Influence of sexual hormones on Chagas disease.

OBJECTIVE: Analyze sex hormone's influence during Chagas´ Disease. METHODS: Male and female BALB/c mice were divided into six groups, four experimental (sham, orchiectomized, orchiectomized and supplemented with estradiol, orchiectomized supplemented with testosterone, oophorectomized, oophorectomized and supplemented with estradiol, and oophorectomized and supplemented with testosterone), and two control (healthy and intraperitoneally with T. cruzi strain NINOA infected). Clinical data were recorded daily, parasitemia was evaluated using a Neubauer chamber during the infection, and heart histopathological analysis was performed using the paraffin embedding technique. To analyze parasitemia curves and the area under the parametric curves, two-way ANOVA test was performed to correlate groups´ data. P-values <0.05 were considered statistically significant. RESULTS: Higher mortality rates, cardiomegaly, hepatomegaly, ascites, edema, higher parasitemia levels, more amastigote nests, and more severe inflammatory infiltrate were found in higher testosterone concentration mice, whereas in higher estradiol concentration groups, paresia, prostration, edema, and necrosis were found. CONCLUSIONS: Our results showed that testosterone increased infection severity, whereas estradiol had the opposite effect. This research improves the understanding of sex hormones´infuence upon this infection to contribute with the handling of Chagas´disease.

Steroid profile in patients with breast cancer and in mice treated with mifepristone.

Progesterone receptors (PRs) are biomarkers used as prognostic and predictive factors in breast cancer, but they are still not used as therapeutic targets. We have proposed that the ratio between PR isoforms A and B (PRA and PRB) predicts antiprogestin responsiveness. The MIPRA trial confirmed the benefit of 200 mg mifepristone, administered to patients with tumors with a high PRA/PRB ratio, but dose-ranging has not been conducted. The aim of this study was to establish the plasma mifepristone levels of patients from the MIPRA trial, along with the resultant steroid profiles, and compare these with those observed in mifepristone-treated mice using therapeutic schemes able to induce the regression of experimental mammary carcinomas with high PRA/PRB ratios: 6 mg pellets implanted subcutaneously, or daily doses of 12 mg/kg body weight. The plasma levels of mifepristone and other 19 plasma steroids were measured by liquid chromatography-tandem mass spectometry. In mifepristone-treated mice, plasma levels were lower than those registered in mifepristone-treated patients (i.e. day 7 after treatment initiation, pellet-treated mice: 8.4 ± 3.9 ng/mL; mifepristone-treated patients: 300.3 ± 31.7 ng/mL (mean ± s.d.; P < 0.001)). The increase in corticoid related steroids observed in patients was not observed in mifepristone-treated mice. The increase in progesterone levels was the most significant side effect detected in mifepristone-treated mice after 14 or 21 days of treatment, probably due to an ovarian compensatory effect not observed in postmenopausal patients. We conclude that in future clinical trials using mifepristone, the possibility of lowering the standard daily dose of 200 mg should be considered.

Progesterone release profile and follicular development in Nelore cows receiving intravaginal progesterone devices.

This study aimed to evaluate the progesterone (P4) release profile provided by four commercially available intravaginal P4 devices, as well as the effect of circulating P4 concentrations exclusively from these devices on the development of the dominant follicle (DF) in Nelore (Bos indicus) cows. Therefore, non-lactating multiparous Nelore cows were enrolled in an experimental design, over three replicates, starting on Day -9 with the insertion of a reused P4 device (2 g - original P4 load) for 7 d, followed by two treatments of cloprostenol sodium (PGF; 0.482 mg), 24 h apart, on Days -3 and -2. Just before device removal, on Day -2, a norgestomet ear implant was inserted and, 2 d later (Day 0), at the time of norgestomet withdrawal, cows were randomly assigned to receive one of the intravaginal devices: Primer (0.5 g); Prociclar (0.75 g); Sincrogest (1 g); or CIDR (1.9 g), and 2 mg of estradiol benzoate (EB) im. Blood samples were collected immediately before P4 device insertion, 12 h later and daily over 15 d (1 d after P4 device removal). Ultrasound examinations were performed on Days 0, 7, 8, 9, 10, 12, and 14 to evaluate ovarian dynamics. Results are presented as mean ± SEM and differences were considered when P ≤ 0.05. Overall, the devices resulted in distinct circulating P4 concentrations over 10 d, varying according to their initial P4 load and P4 impregnated surface area. Primer provided the lowest circulating P4 concentrations over time, whereas, CIDR had the greatest concentration. Sincrogest and Prociclar were similar, producing intermediary circulating P4. There was no effect of treatment on the DF diameter on any specific day, nor on follicular growth rate from Day 7-10. However, the Primer device resulted in a greater mean DF diameter over time. Additionally, greater circulating P4 concentrations, mainly during the first 3 d of device insertion, were associated with smaller DF diameters regardless of the treatment. In conclusion, results from this study provided a better understanding of the P4 profile of intravaginal P4 devices, as well as, their effect on DF development in Bos indicus cows. These data contribute to optimize the use of P4 devices in the reproductive management of beef cattle.

Progesterone release profile and follicular development in Holstein cows receiving intravaginal progesterone devices.

The aim of this study was to evaluate the progesterone (P4) release profile provided by eight commercial intravaginal P4 devices, as well as the effect of circulating P4 concentrations produced exclusively by these devices on the development of the dominant follicle (DF) in non-lactating multiparous Holstein cows. All cows were submitted to the same experimental design starting with the insertion of a reused P4 device (2 g - original P4 load) for 7 d, followed by two treatments of cloprostenol sodium (PGF; 0.482 mg), 24 h apart, 6 and 7 d after device insertion. Just before device removal, a Norgestomet ear implant was inserted and, 2 d later (Day 0), simultaneously to Norgestomet withdrawal, cows received one of the tested intravaginal devices and 2 mg of estradiol benzoate (EB) im. In Exp.1 (n = 22; three replicates), cows were randomized to receive: CIDR (1.38 g); PRID-Delta (1.55 g); Prociclar (0.75 g); or Repro sync (2 g). In Exp. 2 (n = 29; four replicates), cows were randomized to receive: Cue-Mate (1.56 g); DIB 0.5 (0.5 g); DIB (1 g); PRID-Delta (1.55 g); or Sincrogest (1 g). Blood samples were collected before P4 device insertion (Day 0), 12 h later and daily over 15 d (1 d after P4 device removal). Ultrasound examinations were performed to evaluate growth of the DF on Days 0, 7, 8, 9, and 10. Results are presented as mean ± SEM and differences were considered when P ≤ 0.05. Overall, the circulating P4 profile and mean circulating P4 over 10 d differed among treatments. However, no effects were observed on the DF diameter and follicular growth rate from Day 7-10 after P4 device insertion. In Exp. 2, devices that provided higher circulating P4 concentrations were associated to a slower DF growth during the treatment period. Finally, this study provided a better understanding of the P4 release profile produced by intravaginal P4 devices as well as their effect on circulating P4 concentrations and DF development in non-lactating Holstein cows.

Training counteracts DEX-induced microvascular rarefaction by improving the balance between apoptotic and angiogenic proteins.

This work investigated the mechanisms induced by exercise training that may contribute to attenuate dexamethasone (DEX)-induced microvascular rarefaction and hypertension. Wistar rats underwent training protocol or were kept sedentary for 8 weeks. Dexamethasone was administered during the following 14-days and hemodynamic parameters were recorded at the end. Capillary density (CD) and capillary-to-fiber ratio (C:F ratio) were obtained in soleus muscle (SOL). Also, vascular endothelial growth factor (VEGF), vascular endothelial growth factor receptor-2 (VEGFR-2), endothelial nitric oxide synthase (eNOS), B-cell lymphoma 2 (Bcl-2), Bcl-2-like protein 4 (Bax), p-BAX and caspase-3 cleaved protein levels were analyzed. DEX treatment significantly increased blood pressure (+14%), which was associated with reduced C:F ratio (-41.0%) and CD (-43.1%). Reduction of vessel density was associated with decreased VEGF (-15.6%), VEGFR-2 (-14.6%), Bcl-2 (-18.4%), Bcl-2/Bax ratio (-29.0%) and p-Bax/Bax (-25.4%), and also with increased caspase-3 cleaved protein level (25%). Training, on the other hand, prevented microvessels loss by mitigating all proteins changes induced by DEX. In addition, angiogenic and apoptotic proteins were significantly correlated with CD, which, in turn, was associated with blood pressure. Therefore, we may point out that exercise training is a good strategy to attenuate DEX-induced microvascular rarefaction in soleus muscle and this response involves a better balance between apoptotic and angiogenic proteins, which may contribute for the attenuation of hypertension.

Eliciting Negative Affects Using Film Clips and Real-Life Methods.

Although audio-visual stimuli are among the most frequently used methods to elicit emotional reactions in experimental conditions, real-life manipulations have increasingly been used in different countries. However, the applicability of such protocols has not yet been tested in Brazilian Portuguese speakers. Thus, we conducted two experiments to investigate the effectiveness of both methods. In the first experiment, we used film clips to induce negative emotions (i.e., anger, fear, or sadness) or an emotionally neutral condition in 321 undergraduate students. After watching one of the online videos, volunteers completed an emotional assessment. As expected, there were significant differences in all groups. Our results corroborate the relatively discrete patterns in emotion elicitation using films. In the second experiment, anger was elicited in 18 male undergraduates through a hostile social interaction with a confederate and measured by the corrugator muscle activity and cortisol responses. Indeed, there was an increase in corrugator activity in the group exposed to anger induction, even after a few minutes from the end of the experimental manipulation. Implications for experiments on the negative emotions are discussed.

Quantitative proteomic profiling of bovine follicular fluid during follicle development.

Bovine follicular fluid (FF) constitutes the microenvironment of follicles and includes various biologically active proteins. We performed a study involving 18 healthy nonlactating Holstein cows to determine the protein expression profile of FF at key stages of follicular development. Follicles were individually aspirated in vivo at predeviation (F1 ∼ 7.0 mm), deviation (F1 ∼ 8.5 mm), postdeviation (F1 ∼ 12.0 mm), and preovulatory stages of follicle development, which were confirmed by measurement of follicular estradiol and progesterone concentrations. The FFs from nine cows were selected for proteomic analysis. After albumin depletion, triplicates of pooled FF were reduced, alkylated, and digested with trypsin. The resulting peptides were labeled with TMTsixplex and quantified using liquid chromatography-mass spectrometry/mass spectrometry. A total of 143 proteins were identified and assigned to a variety of biological processes, including response to stimulus and metabolic processes. Twenty-two differentially (P < 0.05) expressed proteins were found between stages indicating intrafollicular changes over development, with expected deviation time critical to modulate the protein expression. For instance, high concentrations of follistatin, inhibin, serglycin, spondin-1, fibrinogen, and anti-testosterone antibody were found during early stages of follicular development. In contrast, apolipoprotein H, alpha-2-macroglobulin, plasminogen, antithrombin-III, and immunoglobulins were increased after deviation. Among the differentially abundant proteins, 19 were found to be associated with steroidogenesis. Pathway analysis identified proteins that were mainly associated with the acute phase response signaling, coagulation system, complement system, liver/retinoid X receptor activation, and biosynthesis of nitric oxide and reactive oxygen. The differentially expressed proteins provide insights into the size-dependent protein changes in the ovarian follicle microenvironment that could influence follicular function.

Distribution and concentration of maternal progesterone in the yolk of Greater Rhea eggs (Rhea americana).

Progesterone is the most concentrated maternal yolk steroid characterized to date in birds; however, no information about it is available in ratite eggs. We collected freshly laid eggs from zoo-housed Greater Rhea females (Rhea americana) bred under similar rearing conditions during two breeding seasons to characterize concentration and distribution of maternal yolk progesterone. After high-performance liquid chromatography analysis, yolk hormone was measured using a commercial electrochemiluminescence immunoassay. Progesterone concentrations were found to vary significantly among the yolk layers, supporting a follicular origin for this steroid in Greater Rhea eggs. Additionally, highly similar mean absolute yolk progesterone concentrations were detected between 2013 and 2015 breeding seasons (1,332.98 ± 82.59 and 1,313.59 ± 85.19 ng/g, respectively). These values are also comparable to those found in some domestic carinate species. Findings suggest that at population level, when rearing conditions are similar, mean absolute yolk maternal progesterone concentrations also appear bounded. Future research on the factors and mechanisms that regulate progesterone deposition in Greater Rhea eggs is needed to better understand whether its levels depend on different rearing conditions.

The trophic effect of ouabain on retinal ganglion cells is mediated by IL-1ß and TNF-α.

Ouabain is a steroid hormone that binds to the enzyme Na(+), K(+) - ATPase and stimulates different intracellular pathways controlling growth, proliferation and cell survival. IL-1ß and TNF-α are pleiotropic molecules, conventionally regarded as pro-inflammatory cytokines with well-known effects in the immune system. In addition, IL-1ß and TNF-α also play important roles in the nervous system including neuroprotective effects. Previous data from our group showed that ouabain treatment is able to induce an increase in retinal ganglion cell survival kept in mixed retinal cell cultures. The aim of this work was to investigate if IL-1ß and TNF-α could be mediating the trophic effect of ouabain on retinal ganglion cells. Our results show that the trophic effect of ouabain on retinal ganglion cell was inhibited by either anti-IL-1ß or anti-TNF-α antibodies. In agreement, IL-1ß or TNF-α increased the retinal ganglion cells survival in a dose-dependent manner. Accordingly, ouabain treatment induces a temporal release of TNF-α and IL-1ß from retinal cell cultures. Interestingly, TNF-α and IL-1ß regulate each other intracellular levels. Our results suggest that ouabain treatment triggers the activation of TNF-α and IL-1ß signaling pathways leading to an increase in retinal ganglion cell survival.

Postnatal development and histofunctional differentiation of the oviduct in the broad-snouted caiman (Caiman latirostris).

Caiman latirostris is a South American crocodilian species characterized as a sentinel of the presence of endocrine-disrupting compounds (EDCs). Evaluating developmental events in hormone-dependent organs, such as the oviduct, is crucial to understand physiological postnatal development, to identify putative periods of exposure sensitive to EDCs, and/or to identify biomarkers useful to evaluate the effects of EDC exposure. In this study, we describe the histomorphological features of C. latirostris oviducts by establishing the ontogeny of changes at cellular, tissue and molecular levels from the neonatal to the pre-pubertal juvenile stages. Since the histological diagnosis of the adenogenic oviduct lies on a group of features, here we defined a histofunctional score system and a cut-off value to distinguish between preadenogenic and adenogenic oviducts. Our results showed that the maturation of the C. latirostris oviduct is completed postnatally and characterized by changes that mimic the pattern of histological modifications described for the mammalian uterus. Ontogenic changes in the oviductal epithelium parallel changes at subepithelial level, and include collagen remodeling and characteristic spatial-temporal patterns of α-actin and desmin. The expression pattern of estrogen receptor alpha and progesterone receptor evidenced that, even at early postnatal developmental stages, the oviduct of C. latirostris is a target organ of endogenous and environmental hormones. Besides, oviductal adenogenesis seems to be an estrogen-dependent process. Results presented here provide not only insights into the histophysiological aspect of caiman female reproductive ducts but also new tools to better characterize caimans as sentinels of endocrine disruption.