Novel Roles of HCG/LH Signalling in the Mouse Mammary Gland and its Tumorigenesis.

Breast cancer is the most common cancer in women, and its development is intimately related to hormonal factors, but how hormones affect breast physiology and tumorigenesis is not sufficiently known. Pregnancy elicits long-term protection from breast cancer, but during the first ten years after pregnancy, breast cancer risk is increased. In previous studies, there has been conflicting data on the role of human chorionic gonadotropin (HCG) and the functionality of its receptor in extragonadal tissues. The aim of this study was to elucidate the role of chronically elevated HCG in mouse physiology. We have created a transgenic (TG) mouse model that overexpresses HCG. HCG is similar to lutenizing hormone (LH), but is secreted almost solely by the placenta during pregnancy. HCG and LH both bind to the LH receptor (LHR). In the current study, mammary gland tumors were observed in HCG TG mice. We elucidated the role of HCG in mammary gland signalling and the effects of LHR mediated signalling in mouse mammary gland gene expression. We also studied the effects of HCG in human breast epithelial cell cultures. Several endocrine disturbances were observed in HCGβ TG female mice, resulting in precocious puberty, infertility, obesity and pituitary and mammary gland tumors. The histology of the mammary gland tumors of HCGβ TG females resembled those observed in mouse models with activated Wnt/β-catenin signalling pathway. Wnts are involved in stem cell regulation and tumorigenesis, and are hormonally regulated in the mammary gland. We observed activated β-catenin signalling and elevated expression of Wnt5b and Wnt7b in TG tumors and mammary glands. Furthermore, we discovered that HCG directly regulates the expression of Wnt5b and Wnt7b in the mouse mammary gland. Pharmacological treatment with HCG also caused upregulation of several Wnt-pathway target genes in ovariectomized wild type (WT) mice in the presence of physiological concentrations of estradiol and progesterone. In addition, differential expression of several metabolic genes was observed, suggesting that HCG affects adipocyte function or glucose metabolism. When WT mice were transplanted with LHR deficient or wild type WT mammary epithelium, differential expression of several genes affecting the Wnt-signalling pathway was observed in microarray analysis. Diminished expression of several genes associated with LHR function in other tissues, such as the ovary, was observed in mammary glands deficient of epithelial LHR. In cultured human mammary epithelial cells HCG upregulated the expression of WNT5B, WNT7B similar to mouse, suggesting that the observations found are relevant in human physiology. These studies suggest that HCG/LHR signalling affects gene expression in non-gonadal tissues, and that Wnt-signalling is regulated by HCG/LH in human and mouse mammary glands.

Epidermal growth factor secreted from submandibular salivary glands interferes with the lipolytic effect of adrenaline in mice

We had described that epidermal growth factor (EGF) interfered with the lipolytic effect of catecholamines in isolated adipocytes. Since catecholamines stimulate the release of EGF from submandibular salivary glands to blood plasma in male mice, we studied whether EGF affected also the lipolytic response to adrenaline in whole animals. We studied the effect of adrenaline in sialoadenectomized and sham-operated mice receiving or not a high dose of EGF following adrenaline injection. There was no difference in plasma EGF concentration between sham-operated and sialoadenectomized animals receiving saline. After adrenaline administration plasma EGF increased by 20-fold in sham-operated but did not increase in sialoadenectomized mice. Indeed, the increase was much higher (more than 100-fold) in mice receiving exogenous EGF. The effect of adrenaline on plasma concentration of both glycerol and nonesterified fatty acids was higher as lower was plasma EGF concentration. Isolated adipocytes obtained from sham-operated or sialoadenectomized mice had identical lipolytic response to adrenaline. The lipolytic response of adipocytes to isoproterenol was decreased by addition of EGF. To study whether the interference with the in vivo lipolytic effect of adrenaline had further metabolic consequences, we measured plasma b-hydroxybutyrate concentration in plasma. There was no difference in the response to adrenaline between sham-operated and sialoadenectomized mice in spite of the difference in plasma nonsterified fatty acid concentration. Studies in isolated hepatocytes indicated that ketogenesis run at near maximal rate in this range of substrate concentration. These results suggest that EGF in the physiological range decreases the lipolytic effect of adrenaline but does not compromise further metabolic events like the enhancement of ketogenesis.

Circadian variation of salivary immunoglobin A, alpha-amylase activity and mood in response to repeated double-poling sprints in hypoxia.

PURPOSE: To assess the circadian variations in salivary immunoglobin A (sIgA) and alpha-amylase activity (sAA), biomarkers of mucosal immune function, together with mood during 2 weeks of repeated sprint training in hypoxia (RSH) and normoxia (RSN). METHODS: Over a 2-week period, 17 competitive cross-country skiers performed six training sessions, each consisting of four sets of five 10-s bouts of all-out double-poling under either normobaric hypoxia (FiO2: 13.8 %, 3000 m) or normoxia. The levels of sIgA and sAA activity and mood were determined five times during each of the first (T1) and sixth (T6) days of training, as well as during days preceding (baseline) and after the training intervention (follow-up). RESULTS: With RSH, sIgA was higher on T6 than T1 (P = 0.049), and sAA was increased on days T1, T6, and during the follow-up (P < 0.01). With RSN, sIgA remained unchanged and sAA was elevated on day T1 only (P = 0.04). Similarly, the RSH group demonstrated reduced mood on days T1, T6, and during the follow-up, while mood was lowered only on T1 with RSN (P < 0.01). CONCLUSIONS: The circadian variation of sIgA and sAA activity, biomarkers of mucosal immune function, as well as mood were similar on the first day of training when repeated double-poling sprints were performed with or without hypoxia. Only with RSH did the levels of sIgA and sAA activity rise with time, becoming maximal after six training sessions, when mood was still lowered. Therefore, six sessions of RSH reduced mood, but did not impair mucosal immune function.

Imaging of the adrenal gland lesions

With the steep increase in the use of cross-sectional imaging in recent years, the incidentally detected adrenal lesion, or "incidentaloma", has become an increasingly common diagnostic problem for the radiologist, and a need for an approach to classifying these lesions as benign, malignant or indeterminate with imaging has spurred an explosion of research. While most incidentalomas represent benign disease, typically an adenoma, the possibility of malignant involvement of the adrenal gland necessitates a reliance on imaging to inform management decisions. In this article, we review the literature on adrenal gland imaging, with particular emphasis on computed tomography, magnetic resonance imaging, and photon-emission tomography, and discuss how these findings relate to clinical practice. Emerging technologies, such as contrast-enhanced ultrasonography, dual-energy computed tomography, and magnetic resonance spectroscopic imaging will also be briefly addressed.

Low-level laser therapy for the prevention of low salivary flow rate after radiotherapy and chemotherapy in patients with head and neck cancer

Abstract Objective: To determine whether low-level laser therapy can prevent salivary hypofunction after radiotherapy and chemotherapy in head and neck cancer patients. Materials and Methods: We evaluated 23 head and neck cancer patients, of whom 13 received laser therapy and 10 received clinical care only. An InGaAlP laser was used intra-orally (at 660 nm and 40 mW) at a mean dose of 10.0 J/cm2 and extra-orally (at 780 nm and 15 mW) at a mean dose of 3.7 J/cm2, three times per week, on alternate days. Stimulated and unstimulated sialometry tests were performed before the first radiotherapy and chemotherapy sessions (N0) and at 30 days after the end of treatment (N30). Results: At N30, the mean salivary flow rates were significantly higher among the laser therapy patients than among the patients who received clinical care only, in the stimulated and unstimulated sialometry tests (p = 0.0131 and p = 0.0143, respectively). Conclusion: Low-level laser therapy, administered concomitantly with radiotherapy and chemotherapy, appears to mitigate treatment-induced salivary hypofunction in patients with head and neck cancer.

In vitro antimicrobial and antioxidant activities of a crude extract and fractions from Buddleja thyrsoides Lam. Leaves

Crude extract and fractions of Buddleja thyrsoides were investigated regarding antioxidant activities by DPPH, total phenolic contents by Folin-Ciocalteau and antimicrobial activity by the broth microdilution method. Total phenolics varied from 214.07 ± 3.6 to 438.4 ± 0.3 mg g-1. Crude extract, ethyl acetate, dichloromethane and butanolic fractions exhibited a weak scavenging activity (SC50=186.04 ± 10.8, 137.70 ± 8.5, 146.89 ± 9.0 and 165.71 ± 3.2 µg mL-1, respectively). A correlation between the antioxidant activities and total phenolic contents could be shown (r=0.857, p<0.01). The lowest value of MIC was observed with butanolic fraction against Saccharomyces cerevisiae (MIC and MFC at 62.5 µg mL-1). Dichloromethane and ethyl acetate fractions were effective against Staphylococcus aureus with MIC value at 250 and 500 µg mL-1 respectively.

Determination of rutin and narcissin in marigold extract and topical formulations by liquid chromatography: applicability in skin penetration studies

A chromatographic technique for determination of rutin and narcissin in marigold extract and topical formulations was developed and validated. The method shows linearity over the concentration range of 0.2 - 6.0 μg/mL of rutin (r = 0.9986) and 0.8 - 12.0 μg/mL of narcissin (r = 0.9951). The values obtained for precision and accuracy are in agreement with ICH guidelines. Both the formulation excipients and the porcine ear skin samples did not interfere with the flavonoids determination. The recovery of rutin and narcissin in skin samples added with marigold extract was 81.41% and 83.35%, respectively, which demonstrate the applicability of this method to perform skin penetration studies.

Seasonal variation of the major secondary metabolites present in the extract of Eremanthus mattogrossensis Less (Asteraceae: Vernonieae) leaves

The species Eremanthus mattogrossensis, known as "veludo do cerrado" (cerrado velvet), is native to the Brazilian Cerrado. Because the amount of metabolites present in plants may be influenced by biological and environmental factors, here we conducted an HPLC-DAD-MS/MS investigation of the metabolite concentrations found in the MeOH/H2O extract of the leaves of this species. The main compounds were identified and quantified, and the metabolites were grouped by chemical class (caffeoylquinic acids, flavonoids, and sesquiterpene lactone). Statistical analysis indicated a straight correlation between the quantity of metabolites and seasonality, suggesting that environmental properties elicit important metabolic responses.

Optimization of solvent mixtures for extraction from bark of Schinus terebinthifolius by a statistical mixture-design technique and development of a UV-Vis spectrophotometric method for analysis of total polyphenols in the extract

A statistical mixture-design technique was used to study the effects of different solvents and their mixtures on the yield, total polyphenol content, and antioxidant capacity of the crude extracts from the bark of Schinus terebinthifolius Raddi (Anacardiaceae). The experimental results and their response-surface models showed that ternary mixtures with equal portions of all the three solvents (water, ethanol and acetone) were better than the binary mixtures in generating crude extracts with the highest yield (22.04 ± 0.48%), total polyphenol content (29.39 ± 0.39%), and antioxidant capacity (6.38 ± 0.21). An analytical method was developed and validated for the determination of total polyphenols in the extracts. Optimal conditions for the various parameters in this analytical method, namely, the time for the chromophoric reaction to stabilize, wavelength of the absorption maxima to be monitored, the reference standard and the concentration of sodium carbonate were determined to be 5 min, 780 nm, pyrogallol, and 14.06% w v-1, respectively. UV-Vis spectrophotometric monitoring of the reaction under these conditions proved the method to be linear, specific, precise, accurate, reproducible, robust, and easy to perform.

HPLC-DAD based method for the quantification of flavonoids in the hydroethanolic extract of Tonina fluviatilis Aubl. (Eriocaulaceae) and their radical scavenging activity

This article describes the isolation and identification of flavonoids in the hydroethanolic extract of the aerial parts from Tonina fluviatilis and evaluation of their antiradical activity. A method based on HPLC-DAD was developed and validated for detecting and quantifying flavonoids in hydroethanolic extracts. The flavonoids identified and quantified in the extract were 6,7-dimethoxyquercetin-3-O-β-D-glucopyranoside (1), 6-hydroxy-7-methoxyquercetin-3-O-β-D-glucopyranoside (2), and 6-methoxyquercetin-3-O-β-D-glucopyranoside (3). The developed method presented good validation parameters, showing that the results obtained are consistent and can be used in ensuring the quantification of these constituents in the extracts. Compounds 2 and 3 showed strong antiradical activity when compared with the positive controls (quercetin and gallic acid).

Evaluation of the effect of the seed extract of Moringa oleifera Lam over the efficiency of organic filters in wastewater treatment of dairy cattle breeding

This study aimed to evaluate the effect of Moringa oleifera Lam extract on the removal of total solids (TS), total suspended solids (TSS) and chemical oxygen demand (COD), in different filter media for treating wastewater of dairy cattle breeding (DCW). The moringa seed extract was obtained by grinding 50 g of seeds in one liter of distilled water and, after passing the solution through a quantitative paper filter of 25 microns, 60 mL of the extract were added to wastewater from cattle breeding before the filtration process in organic filters made of thin coal, bamboo leaves, eucalyptus leaves, gliricidia branches and sawdust. This was followed by the completely randomized experimental design, adopting a factorial of 5 x 2. Aliquots of filtered effluent were collected and the total solids (TS) concentrations, total suspended solids (TSS) and chemical oxygen demand (COD) were determined. It was found that the increase in the efficiency of removal of COD and total solids can be attributed to the coagulating power of the moringa seed extract, wherein the filter medium with bamboo leaves presented the best performance, showing potential for use as alternative filter material in the primary treatment of DCW.

Use of raw Euphorbia tirucalli extract for inhibition of ascitic Ehrlich tumor

Objective: to evaluate the effect of the Euphorbia tirucalli hydroalcoholic extract (ETHE) on the development of Ehrlich Tumor, in its ascitic form. Methods: we intraperitoneally inoculated 15 Swiss mice with 10.44 x 107 cells of Ehrlich Tumor and divided them in two groups one day after: ETHE Group (eight mice), treated with a dosage of 125 mg/kg/day of EHTE for five days; and Control Group (seven mice), treated only with 0.9% isotonic saline solution over the same period. The treatment was done by gavage. Ten days after inoculation, four mice from each group were sacrificed for quantification of tumor cell number, ascitic fluid volume and bone marrow cell number. The remaining animals were maintained to evaluate survival. Results: The ascitic fluid volume and the tumor cell number were decreased in the ETHE group when compared with the control group, but with no statistical significance. On the other hand, survival was higher in the ETHE group, as well as the number of bone marrow cells. Conclusion: Treatment with ETHE after inoculation of Ehrlich Tumor decreases its development and increases survival and the bone marrow cellularity, thus reducing the myelosuppression present in the Ehrlich Tumor bearing mice.

Bartholin's gland cysts: management with carbon-dioxide laser vaporization

PURPOSE: To evaluate the effectiveness, recurrence rate, and complications of carbon-dioxide laser vaporization in the treatment of Bartholin's gland cysts. METHODS: A retrospective study including 127 patients with symptomatic Bartholin' gland cysts submitted to carbon-dioxide laser vaporization at our institution from January 2005 to June 2011. Patients with Bartholin's gland abscesses and those suspected of having neoplasia were excluded. All procedures were performed in an outpatient setting under local anaesthesia. Clinical records were reviewed for demographic characteristics, anatomic parameters, intraoperative and postoperative complications, and follow-up data. Data were stored and analyzed in Microsoft Excel® 2007 software. A descriptive statistical analysis was performed, and its results were expressed as frequency (percentage) or mean±standard deviation. Complication, recurrence, and cure rates were calculated. RESULTS: The mean age of the patients was 37.3±9.5 years-old (range from 18 to 61 years-old). Seventy percent (n=85) of them were multiparous. The most common symptom was pain and 47.2% (n=60) of patients had a history of previous medical and/or surgical treatment for Bartholin's gland abscesses. Mean cyst size was 2.7±0.9 cm. There were three (2.4%) cases of minor intraoperative bleeding. Overall, there were 17 (13.4%) recurrences within a mean of 14.6 months (range from 1 to 56 months): ten Bartholin's gland abscesses and seven recurrent cysts requiring reintervention. The cure rate after single laser treatment was 86.6%. Among the five patients with recurrent disease that had a second laser procedure, the cure rate was 100%. CONCLUSIONS: At this institution, carbon-dioxide laser vaporization seems to be a safe and effective procedure for the treatment of Bartholin's gland cysts.

Salivary and serum cortisol levels, salivary alpha-amylase and unstimulated whole saliva flow rate in pregnant and non-pregnant

PURPOSE: To compare salivary and serum cortisol levels, salivary alpha-amylase (sAA), and unstimulated whole saliva (UWS) flow rate in pregnant and non-pregnant women. METHOD: A longitudinal study was conducted at a health promotion center of a university hospital. Nine pregnant and 12 non-pregnant women participated in the study. Serum and UWS were collected and analyzed every trimester and twice a month during the menstrual cycle. The salivary and serum cortisol levels were determined by chemiluminescence assay and the sAA was processed in an automated biochemistry analyzer. RESULTS: Significant differences between the pregnant and non-pregnant groups were found in median [interquartile range] levels of serum cortisol (23.8 µL/dL [19.4-29.4] versus 12.3 [9.6-16.8], p<0.001) and sAA (56.7 U/L [30.9-82.2] versus 31.8 [18.1-53.2], p<0.001). Differences in salivary and serum cortisol (µL/dL) and sAA levels in the follicular versus luteal phase were observed (p<0.001). Median UWS flow rates were similar in pregnant (0.26 [0.15-0.30] mL/min) and non-pregnant subjects (0.23 [0.20-0.32] mL/min). Significant correlations were found between salivary and serum cortisol (p=0.02) and between salivary cortisol and sAA (p=0.01). CONCLUSIONS: Serum cortisol and sAA levels are increased during pregnancy. During the luteal phase of the ovarian cycle, salivary cortisol levels increase, whereas serum cortisol and sAA levels decline.