Leonurine hydrochloride-a new drug for the treatment of menopausal syndrome: Synthesis, estrogen-like effects and pharmacokinetics.

This research aimed to investigate the estrogen-like effects of Leonurine hydrochloride (Leo). First, we developed a total synthesis of Leo from 3,4,5-trimethoxy-benzoic acid and the structure was confirmed through 1H NMR and mass spectrometry (MS). Then the estrogenic activity of Leo in vitro and in vivo was studied. The proliferation and proliferation inhibitory effects of Leo on MCF-7 cells and MDA-MB-231 cells indicate that Leo exerts estrogen-like effects through estrogen receptor α (ERα) and estrogen receptor ß((ERß) in vitro. Uterotrophic assay in juvenile mice showed that Leo has an estrogen-like effect in vivo, as it can promote the development of the uterus of juvenile mice, increase its uterine coefficient and the size of the uterine cavity, as well as the increased number of uterine glands and the thickened uterine wall. For further research, cyclophosphamide (CTX) was used to establish a mouse model of ovarian function decline. Through this model, we found that Leo can restore the estrous cycle of mice, increase the number of primordial and primary follicles in the ovaries of mice, and regulate the disordered hypothalamic-pituitary-ovarian (HPOA) axis of mice. Finally, the pharmacokinetics of Leo was studied and oral bioavailability of Leo was calculated to be 2.21%. Leo was synthesized and the estrogen-like effect in vitro and in vivo was confirmed as well as its pharmacokinetics.

Insight on the Intracrinology of Menopause: Androgen Production within the Human Vagina.

In this study, we investigated steroidogenic gene mRNA expression in human vaginas and verified the ability of human vagina smooth muscle cells (hvSMCs) to synthesize androgens from upstream precursor dehydroepiandrosterone (DHEA). As a readout for androgen receptor (AR) activation, we evaluated the mRNA expression of various androgen-dependent markers. hvSMCs were isolated from vagina tissues of women undergoing surgery for benign gynecological diseases. In these cells, we evaluated mRNA expression of several steroidogenic enzymes and sex steroid receptors using real time reverse transcription-polymerase chain reaction. Androgen production was quantified with liquid chromatography tandem-mass spectrometry (LC-MS/MS). In vaginal tissues, AR mRNA was significantly less expressed than estrogen receptor α, whereas in hvSMCs, its mRNA expression was higher than progestin and both estrogen receptors. In hvSMCs and in vaginal tissue, when compared to ovaries, the mRNA expression of proandrogenic steroidogenic enzymes (HSD3ß1/ß2, HSD17ß3/ß5), along with 5α-reductase isoforms and sulfotransferase, resulted as being more abundant. In addition, enzymes involved in androgen inactivation were less expressed than in the ovaries. The LC-MS/MS analysis revealed that, in hvSMCs, short-term DHEA supplementation increased Δ4-androstenedione levels in spent medium, while increasing testosterone and DHT secretion after longer incubation. Finally, androgenic signaling activation was evaluated through AR-dependent marker mRNA expression, after DHEA and T stimulation. This study confirmed that the human vagina is an androgen-target organ with the ability to synthesize androgens, thus providing support for the use of androgens for local symptoms of genitourinary syndrome in menopause.

Protection and immune modulation of activated human vaginal epithelial cells by Aurea helianthus extract.

Aurea helianthus extract is associated with various properties including anti-melanogenesis, anti-oxidation, tumorigenic suppression, and immunoregulation; however, the mechanism by which it executes the immunomodulation of human vaginal epithelial cells (HVECs) remains elusive. We established three immunological functions of the extract. First, it mediated tumorigenic suppression in HVECs. Expression of cytokeratin 8, cancer antigen-125, and vimentin was dramatically downregulated in HVECs exposed to the extract under oxidative and fungal stresses. Second, the extract activated dendritic cells and macrophages. On exposing progenitor dendritic cells to the extract, the number of CD304+ cells increased by 40%; further, under oxidative and fungal stresses, this number was approximately 1.8 and 1.3 times lower, respectively, compared to that in the stressed cells. In monocytic differentiation, the number of dendritic cells and macrophages increased 9 and 6 times, respectively, compared to that in the control. Additionally, the extract enhanced and recovered polarisation by approximately 1.5 and 2 times, respectively, than that under stressed conditions. Third, the phagocytic activity of macrophages, against HPV16, 18, and 33 peptides, was enhanced by 12-35 times compared with that under stressed conditions. Thus, A. helianthus extract is a strong stimulator of the immune system and tumorigenic suppression under stress conditions.

[Effect of butyl alcohol extract of Baitouweng Decoction on vaginal mucosal neutrophil chemotaxis in vulvovaginal candidiasis mice].

To investigate the effects of butyl alcohol extract of Baitouweng Decoction(BAEB) on neutrophil chemotaxis in vaginal mucosa of mice with vulvovaginal candidiasis(VVC). Seventy-two SPF female Kunming mice were randomly divided into normal control group, model group, fluconazole group, BAEB low-dose group, middle-dose group and high-dose group. Subcutaneous injection of estradiol benzoate was conducted to induce pseudo-estrus, and then 2×10~6 CFU·mL~(-1)of Candida albicans was inoculated into vaginal lumen, followed by drug treatment for 7 days. Gram staining was used to observe the morphological changes of C. albicans in vagina; vaginal fungal load was detected on agar plate. Histological changes of vaginal tissues in mice were observed by HE staining. Lactate dehydrogenase(LDH), interleukin-6(IL-6) and tumor necrosis factor(TNF-α) levels in mouse lavage fluid were detected by enzyme-linked immunosorbent assay(ELISA). Neutrophils in vaginal lavage fluid was observed and counted by using Pap smear. The levels of IL-8 and MIP-2 in vaginal mucosa were detected by ELISA. IL-8 and MIP-2 mRNA levels in vaginal mucosa of mice were detected by qRT-PCR. The results showed that as compared with the normal group, VVC model group had a large number of hyphae and a high level of fungal loadinvagina. The vaginal mucosa was completely destroyed, the number of neutrophils increased, and the protein and mRNA levels of IL-8 and MIP-2 were up-regulated. After BAEB treatment, the hyphae of the treatment group was decreased, the fungal load was decreased, the impaired mucosa showed different degrees of improvement, the inflammatory factors were decreased to varying degrees, and the protein and mRNA levels of chemokine IL-8 and MIP-2 were down-regulated. In conclusion, BAEB may be used to treat VVC by inhibiting vulvovaginal candidiasis via blocking neutrophils recruitment into vagina.

Sex-Dependent Mechanisms of Glucocorticoid Regulation of the Mouse Hypothalamic Corticotropin-Releasing Hormone Gene.

To limit excessive glucocorticoid secretion following hypothalamic-pituitary-adrenal (HPA) axis stimulation, circulating glucocorticoids inhibit corticotropin-releasing hormone (CRH) expression in paraventricular nucleus (PVN) neurons. As HPA function differs between sexes and depends on circulating estradiol (E2) levels in females, we investigated sex/estrous stage-dependent glucocorticoid regulation of PVN Crh. Using NanoString nCounter technology, we first demonstrated that adrenalectomized (ADX'd) diestrous female (low E2), but not male or proestrous female (high E2), mice exhibited a robust decrease in PVN CRH mRNA following 2-day treatment with the glucocorticoid receptor (GR) agonist RU28362. Immunohistochemical analysis of PVN CRH neurons in Crh-IRES-Cre;Ai14 mice, where TdTomato fluorescence permanently tags CRH-expressing neurons, showed similarly abundant co-expression of GR-immunoreactivity in males, diestrous females, and proestrous females. However, we identified sex/estrous stage-related glucocorticoid regulation or expression of GR transcriptional coregulators. Out of 17 coregulator genes examined using nCounter multiplex analysis, mRNAs that were decreased by RU28362 in ADX'd mice in a sex/estrous stage-dependent fashion included: GR (males = diestrous females > proestrous females), signal transducer and activator of transcription 3 (STAT3) (males < diestrous = proestrous), and HDAC1 (males < diestrous > proestrous). Steroid receptor coactivator 3 (SRC-3), nuclear corepressor 1 (NCoR1), heterogeneous nuclear ribonucleoprotein U (hnrnpu), CREB binding protein (CBP) and CREB-regulated transcription coactivator 2 (CRTC2) mRNAs were lower in ADX'd diestrous and proestrous females versus males. Additionally, most PVN CRH neurons co-expressed methylated CpG binding protein 2 (MeCP2)-immunoreactivity in diestrous female and male Crh-IRES-Cre;Ai14 mice. Our findings collectively suggest that GR's sex-dependent regulation of PVN Crh may depend upon differences in the GR transcriptional machinery and an underlying influence of E2 levels in females.

Antiviral effect of Chinese herbal prescription JieZe-1 on adhesion and penetration of VK2/E6E7 with herpes simplex viruses type 2.

ETHNOPHARMACOLOGICAL RELEVANCE: The Chinese Herbal Prescription JieZe-1(JZ-1), added and subtracted from Yihuang Decoction, a famous formula in the 12th year of Kangxi in Qing Dynasty, has a clear effect on Genital Herpes (GH) and no obvious adverse reactions occur clinically. JZ-1 also has preventive and therapeutic effects on Trichomonas vaginitis, Candida albicans vaginitis and GH in vitro and in vivo experiments. AIM OF STUDY: The effect and mechanism of JZ-1 on anti-herpes simplex virus type 2(HSV-2) in vitro focusing on adhesion and penetration stages were investigated. MATERIALS AND METHODS: A model of HSV-2 infection of VK2/E6E7 was developed. In order to explore JZ-1's anti-HSV-2 effect in vitro, cell morphology, ultrastructural pathology, cell viability and expression of viral glycoprotein D (gD) were assessed at 6 h, 12 h, 18 h, and 24 h of JZ-1 treatment. Then we measured the exact time required for adhesion and penetration of HSV-2 into VK2/E6E7 among a series of times at room temperature and under temperature control techniques. We treated VK2/E6E7 with JZ-1, penciclovir, or berberine and explored the mechanism of JZ-1 in blocking HSV-2 adhesion and penetration of host cells by assessing the cell ultrastructural pathology, viability, viral proteins gB, gD, VP16, ICP5, and ICP4 and host cell proteins HVEM, Nectin-1, and Nectin-2. RESULTS: HSV-2 can fully adhere and penetrate into VK/E6E7 within 5 mins at room temperature while it takes 60mins under temperature control techniques. JZ-1 and penciclovir showed significant anti-HSV-2 effects, with improved host cell morphologies and increased host cell viabilities observed after treatment for 24 h. The anti-HSV-2 effect of JZ-1 can be detected after treatment for 6 h while that of penciclovir was not obvious until treatment for 12 h. JZ-1 showed distinct effect on HSV-2 adhesion and penetration stages by significantly reducing the expression of viral proteins gB, gD, VP16, ICP5, and ICP4, improving cell morphology and increasing cell viability. However, these effects were not exerted via downregulated expression of membrane fusion-related proteins such as HVEM, Nectin-1, or Nectin-2. The specific anti-HSV-2 mechanism of JZ-1 need to be further explored. CONCLUSION: The anti-HSV-2 effect of JZ-1 was superior to that of penciclovir and berberine in vitro, and was mainly mediated by enhancing host cell defense and blocking adhesion and penetration of HSV-2.

Endometriosis de pared abdominal. Caso clínico y revisión / Abdominal wall endometriosis. Clinical case and review

Se presenta un caso de endometriosis de pared abdominal, en una mujer de 33 años, G1C1, con cesárea 13 años antes. Apareció nódulo subcutáneo cerca de la cicatriz de Pfannenstiel de la cesárea 6 años antes, y presentó dolor cíclico en la zona, desde hacía 6 meses. En la ecografía abdominal tenía nódulo irregular de 12x25 mm, que ya se había visto en una tomografía computadorizada. El diagnóstico clínico de edema pulmonar agudo, se confirmó con la anatomía patológica de biopsia con anestesia local, viendo tejido adiposo y fragmentos de endometriosis, y posteriormente en la resección del nódulo (AU)

We present a rare case of abdominal wall endometriosis in a 33 year-old woman, G1C1, with cesarean section 13 years before. She had a subcutaneous mass near to Pfannenstiel surgical scar 6 years before, and she presented cyclic pain in the area from 6 months back. In the transabdominal sonography there was a hypoechoic mass of 12x25 mm, déjà vu in the computed tomography. The clinical diagnostic of abdominal wall endometriosis was confirmed in the Pathology of local anesthesic biopsy, showing adipose tissue and endometriosis fragments, and was confirmed after resection of the mass (AU)

Role of the medial preoptic area in electroacupuncture treatment of reproductive dysfunction in ovariectomised rats.

OBJECTIVE: To investigate the role of the medial preoptic area (MPOA) of the hypothalamus in electroacupuncture (EA) treatment of reproductive dysfunction in ovariectomised (OVX) rats and to examine the underlying mechanisms of action. METHODS: 62 female Sprague-Dawley rats (200-220 g) underwent bilateral ovariectomy (OVX). Three weeks later, 32 rats assigned to experiment 1 were randomly divided into four groups: two groups underwent MPOA lesioning (n=16) followed by EA at CV4, CV3, SP6 and bilateral Zigong (30 min per day) for 3 days (OVX+MPOA+EA group, n=8) or no treatment (OVX+MPOA group, n=8); and two groups underwent sham (S) surgery (n=16) followed by EA (OVX+SMPOA+EA group, n=8) or no treatment (OVX+SMPOA group, n=8). Plasma oestradiol and luteinising hormone (LH) were measured by radioimmunoassay and mature vaginal epithelial cells stained and quantified post-treatment. 30 rats assigned to experiment 2 were randomised to receive EA (OVX+EA group, n=15) or remain untreated (OVX group, n=15), following which push-pull perfusion in combination with high pressure liquid chromatography/electrochemical (HPLC-EC) detection was used to measure neurotransmitter release in the MPOA. RESULTS: EA significantly increased the percentage of mature vaginal epithelial cells and plasma oestradiol levels in sham MPOA-lesioned rats compared with MPOA-lesioned OVX rats (p<0.05). In sham MPOA-lesioned OVX rats, EA significantly decreased plasma LH levels compared with no treatment (p<0.01). HPLC-EC results showed that γ-aminobutyric acid (GABA) and ß-endorphin levels in the MPAO were significantly increased in OVX rats following EA (p<0.01). CONCLUSIONS: The present study suggests that the MPOA plays a crucial role in EA treatment of abnormal reproductive function in OVX rats.

Effect of metronidazole supplemented with hydroquinone on the adhesion of Lactobacillus acidophilus in ovine vaginal cells.

This work demonstrates that the addition of metronidazole together with a ubiquitous quinone compound reduces adherence of Lactobacillus acidophilus to ovine vaginal cells. Spectrophotometric and voltammetric studies have shown that neoformed compounds were observed in these systems; there were also changes in their electroactive composition, and the oxidant status had a significantly higher value compared to the control (p<0.05). Based on reduction potential (E; mV), the distribution of electroactive compound concentrations suggests that the compounds with low reduction potential induce this behavior, which would indicate that the addition of metronidazole with a ubiquitous quinone compound to the vaginal system might increase the reductive capacity of these systems. This work shows that the study of behavior and fluctuations of the redox compounds that compose the vaginal environment, in terms of concentration and species of redox molecules, must be hierarchized in order to better understand the early stages of colonization by microorganisms.

Estrogenic and cytotoxic potentials of compounds isolated from Millettia macrophylla Benth (Fabaceae): towards a better understanding of its underlying mechanisms.

BACKGROUND: Millettia macrophylla was previously reported to have estrogenic effects and to prevent postmenopausal osteoporosis in Wistar rats. So, the study deals with the identification of its secondary metabolites and the evaluation of their estrogenicity and cytotoxicity toward tumoural cells. Thus, 13 known compounds were obtained from successive chromatographic columns and identified by NMR data compared to those previously reported. METHODS: In vitro estrogenicity of the isolates and the phenolic fraction (PF) of M. macrophylla were performed by E-screen and reporter gene assays, while their cytotoxicity was evaluated by Alamar Blue (resazurin) assay. A 3-days uterotrophic assay and the ability of PF to alleviate hot flushes in ovariectomized adult rats were tested in vivo. RESULTS: Seven of the 13 secondary metabolites turned to be estrogenic. Only two exhibited cytotoxic effects on MCF-7 and MDA-MB-231 with CC50 values of 110 µM and 160 µM, respectively. PF induced a significant (p < 0.01) MCF-7 cells proliferation and transactivated both ERα and ERß in the reported gene assay at 10-2 µg/mL. In vivo, PF acted more efficiently than the methanol crude extract, resulting to a significant (p < 0.01) increase in the uterine wet weight, uterine protein level, uterine and vaginal epithelial height at the dose of 10 mg/kg BW. In addition, PF reduced the average duration and frequency of hot flushes induced in rat. CONCLUSION: These aforementioned results indicate that PF is a good candidate for the preparation of an improved traditional medicine able to alleviate some menopausal complaints such as vaginal dryness and hot flushes. Estrogenic and cytotoxic potentials of compounds isolated from Millettia macrophylla Benth. (Fabaceae): towards a better understanding of its underlying mechanism.

Estrogenic activity of isoflavonoids from the stem bark of the tropical tree Amphimas pterocarpoides, a source of traditional medicines.

Various preparations of the African tree Amphimas pterocarpoides Harms are traditionally used to treat endocrine- related adverse health conditions. In the ovariectomized rat, the enriched in phenolics fraction of the methanol extract of stem bark of A. pterocarpoides acted as vaginotrophic agent of considerably weaker uterotrophic activity compared to estradiol. Evaluation of the fraction and 11 isoflavonoids isolated therefrom using Ishikawa cells and estrogen receptor (ER) isotype-specific reporter cells suggested that the estrogenic activity of the fraction could be attributed primarily to daidzein and dihydroglycitein and secondarily to glycitein. The potency-based selectivity of daidzein, dihydroglycitein and glycitein for gene expression through ERß versus ERα, expressed relative to estradiol, was 37, 27 and 20, respectively. However, the rank order of relative-to-estradiol potencies of induction of alkaline phosphatase in Ishikawa cells, a reliable marker of estrogenic activity, was daidzein>dihydroglycitein>>glycitein. The considerably higher estrogenic activity of dihydroglycitein compared to glycitein could be attributed to the partial agonist/antagonist activity of dihydroglycitein through ERß. Calculation of theoretical free energies of binding predicted the partial agonism/antagonism of dihydroglycitein through ERß. The fraction and the isolated isoflavonoids promoted lactogenic differentiation of HC11 mammary epithelial cells at least as effectively as premenopausal levels of estradiol. This data suggests that the estrogenic activity of the fraction likely depends on the metabolism of glycitein to dihydroglycitein; that the fraction could exert vaginotrophic activity likely without challenging endocrine cancer risk more than estrogen-alone supplementation; and that the fraction's safety for the reproductive track warrants a more detailed evaluation.

Gene Expression Profiling of Human Vaginal Cells In Vitro Discriminates Compounds with Pro-Inflammatory and Mucosa-Altering Properties: Novel Biomarkers for Preclinical Testing of HIV Microbicide Candidates.

BACKGROUND: Inflammation and immune activation of the cervicovaginal mucosa are considered factors that increase susceptibility to HIV infection. Therefore, it is essential to screen candidate anti-HIV microbicides for potential mucosal immunomodulatory/inflammatory effects prior to further clinical development. The goal of this study was to develop an in vitro method for preclinical evaluation of the inflammatory potential of new candidate microbicides using a microarray gene expression profiling strategy. METHODS: To this end, we compared transcriptomes of human vaginal cells (Vk2/E6E7) treated with well-characterized pro-inflammatory (PIC) and non-inflammatory (NIC) compounds. PICs included compounds with different mechanisms of action. Gene expression was analyzed using Affymetrix U133 Plus 2 arrays. Data processing was performed using GeneSpring 11.5 (Agilent Technologies, Santa Clara, CA). RESULTS: Microarraray comparative analysis allowed us to generate a panel of 20 genes that were consistently deregulated by PICs compared to NICs, thus distinguishing between these two groups. Functional analysis mapped 14 of these genes to immune and inflammatory responses. This was confirmed by the fact that PICs induced NFkB pathway activation in Vk2 cells. By testing microbicide candidates previously characterized in clinical trials we demonstrated that the selected PIC-associated genes properly identified compounds with mucosa-altering effects. The discriminatory power of these genes was further demonstrated after culturing vaginal cells with vaginal bacteria. Prevotella bivia, prevalent bacteria in the disturbed microbiota of bacterial vaginosis, induced strong upregulation of seven selected PIC-associated genes, while a commensal Lactobacillus gasseri associated to vaginal health did not cause any changes. CONCLUSIONS: In vitro evaluation of the immunoinflammatory potential of microbicides using the PIC-associated genes defined in this study could help in the initial screening of candidates prior to entering clinical trials. Additional characterization of these genes can provide further insight into the cervicovaginal immunoinflammatory and mucosal-altering processes that facilitate or limit HIV transmission with implications for the design of prevention strategies.

Studies and methodologies on vaginal drug permeation.

The vagina stands as an important alternative to the oral route for those systemic drugs that are poorly absorbed orally or are rapidly metabolized by the liver. Drug permeation through the vaginal tissue can be estimated by using in vitro, ex vivo and in vivo models. The latter ones, although more realistic, assume ethical and biological limitations due to animal handling. Therefore, in vitro and ex vivo models have been developed to predict drug absorption through the vagina while allowing for simultaneous toxicity and pathogenesis studies. This review focuses on available methodologies to study vaginal drug permeation discussing their advantages and drawbacks. The technical complexity, costs and the ethical issues of an available model, along with its accuracy and reproducibility will determine if it is valid and applicable. Therefore every model shall be evaluated, validated and standardized in order to allow for extrapolations and results presumption, and so improving vaginal drug research and stressing its benefits.

UVA-photoactivated riboflavin treatment of vaginal cells derived from pelvic organ prolapse cases.

BACKGROUND: The pathophysiology of pelvic organ prolapse (POP) involves vaginal collagen degradation. Strengthening collagen by UVA-photoactivated cross-linking has been demonstrated and suggested target applications include the vaginal wall. AIM: To identify UVA irradiation and riboflavin effects on vaginal cells. MATERIALS AND METHODS: Vaginal cells were incubated for 24 h (DMEM/F-12 Ham's media) and were exposed to riboflavin (0, 0.1 and 10%) for 30 min before UVA photoactivation. Percentages of live, apoptotic and necrotic cells were determined by propidium iodide/Hoechst 33342 stains. RESULTS: UVA decreased vaginal cell viability [mean ± standard error of the mean: 26.2 ± 0.5% vs. control (43.9 ± 3.8%)], but riboflavin blocked UVA-induced damage (57.9 ± 2.7 and 56.7 ± 2.1% at 0.1 and 10% riboflavin, respectively). Cells treated with low- and high-dose riboflavin had lower apoptosis (32.9 ± 1.0 and 35.5 ± 0.9%, respectively). Furthermore, riboflavin-treated cells had reduced necrosis (9.3 ± 1.7, 7.8 ± 3.0%) versus UVA-only (32.4 ± 5.5%) or control (17.1 ± 2.8%). Viability was similar for cells from the cervical and hymenal segments. CONCLUSION: The results demonstrated that riboflavin attenuated UVA damage in vaginal cells by inhibiting necrosis. Cervical and hymenal end vaginal cells were equally affected by UVA. UVA phototoxicity was reduced by the presence of riboflavin.

Germinated brown rice and its bioactives modulate the activity of uterine cells in oophorectomised rats as evidenced by gross cytohistological and immunohistochemical changes.

BACKGROUND: Germinated brown rice (GBR) is gaining momentum in the area of biomedical research due to its increased use as a nutraceutical for the management of diseases. The effect of GBR on the reproductive organs of oophorectomised rats was studied using the gross, cytological, histological and immunohistochemical changes, with the aim of reducing atrophy and dryness of the genital organs in menopause. METHODS: Experimental rats were divided into eight groups of six rats per group. Groups 1, 2 and 3 (sham-operated (SH), oophorectomised without treatment (OVX) and oophorectomised treated with 0.2 mg/kg oestrogen, respectively) served as the controls. The groups 4,5,6,7 and 8 were treated with 20 mg/kg Remifemin, 200 mg/kg of GBR, ASG, oryzanol and GABA, respectively. All treatments were administered orally, once daily for 8 weeks. Vaginal smear cytology was done at the 7th week on all the rats. The weight and dimensions of the uterus and vagina were determined after sacrifice of the rats. Uterine and vaginal tissues were taken for histology and Immunohistochemical examinations. RESULTS: GBR and its bioactives treated groups significantly increased the weight and length of both the uterus and the vagina when compared to Oophorectomised non-treated group (OVX-non-treated) (p < 0.05). Significant changes were observed in the ratio of cornified epithelial cells and number of leucocytes in the vaginal cytology between the oophorectomised non-treated and treated groups. There was also an increase in the luminal and glandular epithelial cells activity in the treated compared with the untreated groups histologically. Immunohistochemical staining showed specific proliferating cell nuclear antigen (PCNA) in the luminal and glandular epithelium of the treated groups, which was absent in the OVX-non-treated group. GBR improved the length and weight of the uterus and also increased the number of glandular and luminal cells epithelia of the vagina. CONCLUSION: GBR and its bioactives could be a potential alternative in improving reproductive system atrophy, dryness and discomfort during menopause.

Prophylactic, therapeutic and neutralizing effects of zinc oxide tetrapod structures against herpes simplex virus type-2 infection.

The attachment of Herpes simplex virus type-2 (HSV-2) to a target cell requires ionic interactions between negatively charged cell surface co-receptor heparan sulfate (HS) and positively charged residues on viral envelop glycoproteins, gB and gC. Effective blocking of this first step of HSV-2 pathogenesis demonstrates significant prophylactic effects against the viral disease; any in vitro therapeutic effects of blocking this interaction, however, are not clear. Here, we provide new evidence that zinc oxide tetrapod micro-nanostructures synthesized by flame transport approach significantly block HSV-2 entry into target cells and, in addition, demonstrate the potential to stop the spread of the virus among already infected cells. The zinc oxide tetrapods (ZnOTs) also exhibit the ability to neutralize HSV-2 virions. Natural target cells such as human vaginal epithelial and HeLa cells showed highly reduced infectivity when infected with HSV-2 virions that were pre-incubated with the ZnOTs. The mechanism behind the ability of ZnOTs to prevent, neutralize or reduce HSV-2 infection relies on their ability to bind the HSV-2 virions. We used fluorescently labeled ZnOTs and GFP-expressing HSV-2 virions to demonstrate the binding of the ZnOTs with HSV-2. We also show that the binding and hence, the antiviral effects of ZnOTs can be enhanced by illuminating the ZnOTs with UV light. Our results provide new insights into the anti-HSV-2 effects of ZnOT and rationalize their development as a HSV-2 trapping agent for the prevention and/or treatment of infection. The observed results also demonstrate that blocking HSV-2 attachment can have prophylactic as well as therapeutic applications.

Mesquite pod extract modifies the reproductive physiology and behavior of the female rat.

Phytoestrogens are non steroidal compounds that can bind to estrogen receptors, mimicking some effects of estradiol (E(2)). These compounds are widespread among legumes, which are used as pasture, and their importance in animal agriculture has increased. Mesquite (Prosopis sp) is a widespread legume, widely used to feed several livestock species in Mexico. The main product of mesquite is the pod, which is considered high quality food. As a legume, it could be assumed that mesquite contains some amounts of phytoestrogens which might induce potential estrogenic effects. However, to our knowledge, there are no reports regarding the possible estrogenic activity of this legume either in livestock or in animal models such as the rat. Therefore, in this study, we evaluated the potential estrogenic effects of mesquite pod extract on several aspects of behavior and reproductive physiology of the female rat. The effects of the extract were compared with those of E(2) and two isoflavones: daidzein (DAI) and genistein (GEN). The following treatments were given to groups of intact and ovariectomized (OVX) female rats: vehicle; mesquite pod extract; E(2); GEN; DAI. Compared to vehicle groups, mesquite pod extract, DAI, GEN, and E(2) increased uterine weight and induced growth in vaginal and uterine epithelia. In intact rats, mesquite pod extract, GEN and DAI altered estrous cyclicity, decreased lordotic quotient and intensity of lordosis. In OVX rats, mesquite pod extract, DAI and GEN induced vaginal estrus, increased vaginal epithelium height, and induced lordosis, although its intensity was reduced, compared with intact rats in estrus and E2-treated rats. These results suggest that mesquite pod extract could have estrogenic activity. However, the presence of phytoestrogens in this legume remains to be confirmed.

Inhibition of adherence of multi-drug resistant E. coli by proanthocyanidin.

Proanthocyanidin is commonly used for inhibiting urinary tract infection (UTI) of sensitive strains of Escherichia coli. The aim of this study was to investigate the effect of proanthocyanidin on adherence of uropathogenic multi-drug resistant E. coli to uroepithelial cells, which has not yet been investigated so far. Extracts of the purified proanthocyanidin were prepared from dried cranberry juice. Purity and structural assignment of proanthocyanidin was assessed using high performance liquid chromatography and (13)C nuclear magnetic resonance spectroscopy, respectively. Subsequently, its affect on multi-drug resistant bacteria as well as quantification of anti-adherence bioactivity on human vaginal and bladder epithelial cells was appraised. Inhibition of adherence to an extent of about 70% with multi-drug resistant E. coli strains was observed on uroepithelial cell. The anti-adherence bioactivity of the proanthocyanidin was detected at concentrations of 10-50 µg/ml with significant bacteriuria. Probable proanthocyanidin through A-type linkages either combines to P-fimbriae of bacterial cells or modifies the structural entity of P-fimbriae and inhibits bacterial adherence to uroepithelial cells. The proanthocyanidin exhibited anti-adherence property with multi-drug resistant strains of uropathogenic P-fimbriated E. coli with in vitro study. Hence proanthocyanidin may be considered as an inhibitory agent for multi-drug resistant strains of E. coli adherence to uroepithelial cells.

Effects of alpinumisoflavone and abyssinone V-4'-methyl ether derived from Erythrina lysistemon (Fabaceae) on the genital tract of ovariectomized female Wistar rat.

Erythrina lysistemon is an African medicinal plant used as a palliative for problems in women. The crude extract of this plant was shown to exhibit estrogenic effects on the female rat reproductive tract and on cell cuture. Using classic chromatographic methods, two compounds have been isolated as major constituents of this extract: alpinumisoflavone (1) and abyssinone V-4'-methyl ether (2). To determine whether both compounds are actives principles accounting for E. lysistemon effects, we applied the classic 3-day uterotrophic assay. We also carried out a ligand binding assay to determine whether the observed effects are estrogen receptor (ER) mediated. This study showed that whereas compound 1 displayed a fourfold preference for ERα, compound 2 bound ERα and ERß with a same affinity. The in vivo study showed that compound 1 increased the uterine wet weight by 182.23% (p < 0.01) and 71.79% (p < 0.05) at doses of 0.1 and 1 mg/kg BW/day respectively. The uterine epithelium thickened dose-dependently. Vaginal epithelial height also increased by 369.97 and 226.76% respectively (p < 0.01). Compound 2 acted only on the vagina and increased vaginal epithelial height by 244.56% (p < 0.01) at 1 mg/kg BW/day. These results suggest that compounds 1 and 2 are endowed with estrogenic properties accounting, at least in part, for E. lysistemon effects.

Histologic and cytologic detection of endocrine and reproductive tract effects of exemestane in female rats treated for up to twenty-eight days.

The objective of this study was to determine the shortest period of time necessary to detect histologic evidence of estrous cycle disruption in Sprague-Dawley rats treated for up to 28 days with the aromatase inhibitor exemestane at 1,000 mg/kg. Rats were evaluated on day 5, 8, 15, or 29. Vaginal mucification, uterine and cervical epithelial atrophy, uterine luminal epithelial vacuolation, decreased uterine granulocytes, and hypertrophy/hyperplasia of mammary ducts and alveoli were noted by day 5 and persisted throughout the study. From day 8 to day 29, absence of recent basophilic corpora lutea, increased atresia of antral follicles, interstitial cell hyperplasia, and increased luteinized follicles were present in the ovaries of treated rats. Vaginal smears detected persistent diestrus, confirming estrous cycle disruption between days 5 and 8. Ovary and uterine weights were largely unaffected. Serum hormone levels were not useful due to the study design employed. Other effects of exemestane included decreased adrenal weights and decreased cell size in both the adrenal zona fasciculata and the pituitary pars distalis. While early histologic changes were evident on day 5, only after 8 days of treatment were findings considered sufficient to clearly identify exemestane-induced estrous cycle disruption using microscopy alone.