Intestinal alterations and mild glucose homeostasis impairments in the offspring born to overweight rats.

The gut plays a crucial role in metabolism by regulating the passage of nutrients, water and microbial-derived substances to the portal circulation. Additionally, it produces incretins, such as glucose-insulinotropic releasing peptide (GIP) and glucagon-like derived peptide 1 (GLP1, encoded by gcg gene) in response to nutrient uptake. We aimed to investigate whether offspring from overweight rats develop anomalies in the barrier function and incretin transcription. We observed pro-inflammatory related changes along with a reduction in Claudin-3 levels resulting in increased gut-permeability in fetuses and offspring from overweight rats. Importantly, we found decreased gip mRNA levels in both fetuses and offspring from overweight rats. Differently, gcg mRNA levels were upregulated in fetuses, downregulated in female offspring and unchanged in male offspring from overweight rats. When cultured with high glucose, intestinal explants showed an increase in gip and gcg mRNA levels in control offspring. In contrast, offspring from overweight rats did not exhibit any response in gip mRNA levels. Additionally, while females showed no response, male offspring from overweight rats did exhibit an upregulation in gcg mRNA levels. Furthermore, female and male offspring from overweight rats showed sex-dependent anomalies when orally challenged with a glucose overload, returning to baseline glucose levels after 120 min. These results open new research questions about the role of the adverse maternal metabolic condition in the programming of impairments in glucose homeostasis, enteroendocrine function and gut barrier function in the offspring from overweight mothers and highlight the importance of a perinatal maternal healthy metabolism.

Maternal Overweight Disrupts the Sexual Maturation of the Offspring.

The aims of the present work were to study the effect of maternal overweight and obesity on the ovarian reserve, follicular development, and ovulation of the offspring and to assess whether this maternal condition alters oocyte integrity. To this end, female offspring from rats fed standard (OSD) or cafeteria (OCD) diet were used. Body weight, vaginal opening, and estrous cycle were recorded and ovaries were obtained on the day of the second estrus. In addition, ovarian weight, ovulation rate (measured by the number of oocytes within oviducts), follicular development (determined by histology), and oocyte integrity were examined. The OCD were divided into 2 groups: offspring from rats with 17% and 28% of overweight (OCD17 and OCD28, respectively). Both OCD groups showed higher body weight, but OCD28 also exhibited early vaginal opening and higher ovarian weight and glycemia at euthanasia compared with OSD. Both OCD17 and OCD28 had lower number of primordial and primary follicles, and only OCD28 exhibited lower number of antral follicles, all compared with OSD rats. In addition, both OCD17 and OCD28 had higher ovulation rate than controls, and OCD28 had lower number of healthy oocytes, which, in turn, exhibited morphological alterations such as larger perivitelline space and zona pellucida than those of control animals. These results suggest that maternal overweight may severely affect the reproductive ability of the offspring, likely as a result of altering the organogenesis.

Seasonal changes in ovarian steroid hormone concentrations in the large hairy armadillo (Chaetophractus villosus) and the crying armadillo (Chaetophractus vellerosus).

Knowledge of armadillo reproductive physiology is essential for developing ex situ and in situ assisted reproductive techniques for propagating and/or controlling populations of these animals. The present study included assessment of fecal sex steroids by radioimmunoassay, determining reproductive status via monitoring ovarian activity (in the wild) and therefore reproductive status, in wild females of the large hairy armadillo (Chaetophractus villosus) and the crying armadillo (Chaetophractus vellerosus) in the southern hemisphere. Plasma and fresh fecal progesterone concentrations were not significantly correlated in either species. However, in both species, there was a significant positive correlation between plasma progesterone and dry fecal progesterone concentrations (r = 0.82, P < 0.05 and r = 0.60, P < 0.05, respectively). Dry fecal progesterone and estradiol concentrations were measured in one captive C. villosus (average baseline progesterone and estradiol concentrations 28.72 ± 11.75 ng/g dry feces and 3.04 ± 0.80 ng/g dry feces, respectively) and one captive C. vellerosus (average baseline progesterone and estradiol concentrations 14.05 ± 3.03 ng/g dry feces and 3.46 ± 1.20 ng/g dry feces, respectively) to detect hormonal peaks over 1 y; these occurred from late fall to early summer. Feces from wild C. villosus and C. vellerosus were also collected over 1 y to determine progesterone peaks, which occurred in winter and spring in both species (with no peaks during the summer or fall). Accordingly, C. villosus and C. vellerosus had a seasonal reproductive pattern. The significant correlations between dry fecal and plasma progesterone concentrations validated this method for monitoring reproductive status in these species.

Daily treatment with α-naphthoflavone enhances follicular growth and ovulation rate in the rat.

The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor and the first protein involved in a variety of physiological and toxicological processes, including those of xenobiotic metabolizing enzymes. AhR has been found in the ovary of many species and seems to mediate the ovarian toxicity of many environmental contaminants, which are AhR ligands. However, the role of AhR in the ovarian function is unknown. Therefore, the aim of this work was to study the action of α-naphthoflavone (αNF), known to be an AhR antagonist, on both follicular growth and ovulation. Immature Sprague-Dawley rats were daily injected intraperitoneally with αNF (0.1-80 mg/kg) or vehicle for 12 days, and primed with gonadotrophins (eCG/hCG) to induce follicular growth and ovulation. Ovaries were obtained 20 h after hCG administration. By means of immunohistochemistry, we found that the numbers of primordial, primary and antral follicles were increased in rats treated with 80 mg/kg αNF and that there were no differences with other doses. Likewise, the ovarian weight and the ovulation rate, measured by both number of oocytes within oviducts and corpora lutea in ovarian sections, were increased when the rats received either 1 or 10mg/kg daily. Although further studies are necessary to know the mechanism of action of αNF, it is possible that the different ovarian processes can be differentially responsive to the presence of different levels of αNF, and that the same or different endogenous AhR ligands can be involved in these ovarian processes in a cell type-dependent manner.

Hyperandrogenism alters intraovarian parameters during early folliculogenesis in mice.

This study aimed to investigate how hyperandrogenism affects early folliculogenesis. Hyperandrogenism was induced in prepuberal female BALB/c mice by daily s.c. injection of dehydroepiandrosterone (60 mg/kg body weight in 0.1 ml sesame oil) for 10 consecutive days. Although hyperandrogenism increased the growth rate of primary follicles, it also increased ovarian oxidative stress (evaluated by the increase in lipid peroxidation, the decrease in superoxide dismutase activity and the fact that glutathione content was not modified). By using the annexin V/cytometry assay it was found that the excess of androgens decreased viable ovarian cells and increased early apoptotic ones. The increased lipid peroxidation induced enhanced ovarian prostaglandin E production. In addition, hyperandrogenism increased the number of T lymphocytes that infiltrate ovarian tissue and modified their phenotype (decreased CD4+ or helper and increased the suppressor/cytotoxic CD8+). The excess of androgens decreased the ovarian expression of the long isoform of leptin receptor (Ob-Rb, the only isoform expressed in the ovarian tissue) when compared with controls. All these alterations increased serum concentrations of oestradiol, a pro-apoptotic agent. It is concluded that the excess of androgens impairs early follicular development by modulating some endocrine and immune parameters that are either directly or indirectly related to follicular atresia.

Prolactin inhibits oocyte release after gonadotropin stimulation in the rat: putative mechanism involving ovarian production of beta-endorphin and prostaglandin.

OBJECTIVE: To evaluate whether prolactin (PRL) is able to inhibit ovulation induced with exogenous gonadotropins in the rat and whether this effect could be mediated by the ovarian production of beta-endorphin, prostaglandin, and nitric oxide (NO). DESIGN: Controlled in vivo and in vitro experiments. SETTING: Academic research laboratories. ANIMAL(S): Immature female rats undergoing ovulation induction with equine gonadotropins and hCG. INTERVENTION(S): Prolactin (100 or 200 microg), PRL + the opioid antagonist naloxone (200 microg each), or placebo were injected SC 4 hours after hCG administration for ovulation induction. In the in vitro experiments, isolated preovulatory ovaries were incubated with or without PRL in a final concentration of 100 or 200 ng/mL. MAIN OUTCOME MEASURES(S): Number of oocytes ovulated in vivo, ovarian beta-endorphin, PGE(2) and NO(2)(-)/NO(3)(-) release, and NO synthase activity in vitro. RESULT(S): Prolactin reduced significantly the number of oocytes ovulated at the doses of 100 and 200 microg, and this effect was partially reversed by naloxone administration together with 200 mug PRL. PRL also induced a twofold increase in the ovarian release of beta-endorphin and a threefold decrease in the ovarian production of PGE(2). Ovarian NO synthase activity and the concentrations of NO(2)(-)/NO(3)(-) in the incubation medium were not modified by PRL. CONCLUSION(S): Prolactin is able to reduce the number of oocytes released and modulate ovarian beta-endorphin and PGE(2) release, which may account for its peripheral anovulatory effects. This local effect of PRL could interfere in the process of ovulation induction by exogenous gonadotropins.

Expression of inducible nitric oxide synthase in tumoral and non-tumoral epithelia from bladder cancer patients.

We have previously demonstrated that nitric oxide (NO) is elevated in the urine from bladder cancer patients. As the inducible nitric oxide synthase (iNOS) produces high NO output, the aim of this study was to examine iNOS expression and activity in tumoral (BT) and non-tumoral bladder tissue (NT). iNOS expression was determined by Western blot in 42 BT, 22 NT, and 4 normal bladders (normal B). iNOS activity was evaluated by conversion of [(14)C]l-arginine to [(14)C]l-citrulline plus NO, in additional 15 BT, 8 NT, and 1 normal B. iNOS tissue localization was studied by immunohistochemistry. iNOS expression and activity were found in almost 50% of bladder cancer patients, in both BT and in NT. A similar positive or negative iNOS expression in each pair of NT and BT tissue compared was observed, suggesting that high urine NO levels could be generated by an active iNOS present not only in the tumor but also in the non-tumoral bladder tissue. By immunohistochemistry, heterogeneous iNOS staining was detected in tumor cells from superficial and invasive tumors, while it was not evident in the normal bladder epithelium. A follow-up of 21 patients during 2 years showed recurrences in 80% with positive iNOS. On the contrary, no recurrences were observed in 73% of iNOS negative patients. Our results suggest that iNOS expression in bladder tissue may predispose to cancer recurrences.

Effect of the pretreatment with prolactin on the distribution of immunoreactive beta-endorphin through different ovarian compartments in immature, superovulated rats.

Beta-endorphin and prolactin (PRL) are natural inhibitors of ovulation via central and peripheral mechanisms, but their possible interactions within the ovary are still unknown. The aims of the present study were to determine the gene expression and the topographic distribution of beta-endorphin, and the possible changes evoked by the pretreatment with PRL on the ovarian beta-endorphin localization in immature, superovulated rats. Prepuberal female Wistar rats weighing 60-70 g were superovulated with 20 IU equine gonadotrophins and, 48 h later, 20 IU human chorionic gonalotropin (hCG). Four hours after the hCG injection, the rats received either 200 microg rat PRL .i.p. (n = 12) or saline vehicle (n = 10). In the following morning the rats were killed and their ovaries were quickly removed. Beta-endorphin localization was assessed by immunohistochemistry and proopiomelanocortin (POMC) mRNA was detected by reverse transcription polymerase chain reaction (RT-PCR). Beta-endorphin was expressed mostly in the corpora lutea and perivascular stroma, but a weak to moderate immunostaining was also present in the theca cells and some granulosa cells of tertiary/antral and preovulatory follicles. The main differences observed in the distribution of ovarian beta-endorphin between the two groups were a more intense immunostaining in the granulosa cells of antral follicles, corpus luteum and stroma of PRL-treated rats. POMC gene transcripts were detected in 2/5 samples from the control group and in 3/7 samples from the PRL-treated group. Thus, the expression of beta-endorphin in tertiary/antral follicles is enhanced by PRL treatment in immature, superovulated rats, providing a putative mechanism by which PRL could inhibit the ovarian response to induced ovulation.