Reliability of the grip strength coefficient of variation for detecting sincerity in normal and blocked median nerve in healthy adults.

In the assessment of hand and upper limb function, detecting sincerity of effort (SOE) for grip strength is of major importance to identifying feigned loss of strength. Measuring maximal grip strength with a dynamometer is very common, often combined with calculating the coefficient of variation (CV), a measure of the variation over the three grip strength trials. Little data is available about the relevance of these measurements in patients with median nerve impairment due to the heterogeneity of patient groups. This study examined the reliability of grip strength tests as well as the CV to detect SOE in healthy subjects. The power distribution of the individual fingers and the thenar was taken into account. To assess reliability, the measurements were performed in subjects with a median nerve block to simulate a nerve injury. The ability of 21 healthy volunteers to exert maximal grip force and to deliberately exert half-maximal force to simulate reduced SOE in a power grip was examined using the Jamar® dynamometer. The experiment was performed in a combined setting with and without median nerve block of the same subject. The force at the fingertips of digits 2-5 and at the thenar eminence was measured with a sensor glove with integrated pressure receptors. For each measurement, three trials were recorded subsequently and the mean and CV were calculated. When exerting submaximal force, the subjects reached 50-62% of maximal force, regardless of the median nerve block. The sensor glove revealed a significant reduction of force when exerting submaximal force (P1 sensor) with (P<0.032) and without median nerve block (P<0.017). An increase in CV at submaximal force was found, although it was not significant. SOE can be detected with the CV at the little finger at using a 10% cut-off (sensitivity 0.84 and 0.92 without and with median nerve block, respectively). These findings suggest low reliability of the power grip measurement with the Jamar® dynamometer, as well as that of the CV for detecting SOE. However, the combination of finger forces including the thenar area and the CV at the little finger could lead to better reliability for detecting feigned reduction of grip strength. The methods were as reliable in subjects with a median nerve block as in healthy subjects.

Very low-dose (femtomolar) 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) disrupts steroidogenic enzyme mRNAs and steroid secretion by human luteinizing granulosa cells.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is the most toxic congener of the polyhalogenated aromatic hydrocarbons (PAH), which causes anatomical abnormalities and developmental defects, impairs ovulation and reduces fertility. TCDD's endocrine-disrupting effects are, in part, caused by a direct action at the ovary. Herein we investigated the in-vitro effects of environmentally relevant doses of TCDD on estradiol-17ß (E2) production by human luteinizing granulosa cells (hLGC) obtained from women stimulated for in-vitro fertilization (IVF). TCDD at all concentrations tested (3.1fM, 3.1pM and 3.1nM) significantly decreased E2 secretion when assayed for by radioimmunoassay (RIA). Herein we confirm that TCDD alters E2 secretion by hLGC in a time-, not dose-dependent fashion and are the first to show decreases in E2 secretion with fM concentrations of TCDD. Using real-time quantitative PCR (RT-qPCR), the decreased E2 secretion correlates with a decrease in the mRNA expression levels two enzymes in the estrogen biosynthesis pathway: CYP11A1 and CYP19A1.

Familiar and novel reproductive endocrine disruptors: xenoestrogens, dioxins and nanoparticles.

Environmental contaminants are known to exert endocrine-disrupting effects on the reproductive axis of animals. Many of these molecules can affect steroid biosynthesis or estrogen-receptor signaling by behaving as estrogen-like molecules ("xenoestrogens"), or by exerting estrogenmodulatory effects. Exposure to some compounds has been correlated with the skewing of sex ratios in aquatic species, feminization and demasculinization of male animals, declines in human sperm counts, and overall diminution in fertility of birds, fish, and mammals. We herein devote space to several classes of endocrine-disrupting compounds (EDCs), including estrogenic substances such as bisphenol A (BPA), molecules that can behave at times anti-estrogenically while activating the aromatic hydrocarbon receptor (AHR), such as dioxins (a known human carcinogen), and novel, ubiquitous molecules such as nanoparticles, particularly gold nanoparticles (GNPs), that appear to alter the sexsteroid biosynthetic pathway.

Environmental toxicants and effects on female reproductive function.

One of the most toxic substances known to humans, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD or dioxin), is also highly pervasive in the environment. It is created naturally in volcanic eruptions and forest fires, and anthropogenically in waste incineration, chlorination processes and certain plastics manufacture. From reports of large industrial and other accidents, or from experimental studies, dioxin exposure has been correlated in animal models and/or humans with chloracne of the skin, organ cancers, hepatotoxicity, gonadal and immune changes, pulmonary and other diseases such as diabetes, skewing of the sex ratio, and infertility. We have demonstrated that the aromatic hydrocarbon receptor (AHR) that binds dioxin in tissues is localized in zebrafish, rat and rhesus monkey (Macaca mulatta) ovaries and in rat and human luteinizing granulosa cells (GC) (among other tissues), that labeled dioxin is specifically localized to granulosa cells of the ovarian follicle as observed by autoradiography, and that incubations of GC or ovarian fragments with environmentally relevant concentrations (fM to nM) of dioxin inhibit estradiol secretion significantly. Our experiments show that in human, non-human primate, rat, trout, and zebrafish ovarian tissues, dioxin inhibits estrogen synthesis at some level of the steroid biosynthetic pathway, most likely by inhibiting transcription of mRNAs for or activity of side-chain cleavage (Cyp11a1 gene) and/or aromatase (Cyp19a1 gene) enzymes, or conceivably other steroidogenic enzymes/factors. Such an untoward effect on estrogen synthesis in females exposed to dioxin environmentally may predispose them to defects in aspects of their fertility.

2,3,7,8-Tetrachlorodibenzo-p-dioxin increases steady-state estrogen receptor-beta mRNA levels after CYP1A1 and CYP1B1 induction in rat granulosa cells in vitro.

Previous in-vitro investigations of rat granulosa cells (GC) have shown that 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) inhibits estrogen secretion and FSH-induced aromatase activity. Although TCDD exerted no effect on basal aromatase enzyme activity, TCDD did reduce steady-state aromatase mRNA levels in GC using competitive RT-PCR. TCDD is hypothesized to induce these changes through aromatic hydrocarbon receptor(AHR)-mediated gene transcription and the modulation of the estrogen receptor (ER)-signaling pathway. In this study we show that rat GC express mRNA for AHR and the AHR nuclear translocator (ARNT) as well as biomarkers of TCDD action, CYP1A1 and CYP1B1 mRNA. Basal CYP1A1 and ER-alpha mRNAs were present only in trace amounts. By relative RT-PCR analysis we showed that CYP1A1 and CYP1B1 mRNA were induced significantly by TCDD at 6 h and that induction of CYP1A1 was maintained throughout the experiment. Using competitive RT-PCR, we observed no significant change in the mRNA levels of ARNT between control and TCDD-treated GC. Both AHR and ER-beta mRNA levels increased significantly at 48 h with TCDD compared with controls. Since ER-beta mRNA was not increased significantly until 48 h in culture, we suggest that in rat GC, the observed ER-beta mRNA increase by TCDD might be a result of CYP1A1/CYP1B1 catalyzed estrogen metabolism and aromatase mRNA inhibition via AHR.

Effects of dioxin, an environmental pollutant, on mouse blastocyst development and apoptosis.

OBJECTIVE: To evaluate the effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; dioxin) on mouse embryo development and apoptosis. DESIGN: Controlled animal study. SETTING: Academic research environment. ANIMAL(S): Female mice (CB6F1) at 3 to 6 weeks of age and proven breeders (C578B46). INTERVENTION(S): Mouse embryos were obtained at the morula stage and cultured to the blastocyst stage in a pharmacologic dose of TCDD (3.1 microM) or a control medium. The morphology was assessed, and staining for apoptosis was performed. Immunohistochemistry for the presence of aromatic hydrocarbon receptor (AhR) was performed in another set of morula-stage embryos. MAIN OUTCOME MEASURE(S): The number of embryos developing from the morula to the blastocyst stage and number of apoptotic blastomeres in control vs. TCDD culture conditions. RESULT(S): No statistically significant differences were observed in the percentage of embryos reaching the blastocyst stage: 80.9% (115 of 142) in the TCDD-treated group, vs. 82.9% (121 of 146) in the control group. There was also no difference in the degree of apoptosis: 22.6 +/- 7.3% apoptotic cells (TCDD) vs. 25.3 +/- 9.7% (controls). Staining indicated the slight presence of aromatic hydrocarbon receptor in the morula-stage mouse embryos. CONCLUSION(S): TCDD at 3.1 microM did not alter the development of early mouse morula to blastocysts and did not significantly induce apoptosis in vitro.

Demonstration of 2,3,7,8-tetrachlorodibenzo-p-dioxin attenuation of P450 steroidogenic enzyme mRNAs in rat granulosa cell in vitro by competitive reverse transcriptase-polymerase chain reaction assay.

We investigated the effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), in prepubertal (PP) and adult (A) rat granulosa cells (GC) in vitro by examining the changes in estrogen secretion, aromatase enzyme activity and mRNAs for steroidogenic enzymes P450scc, 3beta-HSDI, P450arom; and for components of the AHR signaling pathway-CYP1A1, aromatic hydrocarbon receptor (AHR), and the AHR nuclear translocator protein (ARNT). In PP and A rat GC, TCDD (3.1 nM) reduced estrogen secretion at 48 h without altering aromatase enzyme activity. Addition of FSH (50 ng/ml) increased aromatase activity in GC with or without TCDD. FSH-induced aromatase activity was significantly reduced by TCDD (3.1 nM) at 48 h. Semi-quantitative RT-PCR showed a significant increase in CYP1A1 mRNA both at 24 and 48 h with TCAP, while a significant reduction in P450scc and P450arom mRNA was observed with competitive RT-PCR. All steroidogenic enzyme mRNAs were significantly lower in adults than in PP GC. We conclude that in rat GC, TCDD modulates the level of cytochrome P450 enzymes involved in the steroid biosynthetic cascade. This effect may be attributable to AHR interaction with dioxin-responsive elements present in the genes encoding these enzymes.

Follicle selection in cyclic guinea pigs with active immunization against inhibin alpha-subunit.

Experiments were conducted to elucidate the mechanisms of active immunization against inhibin on ovarian follicular development and selection in guinea pigs. Estrous cycle was synchronized in experimental guinea pigs by implanting progesterone containing tubes. Antibodies that bound 125I-labeled bovine inhibin were produced by all guinea pigs receiving the inhibin vaccine (recombinant ovine alpha-subunit in oil emulsion) without any effects on duration of the estrous cycle. Active immunization against inhibin increased the plasma concentrations of progesterone during the luteal phase and the plasma concentrations of estradiol but failed to increase the plasma concentration of follicle-stimulating hormone (FSH) during preovulatory period. The treatment also increased the number of corpora lutea (from 1.3+/-0.3 to 7.0+/-1.6 per each ovary), and preovulatory sized follicles (from 1.8+/-0.6 to 7.0+/-1.6 per each ovary), and follicles stained positively for inhibin alpha-subunit (from 2.3+/-0.5 to 6.3+/-1.3 per each ovary) significantly. The results indicate that active immunization against inhibin enhances ovulation rate by affecting the follicle selection and only dominant follicle can be stained for inhibin alpha-subunit in guinea pigs. This study is firstly to provide direct evidence that inhibins play important role in follicle selections in guinea pigs.

Estrous cycle-dependent changes in the expression of aromatic hydrocarbon receptor (AHR) and AHR-nuclear translocator (ARNT) mRNAs in the rat ovary and liver.

The aromatic hydrocarbon receptor (AHR) and AHR nuclear translocator protein (ARNT) mediate the toxic effects of a wide variety of halogenated and polycyclic aromatic hydrocarbons. While it can be assumed that AHR has an endogenous function, its role in reproduction is currently undefined. The present study seeks to examine the regulation of AHR and ARNT mRNAs in liver and ovarian tissues across the rat estrous cycle. Message for hepatic AHR was increased significantly on the morning of proestrus, and decreased dramatically by the evening of proestrus; while hepatic ARNT mRNA was significantly decreased between diestrus and the morning of proestrus, and between the evening of proestrus and the morning of estrus. Ovarian AHR mRNA was unchanged from diestrus to proestrus, and was decreased on the evening of proestrus. Changes in the expression of ARNT mRNA mirrored changes in the liver. To assess interaction between the AHR- and estrogen-receptor (ER)-signaling pathways and to test the hypothesis that estrogen regulates AHR mRNA, 25-day-old female rats were injected with either 17beta-estradiol, the ER antagonist ICI 182 780, or with vehicle, and hepatic AHR mRNA was measured. Treatment with estrogen or the estrogen antagonist did not alter the abundance of AHR mRNA in the liver. These data suggest that while estrogen may not be the key regulator of AHR mRNA expression, a factor associated with the rat reproductive cycle may be important in regulating the expression of both the AHR and ARNT genes in the ovary and liver.

Minimal endocrine alterations in rodents after consumption of lake trout (Salvelinus namaycush).

Polychlorinated biphenyls (PCBs) and dioxins are known to cause disruptions in circulating hormone concentrations, which may influence fertility and normal fetal development. Structure activity relationships have been determined for individual congeners, but it is unclear what impacts occur due to exposure to complex mixtures of chemicals found in the environment. Most laboratory studies of PCB exposure have used commercial mixtures in high doses, which may not be representative of environmental concentrations of individual congeners, nor accurately represent complex interactions of multiple contaminants. The present study investigated endocrine alterations in rats associated with the consumption of lake trout collected from three specific locations in the Great Lakes. Composite fish samples were analyzed for PCBs, organochlorines, and mercury and ranged from 415 ppb to 1,275 ppb for individual contaminants. Fillet composites were fed to timed-pregnant Long-Evans rats as 30% of their diet. Concentrations of total thyroxine and estrogen were not significantly different in offspring of dosed dams from that of controls. However, aromatase activity was lowered in all dosed groups as compared with controls. This may represent a lowered expression of the CYP 19 gene in exposed rats or may be due to the presence of one or more substances in the contaminants that are capable of altering the affinity of the aromatase enzyme for its normal endogenous substrate. It is also possible that the number of maturing follicles in the lake trout-fed rats may be fewer than controls, which would result in an overall reduction in the enzyme activity. Data regarding the endocrine effects of environmental contaminant mixtures found in fish from the Great Lakes Basin are still controversial. Additionally, information is scarce with respect to the F1 generation of laboratory animals following environmental maternal exposures, therefore, we investigated the reproductive-endocrine alterations in rat offspring associated with the consumption of lake trout (Salvelinus namaycush) collected from three areas in the Great Lakes.

Lack of effect of beta-naphthoflavone on induction of Nramp genes in adult rainbow trout Oncorhynchus mykiss.

Natural resistance-associated macrophage protein (Nramp) genes in rainbow trout, Oncorhynchus mykiss, were identified and characterized. The greatest mRNA level encoding these genes was in the developing ovary of rainbow trout. We evaluated the response of these genes to a certain aromatic hydrocarbon receptor (AHR) agonist. Adult rainbow trout were treated with beta-naphthoflavone (BNF) (50 and 100 mg/kg) for 48 h. Using reverse-transcriptase polymerase chain reaction with ovary and head kidney RNA and specific alpha and beta Nramp primers, a 400 bp Nramp-alpha- and a 400 bp Nramp-beta-specific cDNA were obtained. There were no changes in the alpha and beta Nramp mRNA levels in the ovary following BNF administration. CYP1A1 mRNA was increased in the ovary and kidney, suggesting the presence of AHR in rainbow trout ovary, while the AHR agonist produced no effect on Nramp mRNAs.

Secretion of ovarian inhibin and its physiologic roles in the regulation of follicle-stimulating hormone secretion during the estrous cycle of the female guinea pig.

To characterize inhibin secretion during the estrous cycle in guinea pigs, the concentrations of plasma inhibin, estradiol, progesterone, and FSH were determined. A significant positive correlation was observed between inhibin and estradiol throughout the estrous cycle. Plasma inhibin and estradiol started to increase a few days before ovulation (Day 0 = day of estimated ovulation), and decreased after ovulation. These two hormones remained low during the luteal phase. The immunoreactivity of inhibin alpha, betaA, and betaB subunits was colocalized in the granulosa cells of one or two healthy large follicles in the ovary before ovulation. There was no positive reaction of inhibin alpha and beta subunits in the corpora lutea or other follicles. Ovariectomy resulted in an abrupt decrease in plasma inhibin and a significant increase in plasma FSH. Injection of anti-inhibin serum into adult female guinea pigs induced an elevation in plasma FSH in a dose-dependent manner. This report presents the first description of sequential changes in plasma inhibin and estradiol during the estrous cycle of guinea pigs. Results suggest that inhibin is secreted mainly by granulosa cells of a few healthy large follicles in the ovary and that it plays an important role in the regulation of FSH secretion during the estrous cycle in guinea pigs.

Dioxin perturbs, in a dose- and time-dependent fashion, steroid secretion, and induces apoptosis of human luteinized granulosa cells.

Dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin; TCDD) is the most toxic congener of a large class of environmental pollutants. Several studies have shown that TCDD exposure reduced fecundity and ovulatory rate in rats and increased the incidence of endometriosis in monkeys. Recent work suggests that TCDD's endocrine-disrupting effects are, at least in part, caused by a direct action at the ovary. Although the factors involved in TCDD-induced toxicity are still under investigation, several studies have shown that TCDD induces programmed cell death, or apoptosis, in various tissues and may act in a similar fashion in the ovary. In the present study, we set out to evaluate the in vitro effects of TCDD on steroid secretion, specifically estradiol-17beta (E2) and progesterone, by human luteinized granulosa cells (LGC), and to further determine whether TCDD is capable of inducing apoptosis in this cell type. Human LGC were obtained from women participating in an in vitro fertilization program. Medium, with or without three different concentrations of TCDD and substrates [androstenedione (A4) or pregnenolone], was added to each culture. The media were collected at 4, 8, 12, 24, 36, and 48 h and were assayed by RIA. At 24 and 48 h, the LGC were fixed for assessment of DNA fragmentation via an in situ immunofluorescence technique. Transmission electron microscopy was also performed on LGC after 24 and 48 h with TCDD. TCDD, at all concentrations tested (3.1 pM, 3.1 nM, and 3.1 microM), significantly reduced E2 accumulation in the media at 8, 12, and 24 h, compared with controls. At 36 and 48 h, TCDD treatment (at 3.1 microM) caused a significant increase in E2, compared with controls. The effect of TCDD on E2 was abolished with the addition of A4. TCDD treatment did not alter progesterone accumulation. Apoptosis increased at 24 h with 3.1 microM TCDD, with no apparent effect at 3.1 nM. By 48 h, however, TCDD increased apoptosis in a dose-dependent manner. Transmission electron microscopy showed ultrastructural differences in LGC with 3.1 microM TCDD at 24 and 48 h. Collectively, the results of the present study suggest that TCDD perturbs E2 secretion by depletion of A4 precursor and increases apoptotic cell death of human LGC in a dose- and time-dependent fashion.

Reduced fertility in female mice lacking copper-zinc superoxide dismutase.

Copper-zinc superoxide dismutase (CuZn-SOD) is believed to play a major role in the first line of antioxidant defense by catalyzing the dismutation of superoxide anion radicals to form hydrogen peroxide and molecular oxygen. Recent studies have shown that missense mutations in this gene contribute, evidently through a gain-of-function mechanism, to about 20% of familial amyotrophic lateral sclerosis. To define further the physiologic role of this enzyme, a model of mice deficient in this enzyme was generated using gene targeting technology. Mice lacking this enzyme were apparently healthy and displayed no increased sensitivity to hyperoxia. However, they exhibited a pronounced susceptibility to paraquat toxicity. Most surprisingly, female homozygous knock-out mice showed a markedly reduced fertility compared with that of wild-type and heterozygous knock-out mice. Further studies revealed that although these mice ovulated and conceived normally, they exhibited a marked increase in embryonic lethality. These data, for the first time, suggest a role of oxygen free radicals in causing abnormality of female reproduction in mammals.

Modulation of ovarian follicle maturation and effects on apoptotic cell death in Holtzman rats exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in utero and lactationally.

Recent reports have described the reproduction-modulating and endocrine-disrupting effects following exposure to toxic substances such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Herein, we set out (1) to determine whether TCDD exposure exerts detrimental effects on follicle maturation in the Holtzman rat ovary and (2) to determine whether the effects of TCDD are mediated in part via apoptotic cell death. In certain species, dioxin exposure is correlated with reduced fecundity, reduced ovulatory rate, an increased incidence of endometriosis, and various reproductive cancers. Although some of the effects of TCDD are mediated via the hypothalamic-pituitary axis, direct effects on the ovary have also been observed. In the present study, an oral dose of 1 microgram TCDD/kg maternal body weight was administered on Day 15 of gestation. Female pups were sacrificed on Postnatal Day 21/22, and the ovaries were excised, fixed for histologic analysis, and analyzed in a double-blind paradigm. The analysis included a count and measurement and classification of preantral and antral follicles throughout the entire ovary. The contralateral ovary from each animal was analyzed for DNA fragmentation indicative of apoptotic cell death. The results indicate that TCDD treatment significantly reduced the number of antral follicles in the size classes 50,000 to 74,999 microns2 and > 100,000 microns2. We also observed a reduction in the number of preantral follicles less than 50,000 microns2. No difference was observed in the degree of apoptotic cell death in antral (50,000 to > 100,000 microns2) and preantral follicles (50,000 microns2 to > 75,000 microns2) between TCDD-treated and control-treated tissues. These data support the hypothesis that TCDD results in a diminution in the number of antral and preantral follicles of certain size classes in animals exposed during critical periods of development, but that apoptosis does not appear to be the underlying mechanism in these particular follicles. This does not preclude apoptosis occurring in pools of smaller precursor follicles.

Alterations to the pituitary-gonadal axis in the peripubertal female rat exposed in utero and through lactation to 2,3,7,8-tetrachlorodibenzo-p-dioxin.

The environmental pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; dioxin) is a potent disrupter of vertebrate endocrine systems. It was shown previously that in utero and lactational (IUL) exposure to TCDD resulted in a reduction in serum estradiol concentrations; however, the mechanism for this remains unknown. In the current study, the effects of perinatal exposure to TCDD on the pituitary-ovarian axis were examined. Pregnant rats were given a single oral dose of 1 microg TCDD/kg or vehicle as control on gestation Day 15, and female pups were killed on postnatal Day 21. Pituitaries were assayed for gonadotropin beta-subunit mRNA; additional pituitaries were cultured for 4 h and the media were assayed for FSH. Gonadotropin receptor mRNAs from vehicle- and TCDD-exposed animals were compared, with some ovaries cultured and the media assayed for estrogen secretion. LH, FSH, progesterone, and androstenedione concentrations were determined in serum. IUL exposure to TCDD resulted in a significant reduction of pituitary FSHbeta mRNA. Although estrogen output was shown to be reduced, neither serum FSH nor LH concentration was increased significantly, and FSH secretion in vitro was not altered. Similarly, serum progesterone and androstenedione were not altered by TCDD exposure, while in vitro estrogen secretion was significantly reduced. These data suggest that TCDD did not act on serum gonadotropin concentrations. The reduction in the concentration of serum estrogen appears to result from direct or indirect actions on the ovary at some point following androstenedione production.

In utero and lactational exposure of female Holtzman rats to 2,3,7,8-tetrachlorodibenzo-p-dioxin: modulation of the estrogen signal.

The environmental pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; dioxin) induces severe reproductive defects in male rats when exposure occurs in utero and during lactation. Yet there is currently a paucity of information regarding the effects of this exposure paradigm in females. In the current study, we examine the effects of TCDD during fetal and perinatal development on the estrogen-signaling system in peripubertal female rats. Pregnant Holtzman rats were given 1 microgram/kg TCDD or vehicle control by gavage on gestational Day 15. Body weights were reduced, though not significantly, on postnatal Day 21. While ovarian and uterine wet weights were not increased by TCDD exposure, the percentage of body weight attributed to the ovary was increased significantly. Through use of ribonuclease protection and gel-shift assays, exposed females were compared with nonexposed counterparts for estrogen receptor (ER) mRNA and DNA-binding activity in the following tissues: hypothalamus, pituitary (mRNA only), uterus, and ovary. ER mRNA levels increased in the hypothalamus, uterus, and ovary, and decreased in the pituitary. The results of the DNA-binding assays paralleled the mRNA results in the uterus, while DNA-binding activity was decreased in the hypothalamus and was unchanged in ovarian protein extracts. Circulating concentrations of estrogen were significantly lower in TCDD-exposed rats than in controls. These data suggest that the decrease in serum estrogen may be a cause of the alterations in ER mRNA; the changes in ER DNA-binding activity may indicate alterations in either translation or posttranslational receptor processing. Overall, this study shows that TCDD may act systemically in this model, and these effects should not necessarily be characterized as antiestrogenic.

Effects of follicle-stimulating hormone administration on oestradiol-induced cystic ovaries in guinea pigs.

A consistent defect in follicle-stimulating hormone (FSH) secretion is seen in humans with Polycystic Ovarian Syndrome (PCOS); therefore, we evaluated whether Metrodin (a highly purified urinary FSH) administration concurrent with cyst induction or following cyst induction inhibits estrogen-induced cyst development and augments ovarian follicular growth in an established guinea pig model. All animals in these studies received subcutaneous implants containing oestradiol-17 beta (E2)-filled Silastic capsules for a 48-hour period. Guinea pigs in study #1 were administered four 0.25 mL injections of FSH or placebo at twelve-hour intervals simultaneously with the E2 treatment; guinea pigs assigned to study #2 were administered four 0.25 mL injections of FSH or placebo at twelve-hour intervals following the induction of the cystic condition by E2. Exogenous FSH appears to negate cyst formation when superimposed upon the cyst-inducing agent (E2). Further, treatment with FSH augmented the number of mid-sized follicles in both paradigms. This study is the first to establish evidence of an anti-cystic effect of FSH in an animal model.

Estrogen receptor and aromatic hydrocarbon receptor in the primate ovary.

We have previously shown by immunocytochemistry and autoradiography the presence of estrogen receptors (ER) in rhesus monkey ovary. Intense chromogen staining showed specific binding for ER in nuclei of germinal epithelium and granulosa cells of antral follicles; and radiolabeled ligand bound specifically to functional corpora lutea (CL). Although it is accepted that the germinal epithelium of the primate ovary contains ER, some controversy still persists regarding the intraovarian localization of this molecule. In addition, no data exist that localize the aromatic hydrocarbon (dioxin) receptor (AHR), which is known to modulate ER, to the primate ovary. In the present study, we show the presence of ER using Western blot analysis, and ER capable of binding DNA within intraovarian compartments in two species of the genusMacaca (rhesus macaque,Macaca mulatta and stumptail macaque,Macaca arctoides); extend these findings to human ovarian granulosa cells (GC) using Western blot, reverse transcription-polymerase chain reaction (RT-PCR), and gel mobility-shift analysis; and localize the AHR to intraovarian compartments of the macaque ovary by Western blots and gel-shift assays. These experiments strongly suggest that estrogens can exert effects on follicle development directly at the ovary, and provide the first direct evidence that AHR-mediated toxicity may be manifested at the ovary to induce possible antifertility effects.

Modulation of growth axis gene expression by in utero and lactational exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the weaning Holtzman rat.

While thein utero and lactational effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on both male and female reproductive systems appear to be severe, little is known about its effects on the developing growth axis. The objective of this study was to describe changes in growth axis gene expression that accompany exposure to TCDD duringin utero and lactational development. Pregnant Holtzman rats were administered 1 µg TCDD/kg maternal body weight or vehicle control on gestational day 15 by gavage. Using ribonuclease protection assays, we compared mRNA levels measured in 21-d-old female pups exposed to TCDD with levels measured in control animals for the following genes: somatostatin, growth hormone-releasing hormone (GHRH), hypothalamic and pituitary galanin (GAL), growth hormone (GH), and insulin-like growth factor-I (IGF-I). Serum GH concentrations measured by radio-immunoassay were significantly increased, although GH mRNA levels were unchanged from controls by TCDD exposure. Hypothalamic GAL mRNA was decreased in TCDD-treated animals, whereas pituitary GAL mRNA in TCDD-treated animals was not altered. GHRH mRNA was increased in hypothalami from TCDD-exposed animals. IGF-I mRNA in the liver was decreased to 67% of controls. These data indicate that the growth axis is sensitive to the effects of TCDD delivered during critical periods of development. The alterations observed in growth axis gene expression with exposure to TCDD add to the body of data demonstrating a potent effect of this compound on the fetal and neonatal endocrine system.