severe spermatogenic damage. The failure of serum concentrations of inhibin to correlate with those of FSH levels or the degree of testicular damage raise questions as to the clinical value of this parameter alone.     

Studies on the testicular source of inhibin and its route of secretion in rams: failure of the Leydig cell to secrete inhibin in response to a human chorionic gonadotrophin/LH stimulus

To determine whether Leydig cells produce inhibin in the ram, Leydig cells were stimulated by administering human chorionic gonadotrophin (hCG) or raising the levels of endogenous LH by an injection of gonadotrophin releasing hormone (GnRH). Plasma concentrations of testosterone increased in the 72 h after either a single injection (P less than 0.05) or two injections (P less than 0.01) of hCG. Plasma concentrations of inhibin were not significantly influenced by either one or two injections of hCG. Administration of GnRH (1 microgram) caused an 11-fold increase in plasma concentrations of LH but did not influence concentrations of inhibin in either the jugular or testicular veins (pampiniform plexus). In contrast, concentrations of testosterone were increased by about fourfold in both jugular (P less than 0.01) and testicular (P less than 0.05) veins. The concentrations of inhibin in the testicular vein were 1.3-fold higher than in the peripheral plasma (P less than 0.05) both before and following treatment with GnRH whereas the concentrations of testosterone were 18- to 21-fold greater than in peripheral concentrations. In view of the difference in concentrations of inhibin between testicular and jugular veins, in a further experiment a sample was taken from the jugular vein, a vein located in the tunica vasculosa of the testis (testicular vein) and from a vein (spermatic vein) and lymph vessels located in the spermatic cord. The mean (+/- S.E.M.) concentrations of inhibin were highest in the testicular lymph (45.93 +/- 4.21 micrograms/l; P less than 0.001) compared with the peripheral (4.14 +/- 0.31 micrograms/l), spermatic (8.0 +/- 1.17 micrograms/l) or testicular (6.4 +/- 0.82 micrograms/l) plasma.(ABSTRACT TRUNCATED AT 250 WORDS)     

Changes in testicular inhibin after a single episode of heating of rat testes

Brief heating (43 C for 15 min) of the scrota of adult rats was used to induce reversible spermatogenic damage to the testes. In these animals the changes in testicular inhibin content and an index of inhibin production rate, measured after efferent duct ligation, were examined and correlated with serum gonadotropin levels. The effect of heating was not evident until after 1 week when testis weight, inhibin content, and inhibin production rate were significantly reduced and both serum FSH and LH were elevated. By 2 weeks, the maximal effects were observed, and, thereafter, all parameters gradually returned to control values (FSH: by 6 weeks; testis and epididymal weight, inhibin content, inhibin production rate, and seminiferous tubule fluid production: by 17 weeks). Throughout the study, serum testosterone levels showed no significant changes. Significant inverse correlations were found between serum FSH levels and inhibin content (r = -0.502, P less than 0.001) or inhibin production rate (r = -0.533, P less than 0.001), and these were taken as supportive evidence for the hypothesis that inhibin is involved in the feedback control of pituitary FSH secretion. Although serum LH levels were also negatively correlated to the corresponding inhibin content (r = -0.669, P less than 0.001) or inhibin production rate (r = -0.420, P less than 0.001), recent findings of Leydig cell dysfunction in these animals led us to relate the transient rise in LH to the altered state of Leydig cell function.     

Inhibin as an oocyte meiotic inhibitor

The effect of inhibin on oocyte maturation division was studied in vitro. Inhibin inhibited spontaneous division in both cumulus enclosed and denuded rate oocytes obtained from immature rats. The inhibition was dose-dependent (15-5000 units/ml) with an ED50 of 45.8 units/ml (4.00 nM). Follicle stimulating hormone (FSH) suppressing protein (FSP) which has inhibin-like activity accelerated the rate of meiotic maturation of rat oocytes in culture (P less than 0.05, at a dose of 0.7 microgram/ml) while activin, which has opposite biological effects to inhibin in many systems did not significantly stimulate oocyte maturation at a dose of 1.12 micrograms/ml. The actions of the three proteins studied were reversible.     

The influence of luteinizing hormone on the Leydig cells of cryptorchid rat testes

The influence of circulating LH levels on Leydig cells from cryptorchid adult rats was examined after ablation of the pituitary. After 2 weeks cryptorchidism, serum FSH and LH levels rose 2-fold while serum testosterone (T) remained unchanged. Leydig cells were hypertrophied and showed an increased response to in vitro hCG stimulation. Two weeks after hypophysectomy (hypox), serum hormone levels (LH, FSH and T), Leydig cell size, cytoplasm, organelle content and in vitro T production were all dramatically reduced. However, when hypophysectomy was combined with cryptorchidism (hypox/crypt), there was an increase in Leydig cell size, compared to hypophysectomy alone, in the presence of very low levels of serum FSH, LH and T. Compared to the hypophysectomised state, the mitochondria were larger and the cytoplasm contained more smooth endoplasmic reticulum. The response of the hypox/crypt testes to in vitro hCG stimulation, though significantly less than the cryptorchid testes, was significantly greater than the hypox testes. These results demonstrate that the changes observed in the Leydig cell after cryptorchidism can occur in the absence of peripheral pituitary hormones and are consistent with the hypothesis that a local feedback loop exists within the testis.