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PubMed1998

Pyruvate dehydrogenase E1 alpha isoform in rat testis: cDNA cloning, characterization, and biochemical comparison of the recombinant testis and liver enzymes.

Previous data indicated a tissue-specific regulation of mitochondrial pyruvate dehydrogenase (PDH) complex, especially in the brain and testis. The lack of biochemical data on the rat testis PDH limits comparative analysis between testis and liver enzymes. Therefore, we have isolated a cDNA clone encoding rat testis PDH E1 alpha isoform, determined its nucleotide sequence, studied the tissue-specific expression, and characterized the recombinant protein produced in bacteria, compared to the liver counterpart. Our cDNA clone (2.2 kb) contained the identical open reading frame (from nt 974 to 2149) with that previously reported (Cullingford et al., 1993 Biochim Biophys Acta 1216:149-153) but contained a long 5' untranslated region, which has little identity to the other clone. Northern blot confirmed testis-specific expression of this isoform. Genomic DNA analyses by PCR amplification suggested this clone is a gene product distinct from its X-linked somatic counterpart. Our biochemical and kinetic analyses revealed that the purified recombinant rat testis PDH E1 (containing both E1 alpha and E1 beta subunits) was enzymatically active and phosphorylated in vitro by purified PDH-kinase p48 or p45, similar to the recombinant human liver enzyme. Our current data thus indicate that the differential regulation of testis PDH observed in the animal model may result from differential modulation of PDH-kinase or -phosphatase in this tissue rather than the presence of functionally different PDH E1 subunit.

Amino Acid Sequence↗
PubMed2005

Differential steroidogenic gene expression in the fetal adrenal gland versus the testis and rapid and dynamic response of the fetal testis to di(n-butyl) phthalate.

The phthalate ester di(n-butyl) phthalate (DBP) causes feminization of male rats upon in utero exposure by repressing expression of genes required for testicular steroidogenesis. Previous work in our laboratory has shown that repression of gene expression and steroidogenesis in the fetal testis is apparent within a few hours of DBP exposure. The purpose of this study was to determine the precise timing of DBP-associated gene expression changes in the fetal testis using transcriptional profiling and to determine whether DBP exerts similar effects on steroidogenesis in the fetal adrenal. A DBP time-course experiment showed that testicular steroidogenesis was decreased within 1 h of DBP exposure and that this decrease preceded the repressed transcription of Star (steroidogenic acute regulatory protein); Scarb1 (scavenger receptor class B, member 1; also know as Sr-b1); Cyp11a1 (cytochrome P450, family 11, subfamily a, polypeptide 1; also known as P450SCC); and Cyp17a1 (cytochrome P450 family 17, subfamily a, polypeptide 1; also known as Cyp17). Gene expression profiling demonstrated rapid (within 1 to 3 h) and transient induction of immediate early genes in the fetal testis after administration of DBP to the pregnant dam. There was a statistically insignificant decrease in corticosterone production by the fetal adrenal after in utero exposure to DBP from Gestation Day 12 to Gestation Day 19. The extent of steroidogenesis diminution was much less in the adrenal than in the testis (approximately 45% decrease in the adrenal versus 87% decrease in the testis) and expression of genes required for steroidogenesis in the adrenal was unaffected by DBP. Together, these studies demonstrate that DBP initiates a rapid and dynamic change in gene expression in the fetal testis that likely plays a role in the reduction in steroidogenesis that is unique to the fetal testis relative to the steroidogenically active fetal adrenal.

Adrenal Glands↗
PubMed2005

Identification of a novel testis-specific gene and its potential roles in testis development/spermatogenesis.

AIM: To identify and characterize a novel gene with potential roles in testis development and spermatogenesis. METHODS: A cDNA microarray was constructed from a human testis large insert cDNA library and hybridized with probes of human or mouse adult and fetal testes. Differentially expressed genes were isolated and sequenced. RT-PCR was used to test the tissue distribution of the genes of interest and in situ hybridization was performed to localize the gene expression in the mouse testis. A range of bioinformatical programs including Gene Runner, SMART, NCBI Blast and Emboss CpGPlot were used to characterize the new gene's feature. RESULTS: A novel testis-specific gene, NYD-SP5, was differentially expressed in fetal and adult testes. The deduced protein structure of NYD-SP5 was found to contain an IQ motif (a short calmodulin-binding motif containing conserved Ile and Gln residues), a Carbamate kinase-like domain, a Zn-dependent exopeptidase domain and a lactate dehydrogenase (LDH) C-terminal-like domain. RT-PCR analysis revealed that NYD-SP5 was predominantly expressed in the testis but not in other 15 tissues examined. In situ hybridization and RT-PCR examinations revealed that the expression of NYD-SP5 was confined in the male germ cell but not present in the somatic cell in the testes. CONCLUSION: NYD-SP5 is a newly found testis-specific gene with potential roles in testis development and spermatogenesis through a calmodulin-activated enzyme.

Amino Acid Sequence↗
PubMed2005

Mice lacking the UBC4-testis gene have a delay in postnatal testis development but normal spermatogenesis and fertility.

Activation of ubiquitination occurs during spermatogenesis and is dependent on the induction of isoforms of the UBC4 family of ubiquitin-conjugating enzymes. The UBC4-testis isoform is testis specific, is induced in round spermatids, and demonstrates biochemical functions distinct from a ubiquitously expressed isoform UBC4-1. To explore further the function of UBC4-testis, mice bearing inactivation of this gene were produced. Homozygous (-/-) mice showed normal body growth and fertility. Although testis weight and morphology were normal in testes from adult mice, examination of young mice during the first wave of spermatogenesis revealed that testes were approximately 10% smaller in weight at 40 and 45 days of age but had become normal at 65 days of age. Overall protein content, levels of ubiquitinated proteins, and ubiquitin-conjugating activity did not differ between wild-type and homozygous (-/-) mice. Spermatid number, as well as the motility of spermatozoa isolated from the epididymis, was also normal in homozygous (-/-) mice. To determine whether the germ cells lacking UBC4-testis might be more sensitive to stress, testes from wild-type and knockout mice were exposed to heat stress by implantation in the abdominal cavity. Testes from both strains of mice showed similar rates of degeneration in response to heat. The lack of an obvious phenotype did not appear to be due to induction of other UBC4 isoforms, as shown by two-dimensional gel immunoblotting. Our data indicate that UBC4-testis plays a role in early maturation of the testis and suggest that the many UBC4 isoforms have mixed redundant and specific functions.

Animals↗
PubMed1994

GLUT3 glucose transporter isoform in rat testis: localization, effect of diabetes mellitus, and comparison to human testis.

Facilitative hexose transporter expression was compared in rat and human testes. In rat testis, only GLUT1 and GLUT3 proteins were expressed. By contrast, human testis expressed GLUT1 and GLUT3 in addition to GLUT5. Immunocytochemical studies showed that GLUT3 was expressed in all cells of the seminiferous epithelium of rat testis, including sperm. In human testis, GLUT3 was expressed exclusively in cells juxtaposed to the lumen of the seminiferous tubule and ejaculate sperm, a pattern of expression that was identical to that of GLUT5. Induction of insulinopenic diabetes mellitus in the rat did not alter the levels or the distribution of GLUT3 protein or mRNA in the testis. Moreover insulin treatment of the diabetic rats did not produce changes in GLUT3 mRNA or protein levels. The results show that rat and human testis express the high-affinity glucose transporter GLUT3, which allows for the efficient uptake of glucose. In addition, the testis may be protected from changes in glucose transporter expression in experimental diabetes.

Animals↗
PubMed2006

Isolation and identification of F-spondin in the boar testis and its production during testis growth.

F-spondin/vascular smooth muscle cell growth-promoting factor (VSGP), purified from the follicular fluid of adult bovine ovaries, has been identified as a promoter of neuronal differentiation and vascular smooth muscle growth. The objectives of the present study were (1) to clarify whether F-spondin is also produced in the testis, which is ontogenically equivalent to the ovary, and (2) to examine whether production of this protein changes with testicular growth. To isolate F-spondin from the testis, testicular homogenates obtained from 8-week-old boars were sequentially subjected to heparin-Sepharose chromatography, diethylaminoethyl (DEAE)-Sepharose chromatography, and reverse-phase high-performance liquid chromatography (RP-HPLC). The isolated protein had a molecular mass of approximately 110 kDa and was cross-reactive with anti-F-spondin antibody by Western blotting. The purified protein was further characterized by amino acid sequence analysis of its internal peptide. The sequence obtained was GEQCNIVPDN VD, and a homology search indicated that the purified protein is a homologue of rat, human, and bovine F-spondin. By fractionation of the same amounts of testis tissue obtained from 1-, 8-, 16-, and 40-week-old boars, we analyzed age-related production of F-spondin in the testis. Western blotting of the fractions obtained from RP-HPLC revealed the presence of a band at approximately 110 kDa, corresponding to F-spondin, in the testes obtained from boars between 1 and 16 weeks old, but this band was not detected at 40 weeks. These results clearly indicate that (1) the porcine testis produces F-spondin and that (2) production of this protein is evident in the immature porcine testis, but not the adult testis.

Amino Acid Sequence↗
PubMed1996

[Retractile testis and gliding testis. Two distinct clinical entities].

The gliding testis is a testicle located below the external ring, can be manipulated to the upper scrotum, but is prone to ascend to its original position. Histologic changes can be detected in these gonads by 7 years of age. We evaluated 427 consecutive prepubertal boys referred for cryptorchidism. One hundred and twenty-three had classical undescended, 71 ectopic, 55 retractile, and 178 (mean age 6 years 2 moths) gliding testes. The gliding testes were smaller than controlaterals in 24% of boys. All gliding testes were unilateral whereas bilaterality was 85% in the retractile group (p < 0.0001), 17.5% in the undescended (p < 0.001), and 10% in the ectopic group (p < 0.01). A history for one or more of the following: orchidopexy (3), hormonal treatment (5), late testicular descent (9), spermatic cord torsion (5), testicular pain (10), actual retractile testes (20) or actual gliding testis (58), was present in 93 (52.2%) of the fathers of the gliding group. Forty-seven (81%) paternal gliding testes were hypotrophic. Seventy-five boys with gliding testis underwent initial hormonal therapy with transient benefit, and 57 were operated on. Two anatomical findings are typical of the gliding testis: the absence of the gubernaculum and a processus vaginalis partially patent from the upper scrotum to the mid groin area. This feature explains the mobility of the gliding testis from the external ring to the upper scrotum. The absence of the gubernaculum may be responsible for a higher incidence of spermatic cord torsion in this population. The gliding testis is a distinct entity, representing the mildest degree of a true undescended testis. As hormonal treatment gives only transient results, orchidopexy should be considered before testicular damage occurs.

Buserelin↗
PubMed1979

Induction of meiosis in fetal mouse testis in vitro by rete testis tissue from pubertal mice and bulls.

To test whether a meiosis-inducing substance (MIS) is responsible for the induction of meiosis in the testis at puberty, pubertal mouse rete testis was grown with (1) fetal undifferentiated mouse testis attached to the other side of a filter and (2) the used medium obtained from culture of the rete testis of a pubertal bull for 2 days. In both systems meiosis was induced in the fetal testis showing that MIS is not species specific. No meiosis-preventing effect was seen and it is concluded that meiosis in the testis is triggered at puberty as a result of the activity of the MIS concomitant with decreased activity of the meiosis-preventing substance.

Animals↗
PubMed2006

Transcription factor RFX2 is abundant in rat testis and enriched in nuclei of primary spermatocytes where it appears to be required for transcription of the testis-specific histone H1t gene.

Previous work in our laboratory revealed upregulated transcription of the testis-specific linker histone H1t gene in pachytene primary spermatocytes during spermatogenesis. Using the H1t X-box as an affinity chromatography probe, we identified Regulatory Factor X2 (RFX2), a member of the RFX family of transcription factors, as a nuclear protein that binds the probe. We also showed that RFX2 activated the H1t promoter in transient expression assays. However, other RFX family members have the same DNA-binding domain and they also may regulate H1t gene expression. Therefore, in this study we examined the distribution of RFX2 and other RFX family members in rat testis germinal cells and in several tissues. Among tissues examined, RFX2 is most abundant in testis. Testis RFX2 is most abundant in spermatocytes where transcription of the H1t gene is upregulated and the steady-state H1t mRNA level is high. RFX2 levels decrease but RFX1 levels increase in early spermatids where H1t gene transcription is downregulated. Antibodies against RFX2 generate a shifted band in electrophoretic mobility shift assays (EMSA) using H1t or testisin X-box DNA probes with nuclear proteins from spermatocytes. These data support the hypothesis that RFX2 expression is upregulated in spermatocytes where it participates in activating transcription of the H1t gene and other testis genes. These data also support the possibility that other RFX family members may bind to the H1t promoter in other testis germinal cell types and in nongerminal cells to downregulate H1t gene transcription.

Animals↗
PubMed2003

Orchidopexy or orchidectomy for preventing tissue hypoxia in contralateral testis following induction of ipsilateral abdominal testis associated with vas deferens obstruction.

INTRODUCTION: An experimental study was planned to evaluate and compare the effects of orchidopexy and orchidectomy on ipsilateral and contralateral testes in rats subjected to ipsilateral abdominal testis and vas deferens obstruction. MATERIALS AND METHODS: Four groups of 12 rats each were established. Sham operation, intra-abdominal testis with vas deferens obstruction and orchidopexy or orchidectomy for prior intra-abdominal testis with vas deferens obstruction were performed in groups 1, 2, 3 and 4, respectively. While testes were maintained for 8 weeks in the same position in groups 1 and 2, orchidopexy or orchidectomy was performed in groups 3 and 4 after the first 4 weeks, and the remaining testes were harvested after an additional 4 weeks. Lactic acid hypoxanthine contents were determined and the groups were compared with the paired t test. RESULTS: Maintaining intra-abdominal testis with vas deferens obstruction for 8 weeks and orchidopexy yielded the highest lactate values. However lactate levels in contralateral testes did not increase. On the other hand, hypoxanthine levels revealed the highest values after the initial 4 weeks. The 8-week study period resulted in increases of ipsilateral and contralateral testicular hypoxanthine levels. Orchidopexy caused a decrease in ipsilateral testicular values and ameliorated the increase in hypoxanthine levels in contralateral testes. CONCLUSIONS: Replacing an intra-abdominal testis with its vas deferens ligated into the scrotum ameliorates the oxidative stress in both ipsilateral and contralateral testes. Since orchidectomy does not result in better contralateral testicular values, orchidopexy should be preferred when treating an undescended testis with vasal obstruction.

Animals↗
PubMed2006

Detection of quantitative trait loci causing abnormal spermatogenesis and reduced testis weight in the small testis (Smt) mutant mouse.

The small testis (Smt) mutant mouse is characterized by a small testis of one third to one half the size of a normal testis, and its spermatogenesis is mostly arrested at early stages of meiosis, although a small number of spermatocytes at the late prophase of meiosis and a few spermatids can sometimes be seen. We performed quantitative trait locus (QTL) analysis of these spermatogenic traits and testis weight using 221 F2 males obtained from a cross between Smt and MOM (Mus musculus molossinus) mice. At the genome-wide 5% level, we detected two QTLs affecting meiosis on chromosomes 4 and 13, and two QTLs for paired testis weight as a percentage of body weight on chromosomes 4 and X. In addition, we found several QTLs for degenerated germ cells and multinuclear giant cells on chromosomes 4, 7 and 13. Interestingly, for cell degeneration, the QTL on chromosome 13 interacted epistatically with the QTL on chromosome 4. These results reveal polygenic participation in the abnormal spermatogenesis and small testis size in the Smt mutant.

Animals↗
PubMed1999

Quantitative evaluation of biopty gun testis needle biopsy. Correlation between biopsy score of varicocele-bearing testis and sperm count.

OBJECTIVE: To investigate the applicability of quantitative evaluation of needle biopsy of the testis and any correlation between biopsy score and sperm parameters in infertile or subfertile men with varicocele. STUDY DESIGN: A total of 45 infertile men with clinical left varicocele were included in the study. All patients underwent left varicocelectomy and bilateral biopty gun needle biopsy of both testes. Spermiograms were obtained before and three months after the operation. The biopsy specimens were evaluated for Johnsen and Agger score, Leydig cell score, germ cell/Sertoli cell ratio, mean tubular diameter, peritubular fibrosis, and tubular and basement membrane hyalinization. RESULTS: Mean sperm count, motility and normally configured motile sperm counts increased 20%, 25% and 60% by month 3, respectively (P < .05). We did not observe any significant change in normally configured sperm counts. A mean of 14 tubuli per testis were obtained with single-pass needle biopsy. Johnsen and Agger scores, Leydig cell scores, mean tubular diameter and germ cell/Sertoli cell ratios of both testes were comparable. However, there was significantly less peritubular fibrosis, tubular hyalinization and basal membrane hyalinization in the right testis when compared to the varicocele-bearing left testis (P < .05). We found positive correlations between Johnsen and Agger score of varicocele-bearing left testis and preoperative normally configured motile sperm counts (Pearson's r = .34 and P < .05 and Pearson's r = .41 and P < .05, respectively). The Leydig cell score of varicocele-bearing testis correlated inversely with sperm counts (Pearson's r = -0.37, P < .05). CONCLUSION: These observations may prove of prognostic value in infertile or subfertile men with varicocele.

Adult↗
PubMedDate not supplied

Unilateral injection of neuropeptide Y decreases blood flow in the injected testis but may also increase blood flow in the contralateral testis.

Neuropeptide Y (NPY) receptors have recently been described in intratesticular arterioles, but the role of NPY in testicular blood-flow regulation has not been examined. To explore this, we administered NPY in various doses (0.01-10 microg) via intratesticular injections and studied testicular microcirculation using a laser Doppler flow meter. NPY injection shows a dose-response pattern, with 1 microg (the most potent dose) causing a decrease (-42.4 +/- 3.7%, P < 0.00005) in blood flow in the ipsilateral testis of all the animals and an increase in blood flow in the contralateral testis (+17.2 +/- 5.6%, P = 0.03, n = 25 animals). The response in the contralateral testis was variable. A clear-cut increase was seen in 19 of the 25 animals examined, whereas either no response or a slight decrease was seen in the remaining six. The contralateral increase, which was not seen in the hindpaw on the same side, did not occur when the neuronal connections to the testes were blocked by injection of local anesthetics into the spermatic cord, either on the NPY-injected side or on the contralateral side. Our results suggest that NPY may serve as a vasoconstrictor in the testis, probably by acting on the NPY-Y1 receptors present on intratesticular arterioles. Local injection of NPY causes a major decrease in blood flow in the injected testis. This decrease is followed in the majority of animals studied by an increase in blood flow in the contralateral testis, an effect that seems to depend on neuronal mechanisms. This observation suggests that the testes may communicate under certain situations. The functional consequences of this remain to be elucidated.

Animals↗
PubMed2001

Stromal testis tumors in children: a report from the prepubertal testis tumor registry.

PURPOSE: Stromal testis tumors are rare and generally exhibit a benign behavior in prepubertal patients. We reviewed the Prepubertal Testis Tumor Registry to elucidate further the behavior of these tumors. MATERIALS AND METHODS: Epidemiological and clinical information on stromal testis tumors was compiled and reviewed from the Prepubertal Testis Tumor Registry. In addition, original pathology reports were requested for all patients registered as having undifferentiated stromal tumors. RESULTS: There were 43 patients registered with stromal tumors. Of the 21 patients with unspecified stromal tumors pathology reports were obtained on 11. Eight patients had truly mixed or undifferentiated stromal tumors. Mean patient age at presentation was 38 months (Leydig cell 70, Sertoli cell 52.5, juvenile granulosa cell 1.5 and mixed/undifferentiated 41.2). No patient with a Leydig cell, Sertoli cell or juvenile granulosa cell tumor had metastases at presentation or metastatic disease during an average 24.6 months of followup. One undifferentiated tumor demonstrated malignant behavior by presenting with metastatic disease. Pathological examination revealed a poorly differentiated tumor with extension into the adjacent tunica and frequent mitotic figures. While other stromal tumors displayed mitotic figures, none showed local invasion. CONCLUSIONS: Stromal testis tumors are rare. Data from the Prepubertal Testis Tumor Registry confirms the benign behavior of most of these tumors. However, undifferentiated stromal tumors may exhibit metastatic behavior. A high index of suspicion is appropriate when there are a large number of mitotic figures, the tumor is poorly differentiated or when local invasion is present in the primary tumor. Metastatic evaluation and close followup are warranted for this select group of patients.

Adolescent↗
PubMed1981

Accuracy of estimation of testis weight from in situ testis measures in ram lambs.

At a mean age of 93 +/- 5 days and a mean weight of 29 +/- 5 kg, 44 crossbred ram lambs were castrated to evaluate the accuracy of the estimation of testis weight from in situ testis measures (scrotal circumference and testis diameter). Means and standard deviations for testis weight (both testes), scrotal circumference and average in situ testis diameter were 134 +/- 57 g, 20.2 +/- 3.1 cm and 3.7 +/- .7 cm, respectively. Testis weight (W) was predicted from scrotal circumference (C) and average in situ diameter (D(o)) as W=.131C(1.90) D(o)(.88) (R(2)=.949). When adjusted to the same scrotal circumference and in situ diameter, testes of 3/4-Finnish Landrace rams were heavier (P<.05) than testes of 7/8-Dorset rams. However, the additional accuracy obtained by using equations specific to each crossbred group was small.

Journal Article↗
PubMed1999

Is hyperdiploidy of immature ejaculated germ cells predictive of testis malignancy? A comparative study in healthy normozoospermic, infertile, and testis tumor suffering subjects.

The possibility of diagnosing neoplastic testis pathologies by studying immature germ cells released from the seminiferous epithelium and present in the semen has been reported. It has been suggested that carcinoma in situ (CIS) of the testis and testis tumor may be identified by studying specific surface antigenic determinants or ploidy of chromosome 1 of malignant germ cells recovered from the semen. A noninvasive diagnostic approach of this type would be of great interest if we consider that CIS is supposed to precede the development of testicular germ cell tumors and that the frequency of that preneoplastic condition is increased in specific andrologic pathologies. To evaluate the reliability of this diagnostic approach, we have quantified the presence of immature hyperdiploid germ cells in the seminal fluid of normal healthy subjects, of infertile oligozoospermic patients affected by maldescended testis or left vancocele, and of patients suffering from CIS or testis tumor. Cell ploidy was evaluated on seminal cell fractions highly enriched in immature germ cells, by means of in situ hybridization with a DNA-probe specific for chromosome 1. Our observations indicate that chromosome 1 hyperdiploidy is not necessarily a predictive parameter of testis tumoral pathologies. The percentage of hyperdiploid immature seminal germ cells is in fact increased in nontumoral pathologies associated with infertility, such as cryptorchidism and left varicocele.

Adult↗
PubMed1981

Mechanism of damage to the contralateral testis in rats with an ischaemic testis.

A degenerating ischaemic rat testis produces damage to its contralateral testis. When ischaemic testes were grafted or injected as a homogenate for a week into littermate rats there was considerable damage to host testes. This damage was caused by an immunological response, demonstrated as an increase in cytotoxicity of host serum which increased immunofluorescence, particularly of the membrana propria, which had become thickened. Manipulation of the testis and its blood supply was sufficient to cause spermatogenic damage and initiate an immunological response. These results indicate that viable early torted testes must have their torsion corrected and be anchored in the scrotum, but if the testis is ischaemic and non-viable it should be removed to avoid damage to the contralateral testis.

Animals↗
PubMed1985

Immunolocalization of clusterin in the ram testis, rete testis, and excurrent ducts.

Clusterin, a glycoprotein that elicits cell aggregation, has previously been isolated from ram rete testis fluid, and has been partially characterized. In experiments reported, we have used monoclonal antibodies against clusterin in combination with indirect immunofluorescence microscopy to investigate the distribution of clusterin in the adult ram testis, rete testis, and excurrent ducts. Tissue blocks (5 mm3) were fixed in periodate/lysine/paraformaldehyde containing 0.1% glutaraldehyde and, after embedding, 5-microM sections were prepared for immunolocalization. In the testis, 2 basic patterns were observed: 1) strong to moderate staining for clusterin in the adluminal region with little staining in the basal region of the seminiferous epithelium and germinal cells; and 2) moderate staining throughout the seminiferous epithelium between germinal cells. In the rete testis, strong clusterin staining was localized intracellularly in the rete epithelial cells, most often associated with the luminal surface. In the epididymis, intracellular clusterin was localized in some principal cells of the caput epididymidis. The luminal surfaces and spermatozoa within the lumen were strongly positive. In the vas deferens, clusterin staining was associated with the luminal surface only. The presence of clusterin was clearly detected in unwashed isolated epididymal spermatozoa, but not in spermatozoa washed with phosphate-buffered saline containing 0.05% Tween 20.

Animals↗