Mort programmée des cellules germinales testiculaires: causes et mécanismes mis en jeu

Spermatogenesis results from a balance between proliferation and apoptosis. An alteration in this balance could lead to testicular diseases such as testicular tumour or infertility. Apoptosis seem to be important in regulating the processes of spermatogenesis since 60 to 75% of germ cells do not reach the spermatozoa stage. The various molecules of the apoptotic cascade have been detected in rodent or human germ cells, such as effector caspases and upstream proteins from cell death receptor or mitochondrial pathways. One or several different pathways may be involved in the germ cell apoptotic process triggered physiologically, by hormonal deprivation, or by chemical or physical inducers. Finally, caspases appear to play a role in various testicular diseases (particularly infertility).     

Germ cell transplantation in an azoospermic Klinefelter bull

Germ cell transplantation is a technique that transfers donor testicular cells into recipient testes. A population of germ cells can colonize the recipient testis, initiate spermatogenesis, and produce sperm capable of fertilization. In the present study, a nonmosaic Klinefelter bull was used as a germ cell recipient. The donor cell suspension was introduced into the rete testis using ultrasound-guided puncture. A pulsatile administration of GnRH was performed to stimulate spermatogenesis. The molecular approach to detect donor cells was done by a quantitative polymerase chain reaction with allele discrimination based on a genetic mutation between donor and recipient. Therefore, a known genetic mutation, associated with coat-color phenotype, was used to calculate the ratio of donor to recipient cells in the biopsy specimens and ejaculates for 10 mo. After slaughtering, meiotic preparations were performed. The injected germ cells did not undergo spermatogenesis. Six months after germ cell transplantation, the donor cells were rejected, which indicates that the donor cells could not incorporate in the testis. The hormone stimulation showed that the testosterone-producing Leydig cells were functionally intact. Despite subfertility therapy, neither the recipient nor the donor cells underwent spermatogenesis. Therefore, nonmosaic Klinefelter bulls are not suitable as germ cell recipients. Future germ cell recipients in cattle could be mosaic Klinefelters, interspecies hybrids, bulls with Sertoli cell-only syndrome, or bulls with disrupted germ cell migration caused by RNA interference.     

Evidence for protein absorption from the lumen of the seminiferous tubule and rete of the rat testis

As luminal fluid moves from the seminiferous tubule and enters the rete testis, its protein concentration declines from approximately 6 mg/ml to 1 mg/ml. It was therefore suggested that protein is either 1) utilized by the spermatozoa, 2) transported across the epithelium of the terminal segment of the seminiferous tubule, the tubuli recti or rete testis, or 3) absorbed and degraded by the epithelium. Horseradish peroxidase (HRP), a protein marker, was microperfused into single seminiferous tubules or perfused directly into the rete. After fixation, the HRP was localized histochemically and the tissue observed under the light- and electron microscope. HRP was taken up via pinocytotic vesicles into the cytoplasm of the Sertoli cells and germ cells but did not permeate extracellularly beyond the tight junctions. Similar results were obtained in the cells lining the terminal segment and the tubuli recti. The rete epithelium showed uptake of HRP into coated and noncoated vesicles, while some cells additionally revealed diffuse cytoplasmic distribution of HRP. The terminal segment, tubuli recti, and rete testis may be important routes by which proteins may leave the testicular fluid either to be degraded or to enter the blood.     

Une nouvelle observation d’une métastase testiculaire d’un adénocarcinome prostatique: Aspects diagnostiques et éthiopathogéniques

Testicular metastases from prostatic adenocarcinoma are rare. They are often asymptomatic and diagnosed incidentally or at autopsy after orchidectomy in more advanced stages of the disease. The authors report a new case and review the diagnosis and aetiopathogenesis of these metastases. A 67-year-old patient with prostatic adenocarcinoma presented with painless right testicular mass for two months with no inflammatory signs. Germ cell tumour serum markers were negative. Scrotal ultrasound showed a large testicular tumour measuring 7×4×3cm. Histological examination revealed a solid non-differentiated tumour, not suggestive of primary testicular tumour. The immunohistochemical panel confirmed the prostate as the primary site due to the positivity of cytokeratins, PSA and PSAP and the negativity of classical markers of germ cell tumours. Testicular metastases from prostatic adenocarcinoma are rare, but their incidence is currently on the increase. They are often asymptomatic and discovered after pulpectomy. This diagnosis must be considered in the case of a testicular mass in patients over the age of 50 with a history of prostatic adenocarcinoma. The diagnosis was established after orchidectomy by histological examination and immunohistochemical tests.     

Histopathology of the male reproductive system induced by the fungicide benomyl

Benomyl is an effective fungicide that has been in use for many years. This chemical and its primary metabolite, carbendazim, are microtubule poisons that are relatively nontoxic to all mammalian organs, except for the male reproductive system. Its primary effects, at moderate to low dosages, are on the testis, where it causes sloughing of germ cells in a stage-dependent manner. Sloughing is caused by the effects of the chemical on microtubules and intermediate filaments of the Sertoli cell. These effects spread to dividing germ cells and also lead to abnormal development of the head of elongating spermatids. At higher dosages, it causes occlusion of the efferent ducts, blocking passage of sperm from the rete testis to epididymis. The mechanism of occlusion appears to be related to fluid reabsorption, sperm stasis, followed by leukocyte chemotaxis, sperm granulomas, fibrosis and often the formation of abnormal microcanals. The occlusion results in a rapid swelling of the testis and ultimately seminiferous tubular atrophy and infertility. In conclusion, studies that reveal long term testicular atrophy following chronic or subchronic exposure to a toxicant should be re-examined for histopathological lesions in the efferent ductules and head of the epididymis. Lesions in the male track that cause blockage may induce permanent testicular damage and a decrease in sperm production.     

Testicular toxicity and mutagenicity of steroidal and non-steroidal estrogens in the male mouse.

The mutagenicity and toxicity of diethylstilbestrol (DES), 17 beta-estradiol and zeranol on the male mouse germ cells were investigated with meiotic micronucleus assays in vivo and in vitro, sperm-head abnormality test and morphometry. Further, the developmental effects of DES on testicular morphology were explored. Micronucleus induction was observed at 10(-7) M concentration of DES and 17 beta-estradiol in vitro, but other treatments yielded negative results. The micronucleus assay in vivo revealed a small number of micronuclei in early haploid spermatids 17 days after a single subcutaneous injection of DES 50 mg/kg, whereas estradiol and zeranol gave negative results. The sperm-head abnormality rates were significantly elevated 5 weeks after treatments with high doses of DES, 17 beta-estradiol and zeranol, and testicular morphometry revealed transient changes in the volume densities of testicular tissue components. Prenatal and neonatal estrogen administration resulted in permanent alterations in seminiferous epithelium and dilatation of the rete testis, but did not affect micronucleus or sperm-head abnormality rates. The mutagenicity and toxicity of hormones in the mouse testis paralleled the hormonal activity of these compounds. Early estrogenization was the most sensitive toxicity test, followed by in vitro meiotic micronucleus induction, whereas the sperm-head abnormality assay and morphological analysis did not reveal subtle changes.     

Histochemistry and ultrastructure of nerve fibres and contractile cells in the tunica albuginea of the rat testis

Whole-mounted preparations of the tunica albuginea of the rat testis were studied using light microscopy techniques for demonstration of cholinergic nerve fibres (Karnovski-Root method), catecholaminergic nerve fibres (De la Torre's method) and actin filaments (avidin-biotin-peroxidase method). An ultrastructural study of different regions of the albuginea was also performed. Cholinergic fibres were seen to penetrate into the albuginea with the testicular artery to form a broad network in the mediastinum testis, many fibres ending beneath the rete testis epithelium. Catecholaminergic fibres penetrated through the middle part of the cauda epididymis and formed a plexus in the albuginea covering the inferior testicular pole. This plexus gave rise to straight fibres which formed varicosities, some of them appeared related with mast cells. Actin-containing cells were only found beneath the rete testis epithelium. These cells were similar to myofibroblasts. The location of both cholinergic fibres and contractile cells among the rete testis channels suggests that these cells may be involved in the pumping of semen towards the ductuli efferentes and that their contractility may be regulated by cholinergic fibres.     

Viability of ram testicular spermatozoa following cryopreservation in rete testis fluid

Ram testicular spermatozoa, collected continuously from the cannulated testis, were frozen in rete testis fluid in straws using the cryoprotective agents egg yolk and glycerol. The effect of cryopreservation on the viability of the spermatozoa was assessed by studying their metabolism, morphology, ultrastructure, and radioiodination patterns. Freeze-thawing significantly depressed the respiration rate and glycolytic activity of testicular spermatozoa. Morphologically, there was little evidence of cryodamage in frozen-thawed testicular spermatozoa. Except for some slightly corrugated acrosomes and a more loosely attached plasma membrane over the sperm head, frozen-thawed testicular spermatozoa were indistinguishable from nonfrozen control spermatozoa. Surface radioiodination of frozen-thawed testicular spermatozoa was highly selective and resulted in a labeling pattern similar to that of the nonfrozen controls. In contrast, the radiolabeling pattern of frozen-thawed electroejaculated spermatozoa was characterized by high background radioactivity and low selectivity. These results confirm previous suggestions that testicular spermatozoa have a greater low-temperature tolerance than do ejaculated spermatozoa and indicate that cryopreservation of immature testicular spermatozoa in rete testis fluid with added egg yolk and glycerol may be a useful approach to extend the availability of these cells.     

Effects of follicle-stimulating hormone on inhibin release by different testicular compartments in the adult ram.

The aim of this study was to determine the bidirectional release of immunoreactive inhibin-alpha (irINH-alpha) by different testicular compartments in the adult ram and to assess the effects of FSH on the distribution of inhibin in the testis. Immunoreactive INH-alpha was measured by RIA in fluid samples collected concurrently from the three testicular compartments--the seminiferous tubules, the interstitium, and the vascular system--through catheters inserted surgically into the rete testis, testicular lymphatic duct system, and spermatic veins, respectively. Overall, the concentration of irINH-alpha in rete testis fluid was 25 times the level in testicular lymph and over 500 times the concentration in peripheral blood. The pattern of irINH-alpha concentration in rete testis fluid was inversely related to that in testicular lymph, but i.v. administration of FSH had a decoupling effect on this relationship by depressing inhibin concentration in testicular lymph without affecting inhibin levels in rete testis fluid. Nevertheless, increased flow of testicular lymph more than compensated for the transient fall in irINH-alpha concentration so that, overall, the total output of inhibin via the testicular lymphatic duct system (and the vascular system) increased significantly. No persistent or significant changes were observed in the flow rate of rete testis fluid or concentration of irINH-alpha in the fluid after administration of FSH. The time frame for the response of the testis to FSH is indicative of the involvement of a mediator. Electrophoretic analysis of serially collected testicular lymph samples consistently revealed an FSH-induced release of a series of proteins in the M(r) range of 30,000-32,000.(ABSTRACT TRUNCATED AT 250 WORDS)     

Localization of the epidermal growth factor (EGF) and epidermal growth factor receptor (EGFR) in the bovine testis

In the last few decades, several growth factors were identified in the testis of various mammalian species. Growth factors are shown to promote cell proliferation, regulate tissue differentiation, and modulate organogenesis. In the present investigation we have studied the localization of EGF and EGFR in the adult bovine testis by means of immunohistochemical method. Our results demonstrated that EGF and EGFR were localized solely to the bovine testicular germ cells (spermatogonia, spermatocytes, and round spermatids). In contrast, the somatic testicular cells (i.e., Sertoli, Leydig, and myofibroblast cells) exhibited no staining affinity. EGF and EGFR were additionally detected in the epithelial lining of straight tubules and rete testis. Interestingly, the distribution of EGF and EGFR in the germ cells was mainly dependent upon the cycle of the seminiferous epithelium since their localization appeared to be preponderant during the spermatogonia proliferation and during the meiotic and spermiogenic processes. In conclusion, such findings may suggest that EGF and EGFR are important paracrine and/or autocrine regulators of spermatogenesis in bovine.     

Comparative study of the effects of pre and post natal administration of a thyroid drug on testicular activity in adult rat

Thyroid hormone is known to play a critical role in growth and development of rat testes with a specific effect on the differentiation of Sertoli cells leading to a normal evolution of germ cells. In the present study, we aimed to compare the effect of induced hypothyroidism during fetal and post-natal life on the structure and function of the testis in adult. Pregnant or lactating mothers were treated with 6-propyl-2-thiouracil (PTU) during 21 days and weight gain of pups was steady until adult age. Plasma hormonal levels were determined by RIA and morphology of testis was studied on sections stained with Masson's trichrome. Pre and early post natal hypothyroidism resulted in an impairment of body development and a diminution of thyroid hormone levels of treated rats. No significant effect on testicular development has been observed when hypothyroidism is induced in fetal life while it was associated with reduction in testis weight, diameter of seminiferous tubules and hormonal levels and delay in maturation of germ cells, when induced during early post natal life.     

Spatial topography of the excurrent duct system in the bovine testis

Summary The rete testis of the bull is situated within an axial mediastinum and consists of approximately 30 longitudinally arranged, anastomosing rete channels. At the cranial testicular pole all rete channels empty into a common space, the area confluens reds, which is subdivided by small septa and narrow chordae retis. The area confluens always contains numerous spermatozoa and is connected with the bulbous initial portions of the efferent ductules by short, often tortuous rete tubules. Since the connection between rete and efferent ductules is situated within the tunica albuginea, the bovine excurrent duct system is not provided with an extratesticular rete as in many other mammals.Straight testicular tubules merge from all directions to connect with superficial rete channels, but the inlets are not evenly distributed. In the periphery each straight tubule begins with a cup-like structure followed by a narrow stalk region and a heavily folded portion opening either immediately into a rete channel or into a tube-like lateral rete extension.In close contiguity to the rete testis lie extremely coiled arterial portions connecting the centripetal and the centrifugal branches of the testicular artery. Since intrinsic musculature is scarcely developed in the mediastinum, and transport of rete content relies primarily on massage due to external pressure changes, the pulsatile blood flow through these coiled arteries may influence conveyance processes within the rete testis.An intimate spatial association between area confluens reds and adjacent large, thin-walled lymph vessels may facilitate a transfer of androgens into the fluid of the rete testis.Supported by the Stiftung zur Förderung der wissenschaftlichen Forschung an der Universität Bern     

Anomalies de position des testicules dans l’enfance: conséquences à l’âge adulte

Non descended testes in the low scrotum is a common anomaly at birth, with about 4% of the newborn males affected. Only one quarter of these newborn babies will still have non descended testes when one year old. However, the testes that will descend within the first year of life seem no more to be considered as normally descended testes. Moreover, the retractile testis, which represents a secondary anomaly of testicular position occuring after the babies are older than one year, is no more to be regarded as a physiological variant of the normally descended testis, since several reports indicate histological and clinical modifications in such cases. The testicular non descent can be associated with two consequences in adult life. Firstly, an history of non descended testis is the only known risk factor for the testicular cancer. Secondly, such an history is a risk factor for the male fertility because of spermatogenesis alterations, as indicated by qualitative and quantitative histological analyses of the testicular tissue, and by depressed spermatozoa output and quality (motility, normal forms); moreover, testicular volumes are reduced, and the time to pregnancy as well as the rate of infertility are increased. Time is arrived for a reappraisal of the consequences in adult life of the abnormal testicular location (either congenital or acquired) during childhood.     

The effects of long-term administration of either a crude inhibin preparation or an antiserum to FSH on serum hormone levels, testicular function and fertility of adult male rats.

Adult male rats were given either daily injections of ram rete testis fluid for periods of up to 70 days or injections of an antiserum against FSH every 3 days for 90 days. Compared with the control groups, the rats injected with ram rete testis fluid had lowered serum FSH levels, but only at treatment periods of 30 days and less. The levels of LH and testosterone in serum, testicular fluid secretion, sperm counts, testis weights and fertility were not affected by rete testis fluid treatment. The rats injected with anti-FSH serum exhibited an impairment of fertility which was never complete and evident only after 49 days of treatment. After 90 days of anti-FSH treatment, testis weight and free serum FSH were reduced, but sperm counts, testicular fluid secretion and serum levels of LH and testosterone were not affected.     

Nestin, a neuroectodermal stem cell marker molecule, is expressed in Leydig cells of the human testis and in some specific cell types from human testicular tumours

The intermediate filament protein nestin is predominantly expressed in some stem/progenitor cells and appears to be a useful molecular tool to characterise tumours originating from precursor cells of neuroectodermal and mesenchymal lineages. Leydig cells originate in the adult testis by differentiation from stem cells and express a variety of neural and neuroendocrine markers. The possible expression of the neural stem cell marker nestin in Leydig cells and testicular tumour cells was determined by analysing the patterns of nestin expression in normal and pathological human testes by Western blot and immunohistochemical methods. In normal testis, nestin was found in some vascular endothelial cells, a subset of peritubular spindle-shaped cells and some Leydig cells; spermatogenic and Sertoli cells were unstained. In normal Leydig cells, nestin was distributed in the perinuclear cytoplasm and accumulated in the crystalloids of Reinke with ageing. In non-tumour pathologies (cryptorchidism, impaired spermatogenesis), the seminiferous tubules were immunonegative, whereas hyperplastic Leydig cells showed cytoplasmic immunolabelling. In testicular malignancies, nestin was localised in the Sertoli cells of the seminiferous tubules affected with intratubular germ cell neoplasia, in the hyperplastic Leydig cells associated with these tumours and in some components (mesenchymal and neuroepithelial cells) of teratomas; spermatocytic and non-spermatocytic seminomas were unstained. Some vascular endothelial cells were immunolabelled in all tumour samples. Thus, nestin is expressed in a population of normal and hyperplastic Leydig cells and in Sertoli cells in the presence of intratubular germ-cell neoplasia. Nestin may be a good marker for identifying components of testicular teratomas.The two first authors participated equally in this workThis work was supported by a grant from the Fondo de Investigaciones Sanitarias (FIS 02/3003 to M.V.T. Lobo)     

Distribution of histopathology and Ia positive cells in actively induced and passively transferred experimental autoimmune orchitis

Histopathology in testes from mice with actively induced experimental orchitis (EAO) (active EAO) and those from recipients of testis-sensitized lymphocytes (passive EAO) had different distributions. In passive EAO, maximum orchitis existed in the straight tubules, rete testis, and ductus efferentes, obstruction of which led to extreme dilatation of seminiferous tubules. Unusual intralymphatic granulomata also resulted in dilated testicular lymphatics. In active EAO, maximum orchitis affected seminiferous tubules under the testicular capsule, away from the rete testes. Vasitis was common and occurred in both active and passive EAO. In normal testes, IA+ F4/80+ cells were sparse but formed a cuff around the straight tubules. After immunization with testis in adjuvant or with adjuvant alone, the number, size, and staining intensity of IA+ cells increased dramatically beginning on day 5, 7 days before disease onset. Simultaneously, epithelial cells confined to the ductus efferentes became Ia+. Although recipients of sensitized lymphocytes also developed epithelial Ia in the ductus efferentes, they did not show changes in testicular interstitial Ia+ cells. Our findings indicate that testicular autoantigens are not completely sequestered, but are accessible to and can react with passively transferred immune lymphocytes in well-defined regions of the germ cell compartment. These regions coincided to a large extent with maximum expression of periductal or epithelial Ia. Changes in Ia+ cells in the testis, which are inducible by adjuvants and precede orchitis, may account in part for the different distribution of histopathology of active EAO.     

Development of the seminiferious tubules and rete testis during the prenatal period of human ontogenesis

The sources of origin and the peculiarities of formation of the seminal ducts and rete testis during the prenatal period of ontogenesis in man were studied. It has been established that the seminal duct sand the ducts of the rete testis form from the cellular cords of the coelomic epithelium and the primordial germ cells which appear simultaneously in the septum of the testis and the primordial germ cells which appear simultaneously in the septum of the testis and central part of its parenchyma in the embryos, 13.0--17.0 mm long. By plastic and graphic reconstruction, as well as by methods of subtle preparation under binocular microscope MBC-I control it was revealed that the seminal ducts anastomosed between themselves both within the limits of one and the adjacent lobules. The ducts of the rete testis do not form anastomoses, but superimpose over one another, creating an impression of a rete. Approaching the tunica albuginea they merge, continuing into the cuctuli efferentes testis.     

Aetiology of testicular cancer: association with congenital abnormalities,age at puberty,infertility, and exercise. United Kingdom Testicular Cancer Study Group.

OBJECTIVE--To determine the risk of testicular cancer associated with undescended testis, inguinal hernia, age at puberty, marital status, infertility, vasectomy, and amount of exercise. DESIGN--A population based case-control study with a questionnaire administered by an interviewer and with relevant supplementary data extracted from general practitioners'' notes. SETTING--Nine health regions within England and Wales. SUBJECTS--794 men, aged 15-49 years, with a testicular germ cell tumour diagnosed between 1 January 1984 and 1 January 1987; each had an age matched (within one year) control selected from the list of their general practitioner. RESULTS--There was a significant association of testicular cancer with undescended testis (odds ratio 3.82; 95% confidence interval 2.24 to 6.52) and inguinal hernia (1.91; 1.12 to 3.23). The excess risk associated with undescended testis was eliminated in men who had had an orchidopexy before the age of 10 years. There were positive associations with early age at voice breaking, early age at starting to shave, and infertility. There was a significant association with a sedentary lifestyle and a moderate protective effect of exercise. There was no association with vasectomy. CONCLUSION--This study confirms previous reports that developmental urogenital abnormalities result in an increased risk of testicular cancer. The trend to perform orchidopexy at younger ages may reduce the risk associated with undescended testis. The increased risks associated with early age at puberty and low amounts of exercise may be related to effects of exposure to endogenous hormones. Changes in both of these factors may partly contribute to the increasing rates of testicular cancer observed in the past few decades.     

Impact of methoxyacetic acid on mouse Leydig cell gene expression

Background  

Methoxyacetic acid (MAA) is the active metabolite of the widely used industrial chemical ethylene glycol monomethyl ether, which is associated with various developmental and reproductive toxicities, including neural toxicity, blood and immune disorders, limb degeneration and testicular toxicity. Testicular toxicity is caused by degeneration of germ cells in association with changes in gene expression in both germ cells and Sertoli cells of the testis. This study investigates the impact of MAA on gene expression in testicular Leydig cells, which play a critical role in germ cell survival and male reproductive function.