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PubMed2005

Molecular cloning and primary functional analysis of a novel human testis-specific gene.

In this study, a new data mining tool called Digital Differential Display (DDD) from the NCBI was used to predict testis-specific expressed genes from the expressed sequence tag (EST) database. DDD (digital differential display) was performed between nine testis libraries and seventy libraries derived from other tissues. We identified a new contig of ESTs (HS. 326528) which was from testis libraries. To validate the use of bioinformatic approaches in gene discovery, the ESTs (HS. 326528), which were predicted to be testis-specific, were chosen for further study. Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) analysis of matched sets of cDNAs from testis and other tissues indicated that the ESTs were specifically expressed in testis. This result was further validated by multi-tissue Northern blot. The full-length cDNA encompassing the entire open reading frame was cloned and, in view of its apparent specificity to testis, the gene was termed homo sapiens spermatogenesis-related gene 8---SRG8 (GenBank accession number: AY489187). The gene whose full cDNA length is 1 044 bp containing 3 exons and 2 introns is located in human chromosome 15q26.2, the cDNA encodes a novel protein of 105 amino acides with a theoretical molecular weight of 11.7 kD and isoelectric point of 10.09 which shares no significant homology with any known proteins in database. Real time PCR analysis of testis of different developmental periods revealed that SRG8 gene is significantly expressed in adult testis. The green fluorescent protein produced by pEGFP-C3/SRG8 was detected in the nucleus of HeLa cells after 24 h post-transfection. Cell cycle analysis showed that SRG8 can accelerate HeLa cells to traverse the S-phase and enter the G2-phase compared with the control without transfection of SRG8, which suggested that this gene plays an important role in the development of testis. The discovery of SRG8 showed that DDD combined with experiments is a feasible, time-saving strategy to identify new candidate genes for testis-specific development

Adult↗

PubMed2005

[Experimental study of in situ electroporation-induced testis injury in specific pathogen-free male Kunming mice].

OBJECTIVE: To observe testis injuries induced by in situ electroporation in specific pathogen-free (SPF) adult male Kunming mice. METHODS: With two sets of parameters of high and low voltages, respectively, in situ electroporation of the testis was performed in vivo by fixing the testis and epididymis of the mice between a pair of rectangular tweezer-type electrodes. Two weeks after electroporation, the mice were killed and the testis and epididymis separated and fixed with 4% paraformaldehyde after recording the weight of the testis. Routine histological sections were prepared and observed under optical microscope after HE staining. The epididymis was transferred into M16 medium for spermatozoa separation, and after dilution of the spermatozoa suspension, the spermatozoa viability was observed under optical microscope. RESULTS: Two weeks after high-voltage electroporation, examination of spermatozoa viability and microscopy of the testis sections revealed irreversible testis injury, and the testis weight was significantly reduced in comparison with that of control mice (P<0.01). Low-voltage electroporation, in contrast, only caused reversible injuries of the testis, and the male mice retained their reproductive capacity after a certain length of recovery period. The testis weight after low-voltage electroporation showed no significant difference from that of the control mice (P>0.05). CONCLUSIONS: Appropriate setting of the parameters for in vivo electroporation may avoid severe impact on the reproductive capacity of the testis in SPF male Kunming mice. This technique also provides a possibility for exogenous gene transfer into the reproductive cells.

Animals↗

PubMed1976

The mammalian rete testis--a morphological examination.

The fine structure of the rete testis was examined in several primates, domestic animals and rodents. The rete testis consists of a series of interconnected wide channels lined with a simple cuboidal to columnar epithelium, resting on a thick basal lamina. Beneath the basal lamina dense bundles of collagen fibrils and a few blood vessels, lymphatics or nerve tissue are found. The epithelial cells are characterized by large, deeply indented nuclei, spherical or short rod-shaped mitochondria, supranuclear Golgi profiles, some cisterns of rough endoplasmic reticulum, free ribosomes and numerous micropinocytotic vesicles in the ectoplasmic regions. Smooth endoplasmic reticulum, secretory granules, lysosomes or other types of dense bodies are rarely seen. The apical surface of the cells bears numerous microvilli and a single very long flagellum which is presumed to be motile. Ajoining lateral cell membranes exhibit a juxtaluminal tight junction, elaborate interdigitations and desmosomes. The basal plasma membrane is highly irregular greatly increasing its surface area of contact with the underlying interstitium. The nuclei of the rete epithelial cells contain pale-staining, spherical structure, 2 mum in diameter, composed of circularly oriented fine filaments. The significance of the nuclear structures remains unknown. Thorotrast was injected into the lumen of the hamster and rat rete testis and 30 minutes later the proximal portion of the excurrent duct system of the testis was prepared for electron microscopy. Whereas the ductuli efferentes and first part of the epididymis possessed numerous apical vesicles filled with the thorotrast, this electron opaque substance was rarely found in the epithelium of the rete testis. Thus, incorporation of particulate matter into the lining cells of the rete from its lumen is apparently less active than in the epithelium of the ductuli and epididymis. Vascularly introduced intercellular tracer compounds such as lanthanum nitrate or horseradish peroxidase did not enter the lumen of the rete testis from the interstitium. The tracer molecules appeared to be blocked by the juxtaluminal tight junction separating adjacent epithelial cells. This latter observation suggests that a blood-testis barrier exists at the level of the rete testis epithelium. Although physiological studies have indicated that the composition of fluid secreted in the seminiferous epithelium is considerably modified in the rete testis, the present morphological study does not provide additional evidence to support a secretory or absorptive function for this region of the excurrent duct system of the testis.

Animals↗

PubMed1998

Ex vivo and in vitro testis and ovary explants: utility for identifying steroid biosynthesis inhibitors and comparison to a Tier I screening battery.

Testis and ovary explants have been proposed as in vitro screens for identifying potential inhibitors of steroid biosynthesis. The goals of the current study were to optimize the conditions of the two assays, to characterize these assays using several compounds with well-defined endocrine activity, and to compare the responses from the explant assays with an in vivo male battery currently undergoing validation using the Crl:CD BR rat in order to evaluate their utility as test systems for screening unknown compounds for possible steroid biosynthesis inhibition activity. There were two components to the testis/ovary assays: ex vivo and in vitro. The ex vivo component used testes/ovaries from animals dosed with the test compounds in vivo, and the in vitro component used testes/ovaries from control animals. For the testis assays, decapsulated testis explants (50 mg) were placed into glass scintillation vials, +/-1.0 IU/ml hCG for 3 h in a shaking water bath (34 degrees C). Following the incubation period, medium was removed, centrifuged, and frozen until assayed for hormone concentrations. A similar procedure was used for the ovary explant assay except that each ovary was incubated separately. The testis explants were evaluated using the following compounds: ketoconazole (KETO), a testosterone biosynthesis inhibitor; aminoglutethimide (AG) (only in vitro) and anastrozole (ANA), aromatase inhibitors; finasteride (FIN), a 5alpha-reductase inhibitor; 17beta-estradiol (17beta-E2), an estrogen receptor agonist; flutamide (FLUT), an androgen receptor antagonist; ICI-182,780 (ICI), an estrogen receptor antagonist; haloperidol (HALO), a D2 receptor antagonist; and reserpine (RES), a dopamine depletor. In the ovary assay, AG (only in vitro), ANA, ICI, and HALO (only in vitro) were evaluated. Addition of fetal calf serum to the medium allowed measurement of estradiol (E2) in the testis assay, but production was not inhibited by ANA or AG. In the ovary explant assay, only AG was identified as inhibiting E2 production in vitro. Hence, both the testis and ovary explant assays appear to have limited utility for detecting aromatase inhibitors. Screening of these nine diverse endocrine-active compounds resulted in all of them being identified as altering the endocrine system when assessed by ex vivo and in vitro testis explants. Using only the in vitro assessment with the criteria of steroid biosynthesis inhibition, four of nine compounds were correctly identified in the testis explant assay (17beta-E2, KETO, FLUT, and HALO). The predictability of both the in vitro and ex vivo ovary assay was 50%, suggesting a 50% false positive or negative rate with unknown compounds. However, of the seven compounds assessed to date (17beta-E2, ICI, ANA, KETO, FLUT, HALO, and RES), all were correctly identified using an in vivo male battery, which also has the capability to detect other endocrine activities. Therefore, the testis and ovary explant assay would not be necessary if one were using an in vivo male battery, since this screen would identify steroid biosynthesis inhibitors and would also identify several other endocrine activities. Because of the difficulties in assessing cytotoxicity and the high false positive/negative rates, the ovary and testis explant assays are not useful as routine screening procedures for detecting steroid biosynthesis inhibitors; however, they may have utility in confirming in vivo findings.

Animals↗

PubMed2003

Impact of laparoscopy on the management of an unusual case of nonpalpable testis in an adult patient.

BACKGROUND: In the past decade, cryptorchidism has generally been treated in the first 36 months of life using an inguinal approach, in the case of palpable testis, or using laparoscopy, in the case of nonpalpable testis. Nevertheless, before this period some children were managed incorrectly. This case report shows how laparoscopy and the collaboration between pediatric surgeons and urologists may lead to optimal results. CASE REPORT: A 19-year-old male was referred to our unit following routine medical examination for enrollment in the military service with a diagnosis of right cryptorchidism. The clinical history showed the anamnesis of right cryptorchidism diagnosed at birth as a right nonpalpable testis, confirmed at 2 years of age with clinical examination and computer tomography. The patient came to our attention to again undergo a clinical examination, ultrasonography, computed tomography, and magnetic resonance imaging. We found a left, well-positioned testis of 20 ml in volume and an empty right scrotum; all the instrumental examinations were negative. The patient was thus scheduled for a diagnostic laparoscopy. A pediatric surgeon with extensive experience in this pathology performed the procedure. The 10-mm 30 degrees optics introduced through the umbilicus showed a closed right inguinal ring, with no evidence of either vas deferens or inner spermatic vessels at the level of the inguinal region or pelvis. Two more trocars were introduced and a testis of 15 ml in volume was found under the ascending colon 10 cm away from the cecum. An orchiectomy was performed via laparoscopy. CONCLUSION: This case clearly shows that some males with nonpalpable testis may have been treated incorrectly in the prelaparoscopic era and may now have an intraabdominal testis. In addition, our experience shows that intraabdominal testis may sometime be in an unusual location, and a laparoscopic surgeon with experience in this pathology is fundamental to finding the testis. In the case of adults with nonpalpable testis, when echography, computed tomography, and magnetic resonance imaging are not useful, the only effective diagnostic procedure is laparoscopy.

Abdominal Cavity↗

PubMed2006

Androgen receptor immunostaining and androgen receptor messenger ribonucleic acid expression are increased in cremaster muscles associated with undescended testis.

OBJECTIVES: To determine and compare androgen receptor (AR) immunostaining and AR messenger ribonucleic acid (mRNA) expression in cremaster muscles associated with descended or undescended testis. METHODS: Eight boys with descended testis but with inguinal hernia and 8 boys with undescended testis were evaluated. Serum follicle-stimulating hormone (FSH), luteinizing hormone (LH), testosterone, and free testosterone levels were determined, and samples of cremaster muscles were immunostained for AR. Groups were compared by unpaired t tests and Fisher's exact tests; P values of <0.05 were considered significant. Samples of cremaster muscles were obtained from another 5 boys with descended testis but with inguinal hernia and 5 boys with undescended testis. The expression of AR mRNA in those samples was determined by semiquantitative reverse transcriptase polymerase chain reaction. RESULTS: Serum FSH, LH, testosterone, and free testosterone levels were similar among groups. None of the samples from boys with descended testis showed positive staining, but 4 of 8 samples from boys with undescended testis stained positive for AR. Androgen receptor mRNA transcript levels were approximately 10 times lower in cremaster muscles of boys with descended testis compared with those in boys with undescended testis. CONCLUSIONS: Despite similar serum hormone levels, more AR expression in cremaster muscles associated with undescended testis might represent evidence of being subjected to a lesser degree of androgenic effects.

Child, Preschool↗

PubMed1996

Percutaneous testis biopsy: an alternative to open testicular biopsy in the evaluation of the subfertile man.

PURPOSE: We evaluated the safety and efficacy of percutaneous testis biopsy by comparing the ultrasound appearance and histological status of testicular parenchyma obtained to those noted after open testis biopsy. MATERIALS AND METHODS: A total of 51 consecutive infertile men with azoospermia or severely impaired semen quality, in whom ductal obstruction was suspected, underwent percutaneous (31) or open (20) testis biopsy, with 58 and 34 procedures performed, respectively. Scrotal ultrasound was performed preoperatively, and at 2 weeks and 1, 3 and 6 months after biopsy. In addition, immunoglobulins G and A antisperm antibody assays were obtained preoperatively and postoperatively. Two biopsy specimens were obtained from each testis for formal histological evaluation. A touch preparation was also performed and examined immediately for mature spermatozoa using phase contrast microscopy. RESULTS: All biopsies yielded adequate tissue for diagnosis and morphometric analysis. Of 58 percutaneous biopsies 4 (7%) demonstrated sonographic evidence of intratesticular bleeding, characterized by a hypoechoic region within the testicular parenchyma, which resolved by 6 months postoperatively. In contrast, 10 of 34 open biopsies (29%) showed evidence of intratesticular bleeding or a new area of increased echogenicity at 1 month after the procedure (intraparenchymatous scar). All intraparenchymatous scars persisted to 6 months postoperatively. No patient undergoing percutaneous or open testis biopsy had antisperm antibodies in the seminal fluid or serum (azoospermia cases) or on sperm postoperatively. Of the 32 and 20 patients undergoing percutaneous and open testis biopsy 3 (9%) and 14 (70%), respectively, required narcotic analgesia. All patients returned to routine activities within 24 hours after percutaneous testis biopsy. No postoperative infections or extratesticular hematomas were noted. Pathological study was diagnostic in all specimens. CONCLUSIONS: Percutaneous testis biopsy is well tolerated by the patient, with fewer apparent complications than and diagnostic value equal to open testis biopsy. Percutaneous testis biopsy should be considered an alternative to open biopsy.

Biopsy↗

PubMed2002

Expression of a novel HsMCAK mRNA splice variant, tsMCAK gene, in human testis.

Identification of specifically expressed genes in the adult or fetal testis is very important for the study of genes related to the development and function of the testis. In this study, a human adult testis cDNA microarray was constructed and hybridized with 33P-labeled human adult and embryo testis cDNA probes, respectively. After differential display analyzing, a number of new genes related to the development of testis and spermatogenesis had been identified. One of these new genes is tsMCAK. tsMCAK was expressed 2.62 folds more in human adult testis than fetal testis. The full length of tsMCAK is 2401 bp and contains a 2013 bp open reading frame, encoding a 671-amino-acid protein. Sequence analysis showed that it has a central kinesin motor domain and is homologous to HsMCAK gene of the somatic cells. Blasting human genome database localized tsMCAK to human chromosome 1P34 and further investigation showed that it is a splice variant of HsMCAK. The tissue distribution of tsMCAK was determined by RT-PCR and it is expressed highly and specifically in the testis. Southern blot studies of its expression in patients with infertility indicated its specific expression in spermatogenic cells and its correlation with male infertility. The above results suggested that tsMCAK is a candidate gene for the testis-specific KRPs and its specific expression in the testis was correlated with spermatogenesis and may be correlated with male infertility.

Adult↗

PubMed1992

Induction of increased testis growth and sperm production in adult rats by neonatal administration of the goitrogen propylthiouracil (PTU): the critical period.

We have previously shown that treating rats with the reversible goitrogen 6-propyl-2-thiouracil (PTU) from birth to Day 25 increases testis size and sperm production in adulthood by up to 80% and 140%, respectively. The purpose of this study was to determine the critical period(s) during development when PTU treatment can increase adult testis size and sperm production. Rats were treated with PTU beginning on Days 0, 8, 16, or 24 for periods of 9, 17, or 25 days. To further define the critical period, additional rats were treated with PTU prenatally or on Days 4-24. PTU treatments of 9, 17, or 25 days beginning at birth increased testis weight 18%, 38%, and 69%, respectively, by 135 days of age, while daily sperm production (DSP) increased 35%, 65%, and 94%, respectively. Efficiency of sperm production (DSP/g testis) also increased by approximately 25% in these rats. There was an inverse relationship between testis and body weights: increasing lengths of PTU treatment increased testis weight but decreased body weight. PTU treatment starting on Day 8 or later did not increase testis weight or DSP regardless of duration. Treatment on Days 4-24 increased adult testis weight and DSP similarly to treatment from birth to Day 24, but prenatal PTU treatment was ineffective. Testosterone concentrations were not altered in treated rats, even those with increased testis weight. These results indicate that the later part of the first postnatal week (Days 4-8) is the crucial period during which PTU treatment must begin in order to increase adult testis size and sperm production.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1994

Photoperiod regulates testis cell apoptosis in Djungarian hamsters.

Reproductive activity in the Djungarian hamster is controlled by seasonal variations in day length. Exposure to long days stimulates testis development, while exposure to short days induces testis regression. We recently found that testis regression after gonadotropin deprivation in rats is associated with increases in apoptosis. Here we sought to determine whether or not apoptosis is associated with the testis regression and/or recrudescence that occurs naturally in seasonally breeding mammals. Newborn male hamsters were maintained on long days (16L:8D) until 3 wk of age before being transferred to short days (8L:16D). Following decreases in serum FSH within 3 days of exposure to short days, testis weight decreased by 52% at Day 10, reaching a 70% decrease after 21 days. Analysis of testis cell DNA fragmentation showed a 4.9-fold increase of low-molecular-weight DNA as early as 5 days after transfer to short days; this was followed by a time-dependent decrease. The observed increases in testis cell apoptosis were correlated with decreases in serum testosterone, but decreases in Leydig cell LH receptor content were delayed. In a second study, 6-wk-old hamsters with regressed testes due to a 3-wk exposure to short days were transferred back to long days. After increases in serum FSH within 3 days of photostimulation, a 2-fold elevation in testis weight was found at Day 5. The increase in testis weight was associated with a 65% decrease of testis apoptosis within 5 days of photostimulation. Also, increases in serum testosterone and LH receptor content were observed after 5 and 10 days of exposure to long days, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1998

Growth hormone receptors in testis and liver during the spermatogenetic cycle in rainbow trout (Oncorhynchus mykiss).

Changes in growth hormone (GH) receptivity in the testis during the reproductive cycle were studied in rainbow trout (Oncorhynchus mykiss). This necessitated setting up a method to characterize GH receptors (GH-R) in crude testis homogenate. Binding of radiolabeled recombinant rainbow trout GH (125I-rtGH) to crude testis preparation was dependent on testicular tissue concentration. The sites were specific to GH, with no affinity for prolactins, gonadotropins, or insulin. Similar high affinities for 125I-rtGH were obtained with crude testis (4.5 x 10(9) + 0.25 M(-1)) and crude liver preparations (5.1 x 10(9) +/- 0.06 M(-1)) at all stages of spermatogenesis, as well as with testicular membrane preparations (2.4 x 10(9) M(-1)). GH-R concentration in testis was high during the immature stage (590 fmol/g testis); it then decreased regularly from the beginning (stage II, spermatogonia proliferation: 706 fmol/g testis) to the end of spermatogenesis (stage VII: 158 fmol/g testis). Conversely, the absolute testicular capacity increased dramatically between stages IV and VI (1.2-16.2 pmol/2 gonads), when testicular growth is extremely rapid. GH-R concentration in the liver was highest in early stages (6.5-6.7 pmol/g liver); this value then significantly decreased during midspermatogenesis (stage VI: 3.5 pmol/g liver) and returned to immature levels at the end of the cycle. No significant correlation was found between GH-R in testis, GH-R in liver, and plasma GH concentration. Preliminary experiments performed on isolated populations of testicular cells revealed that binding of 125I-rtGH was detectable in Sertoli cell-enriched fractions but not in germ cell populations. We conclude that GH-R are present in the testis during the entire reproductive cycle in rainbow trout, probably in somatic cells and in particular in Sertoli cells.

Animals↗

PubMed2000

Changes in the testis interstitium of Sprague Dawley rats from birth to sexual maturity.

Changes in the rat testis interstitium from birth to adulthood were studied using Sprague Dawley rats of 1, 7, 14, 21, 28, 40, 60, and 90 days of age. Our objectives were 1) to understand the fate of the fetal Leydig cells (FLC) in the postnatal rat testis, 2) to determine the volume changes in testicular interstitial components and testicular steroidogenic capacity in vitro with age, 3) to differentially quantify FLC, adult Leydig cells (ALC), and different connective tissue cell types by number and average volume, and 4) to investigate the relationship between mesenchymal and ALC numbers during testicular development. FLC were present in rat testes from birth to 90 days, and they were the only steroidogenic cells in the testis interstitium at Days 1 and 7. Except for FLC, all other interstitial cell numbers and volumes increased from birth to 90 days. The average volume of an FLC and the absolute volume of FLC per testis were similar at all ages except at Day 21, when lower values were observed for both parameters. FLC number per testis remained constant from birth through 90 days. The observations suggested that the significance of FLC in the neonatal-prepubertal rat testis is to produce testosterone to activate the hypothalamo-hypophyseal-testicular axis for the continued development of the male reproductive system. ALC were the abundant Leydig cell type by number and absolute volume per testis from Day 14 onwards. The absolute numbers of ALC and mesenchymal cells per testis increased linearly from birth to 90 days, with a slope ratio of 2:1, respectively, indicating that the rate of production of Leydig cells is 2-fold greater than that of mesenchymal cells in the postnatal rat testis through 90 days. In addition, this study showed that the mesenchymal cells are an active cell population during testis development and that their numbers do not decrease but increase with Leydig cell differentiation and testicular growth up to sexual maturity (90 days).

11-beta-Hydroxysteroid Dehydrogenases↗

PubMed1981

Changes in Leydig cells and luteinizing hormone receptors in porcine testis during postnatal development.

LH receptors have been characterized in porcine testis (Sus crofa L.) from birth to 220 days of age and have been related to interstitial tissue development (especially Leydig cells). The mean association constant (Ka) of ovine [3H]LH, was 7 +/- 6 X 10(9) M-1, with no apparent age-dependent variation but with some significant individual variations. The concentration of specific LH receptor sites reached a maximum of 9 X 10(-12) M/g testis between days 20-70, decreased to 3 X 10(-12) M/g testis at the onset of puberty (up to 100 days), and remained stable at the same level in the adult. The total number of sites per testis essentially reflected the growth of the testis. Interstitial tissue occupied up to 80% of the volume of the whole testis during the occupied up to 80% of the volume of the whole testis during the first 30 neonatal days. It decreased to about 25% after 120 days. The Leydig cells in this tissue occupied the same proportion (70% of the volume of the whole testis) regardless of the age of the animals. The mean Leydig cell diameter reached a maximum of 20 micrometer at 30 days of neonatal life, decreased to a minimum of 10 micrometer at 90 days, and then increased to a stable value of 15 micrometer after puberty. The number of Leydig cells per unit volume varied with age, with a maximum of 3 X 10(8) cells/ml testis at 90 days, reaching a constant value of 6 X 10(7) cells/ml testis after puberty. These data suggest that each Leydig cell contains 80,000 specific LH-binding sites/cell 30 days after birth and 35,000 in the adult, with no observed periods without receptor sites. The number of receptors per cell is correlated to cell size rather than stage of sexual maturation. (Endocrinology 108: 625, 1981)

Aging↗

PubMed1992

Characterization of epidermal growth factor in mouse testis.

Considerable evidence exists to suggest that epidermal growth factor (EGF) influences spermatogenesis directly. The tissue source of this EGF, however, is not yet clear. In this study we examine whether the testis itself can serve as a source of EGF. Gel filtration fractions of acid extracted testes exhibited the ability to displace 125I-EGF from testis membranes. The testicular fractions containing the 125I-EGF displacement activity coeluted within the same range as those of submandibular gland (SG) fractions containing mature EGF and prepared in an identical fashion. Next, we employed specific antisera probes to investigate first, whether the testis synthesizes this EGF displacement activity and second, to determine the cell distribution of the testicular EGF. Two types of antisera probes were employed: 1) commercially available antisera to mature EGF (EGFm), i.e. the 6,000 M(r) peptide, and 2) polypeptide specific antisera to the C-terminus of the EGF precursor (EGFp), i.e. the 140,000 M(r) integral membrane molecule which exhibits seven EGF-like repeats in addition to the EGFm. Metabolic labeling of testis with 35S-methionine was performed, followed by immunoprecipitation with the anti-EGFm antisera. Parallel studies using kidney and SG were used as positive controls. Fluorograms exhibited a prominent band at M(r) 140,000 for testis and kidney, corresponding to the EGFp. There was, in addition, a M(r) 50,000 band present for the testis. In SG, a band at M(r) 6,000, corresponding to EGFm, in addition to bands at M(r) 21,000 and 46,000 were observed also. Immunoblotting of testis, kidney, and SG membrane preparations with the specific antisera to either the EGFm or EGFp also resulted in identifying the EGFp at M(r) 140,000, as well as other lower mol wt bands. Preadsorption of anti-EGFm antisera with excess EGFm eliminated all of the specific bands that were immunoblotted. Peroxidase immunocytochemistry of testis, kidney, and SG was also performed using the specific antisera to either EGFm or EGFp. EGFp and EGFm staining in SG and kidney was identical to previously published results in which the distribution of EGFm in these tissues was established. In testis, EGFm immunostaining showed positive results in Sertoli cells, pachytene spermatocytes and round spermatids. In contrast, EGFp immunostaining was limited to pachytene spermatocytes and round spermatids. These results suggest that the testis must now be included in the list of tissues capable of synthesizing EGFp. Specifically, EGFp synthesis appears limited to the post meiotic germ cells.

Animals↗

PubMed2003

Dax1 regulates testis cord organization during gonadal differentiation.

Mutations of the DAX1 nuclear receptor gene cause adrenal hypoplasia congenita, an X-linked disorder characterized by adrenal insufficiency and hypogonadotropic hypogonadism. Targeted deletion of Dax1 in mice also reveals primary testicular dysgenesis, which is manifest by obstruction of the rete testis by Sertoli cells and hyperplastic Leydig cells, leading to seminiferous tubule dilation and degeneration of germ cells. Because Dax1 is expressed early in gonadal development, and because Sertoli and Leydig cells are located ectopically in the adult, we hypothesized that these testis abnormalities are the result of an early defect in testis development. In Dax1(-/Y) males, the gonad develops normally until 12.5 dpc. However, by 13.5 dpc, the testis cords are disorganized and incompletely formed in Dax1-deficient mice. The number of germ and Sertoli cells is unchanged, and the expression of Sertoli-specific markers appears to be normal. However, the number of peritubular myoid cells, which normally surround the testis cords, is reduced. BrdU labeling of peritubular myoid cells is low, consistent with decreased proliferation. The basal lamina produced by peritubular myoid and Sertoli cells is disrupted, leading to open and incompletely formed testis cords. Leydig cells, which normally reside in the peritubular space and extend from the coelomic surface to the dorsal surface of the gonad, are restricted to the coelomic surface of Dax1-deficient testis. We conclude that Dax1 plays a crucial role in testis differentiation by regulating the development of peritubular myoid cells and the formation of intact testis cords. The developmental abnormalities in the Dax1-deficient testis lay the foundation for gonadal dysgenesis and infertility in adult mice and, potentially in humans with DAX1 mutations.

Animals↗

PubMedDate not supplied

Peptide and nonpeptide reactive oxygen scavengers provide partial rescue of the testis after torsion.

Ischemia-reperfusion (IR) of the testis results in germ cell-specific apoptosis, followed by a reduction in testis weight and daily sperm production (DSP). This has been associated with an increase in the adhesion of neutrophils to testicular subtunical venules and an increase in reactive oxygen species (ROS). The present study investigated: 1) the effects of a direct, non-IR-related ROS insult to the testis and 2) the effects of catalase, superoxide dismutase (SOD), and a novel nonpeptide mimic of SOD, M40403, on neutrophil recruitment, ROS production, testis weight, and DSP following IR of the rat testis. Results revealed that the infusion of H2O2 increased testicular lipid peroxidation 1 hour after administration and increased germ cell apoptosis within 24 hours of administration. Four hours after the repair of torsion plus vehicle infusion, there was a significant increase in myeloperoxidase (MPO) activity, an indicator of neutrophil accumulation, and thiobarbituric acid reactive substances (TBARS), a measure of ROS production, compared to equivalent data in sham-treated testes. Animals sacrificed 30 days after the torsion plus vehicle infusion revealed a significant decrease in testis weight and DSP compared to the same parameters in sham-operated animals. The treatment of animals with catalase plus SOD or M40403 showed a significant decrease in MPO activity and TBARS 4 hours after IR of the testis. Animals treated with SOD, SOD plus catalase, and M40403 provided a partial rescue of DSP 30 days after IR of the testis. These results demonstrate that oxidative stress can directly cause germ cell apoptosis, even outside the IR model, and confirm the importance of oxidative stress in testicular IR injury. Also, following testicular IR, there is a recruitment of neutrophils and an increase in ROS production in the testis. The administration of ROS scavengers significantly reduced the IR-induced responses. Interestingly, the administration of all ROS scavengers also blocked neutrophil recruitment to the testis. The mechanism by which ROS modulates neutrophil adhesion to venules is presently under investigation, as are additional therapeutic regimens to block oxidative stress.

Animals↗

PubMed1990

[Vascular morphology of the bovine testis. Light and scanning electron microscopic studies].

The testicular artery of the bull-testis shows a straight course from the end of the pampiniform plexus to the caudal extremity of testis. There it branches off in Rami tunicales, which lie as stratum arteriosum superficially to the albugineal veins of testis: a multi-layered stratum venosum. Arterial Rami parenchymales centripetales run directly to the mediastinum testis, form coils and then divide in approximately 10 or more thinner Rami parenchymales centrifugales, which extend from the coils into the parenchyma of the gonad. The three-dimensional microvasculature of the bull-testis is strikingly different from that of rodents: The peritubular network of capillaries in the interstitial space is positioned in a more irregular way. Only here and there is discernible a rope-ladder-like or polygonal arrangement of capillaries. A subalbugineal plexus does not exist in the bovine testis. Parenchymal veins drain in albugineal veins and these empty in the venous networks of the pampiniform plexus. Valves are a rare finding in testicular veins. Already low perfusion-pressure easily forces the corrosion-compound to leave the capillary bed and form typical extravasations as bent shovel-like plates, thus filling the clefts of peritubular spaces. Arteries and veins are directly embedded in the parenchyma of testis, surrounded only by a relatively thin margin of perivascular connective tissue. There are no septula testis and therefore a lobular organisation of bovine testis does not exist. The angioarchitecture of the testis plays an important role in thermoregulatory and androgen-transfer mechanisms as well as in the transport of rete-fluid to the epididymis.

Animals↗

PubMed1996

[Acquired non-scrotal testis in boys: an underrated phenomenon].

OBJECTIVE: To differentiate undescended testis into a congenital and an acquired form using earlier information on testis position. DESIGN: Descriptive. SETTING: Paediatric outpatients' clinic, Medical Centre Alkmaar, the Netherlands. METHOD: In a 3-year period (1991-1994), 77 boys were referred to the paediatric outpatients' clinic for non-descended testis. The testis positions in their earlier years were documented. The undescended testis was defined according to these data as a congenital or an acquired condition. RESULTS: In 23 boys (age: 2.2-12.7; mean: 7.6 years) the testis turned out to be retractile. In 25 boys (0.1-14.3 years; mean: 3.1) the diagnosis was congenital undescended testis; 21 of these underwent orchidopexy. In 29 boys (1.9-14.3 years: mean: 9.9) the non-descent was an acquired condition. Ten of these boys were treated with orchidopexy as initial therapy. In eleven hormonal therapy was given (human chorionic gonadotrophin administered by intramuscular injection) resulting in a fully descended position of the testis in 8 boys. CONCLUSION: The phenomenon of acquired non-descended testis is frequent. Incidence and aetiology are insufficiently known. There is no general agreement whether the condition should be treated and which treatment should be favoured. Very likely, the relatively high incidence of orchidopexy operations in the Netherlands is mainly due to operative treatment of the acquired undescended testis.

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