Gonadal Levels Research Articles

The Effects of Triiodothyronine on Rat Testis: A Morphometric and Immunohistochemical Study

The aim of present study was to investigate the effects of 3,3',5-triiodothyronine (T(3)) on rat testis both morphometrically and immunohistochemically with determining of insulin-like growth factor I (IGF-I) expression. Adult male Wistar-albino rats used in the study were divided into two groups; control and T(3)-treated groups. After T(3) treatment there was observed to be a decrease in testicular weights, diameters of seminiferous tubules and the number of sertoli cells, and an increase in the number of leydig cells (P<0.05). Some of the seminiferous tubule lumens of T(3) administrated rats had cellular debris. IGF-I was localized in sertoli cells, late spermatids and leydig cells of all groups. IGF-I immunoreactivity in T(3) treated rats was higher than in controls in all stages of the cycle of rat seminiferous epithelium, but the staining intensity of leydig cells were similar in both groups. In conclusion, the present results suggest that T(3) may modulate the testicular function by affecting IGF-I activity at the gonadal level.

Male or Female? It Depends on the Dose

Male or Female? It Depends on the Dose

Occupational Radiation Exposure from Fluoroscopically Guided Percutaneous Transhepatic Biliary Procedures

The aim of this study was to determine occupational dose levels for projections commonly used in fluoroscopically guided percutaneous transhepatic biliary (PTB) drainage and stent placement procedures. Exposure data from 71 consecutive PTB examinations were analyzed to determine average examination parameters for biliary drainage and stent placement procedures. An anthropomorphic phantom was exposed at three projections common in PTB interventions according to the actual geometric parameters recorded in the patient study. Scattered air-kerma dose rates were measured for neck, waist, and gonad levels at various sites in the interventional radiology laboratory. To produce technique- and instrumentation-independent data, dose rate values were converted to dose-area product (DAP)-normalized air-kerma values. In addition, sets of thermoluminescent dosimetry crystals were placed in both hands of the interventional radiologist to monitor doses during all PTB procedures. Isodose maps of DAP-normalized air-kerma doses in the interventional laboratory for projections commonly used in PTB procedures are presented. To facilitate effective dose estimation, normalized dosimetric data at the interventional radiologist's position are presented for left and right access drainage procedures, metallic stent placement only, and drainage and metallic stent placement in one-session procedures with and without under-couch shielding. Doses to the hands of interventional radiologists are presented for left and right transhepatic biliary access and metallic stent placement. Body level-specific normalized air-kerma distributions from commonly used projections in PTB procedures may be useful to accurately quantify dose, maximum workloads, and possible radiogenic risks delivered to medical personnel working in the interventional radiology laboratory. Normalized dose data presented will enable occupational exposure estimation from other institutions.

The Influence of Depression, Body Mass Index, and Smoking on Serum Inhibin B Levels in Late Reproductive-Aged Women

Women experiencing depression have difficult psychosocial functioning, and recent data suggest an earlier onset of menopause. Understanding the biological mechanism for the impairment of reproductive function associated with depression is important. The objective of the study was to determine whether a lifetime history of depression is associated with reduced ovarian reserve as reflected in serum levels of the granulosa cell product, inhibin B. Residual serum samples from a subset of patients in the Harvard Study of Cycles and Moods were collected. Patients were recruited from seven Boston-area communities. Women with or without a history of major depression, based on structured clinical interviews for Diagnostic and Statistical Manual of Mental Disorders, fourth edition, were enrolled. A subset of patients who had provided an early follicular phase blood specimen at study enrollment and two or more other samples over the first 18-month period of follow-up were included. There were no interventions. Serum inhibin B levels were measured. Serum FSH levels were higher in women with a history of depression, whereas inhibin B levels did not differ between groups. Body mass index and age were significantly and inversely related to serum inhibin B levels. Smoking history was noted, for the first time, to have a significant negative association with inhibin B levels. Smoking has a direct negative effect on ovarian reserve, as suggested by decreased serum inhibin B levels. In contrast, effects of depression on the reproductive axis may occur at the level of the pituitary and/or hypothalamus rather than at the gonadal level, as suggested by increased serum FSH levels.

Alteration of Steroid Hormone Balance in Juvenile Turbot (Psetta maxima) Exposed to Nonylphenol, Bisphenol A, Tetrabromodiphenyl Ether 47, Diallylphthalate, Oil, and Oil Spiked with Alkylphenols

In recent years, concern has been raised about the ability of some classes of environmental contaminants to disrupt the endocrine system of both humans and wildlife. In this study, juvenile turbots (Psetta maxima) were exposed under laboratory conditions to selected waterborne contaminants: oil, oil spiked with alkylphenols, bisphenol A, diallylphthalate, tetrabrominated diphenyl ether 47, and p-nonylphenol as a positive control for "estrogenic-type" effects. This work focused on sex steroids, because these hormones play a key role in the reproduction process. Analytical procedures, involving the off-line coupling of solid phase extraction and gas chromatography/mass spectrometry, were developed for the determination of 12 endogenous sex steroids levels in fish plasma, bile, and gonads. Because of the sexual immaturity of the fish used in this study, however, only six steroids could be detected in juvenile turbots. Bisphenol A and p-nonylphenol exhibited the highest potency towards steroids dynamics, lowering the ratio of androgens to estrogens in all three studied matrices. However, these two chemicals had different modes of action, because p-nonylphenol induced a decrease of androstenedione and 11-ketotestosterone levels, whereas bisphenol A exposure led to an elevation of estrone level. Overall, these two chemicals seemingly disrupted the activity of some steroidogenesis enzymes, leading to serious hormonal imbalance in juvenile turbot.

Testatin transgenic and knockout mice exhibit normal sex-differentiation

Testatin is identified as a member of the Cystatin family and expressed in germ cells and somatic cells in reproductive tissues. Testatin transcription detectable in males and females at 9.5 days post coitum, before sex-differentiation, is up-regulated just after the onset of sry expression in the male gonads, while is down-regulated to undetectable levels in the female gonads. These expression patterns suggest that Testatin might have some roles in sex-differentiation. To address Testatin function in sex-differentiation, we analyzed the effects of ectopic-expression in females and null-expression in males with testatin transgenic and knockout mice. In the transgenic females, testatin expression was constitutively elevated from embryonic gonad to adult ovary, and its expression was as high as the wild-type male gonads. However, both types of mice were fertile and did not exhibit detectable abnormalities. This suggests that the decrease of endogenous testatin in female gonad is not critical, and the increase of testatin in male gonad is dispensable for sex-differentiation.

Promoter Activity and Chromosomal Location of the Rana rugosa P450 Aromatase (CYP19) Gene

Sex is determined genetically in amphibians, but is reversed occasionally by steroid hormones. The phenotypic sex of some amphibian species can be reversed from male to female by estrogens. Estrogens, which are synthesized from testosterone irreversibly by the enzyme P450 aromatase (CYP19), are essential for ovarian development in vertebrates. CYP19 expression is reportedly regulated by steroidogenic factor-1 (SF-1), also designated as Ad4BP, in fish and mammals. However, it is unknown if this is also the case in amphibians. Thus, to elucidate the role of SF-1 in CYP19 gene expression in the gonad of amphibians, it is necessary to isolate and characterize the promoter region of the CYP19 gene of amphibians. For this purpose, we first cloned the promoter region of CYP19 from genomic DNA fragments of the frog Rana rugosa. As a result, a potential binding site of SF-1 was found in the region. When a luciferase promoter assay in HEK 293 cells was carried out to examine the ability of SF-1 as a transcriptional regulator, we found that R. rugosa SF-1 stimulated the expression of the CYP19 gene of the tilapia Oreochromis niloticus, but not that of the frogs R. rugosa and Xenopus laevis. RT-PCR analysis revealed that CYP19 mRNA was expressed at a higher level in the indifferent gonads of females than in those of males. This was also true to SF-1 mRNA In addition, FISH analysis showed that the CYP19 gene was located on chromosome 3 of R. rugosa. Taken together, our data suggest that CYP19, an autosomal gene, is expressed in the undifferentiated gonads to an extent greater in females than in males, but its expression probably is not regulated by SF-1 alone. Another factor(s) may be required if SF-1 promotes the expression of the CYP19 gene in R. rugosa as it does in fish and mammals.

Pisrt1, a gene implicated in XX sex reversal, is expressed in gonads of both sexes during mouse development

XX sex reversal syndromes not involving Sry provide an opportunity to identify and study genes important for sexual development. The polled intersex syndrome (PIS) in goats, which shares some features with blepharophimosis, ptosis, epicanthus inversus syndrome (BPES) in humans, exemplifies such syndromes. BPES is caused by defects in the forkhead transcription factor gene FOXL2, while PIS is caused by a large deletion of goat chromosome 1q43 that affects transcription of the genes Pisrt1 and Foxl2. Pisrt1 is a non-translated gene that has a sexually dimorphic expression pattern in goats. Here, we describe the structure and expression of the mouse Pisrt1 locus, to investigate its likely role in ovarian development more broadly in mammals. This gene showed some sequence similarity, and was found in a similar genomic context, to its goat and human orthologues. Expression analyses indicated that Pisrt1 is transcribed, and its mRNA polyadenylated and exported to the cytoplasm, but no significant open reading frames were found in a 1.5 kb mouse genomic region corresponding to goat Pisrt1. Pisrt1 transcripts were expressed very broadly among tissues of the developing mouse embryo, and at similar levels in male and female gonads at each stage examined, as determined by in situ hybridisation and RT-PCR. This profile of expression suggests that Pisrt1 is unlikely to contribute to sex-specific events during gonadal development in mice and that divergent pathways of ovarian development operate among different mammalian species.

Genomic organisation and expression of the natural killer cell enhancing factor (NKEF) gene in channel catfish, Ictalurus punctatus (Rafinesque)

The gene encoding Natural Killer Cell Enhancing Factor (NKEF) was identified in clones from a gynogenetic channel catfish BAC library. NKEF gene sequence (5.2 kb) was obtained by direct sequencing of BAC clones. The catfish NKEF gene contains one non-coding exon, five coding exons and five introns. A putative TATA box and other transcription factor binding sites were identified in the promoter region. The NKEF amino acid sequence is highly conserved between fish and mammals. Gene expression, measured by real-time quantitative PCR, was detected in all major tissues with the highest level of expression in stomach and heart and lowest levels in gonad and pituitary gland. Catfish NKEF mRNA levels were slightly upregulated 8 and 24 h after injection of lipopolysaccharide. A TAA-repeat microsatellite was identified in a BAC clone containing the NKEF gene, and this locus contained at least 12 alleles in a random-bred catfish population. Multipoint linkage analysis in two reference families placed the NKEF gene on linkage group U18, 5.1 cM from locus IpCG0177 ( r = 0.05; LOD = 45.8), 6.0 cM from a novel immune type receptor, syntenic (40 cM) with T-cell receptor α, and not linked with MHC loci.

Effect of tributyltin, benzo( a)pyrene and their mixture exposure on the sex hormone levels in gonads of cuvier ( Sebastiscus marmoratus)

Tributyltin (TBT), an organometal used as an antifouling biocide, has been reported to induce masculinization of fish. Benzo( a)pyrene (BaP), a widespread carcinogenic polycyclic aromatic hydrocarbon, has been reported that its microsomal metabolites can produce an estrogenic response when tested in vitro. This study was therefore designed to examine the potential in vivo influence of TBT, BaP and their mixture on sex hormone levels in gonads of Sebastiscus marmoratus, which were given eight separate i.p. injections (a single injection every 7 days) of TBT (0.5, 1, 5 and 10 mg/kg), BaP (0.5, 1, 5 and 10 mg/kg), or both in combination (0.5, 1, 5 and 10 mg/kg); control fish received olive oil vehicle only. Six days after the first (week 1), second (week 2), fourth (week 4) and eighth (week 8) injection, gonads samples were collected and analyzed for sex hormone levels. TBT treatment alone was found to be ineffective at week 1, but significantly elevated the testosterone level in testicle of the male fish at week 4 compared to the corresponding controls. TBT treatment significantly reduced the ovarian testosterone level of the female fish at week 2 in dose-dependent manner. It was observed that TBT, BaP and their mixture significantly reduced the ovarian 17β-estradiol level of the female fish at weeks 2 and 8 in dose-dependent manner, however, the ratios of testosterone to 17β-estradiol in the ovary were elevated. This change of sex hormones levels would be one of the reasons to interpret the masculinization of fish by TBT. The present study demonstrates that BaP could influence in vivo ovarian sex hormone level of the female fish. The elevation of the ratios of testosterone to 17β-estradiol in the female fish exposed to BaP implies that BaP would have an androgenic effect on the fish in vivo, which should be deserving of further study. The joint effect of TBT and BaP at 1:1 concentration ratio on the level of 17β-estradiol in S. marmoratus was antagonism. TBT can antagonize bioactivation of BaP, and BaP can stimulate the Phase II metabolism of TBT and/or its biliary excretion, which were reported in previous studies, would be one of the causes that TBT and BaP had a antagonism on the level of ovarian 17β-estradiol in the present study.

Plasma vitellogenin levels and gonadal morphology of wild carp ( Cyprinus carpio L.) in a receiving rivers downstream of Sewage Treatment Plants

A field study was carried out in order to assess, in a wild cyprinid fish, the biological effects related to the exposure to estrogenic compounds. The gonadosomic index (GSI), plasma vitellogenin (VTG) level and histological observations of gonads were performed to evaluate the reproductive health in common carp ( Cyprinus carpio) from three rivers receiving sewage treatment plant (STP) effluents (Guadarrama, Henares and Jarama rivers, Spain) and from a reference site (Lozoya river). No significative differences in male GSI between sites were described. Only in the 18% of fish from Guadarrama river were elevated plasma VTG levels detected. The presence of oocytes in the testis was not found in any of males examined. The results suggest that the released estrogenic compounds by STPs effluents are not enough capable to adversely affect the gonadal development of the fish studied. Additional studies will be required to assess the risk of estrogenic compounds on fish from other sites of this area.

Puberty and recrudescence in cultured male yellowtail flounder: plasma androgens and spermatogenesis

Two year classes of cultured male yellowtail flounder Limanda ferruginea were followed in order to examine puberty and recrudescence at the gonadal level, and to assess the impacts of culture conditions on male reproduction. Cultured males demonstrated a propensity for maturation as 1 year‐old fish. Pubertal activation of testicular steroid production was associated with the appearance of primary spermatocytes, but a single case of a premeiotic endocrine activation was detected. The onset of puberty occurred during September and October, when adults were post‐spawned or in early recrudescence. Once initiated, pubertal development proceeded in an uninterrupted progression to full maturity by June at 22 months of age. Spermatogenetic activity during the autumn led to a peak in the gonado‐somatic index and total testicular mass by December. Milt could be first expressed from some pubertal males at this time, well in advance of the May to August spawning period in captivity. Meiotic activity could be absent from the testes by the spring, but residual areas of meiotic activity were found in pubertal males of one year class which may indicate a slower spermatogenetic rate. Androgen levels peaked during prespawning and early spawning; 11‐ketotestosterone (11‐KT) was the dominant androgen throughout spermatogenesis. Recrudescing 2 year‐old males showed greater testicular development and androgen production. Spermatogenetic patterns were similar to those seen in pubertal males, with the distinction that recrudescing males had a more discernable mitotic phase prior to meiotic activity; mitotic and meiotic activity overlapped considerably during early puberty. Testicular development well in advance of spawning may confer an evolutionary advantage to males of this cold‐water marine flatfish.

Sox8 is expressed at similar levels in gonads of both sexes during the sex determining period in turtles.

A critical gene involved in mammalian sex determination and differentiation is the Sry-related gene Sox9. In reptiles, Sox9 resembles that of mammals in both structure and expression pattern in the developing gonad, but a causal role in male sex determination has not been established. A closely related gene, Sox8, is conserved in human, mouse, and trout and is expressed in developing testes and not developing ovaries in mouse. In this study, we tested the possibility of Sox8 being important for sex determination or sex differentiation in the red-eared slider turtle Trachemys scripta, in which sex is determined by egg incubation temperature between stages 15 and 20. We cloned partial turtle Sox8 and anti-Müllerian hormone (Amh) cDNAs, and analyzed the expression patterns of these genes in developing gonads by reverse transcriptase-polymerase chain reaction and whole-mount in situ hybridization. While Amh is expressed more strongly in males than in females at stage 17, Sox8 is expressed at similar levels in males and females throughout the sex-determining period. These observations suggest that differential transcription of Sox8 is not responsible for regulation of Amh, nor responsible for sex determination in turtle.

Identification of novel genes of the bone-specific transcription factor Runx2.

The transcription factor Runx2 is a key regulator of osteoblast development and plays a role in chondrocyte maturation. The identification of transcriptional target genes of Runx2 may yield insight into how osteoblastic differentiation is achieved on a molecular level. Using a differential hybridization technique (selective amplification through biotin and restriction-mediated enrichment [SABRE]) and cDNA microarray analysis, 15 differentially expressed genes were identified using mRNA from C3H 10Tl/2 cells with constitutive and inducible overexpression of Runx2. Among the 15 genes identified, 4 encode the extracellular matrix proteins Ecml, Mgp, Fbn5, and Osf-2, three represent the transcription factors Esxl, Osrl, and Sox9, whereas others were Ptn, Npdc-1, Higl, and Tem l. The gene for Pttg1ip was upregulated in Runx2-expressing cells. Pttg1ip is widely expressed during development, but at highest levels in limbs and gonads. The Pttg1ip promoter binds Runx2 in a sequence specific manner, and Runx2 is able to transactivate the Pttg lip promoter in MC3T3-El cells. Therefore, Pttg1ip is likely tobe a novel direct transcriptional target gene of Runx2. In conclusion, the genes identified in this study are important candidates for mediating Runx2 induced cellular differentiation.

Sterol composition of gonad, muscle and digestive gland of Pecten maximus from Málaga (South Spain)

Sterol composition and content and their seasonal variations over 18 months were investigated in adductor muscle, digestive gland and gonads of Pecten maximus. Sterols were isolated by Silicagel 60 thin layer chromatography and identified by gas chromatography/mass spectrometry. Eleven sterols were identified, with cholesterol, brassicasterol, 24-methylenecholesterol and 22- trans-dehydrocholesterol being the principal components. The same sterols were found in all three tissues independent of season. The relative amounts of each sterol present in each tissue differed. Total sterol levels in gonad and muscle were higher than in digestive gland. Statistically significant differences ( P<0.05) were found between the concentrations of each of the sterols isolated from the gonad or muscle and digestive gland. The seasonal variations in the sterol content of the gonad seem be related to the reproductive cycle, while the sterol content of the digestive gland appears to be linked to diet, mainly diatoms or dinoflagellates. The muscle sterol content showed minor changes throughout the year.

Sexually dimorphic expression of multiple doublesex-related genes in the embryonic mouse gonad

The only molecular similarity shown so far for sexual regulatory genes among different phyla involves doublesex ( dsx) of Drosophila, mab-3 and mab-23 of Caenorhabditis elegans, and Dmrt1 of vertebrates. These genes encode DM domain transcription factors (DM= dsx and mab-3) and are required for sexual differentiation. In the case of dsx and mab-3, the two genes control analogous aspects of sexual development, bind similar DNA sequences, and are capable of functional substitution in vivo. All three phyla have multiple DM domain genes, but it is unknown how many of these are involved in sexual development. Mammals, for example, have at least seven DM domain genes, but embryonic expression has only been examined in detail for Dmrt1( dsx- and mab-3 related transcription factor 1). We have identified additional murine DM domain genes and have examined their expression in the mouse embryo, with emphasis on the developing gonad. At least three murine DM domain genes in addition to Dmrt1 are expressed in the embryonic gonad: Dmrt4 is expressed at similar levels in gonads of both sexes; Dmrt3 is more highly expressed in males; and Dmrt7 is more highly expressed in females. Expression of three other genes is low or absent in the embryonic gonad. Two of these, Dmrt5 and Dmrt6, are expressed primarily in the brain, and the third, Dmrt2, is expressed in presomitic mesoderm and developing somites. Our data suggest that multiple DM domain genes may be involved in mammalian sexual development, and that they may function in both testis and ovary development.

Treatment with an aromatase inhibitor induces complete sex change in the protogynous honeycomb grouper (Epinephelus merra)

To study the mechanism of protogynous sex change in honeycomb grouper, the wild fish of both sexes and transitionals were collected for one year, and changes in gonadal structures and serum levels of sex hormones in each individual were examined. The onset of sex change was associated with low serum estradiol - 17β (E2) levels. In order to clarify whether E2 deprivation causes sex change, maturing females were implanted with the non-steroidal aromatase inhibitor (Fadrozole, AI). AI caused complete sex change from female to functional males in two and half months. The present results suggest that E2 is an important endogenous factor which regulates protogynous sex change in the honeycomb grouper.

Sexually dimorphic gene expression in the developing mouse gonad

Over the course of a few days, the bipotential embryonic mouse gonad differentiates into either a testis or an ovary. Though a few gene expression differences that underlie gonadal sex differentiation have been identified, additional components of the testicular and ovarian developmental pathways must be identified to understand this process. Here we report the use of a PCR-based cDNA subtraction to investigate expression differences that arise during gonadal sex differentiation. Subtraction of embryonic day 12.5 (E12.5) XY gonadal cDNA with E12.5 XX gonadal cDNA yielded 19 genes that are expressed at significantly higher levels in XY gonads. These genes display a variety of expression patterns within the embryonic testis and encode a broad range of proteins. A reciprocal subtraction (of E12.5 XX gonadal cDNA with E12.5 XY gonadal cDNA) yielded two genes, follistatin and Adamts19, that are expressed at higher levels in XX gonads. Follistatin is a well-known antagonist of TGFβ family members while Adamts19 encodes a new member of the ADAMTS family of secreted metalloproteases.

Effects of chronic valproate treatment on reproductive endocrine hormones in female and male Wistar rats

Valproate (VPA) has been claimed to induce endocrine disorders in both sexes in humans. There is sparse information regarding the mechanisms behind these disturbances. By using an animal model, we wanted to study the effect of valproate on hormonal function in non-epileptic rats. Female rats were given 0 (vehicle control, n=15), 200 mg/kg ( n=15), or 300 mg/kg ( n=20) valproate twice daily by gavage for 90 days, resulting in mean valproate concentrations within the therapeutic range 4–6 h after the last dose given. Serum testosterone concentrations remained unchanged, while estradiol levels were significantly reduced in both treatment groups, leading to significantly increased testosterone/estradiol ratios. Follicle stimulating hormone (FSH) levels remained unaltered in valproate treated rats, whereas the luteinizing hormone (LH) concentrations were reduced at the lowest valproate dose. Male rats received 0 (vehicle control, n=15), 200 mg/kg ( n=15), or 400 mg/kg ( n=20) valproate twice daily by gavage for 90 days, resulting in mean valproate concentrations within the therapeutic range 4–6 h after the last dose. Serum testosterone levels were not significantly changed, but there was a highly significant increase in FSH and LH concentrations at the high dose. In conclusion, the study demonstrates a drug-induced effect of valproate on endocrine function in both male and female rats. The results indicate that the drug exerts its effect primarily at the gonadal level, although a centrally mediated effect cannot be ruled out.

Evaluation of Heavy Metals in Fish of the Sepetiba and Ilha Grande Bays, Rio de Janeiro, Brazil

Muscles, gonads, and liver tissues of fish caught in Sepetiba and Ilha Grande Bay were analyzed to assess levels of heavy metal concentrations of Cd, Fe, Zn, Pb, Ni, Cu, and Cr. Levels of Cr surpassed the maximum permissible concentration in muscle, followed by Zn in some species. Pb, Cu, and Cd presented concentrations above maximum permissible levels in gonads and liver only. Fe presented concentrations in viscera comparable to those of highly polluted areas, although there are no standards available for this metal. Ni was the only metal that did not present contamination in the fish tissues examined. Overall, fish caught in Sepetiba Bay showed higher metal concentrations than those from Ilha Grande Bay, but the latter also presented high metal concentrations in several samples.