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Sample records for antral follicle counts

  1. Comparison of antral and preantral ovarian follicle populations between Bos indicus and Bos indicus-taurus cows with high or low antral follicles counts.

    PubMed

    Silva-Santos, K C; Siloto, L S; Santos, G M G; Morotti, F; Marcantonio, T N; Seneda, M M

    2014-02-01

    The objective was to compare populations of antral and pre-antral ovarian follicles in Bos indicus and Bos indicus-taurus cows with high and low antral follicle counts. Nelore (Bos indicus, n = 20) and Nelore X Angus (1/2 Bos indicus-taurus, n = 20) cows were subjected to follicular aspiration without regard to the stage of their oestrous cycle (day of aspiration = D0) to remove all follicles ≥3 mm and induce growth of a new follicular wave. Ovaries were examined by ultrasonography on D4, D19, D34, D49 and D64, and antral follicles ≥3 mm were counted. Thereafter, cows were assigned to one of two groups: high or low antral follicular count (AFC, ≥30 and ≤15 antral follicles, respectively). After D64, ovaries were collected after slaughter and processed for histological evaluation. There was high repeatability in the numbers of antral follicles for all groups (range 0.77-0.96). The mean (±SD) numbers of antral follicles were 35 ± 9 (Bos indicus) and 38 ± 6 (Bos indicus-taurus) for the high AFC group and 10 ± 3 (Bos indicus) and 12 ± 2 (Bos indicus-taurus) follicles for the low AFC. The mean number of preantral follicles in the ovaries of Bos indicus-taurus cows with high AFC (116 226 ± 83 156 follicles) was greater (p < 0.05) than that of Bos indicus cows (63 032 ± 58 705 follicles) with high AFC. However, there was no significant correlation between numbers of antral and preantral follicles. © 2013 Blackwell Verlag GmbH.

  2. Applying ultrasonographic evaluation of antral follicle count to improve reproductive management in heifers

    USDA-ARS?s Scientific Manuscript database

    Ultrasonography is a powerful technology that can be used to improve reproductive management in heifers. By counting the number of antral follicles observed on an ultrasound screen the practitioner can gather additional information when reproductive tract scoring, because the number of antral folli...

  3. Effect of antral follicle count in beef heifers on in vitro fertilization/production

    USDA-ARS?s Scientific Manuscript database

    Our objective has been to compare the in vitro fertilization (IVF) and production (IVP) of embryos from low and high antral follicle count (AFC) heifers. This is the 4th year of the study with years 1 to 3 reported individually. For this report, we add data for the 4th year and present a combined an...

  4. Effect of high and low antral follicle count in pubertal beef heifers on IVF

    USDA-ARS?s Scientific Manuscript database

    Pubertal heifers can be classified between those with high (n = 25) or low (n = 15) antral follicle counts (AFC). The objective of this study was to determine oocyte development and maturation (e.g. fertility) in an IVF system for high- and low-AFC heifers. From a pool of 120 heifers, 10 high- and 1...

  5. Heritability and impact of environmental effects during pregnancy on antral follicle count in cattle.

    PubMed

    Walsh, S W; Mossa, F; Butler, S T; Berry, D P; Scheetz, D; Jimenez-Krassel, F; Tempelman, R J; Carter, F; Lonergan, P; Evans, A C O; Ireland, J J

    2014-07-01

    Previous studies have documented that ovarian antral follicle count (AFC) is positively correlated with number of healthy follicles and oocytes in ovaries (ovarian reserve), as well as ovarian function and fertility in cattle. However, environmental factors (e.g., nutrition, steroids) during pregnancy in cattle and sheep can reduce AFC in offspring. The role that genetic and environmental factors play in influencing the variability in AFC and, correspondingly, the size of the ovarian reserve, ovarian function, and fertility, are, however, poorly understood. The present study tests the hypothesis that variability in AFC in offspring is influenced not only by genetic merit but also by the dam age and lactation status (lactating cows vs. nonlactating heifers) and milk production during pregnancy. Antral follicle count was assessed by ultrasonography in 445 Irish Holstein-Friesian dairy cows and 522 US Holstein-Friesian dairy heifers. Heritability estimates for AFC (± standard error) were 0.31 ± 0.14 and 0.25 ± 0.13 in dairy cows and heifers, respectively. Association analysis between both genotypic sire data and phenotypic dam data with AFC in their daughters was performed using regression and generalized linear models. Antral follicle count was negatively associated with genetic merit for milk fat concentration. Also, AFC was greater in offspring of dams that were lactating (n=255) compared with nonlactating dams (n=89) during pregnancy and was positively associated with dam milk fat concentration and milk fat-to-protein ratio. In conclusion, AFC in dairy cattle is a moderately heritable genetic trait affected by age or lactation status and milk quality but not by level of dam's milk production during pregnancy. Copyright © 2014 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  6. The correlation of the antral follicle count and Serum anti-mullerian hormone

    PubMed Central

    Göksedef, Behiye Pınar; Idiş, Nurettin; Görgen, Hüsnü; Asma, Yaprak Rüstemoğlu; Api, Murat; Çetin, Ahmet

    2010-01-01

    Objective To compare the value of the basal serum anti-Müllerian hormone (AMH) level with most of the established ovarian reserve tests. Material and Methods A total of 141 infertile women was studied prospectively. On cycle day 3, serum levels of AMH, inhibin B, estradiol (E), FSH and LH levels were measured, and the number of early antral follicles (2–6 mm in diameter) estimated at ultrasound scanning to compare the strengths of hormonal-follicular correlations. Results The mean age of the participants was 29.18±5.54. The mean AMH and total AFC on day 3 were 2.23±1.90 ng/ml and 8.35±2.83, respectively. Serum AMH levels were more tightly correlated (p<0.001) with number of the early antral follicle count (r=0.467, p<0.0001) than age and serum levels of FSH (r=−0.400, p<0.001; r=−0.299, p<0.001 respectively). No correlation was detected between serum levels of inhibin B, E2, and LH (r=0.154, p=0.06; p=0.31; r=−0.085 and r=0.067, p=0.42) and AFC. Conclusion Serum AMH levels showed a strong correlation with AFC, and also this correlation is stronger than the other ovarian reserve parameters. PMID:24591939

  7. Effect of high and low antral follicle count in pubertal beef heifers on in vitro fertilization (IVF)

    USDA-ARS?s Scientific Manuscript database

    Pubertal heifers can be classified between those with high (= 25) and low (= 15) antral follicle counts (AFC). The objective of this study was to determine oocyte development and maturation (e.g., fertility) in an in vitro fertilization (IVF) system for high and low AFC heifers. From a pool of 120...

  8. In vitro fertilization (IVF) using semi-defined culture conditions from low or high antral follicle count pubertal beef heifers

    USDA-ARS?s Scientific Manuscript database

    To compare the in vitro fertilization (IVF) and production (IVP) of embryos from low and high antral follicle count (AFC) heifers, AFC were determined on 106 heifers using transrectal ultrasonography. Ten heifers with the lowest AFC (avg. 13.2) and 10 heifers with the highest AFC (avg. 27.4) with ev...

  9. In vitro fertilization (IVF) from low or high antral follicle count pubertal beef heifers using semi-defined culture conditions

    USDA-ARS?s Scientific Manuscript database

    Antral follicle counts (AFC) vary among pubertal beef heifers. Our objective was to compare the in vitro maturation and fertilization of oocytes collected from low and high AFC heifers. Previously we reported results using serum-based IVF media and in this study report results using semi-defined m...

  10. Heifers with low antral follicle counts have low birth weights and produce progeny with low birth weights

    USDA-ARS?s Scientific Manuscript database

    Research has demonstrated that heifers with increased numbers of ovarian antral follicle count (AFC) have improved hormonal profiles and improved fertility. Interestingly, heifers with low AFC had lower birth weights, suggesting that genes influencing growth and development also influence the devel...

  11. Heifers with low antral follicle counts have low birth weights and produce progeny with low birth weights

    USDA-ARS?s Scientific Manuscript database

    Research has demonstrated that heifers with increased numbers of ovarian antral follicle counts (AFC) have improved hormonal profiles and improved fertility. Interestingly, heifers with low AFC had lower birth weights, suggesting that genes influencing growth and development also influence the deve...

  12. Interrelationships among reproductive hormones and antral follicle count in human menstrual cycles

    PubMed Central

    Lee, Chel Hee; Baerwald, Angela

    2016-01-01

    It is recognised that ovarian factors, including steroid and protein hormones, are critical in the feedback regulation of pituitary gonadotropins; however, their individual contributions are less defined. The aim of this study was to explore the reciprocal relationships between ovarian and pituitary hormones across the normal ovulatory menstrual cycle as women age. FSH, LH, oestradiol, progesterone, inhibin A, inhibin B and anti-mullerian hormone (AMH) were measured in serum collected every 1–3 days across one interovulatory interval (IOI) from 26 healthy women aged 18–50 years. The antral follicle count (AFC) for follicles 2–5 mm, >6 mm and 2–10 mm were tabulated across the IOI. Independent associations between ovarian hormones/AFC vs pituitary follicle-stimulating hormone (FSH) and luteinising hormone (LH) were investigated using multivariate regression analysis. The data were sub-grouped based on the presence or absence luteal phase-dominant follicles (LPDF). Serum oestradiol and AMH were inversely correlated with FSH in both follicular and luteal phases. Inhibin B correlated inversely with FSH and LH in the late follicular phase and directly in the luteal phase. AFC, inhibin A and progesterone were not key predictors of either FSH or LH. The strong association between AMH and FSH with age implies that AMH, as well as oestradiol and inhibin B are important regulators of FSH. The change in feedback response of inhibin B with both FSH and LH across the cycle suggests two phases of the negative feedback. PMID:27856496

  13. Individualization of the FSH starting dose in IVF/ICSI cycles using the antral follicle count

    PubMed Central

    2013-01-01

    Background The FSH starting dose is usually chosen according to women’s age, anamnesis, clinical criteria and markers of ovarian reserve. Currently used markers include antral follicle count (AFC), which is considered to have a very high performance in predicting ovarian response to FSH. The objective of the present study to elaborate a nomogram based on AFC for the calculation of the appropriate FSH starting dose in IVF cycles. Methods This is a retrospective study performed at the Mother-Infant Department of Modena University Hospital. IVF patients (n=505) were subjected to blood sampling and transvaginal ultrasound for measurement of serum day3 FSH, estradiol and AFC. The variables predictive of the number of retrieved oocytes were assessed by backwards stepwise multiple regression. The variables reaching the statistical significance were then used in the calculation for the final predictive model. Results A model based on age, AFC and FSH was able to accurately predict the ovarian sensitivity and accounted for 30% of the variability of ovarian response to FSH. An FSH dosage nomogram was constructed and overall it predicts a starting dose lower than 225 IU in 50.2% and 18.1% of patients younger and older than 35 years, respectively. Conclusions The daily FSH dose may be calculated on the basis of age and two markers of ovarian reserve, namely AFC and FSH, with the last two variables being the most significant predictors. The nomogram seems easily applicable during the daily clinical practice. PMID:23388048

  14. Individualization of the FSH starting dose in IVF/ICSI cycles using the antral follicle count.

    PubMed

    La Marca, Antonio; Grisendi, Valentina; Giulini, Simone; Argento, Cindy; Tirelli, Alessandra; Dondi, Giulia; Papaleo, Enrico; Volpe, Annibale

    2013-02-06

    The FSH starting dose is usually chosen according to women's age, anamnesis, clinical criteria and markers of ovarian reserve. Currently used markers include antral follicle count (AFC), which is considered to have a very high performance in predicting ovarian response to FSH. The objective of the present study to elaborate a nomogram based on AFC for the calculation of the appropriate FSH starting dose in IVF cycles. This is a retrospective study performed at the Mother-Infant Department of Modena University Hospital. IVF patients (n=505) were subjected to blood sampling and transvaginal ultrasound for measurement of serum day3 FSH, estradiol and AFC. The variables predictive of the number of retrieved oocytes were assessed by backwards stepwise multiple regression. The variables reaching the statistical significance were then used in the calculation for the final predictive model. A model based on age, AFC and FSH was able to accurately predict the ovarian sensitivity and accounted for 30% of the variability of ovarian response to FSH. An FSH dosage nomogram was constructed and overall it predicts a starting dose lower than 225 IU in 50.2% and 18.1% of patients younger and older than 35 years, respectively. The daily FSH dose may be calculated on the basis of age and two markers of ovarian reserve, namely AFC and FSH, with the last two variables being the most significant predictors. The nomogram seems easily applicable during the daily clinical practice.

  15. Anti-Müllerian hormone versus antral follicle count for defining the starting dose of FSH.

    PubMed

    Lan, Vuong Thi Ngoc; Linh, Nguyen Khanh; Tuong, Ho Manh; Wong, P C; Howles, Colin M

    2013-10-01

    This pilot study compared the efficacy and safety of two simple dosing algorithms, one based on anti-Müllerian Hormone (AMH) and the other on the antral follicle count (AFC), to determine the starting dose of recombinant FSH (rFSH) for ovarian stimulation in 348 women. Patients were randomized to a predefined AMH- or AFC-based algorithm. The proportion of cycles with the desired response was similar when rFSH dose was determined using AMH or AFC (35.2% versus 28.4%). There was a significant difference between the groups in the proportion of cycles with a hyperresponse (8.6% and 17.4%, but the incidence of ovarian hyperstimulation syndrome was similar (1.1% and 4.6%). There were no significant differences between two groups in outcomes, including implantation (19.3% versus 19.0%), clinical pregnancy (38.0% versus 46.9%), multiple pregnancy (16.5% versus 15.2%) and miscarriage (7.0% versus 8.3%). However, statistically significant differences in ovarian response were evident among the AMH and AFC subgroups: for AMH, Desired and Hypo; for AFC, Hypo and Hyper. This pilot study provides information for developing protocols to further validate the use of either AMH or AFC to guide the starting dose of rFSH in ovarian stimulation. The ideal outcome for couples undergoing IVF treatment is the birth of a healthy baby. One factor that might influence this is retrieving an adequate number of eggs, which are obtained using various treatment protocols. A group of drugs called gonadotrophins have been used for more than 20years to stimulate the ovaries to produce eggs. However, the dose to start treatment has not been clearly defined. A few studies have looked at ways to use the best gonadotrophin dose for each woman, but to be useful in the clinic any approach needs to be simple and easy to use. This study compared the effectiveness and safety of two simple approaches to determining the starting dose of recombinant FSH (rFSH) for ovarian stimulation in women undergoing IVF

  16. Antral Follicles Count and Anti-Müllerian Hormone Levels after Gonadotoxic Chemotherapy in Patients with Breast Cancer: Cohort Study.

    PubMed

    D'Avila, Ângela Marcon; Capp, Edison; Corleta, Helena von Eye

    2017-04-01

    Aim To assess ovarian reserve (OVR) by means of follicle-stimulating hormone (FSH), anti-Müllerian hormone (AMH), and antral follicle count (AFC) measurement in eumenorrheic women with breast cancer, exposed to gonadotoxic chemotherapy. Method Fifty-two women (35.3 ± 3.8 years old) with breast cancer and undergoing cyclophosphamide-containing chemotherapy were enrolled. The assessment was performed before chemotherapy (T1) and after 2 (T2) and 6 months (T3). Results Six months after chemotherapy, the prevalence of regular cycles was 60%. Anti-Müllerian hormone decreased down to undetectable levels at T2 and T3 (T1: 2.53 [1.00-5.31]; T2 < 0.08; T3: < 0.08 [< 0.08-1.07] ng/mL), (p < 0.0001). Antral follicle count was 11 [8.0-13.5] follicles at T1 and lower at T2 (5.50 [3.75-8.0] and T3 (5.0 [2.5-7.0]) (p < 0.0001). In patients who remained with regular cycles during chemotherapy or resumed normal menses, FSH and estradiol levels remained unchanged. Conclusion Anti-Müllerian hormone and AFC are useful as markers of OVR decline in women exposed to chemotherapy. Follicle-stimulating hormone is only adequate in women who become amenorrheic. Thieme-Revinter Publicações Ltda Rio de Janeiro, Brazil.

  17. Predictive value of antral follicle count and anti-Müllerian hormone for follicle and oocyte developmental competence during the early prepubertal period in a sheep model.

    PubMed

    Torres-Rovira, Laura; Gonzalez-Bulnes, Antonio; Succu, Sara; Spezzigu, Antonio; Manca, Maria E; Leoni, Giovanni G; Sanna, Marina; Pirino, Salvatore; Gallus, Marilia; Naitana, Salvatore; Berlinguer, Fiammetta

    2014-10-01

    Circulating anti-Müllerian hormone (AMH) and antral follicle count (AFC) are addressed as suitable markers of oocyte quantity and quality during adulthood. To investigate whether AFC and circulating AMH could predict follicle development and oocyte quality during the prepubertal period we used 40-day-old ewe lambs with high, intermediate and low AFC (≥30, 16-29 and≤15 follicles respectively). The analysis of the response to the exogenous FSH ovarian reserve test showed a positive correlation between AFC, AMH plasma levels, total follicle number and the number of large follicles (≥3mm) grown after exogenous FSH administration. The incorporation of abattoir-derived oocytes collected from ovaries with different AFC in an in vitro embryo production system showed that a high AFC can predict oocyte quality in prepubertal ovaries, reflecting an ovarian status suitable for follicular development. The histological quantification of the ovarian reserve evidenced that AFC was not predictive of differences in either the number of healthy follicles or the size of the primordial follicle pool in prepubertal ovaries. Further studies are needed to investigate the implication on the reproductive performance of the significant inter-individual differences found in the present study in AFC and circulating AMH in the early prepubertal period.

  18. Differences in antral follicle counts in pubertal Angus heifers are associated with differences in the uterine transcriptome during the late luteal phase

    USDA-ARS?s Scientific Manuscript database

    Heifers with high numbers of antral follicles conceive earlier in the breeding season, have greater follicular fluid hormone concentrations and greater uterine protein secretion. Therefore, we hypothesized that increased numbers of antral follicles associate with differentially expressed genes in t...

  19. Anti-Müllerian hormone concentrations and antral follicle counts for the prediction of pregnancy outcomes after intrauterine insemination.

    PubMed

    Moro, Francesca; Tropea, Anna; Scarinci, Elisa; Leoncini, Emanuele; Boccia, Stefania; Federico, Alex; Alesiani, Ornella; Lanzone, Antonio; Apa, Rosanna

    2016-04-01

    To evaluate anti-Müllerian hormone (AMH) concentrations and antral follicle counts (AFCs) in the prediction of pregnancy outcomes after controlled ovarian stimulation among women undergoing intrauterine insemination. A retrospective study included women with unexplained infertility aged 41years or younger who attended a fertility clinic in Italy between December 2009 and May 2014. Ovarian stimulation was achieved with recombinant follicle-stimulating hormone or highly purified human menopausal gonadotropin. Receiver operating characteristic curves were generated to predict ongoing pregnancy. The primary outcome was the association between AMH/AFC and ongoing pregnancy, and was assessed by logistic regression. Overall, 276 women were included, of whom 43 (15.6%) achieved ongoing pregnancy. Multivariate analysis showed that women with a serum day-3 concentration of AMH higher than 2.3ng/mL were more likely to have ongoing pregnancy than were those with a concentration lower than 2.3ng/mL (odds ratio 5.84, 95% confidence interval 2.38-14.31; P<0.001). No associations were recorded for AFCs. AMH should be used to predict the pregnancy outcome of intrauterine insemination. Copyright © 2015 International Federation of Gynecology and Obstetrics. Published by Elsevier Ireland Ltd. All rights reserved.

  20. Inter-cycle and inter-observer variability of the antral follicle count in routine clinical practice.

    PubMed

    Subirá, Jessica; Alberola-Rubio, Jose; Núñez, María Jose; Escrivá, Alicia Marzal; Pellicer, Antonio; Montañana, Vicente; Díaz-García, César

    2017-07-01

    Antral follicle count (AFC) is a reliable predictor of ovarian response to stimulation and its inter-cycle and inter-observer variability has been extensively studied on in vitro fertilization (IVF), mostly in highly selected populations within studies not originally designed for this purpose. In this retrospective cohort study, we assess the inter-cycle variation of AFC in a setting similar to that of the daily practice. We included only patients undergoing mild stimulation for intrauterine insemination (IUI). One hundred and forty-eight patients had two (62 patients, group A), three (49 patients, group B) or four (37 patients, group C) IUI cycles and AFC was measured on early follicular phase of each cycle by one of the members of the medical team within daily practice. Intra-class correlation coefficients were used to estimate variability. Inter-cycle variability rendered ICCs above 0.70 in all groups improving along with the number of cycles [Group A ICC 0.78 (95%CI 0.66-0.86), Group B ICC 0.87 (95%CI 0.80-0.92) and Group C ICC 0.91 (95%CI 0.85-0.95)]. Inter-observer variability showed a high degree of concordance with ICCs above 0.95. We provide the closest approximation to real inter-cycle and inter-observer AFC variability expected in routine clinical practice.

  1. The association of protein intake (amount and type) with ovarian antral follicle counts among infertile women: results from the EARTH prospective study cohort.

    PubMed

    Souter, I; Chiu, Y-H; Batsis, M; Afeiche, M C; Williams, P L; Hauser, R; Chavarro, J E

    2017-09-01

    To evaluate the association between protein intake (amount and type) and antral follicle count (AFC). Prospective cohort. Academic fertility centre. Two hundred and sixty-five women undergoing fertility treatments at an academic fertility centre and participating in an ongoing study on environment and reproductive health. We measured AFC in ultrasonographic evaluation among women undergoing infertility treatments. Women completed a previously validated semi-quantitative food frequency questionnaire. We used Poisson regression to evaluate the relation between protein intake and AFC while adjusting for age, body mass index, race, smoking status, and total energy intake. Antral follicle count. Among 265 women (mean age: 35.0 ± 3.9 years, 85% Caucasian), total protein intake (% energy) was unrelated to AFC. When protein from different food sources was considered separately, we found a negative association between dairy protein intake and AFC. The mean AFC was 14.4% (3.9-23.7%) lower for women in the highest quintile of dairy protein intake than for women in the bottom quintile after adjusting for potential confounders (P-trend = 0.04). This association was stronger among women who had never smoked (P-trend = 0.002) but was not observed among previous smokers (P-trend = 0.36). There were no associations between protein intake from either non-dairy animal or vegetable sources and AFC. Higher dairy protein intake (≥5.24% of energy) was associated with lower antral follicle counts among women presenting for infertility treatment. These findings should be further investigated in prospective studies also designed to clarify the biology underlying the observed associations. Higher dairy protein intake was associated with lower antral follicle counts in an infertile population. © 2017 Royal College of Obstetricians and Gynaecologists.

  2. Antral Follicle Count Predicts Natural Menopause in a Population-Based Sample: The CARDIA Women’s Study

    PubMed Central

    Wellons, Melissa F.; Bates, Gordon Wright; Schreiner, Pamela J.; Siscovick, David S.; Sternfeld, Barbara; Lewis, Cora E.

    2013-01-01

    Objective The timing of menopause is associated with multiple chronic diseases. Tools to predict this milestone have relevance for clinical and research purposes. Among infertile women, a positive relationship exists between antral follicle count (AFC) and response to controlled ovarian hyperstimulation, a marker of ovarian reserve. However, a relationship between AFC and menopause that is age-independent has not been demonstrated. Thus, our objective was to evaluate the relationship between AFC measured in women at ages 34–49 and incident natural menopause over 7-years of follow-up. Methods The Coronary Artery Risk Development in Young Adults (CARDIA) study is a longitudinal community-based study (Chicago, Illinois; Birmingham, Alabama; Minneapolis, Minnesota; and Oakland, California) begun in 1985–1986. In 2002–03, the CARDIA Women’s Study measured FSH levels and performed a transvaginal ultrasound protocol that included AFC (2mm–10mm follicles on both ovaries). Incident natural menopause was assessed by survey in 2005–06 and 2009–10. Results In our sample (n=456), median AFC and FSH were 5 (IQR 2–9) and 7.8 mIU/mL (IQR 5.6–11.0), respectively, at a mean age of 42 (range 34–49) in 2002–03. 101 women reported natural menopause by 2009–10. In Cox models, current smoking, stable menses, FSH>13, and AFC ≤4 were independently associated with incident natural menopause. Compared to AFC >4, those with AFC ≤4 were nearly twice as likely to have undergone menopause over 7-years of follow-up (HR 1.89, 95% CI 1.19–3.02) after adjustment for covariates. Conclusion AFC is independently associated with natural menopause over 7-years of follow-up after controlling for other markers of ovarian aging. PMID:23422869

  3. Suitability of antral follicle counts and computer-assisted analysis of ultrasonographic and magnetic resonance images for estimating follicular reserve in porcine, ovine and bovine ovaries ex situ.

    PubMed

    Warren, Lisa; Murawski, Maciej; Wilk, Karolina; Zieba, Dorota A; Bartlewski, Pawel M

    2015-05-01

    This study was conducted to determine if correlations exist between the numbers of microscopic follicles comprising ovarian follicular reserve (OFR) and antral follicle counts (AFCs), and to assess the usefulness of computerized analyses of ovarian ultrasonograms and magnetic resonance (MR) images for estimating OFR in excised porcine, ovine and bovine ovaries. As a pre-requisite to these analyses, we characterized and compared ovarian cortical histomorphology and follicle populations in the three species varying in prolificacy and overall reproductive longevity, and hence the total number of microscopic and antral follicles. Ultrasonographic and MR images were obtained at the scanner settings optimized to provide opposing contrasts between antral follicles and the ovarian stroma. Commercially available ImageProPlus® analytical software was used to calculate numerical pixel values (NPVs) and pixel heterogeneity (standard deviation of the pixel values) along the computer-generated lines (4-6) placed in the area corresponding to the ovarian cortex. The numbers of primordial (r = 0.38, P < 0.01) and intermediate follicles (r = 0.37, P < 0.01) were correlated with the numbers of antral follicles in bovine ovarian sections. The numbers of primordial (r = 0.28, P < 0.05), intermediate (r = 0.31, P < 0.01) and primary follicles (r = 0.27, P < 0.05) correlated directly with mean NPVs of the ultrasonographic ovarian images in cattle. There was a negative correlation between primary follicle numbers and NPVs of MR images (3D FAST-SPOILED GRADIENT ECHO) of the porcine ovarian cortex (r = -0.31, P < 0.05). To summarize, the numbers of primordial and intermediate follicles could only be estimated from AFCs in cows. Using ultrasound NPVs, the numbers of primordial, intermediate and primary follicles could be directly estimated in bovine ovaries and the quantitative image attributes of MR images were useful for quantifying porcine primary follicles. The bovine ovarian model is

  4. Suitability of antral follicle counts and computer-assisted analysis of ultrasonographic and magnetic resonance images for estimating follicular reserve in porcine, ovine and bovine ovaries ex situ

    PubMed Central

    Warren, Lisa; Murawski, Maciej; Wilk, Karolina; Zieba, Dorota A

    2015-01-01

    This study was conducted to determine if correlations exist between the numbers of microscopic follicles comprising ovarian follicular reserve (OFR) and antral follicle counts (AFCs), and to assess the usefulness of computerized analyses of ovarian ultrasonograms and magnetic resonance (MR) images for estimating OFR in excised porcine, ovine and bovine ovaries. As a pre-requisite to these analyses, we characterized and compared ovarian cortical histomorhpology and follicle populations in the three species varying in prolificacy and overall reproductive longevity, and hence the total number of microscopic and antral follicles. Ultrasonographic and MR images were obtained at the scanner settings optimized to provide opposing contrasts between antral follicles and the ovarian stroma. Commercially available ImageProPlus® analytical software was used to calculate numerical pixel values (NPVs) and pixel heterogeneity (standard deviation of the pixel values) along the computer-generated lines (4–6) placed in the area corresponding to the ovarian cortex. The numbers of primordial (r = 0.38, P < 0.01) and intermediate follicles (r = 0.37, P < 0.01) were correlated with the numbers of antral follicles in bovine ovarian sections. The numbers of primordial (r = 0.28, P < 0.05), intermediate (r = 0.31, P < 0.01) and primary follicles (r = 0.27, P < 0.05) correlated directly with mean NPVs of the ultrasonographic ovarian images in cattle. There was a negative correlation between primary follicle numbers and NPVs of MR images (3D FAST-SPOILED GRADIENT ECHO) of the porcine ovarian cortex (r = −0.31, P < 0.05). To summarize, the numbers of primordial and intermediate follicles could only be estimated from AFCs in cows. Using ultrasound NPVs, the numbers of primordial, intermediate and primary follicles could be directly estimated in bovine ovaries and the quantitative image attributes of MR images were useful for quantifying

  5. Specific ranges of anti-Mullerian hormone and antral follicle count correlate to provide a prognostic indicator for IVF outcome.

    PubMed

    Keane, Kevin; Cruzat, Vincius Fernandes; Wagle, Susbin; Chaudhary, Nitin; Newsholme, Philip; Yovich, John

    2017-03-01

    Advanced female age correlates with reduced ovarian reserve (OR) and is the primary factor underlying the limitation of success rates in Assisted Reproductive Technology (ART). Currently, predicted ovarian response to gonadotrophin stimulation is determined using transvaginal ultrasound to measure antral follicle count (AFC), an indirect measure of OR. However, assessing the level of anti-Mullerian hormone (AMH) has more recently been shown to correlate with OR, and its application has been adopted widely. This retrospective study was designed to determine the relationship between novel ranges of AMH and AFC in patients undergoing ART. There was a positive correlation between AMH and AFC category (r=0.458, P<0.01), with an 87% linear concordance observed for specific AFC ranges and mean serum AMH levels. Both OR markers were inversely correlated with female age (r=-0.428 and r=-0.392, respectively). Pregnancy and live birth rates were influenced by both AMH (P<0.05) and AFC categories (P<0.05). Conversely, miscarriage rates appeared to be more dependent on AFC categories (P >0.05), but even more reliant on female age. Finally, the number of oocytes collected was positively correlated with AMH and AFC grading, while oocyte and embryo utilization rates were negatively correlated. Overall, both OR markers were positively and strongly related with each other, and when individual AMH readings were categorised into specific novel ranges, they demonstrated a more robust correlation with AFC groupings. Taken together, applying patient AMH within specific ranges may lead to a better estimation of OR and IVF outcomes.

  6. Antral follicle count as a marker of ovarian biological age to reflect the background risk of fetal aneuploidy.

    PubMed

    Grande, Maribel; Borobio, Virginia; Jimenez, Jose Miguel; Bennasar, Mar; Stergiotou, Iosifina; Peñarrubia, Joana; Borrell, Antoni

    2014-06-01

    Can antral follicle count (AFC) measured during pregnancy be used as a marker of ovarian age to assess the background risk of fetal aneuploidy? AFC was lower than expected according to maternal chronological age in trisomic pregnancies; therefore ovarian age could potentially reflect a more precise background risk of fetal aneuploidy screening. The decline in a woman's reproductive function is determined by a decline in the ovarian follicle pool and the quality of oocytes. The quantitative status of ovarian reserve can be indirectly assessed by AFC, but the role of AFC as an aneuploidy risk marker in pregnant women has not been assessed yet. Our study comprised a prospective cohort including 1239 singleton pregnancies scanned before 14 weeks in our center during a 14-month period. Reference ranges for AFC were constructed using 812 spontaneously conceived, chromosomally normal singleton ongoing pregnancies using the Lambda-Mu-Sigma method. The study population (n = 934) included 19 pregnancies with viable autosomal trisomies (trisomies 21, 18 and 13), 17 non-viable autosomal trisomies (other than 21, 18 or 13), 7 monosomies X, 1 sex trisomy and 3 triploidies (total n = 47 with chromosomal abnormalities). AFC in chromosomally abnormal pregnancies was plotted against the reference ranges. AFC multiple of the median was calculated according to the median AFC obtained by each year of age. Sixty-eight percent of women carrying a pregnancy with viable trisomies and 65% with non-viable trisomies presented an AFC below the 50th percentile. The median ovarian age in viable trisomies and non-viable trisomies was estimated to be 3 and 6 years above than median maternal age, respectively. However, the median ovarian age in monosomies X and triploidies was not higher than median maternal age. We did not assess the intra- and inter-observer reliability, or use specific three-dimensional analysis which may have advantages over our two-dimensional study. In clinical practice, a

  7. The association of bisphenol-A urinary concentrations with antral follicle counts and other measures of ovarian reserve in women undergoing infertility treatments.

    PubMed

    Souter, Irene; Smith, Kristen W; Dimitriadis, Irene; Ehrlich, Shelley; Williams, Paige L; Calafat, Antonia M; Hauser, Russ

    2013-12-01

    In this prospective cohort of women undergoing infertility treatments, we measured specific-gravity adjusted urinary BPA (SG-BPA) concentrations and used regression models to evaluate the association of BPA with antral follicle count (AFC), day-3 serum follicle stimulating hormone levels (FSH), and ovarian volume (OV). BPA, detected in >80% of women, had a geometric mean (±GSD) of 1.6±2.0, 1.7±2.1, and 1.5±1.8μg/L for the women contributing to the AFC (n=154), day-3 FSH (n=120), and OV (n=114) analyses, respectively. There was an average decrease in AFC of 12% (95% CI: -23%, -0.6%), 22% (95% CI: -31%, -11%), and 17% (95% CI: -27%, -6%), in the 2nd, 3rd, and 4th SG-BPA quartile compared to the 1st quartile, respectively (p-trend: <0.001). No association of SG-BPA with FSH or OV was observed. Among women from an infertility clinic, higher urinary BPA concentrations were associated with lower AFC, raising concern for possible accelerated follicle loss and reproductive aging.

  8. The Association of Bisphenol-A Urinary Concentrations with Antral Follicle Counts and Other Measures of Ovarian Reserve in Women Undergoing Infertility Treatments.☆

    PubMed Central

    Souter, Irene; Smith, Kristen W; Dimitriadis, Irene; Ehrlich, Shelley; Williams, Paige L; Calafat, Antonia M; Hauser, Russ

    2015-01-01

    In this prospective cohort of women undergoing infertility treatments, we measured specific-gravity adjusted urinary BPA (SG-BPA) concentrations and used regression models to evaluate the association of BPA with antral follicle count (AFC), day-3 serum follicle stimulating hormone levels (FSH), and ovarian volume (OV). BPA, detected in >80% of women, had a geometric mean (±GSD) of 1.6±2.0, 1.7±2.1, and 1.5±1.8µg/L for the women contributing to the AFC (n=154), day-3 FSH (n=120), and OV (n=1 14) analyses, respectively. There was an average decrease in AFC of 12% (95% CI: −23%, −0.6%), 22% (95% CI: −31%, −11%), and 17% (95% CI: −27%, −6%), in the 2nd, 3rd, and 4th SG-BPA quartile compared to the 1st quartile, respectively (p-trend: <0.001). No association of SG-BPA with FSH or OV was observed. Among women from an infertility clinic, higher urinary BPA concentrations were associated with lower AFC, raising concern for possible accelerated follicle loss and reproductive aging. PMID:24100206

  9. Evaluation of ovarian reserve using anti-müllerian hormone and antral follicle count in Sjögren's syndrome: Preliminary study.

    PubMed

    Karakus, Savas; Sahin, Ali; Durmaz, Yunus; Aydin, Huseyin; Yildiz, Caglar; Akkar, Ozlem; Dogan, Mansur; Cengiz, Ahmet; Cetin, Meral; Cetin, Ali

    2017-02-01

    The aim of this study was to determine ovarian reserve status using anti-müllerian hormone (AMH) level and antral follicle count (AFC) in patients with Sjögren's syndrome (SS). Twenty-four women with SS diagnosed according to the classification criteria proposed by the American-European Consensus Group and 25 healthy women as controls were enrolled in this study. Ovarian reserve was assessed on clinical findings, AFC, and serum AMH and reproductive hormone levels. Compared with the healthy controls, in the SS patients, the duration of menstrual cycle was significantly shorter (P = 0.043); serum AMH (P = 0.001) and AFC (P = 0.001) were significantly lower, and serum luteinizing hormone (LH) was significantly higher (P = 0.019). The right (P = 0.555) and left ovarian (P = 0.386) volumes were also lower but this did not reach statistical significance. Serum follicle-stimulating hormone (P = 0.327), estradiol (P = 0.241), and prolactin (P = 0.55) were similar between the two groups. Ovarian reserve may be reduced in SS patients. For the assessment of ovarian reserve, serum AMH and ovarian AFC with serum LH may be useful. Further studies with long-term follow-up are required to determine the course of ovarian reserve abnormalities and best possible biomarkers of reduced ovarian reserve in SS patients. © 2016 Japan Society of Obstetrics and Gynecology.

  10. Evaluation of Antral Follicle Count and Ovarian Morphology in Crossbred Beef Cows: Investigation of Influence of Stage of the Estrous Cycle, Age, and Birth Weight

    USDA-ARS?s Scientific Manuscript database

    Depletion of the ovarian reserve is associated with reproductive senescence in mammalian females, and there is a positive relationship between the size of the ovarian reserve and the number of antral follicles on the surface of the ovary. Therefore, we conducted a series of experiments to investiga...

  11. Association between antral follicle count and reproductive measures in New Zealand lactating dairy cows maintained in a pasture-based production system.

    PubMed

    Martinez, Marcelo F; Sanderson, Neil; Quirke, Laurel D; Lawrence, Stephen B; Juengel, Jennifer L

    2016-02-01

    The antral follicle count (AFC) in cattle is consistent throughout the estrous cycle of individual cows, and cows with a lower AFC have lower fertility. We assessed the AFC at random stages of the estrous cycle, examined the correlation between AFC classifications, and determined the relationship between the most rapid and practical laboratory-based AFC classification (AFC of follicles of ≥ 2 mm in diameter) and fertility measures in New Zealand lactating dairy cows. Cows detected in estrus (n = 202) or not (n = 239) during the first 4 weeks of the breeding season were subjected to ultrasonography and classified as having a high, medium, or low AFC at the time of scanning (on-site classification). Images from ultrasound scanning were recorded onto video for accurate follicle counting in an imaging laboratory. A strong association (P < 0.05) between the AFC of follicles with a diameter of 2 mm or greater and fertility was observed. Cows with a high AFC had a shorter (P < 0.05) interval from calving to conception by artificial insemination (AI; 82.4 ± 1.6 vs. 87.3 ± 1.2 days) and greater pregnancy rates (PRs; i.e., PR to the first AI [68.1% vs. 45.3%], 6-week PR [81.9% vs. 67.3%], and overall PR [91.3% vs. 79.7%]) than cows with a low AFC. The AFC was positively associated (P < 0.0001) with age. Progesterone concentrations during diestrus were greater (P < 0.05) in high-AFC cows (7.6 ± 0.3 ng/mL) than in low-AFC cows (6.5 ± 0.3 ng/mL), whether these were pregnant (7.7 ± 0.3 ng/mL) or not (6.3 ± 0.2 ng/mL). A rapid on-site scoring system determined that cows classified as having a high AFC had a shorter (P < 0.05) interval from calving to the first AI (76.5 ± 1.7 vs. 82.3 ± 1.9 days) and were more likely to show estrus (P < 0.01; 56.8% vs. 36.4%) and have a CL at the beginning of the breeding season (P < 0.01; 93.4% vs. 79.6%) than cows with a low on-site AFC. Collectively, we have confirmed an association between AFC2 and fertility, and these results

  12. Antral follicle count predicts natural menopause in a population-based sample: the Coronary Artery Risk Development in Young Adults Women's Study.

    PubMed

    Wellons, Melissa F; Bates, Gordon Wright; Schreiner, Pamela J; Siscovick, David S; Sternfeld, Barbara; Lewis, Cora E

    2013-08-01

    The timing of menopause is associated with multiple chronic diseases. Tools that predict this milestone have relevance for clinical and research purposes. Among infertile women, a positive relationship exists between antral follicle count (AFC) and response to controlled ovarian hyperstimulation, a marker of ovarian reserve. However, an age-independent relationship between AFC and menopause has not been demonstrated. Thus, our objective was to evaluate the relationship between AFC measured in women aged 34 to 49 years and incident natural menopause during 7 years of follow-up. The Coronary Artery Risk Development in Young Adults Study is a longitudinal community-based study (Chicago, IL; Birmingham, AL; Minneapolis, MN; and Oakland, CA) begun in 1985-1986. In 2002-2003, the Coronary Artery Risk Development in Young Adults Women's Study measured follicle-stimulating hormone (FSH) levels and performed a transvaginal ultrasound protocol that included AFC (2-10 mm follicles on both ovaries). Incident natural menopause was assessed by surveys in 2005-2006 and 2009-2010. In our sample (n = 456), the median (interquartile range) AFC and FSH level were 5 (2-9) and 7.8 (5.6-11.0) mIU/mL, respectively, at a mean (range) age of 42 (34-49) years in 2002-2003. One hundred one women reported natural menopause by 2009-2010. In Cox models, current smoking, stable menses, FSH level higher than 13 mIU/mL, and AFC of 4 or less were independently associated with incident natural menopause. Compared with AFC higher than 4, those with an AFC of 4 or less were nearly twice as likely to have undergone menopause during 7 years of follow-up (hazard ratio, 1.89; 95% CI, 1.19-3.02) after adjustment for covariates. AFC has been found to be independently associated with natural menopause during 7 years of follow-up after controlling for other markers of ovarian aging.

  13. Relationship of antral follicle counts to fertility and ovarian AMH mRNA levels in beef cows

    USDA-ARS?s Scientific Manuscript database

    In mammalian females, reproductive senescence is associated with decreased numbers of follicles in the ovary. Anti-Mullerian Hormone (AMH) of ovarian origin has been proposed to be a biomarker of the number of follicles in the ovary and of fertility in women, as well as to control activation of pri...

  14. Predictive values of anti-müllerian hormone, antral follicle count and ovarian response prediction index (ORPI) for assisted reproductive technology outcomes.

    PubMed

    Ashrafi, Mahnaz; Hemat, Mandana; Arabipoor, Arezoo; Salman Yazdi, Reza; Bahman-Abadi, Akram; Cheraghi, Rezvaneh

    2017-01-01

    This prospective study was performed from June 2012 to June 2014 at Royan Institute to compare the predictive values of serum anti-müllerian hormone (AMH), antral follicle count (AFC) and ovarian response prediction index (ORPI) ([AFC × AMH]/age) for in vitro fertilisation/intracytoplasmic sperm injection (IVF/ICSI) cycle outcomes. Five hundred and fifty women included in the study with male factor and unexplained infertility were the first candidates for IVF/ICSI cycles. Serum AMH level was measured by a commercial ELISA kit and AFC was calculated by a transvaginal ultrasonography on day 2-3 of the menstrual cycle before starting ovarian stimulation. All women underwent IVF/ICSI cycles using a long standard protocol with gonadotropin-releasing hormone agonist. The receiver operating characteristic curves (ROC) analysis showed that both AMH and AFC were good predictors of ovarian response with an area under the curves (AUC) > 0.75; even it seems that AFC was being a better predictor. Combining these variables is necessary as ORPI will not improve the prediction value. All the variables had poor predictive ability (AUC <0.60) for clinical pregnancy and live birth rates. Logistic regression analysis showed the AMH less than 0.4 ng/ml and quality of transferred embryos were significant predictors for clinical pregnancy rate.

  15. Antimüllerian hormone levels and antral follicle count as prognostic indicators in a personalized prediction model of live birth.

    PubMed

    Nelson, Scott M; Fleming, Richard; Gaudoin, Marco; Choi, Bokyung; Santo-Domingo, Kenny; Yao, Mylene

    2015-08-01

    To compare antimüllerian hormone (AMH) and antral follicle count (AFC) separately and in combination with clinical characteristics for the prediction of live birth after controlled ovarian stimulation. Retrospective development and temporal external validation of prediction model. Outpatient IVF clinic. We applied the boosted tree method to develop three prediction models incorporating clinical characteristics plus AMH or AFC or the combination on 2,124 linked IVF cycles from 2006 to 2010 and temporally externally validated predicted live-birth probabilities with an independent data set comprising 1,121 cycles from 2011 to 2012. None. Predictive power (posterior log of odds ratio compared to age, or PLORA), reclassification, receiver operator characteristic analysis, calibration, dynamic range. Predictive power, was highest for the AMH model (PLORA = 29.1), followed by the AMH-AFC model (PLORA = 28.3) and AFC model (PLORA = 22.5). The prediction errors were 1% to <5% in each prognostic tier for all three models, except for the predicted live-birth probabilities of <10% in the AFC model, where the prediction error was 8%. The improvement in predictive power was highest for the AMH model: 76.2% improvement over age alone relative to 59% improvement for AFC and 73.3% for the combined model. Receiver operating characteristic analysis demonstrated that the AMH and the combined model had comparable discrimination (area under the curve = 0.716) and similar prediction error for high and low strata of live-birth prediction, with an improvement of 6.3% over age alone. The validated prediction model confirmed that AMH when combined with clinical characteristics can accurately identify the likelihood of live birth with a low prediction error. AFC provided no added predictive value beyond AMH. Copyright © 2015 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  16. Role of Baseline Antral Follicle Count and Anti-Mullerian Hormone in Prediction of Cumulative Live Birth in the First In Vitro Fertilisation Cycle: A Retrospective Cohort Analysis

    PubMed Central

    Li, Hang Wun Raymond; Lee, Vivian Chi Yan; Lau, Estella Yee Lan; Yeung, William Shu Biu; Ho, Pak Chung; Ng, Ernest Hung Yu

    2013-01-01

    Objective This retrospective study determined for the first time the role of baseline antral follicle count (AFC) and serum anti-Mullerian hormone (AMH) level in the first in-vitro fertilisation (IVF) cycle in predicting cumulative live birth from one stimulation cycle. Methods We studied 1,156 women (median age 35 years) undergoing the first IVF cycle. Baseline AFC and AMH level on the day before ovarian stimulation were analysed. The main outcome measure was cumulative live birth in the fresh plus all the frozen embryo transfers after the same stimulation cycle. Results Serum AMH was significantly correlated with AFC. Both AMH and AFC showed significant correlation with age and ovarian response in the stimulated cycle and total number of transferrable embryos. Baseline AFC and serum AMH were significantly higher in subjects attaining a live birth than those who did not in the fresh stimulated cycle, as well as those attaining cumulative live birth. There was a significant trend of higher cumulative live birth rate in women with higher AMH or AFC. However, logistic regression revealed that both AMH and AFC were not significant predictors of cumulative live birth after adjusting for age and number of embryos available for transfer. Considering only one single predictor, the areas under the ROC curves for AMH (0.646, 95% CI 0.616–0.675) and age (0.648, 95% CI 0.618–0.677) were slightly higher than that for AFC (0.617, 95% CI 0.587–0.647) in predicting cumulative live birth. However, a model combining AMH (with or without AFC) and age of the women only classified an addition of less than 2% of subjects correctly compared to the model with age alone. Conclusion Baseline AFC and serum AMH have only modest predictive performance on the occurrence of cumulative live birth, and may not give additional value on top of the women's age. PMID:23637787

  17. Di(2-ethylhexyl) phthalate inhibits antral follicle growth, induces atresia, and inhibits steroid hormone production in cultured mouse antral follicles

    SciTech Connect

    Hannon, Patrick R. Brannick, Katherine E. Wang, Wei Gupta, Rupesh K. Flaws, Jodi A.

    2015-04-01

    Di(2-ethylhexyl) phthalate (DEHP) is a ubiquitous environmental toxicant found in consumer products that causes ovarian toxicity. Antral follicles are the functional ovarian units and must undergo growth, survival from atresia, and proper regulation of steroidogenesis to ovulate and produce hormones. Previous studies have determined that DEHP inhibits antral follicle growth and decreases estradiol levels in vitro; however, the mechanism by which DEHP elicits these effects is unknown. The present study tested the hypothesis that DEHP directly alters regulators of the cell cycle, apoptosis, and steroidogenesis to inhibit antral follicle functionality. Antral follicles from adult CD-1 mice were cultured with vehicle control or DEHP (1–100 μg/ml) for 24–96 h to establish the temporal effects of DEHP on the follicle. Following 24–96 h of culture, antral follicles were subjected to gene expression analysis, and media were subjected to measurements of hormone levels. DEHP increased the mRNA levels of cyclin D2, cyclin dependent kinase 4, cyclin E1, cyclin A2, and cyclin B1 and decreased the levels of cyclin-dependent kinase inhibitor 1A prior to growth inhibition. Additionally, DEHP increased the mRNA levels of BCL2-associated agonist of cell death, BCL2-associated X protein, BCL2-related ovarian killer protein, B-cell leukemia/lymphoma 2, and Bcl2-like 10, leading to an increase in atresia. Further, DEHP decreased the levels of progesterone, androstenedione, and testosterone prior to the decrease in estradiol levels, with decreased mRNA levels of side-chain cleavage, 17α-hydroxylase-17,20-desmolase, 17β-hydroxysteroid dehydrogenase, and aromatase. Collectively, DEHP directly alters antral follicle functionality by inhibiting growth, inducing atresia, and inhibiting steroidogenesis. - Highlights: • DEHP inhibits antral follicle growth by dysregulating cell cycle regulators. • DEHP induces antral follicle atresia by dysregulating apoptosis regulators. • DEHP

  18. A prediction model to select PCOS patients suitable for IVM treatment based on anti-Mullerian hormone and antral follicle count.

    PubMed

    Guzman, L; Ortega-Hrepich, C; Polyzos, N P; Anckaert, E; Verheyen, G; Coucke, W; Devroey, P; Tournaye, H; Smitz, J; De Vos, M

    2013-05-01

    Which baseline patient characteristics can help assisted reproductive technology practitioners to identify patients who are suitable for in-vitro maturation (IVM) treatment? In patients with polycystic ovary syndrome (PCOS) who undergo oocyte IVM in a non-hCG-triggered system, circulating anti-Müllerian hormone (AMH), antral follicle count (AFC) and total testosterone are independently related to the number of immature oocytes and hold promise as outcome predictors to guide the patient selection process for IVM. Patient selection criteria for IVM treatment have been described in normo-ovulatory patients, although patients with PCOS constitute the major target population for IVM. With this study, we assessed the independent predictive value of clinical and endocrine parameters that are related to oocyte yield in patients with PCOS undergoing IVM. Cohort study involving 124 consecutive patients with PCOS undergoing IVM whose data were prospectively collected. Enrolment took place between January 2010 and January 2012. Only data relating to the first IVM cycle of each patient were included. Patients with PCOS underwent oocyte retrieval for IVM after minimal gonadotrophin stimulation and no hCG trigger. Correlation coefficients were calculated to investigate which parameters are related to immature oocyte yield (patient's age, BMI, baseline hormonal profile and AMH, AFC). The independence of predictive parameters was tested using multivariate linear regression analysis. Finally, multivariate receiver operating characteristic (ROC) analyses for cumulus oocyte complexes (COC) yield were performed to assess the efficiency of the prediction model to select suitable candidates for IVM. Using multivariate regression analysis, circulating baseline AMH, AFC and baseline total testosterone serum concentration were incorporated into a model to predict the number of COC retrieved in an IVM cycle, with unstandardized coefficients [95% confidence interval (CI)] of 0.03 (0

  19. Di(2-ethylhexyl) phthalate inhibits antral follicle growth, induces atresia, and inhibits steroid hormone production in cultured mouse antral follicles

    PubMed Central

    Hannon, Patrick R.; Brannick, Katherine E.; Wang, Wei; Gupta, Rupesh K.; Flaws, Jodi A.

    2015-01-01

    Di(2-ethylhexyl) phthalate (DEHP) is a ubiquitous environmental toxicant found in consumer products that causes ovarian toxicity. Antral follicles are the functional ovarian units and must undergo growth, survival from atresia, and proper regulation of steroidogenesis to ovulate and produce hormones. Previous studies have determined that DEHP inhibits antral follicle growth and decreases estradiol levels in vitro; however, the mechanism by which DEHP elicits these effects is unknown. The present study tested the hypothesis that DEHP directly alters regulators of the cell cycle, apoptosis, and steroidogenesis to inhibit antral follicle functionality. Antral follicles from adult CD-1 mice were cultured with vehicle control or DEHP (1-100μg/ml) for 24-96 hr to establish the temporal effects of DEHP on the follicle. Following 24-96 hr of culture, antral follicles were subjected to gene expression analysis, and media were subjected to measurements of hormone levels. DEHP increased the mRNA levels of cyclin D2, cyclin dependent kinase 4, cyclin E1, cyclin A2, and cyclin B1 and decreased the levels of cyclin-dependent kinase inhibitor 1A prior to growth inhibition. Additionally, DEHP increased the mRNA levels of BCL2-associated agonist of cell death, BCL2-associated X protein, BCL2-related ovarian killer protein, B-cell leukemia/lymphoma 2, and Bcl2-like 10, leading to an increase in atresia. Further, DEHP decreased the levels of progesterone, androstenedione, and testosterone prior to the decrease in estradiol levels, with decreased mRNA levels of side-chain cleavage, 17α-hydorxylase-17,20-desmolase, 17β-hydroxysteroid dehydrogenase, and aromatase. Collectively, DEHP directly alters antral follicle functionality by inhibiting growth, inducing atresia, and inhibiting steroidogenesis. PMID:25701202

  20. The correlation between the number of antral follicles and ovarian reserves (preantral follicles) in purebred Bos indicus and Bos taurus cows.

    PubMed

    Silva-Santos, Katia Cristina; Santos, Gustavo Martins Gomes Dos; Siloto, Letícia Schmidt; Santos, Joabel Tonellotto Dos; Oliveira, Eduardo Raele de; Machado, Fernanda Zandonadi; Rosa, Camila Oliveira; Seneda, Marcelo Marcondes

    2014-12-30

    The objectives of this study were to compare populations of preantral follicles between purebred Bos indicus and Bos taurus cows with high or low antral follicle counts (AFC) and to correlate the number of preantral follicles with the population of antral follicles. Nelore (Bos indicus, n=100) and Angus (Bos taurus, n=100) cow ovaries were collected at abattoirs and examined using ultrasonography. Antral follicles ≥3mm were counted, and the cows ovaries were assigned to high (G-High) or low (G-Low) AFC groups based on the mean number (±1 SD) of ovarian antral follicles: Bos indicus with high AFC (≥57 follicles, n=8) or low AFC (≤21 follicles, n=8) and Bos taurus with high (≥45 follicles, n=10) or low AFC (≤13 follicles, n=10). The ovaries were processed, and the number of preantral follicles was estimated. Between-groups comparisons were performed using a Kruskal-Wallis test, and the correlation between preantral and antral follicles was evaluated using a Pearson's correlation test (P≤0.05). A large variation in the number of preantral follicles was observed among the animals. Although there was a correlation between the population of preantral follicles and the number of antral follicles, there was no difference between the mean number of preantral follicles in the Bos indicus G-High (48,349±30,149) and G-Low groups (33,037±31,710) or between the Bos taurus G-High (35,050±36,060) and G-Low groups (30,481±43,360). Therefore, the preantral follicle population did not differ between purebred Bos indicus and Bos taurus cattle with high or low AFC but was correlated with the number of antral follicles. In addition to the large within-groups variation in the number of preantral follicles, some cows with high AFC had lower populations of preantral follicles compared to the low AFC group, and the highest population of preantral follicles was observed in both Bos indicus and Bos taurus with low AFC. Copyright © 2014 Elsevier B.V. All rights reserved.

  1. Abnormal growth of ovarian antral follicles in breast cancer patients.

    PubMed

    Byskov, A G; McNatty, K P; Westergaard, L; Larsen, J K; Grinsted, J; Peters, H

    1983-07-01

    Ovarian antral follicles from patients with breast cancer were compared with follicles from healthy women. Steroid levels in the follicular fluid and the health status of the follicles were evaluated. Follicles were judged to be healthy or atretic by flow cytometric determinations of the deoxyribonucleic acid content of aspirated granulosa cell nuclei. Fifteen of the 25 follicles (60%) from the cancer patients contained unmeasurable or abnormally low steriod levels (i.e., less than 100 ng/ml) which were significantly (P less than 0.001) lower than in follicles of the same health status from healthy women (500 to 1000 ng/ml). It is speculated whether substances other than the usual follicular steriods are produced by the cancer patients, which stimulate mitotic activity of the granulosa cells.

  2. The significance of antral follicle size prior to stimulation in predicting ovarian response in a multiple dose GnRH antagonist protocol.

    PubMed

    Lai, Qiaohong; Chen, Cai; Zhang, Zhijun; Zhang, Shu; Yu, Qilin; Yang, Ping; Hu, Jun; Wang, Cong-Yi

    2013-01-01

    Prediction of ovarian responses prior to stimulation is not only useful for patient counseling, but also important in tailoring the optimal dosage of gonadotrophin for individual patients. By prospectively study of 214 women undergoing in vitro fertilization and embryo transfer (IVF-ET) treatment, we obtained data supporting that antral follicle size could be an additional valuable predictive marker other than the antral follicle count (AFC) in predicting ovarian response. Our studies revealed that AFC achieved the best predictive value in relation to the number of oocyte obtained, followed by antral follicle size, basal follicle stimulating hormone (FSH) and body mass index (BMI). Unlike AFC, antral follicle size was noted to be negatively correlated with the dosage (R = -0.493) and duration (R = -0.465) of rFSH stimulation. Antral follicle size was also found with higher negative regression coefficient (B = -0.661) as compared with that of basal FSH concentration (B = -0.326) and BMI (b = -0.281). More importantly, women with antral follicle size 6-7 mm showed significantly higher AFC, oocytes retrieved, fertilized oocytes and grade I/II embryos along with much lower transfer cycle cancellation rate (7.5% vs. 16-17%). Together, our data suggest that basal antral follicle size could be a valued predictive marker in women with IVF-ET treatment, in which women with antral follicle size 6-7 mm are likely predisposed to better IVF-ET outcomes.

  3. Antral follicle count (AFC) can be used in the prediction of ovarian response but cannot predict the oocyte/embryo quality or the in vitro fertilization outcome in an egg donation program.

    PubMed

    Melo, Marco Antonio Barreto; Garrido, Nicolás; Alvarez, Claudio; Bellver, José; Meseguer, Marcos; Pellicer, Antonio; Remohí, José

    2009-01-01

    To verify whether the antral follicle count (AFC) could predict ovarian response, oocyte/embryo quality, and IVF outcome. Prospective study. Instituto Universitario-Instituto Valenciano de Infertilidad, Valencia, Spain. One thousand seventy-four donors and 975 oocyte recipient cycles. Controlled ovarian hyperstimulation (COH), endometrial preparation, IVF/intracytoplasmic sperm injection, ET. COH and oocyte/embryo quality parameters and IVF outcome. We observed lower E(2) levels and fewer mature retrieved oocyte numbers among donors who showed an AFC that was <10. These donors also showed significantly higher cancellation and no-donation rates; poor and/or insufficient response was the principal cause (82%). However, there were no differences among the groups regarding embryo development parameters and IVF outcome. AFC is a noninvasive and simple tool that can improve the oocyte donor's selection of an egg donation program. This study suggests that AFC is a good predictor of ovarian response but cannot be used to predict oocyte/embryo quality or IVF outcome.

  4. Beef heifers with diminished numbers of antral follicles have decreased uterine protein concentrations

    USDA-ARS?s Scientific Manuscript database

    Previous research demonstrated a favorable relationship between the number of follicles detectable in the bovine ovary by ultrasonography and fertility, and bovine females with diminished numbers of antral follicles had smaller reproductive tracts. Therefore, we hypothesized that uterine function w...

  5. Morphometric characteristics of preantral and antral follicles and expression of factors involved in folliculogenesis in ovaries of adult baboons (Papio anubis).

    PubMed

    Amorim, Christiani A; Moya, Cristina Fortuño; Donnez, Jacques; Dolmans, Marie-Madeleine

    2016-05-01

    Baboons are commonly utilized as an animal model for studies of human reproduction. However, folliculogenesis in this species has not been fully documented. The aim of this study was to assess follicle morphometry and expression of essential proteins involved in folliculogenesis in baboons. Ovaries were recovered from four adult baboons and processed for histological evaluation and immunohistochemical analyses. Follicle proportion, follicle and oocyte diameter, theca layer thickness, number of granulosa cells, and follicle density were calculated. Immunohistochemical staining was also carried out for connexin 43 (Cx43), aromatase, and zona pellucida 3 (ZP3). A total of 2221 follicles were counted and measured. Proportions of primordial, primary, secondary, small antral, and large antral follicles were 49, 26, 23, 1, and 1 %, respectively. The increase in follicle diameter was due not only to the increase in oocyte diameter but also to granulosa cell proliferation. Almost all antral follicles were positive for Cx43 (89.8 %), aromatase (84.8 %), and ZP3 (100 %). Most secondary follicles were positive for Cx43 (65 %) and ZP3 (64.5 %), and some primary follicles were positive only for Cx43. No primordial follicles stained positive in any of these immunohistochemical analyses. Only antral follicles showed aromatase activity. On the basis of these results, we can conclude that folliculogenesis in baboons appears to be similar to that in humans, and this animal therefore constitutes a valuable model.

  6. Anti-Müllerian hormone and antral follicle count reveal a late impairment of ovarian reserve in patients undergoing low-gonadotoxic regimens for hematological malignancies.

    PubMed

    Di Paola, Rossana; Costantini, Claudio; Tecchio, Cristina; Salvagno, Gian Luca; Montemezzi, Rachele; Perandini, Alessio; Pizzolo, Giovanni; Zaffagnini, Stefano; Franchi, Massimo

    2013-01-01

    The impact of cancer therapy on the reproductive potential of patients is increasingly recognized because survival rates of patients have clearly improved in recent years. Different fertility preservation methods, either generally accepted or still experimental, are currently available, and counseling of patients requires a delicate balance between the efficacy and side effects of the proposed method and the characteristics of both the tumor and the therapy. Deeper knowledge of the effects of cancer therapy on the reproductive potential of patients over time is required to identify the most appropriate fertility preservation method. In this paper, we report a case-control study in which female patients who were diagnosed with hematological malignancies and treated with chemotherapy and/or radiotherapy were compared with age-matched controls in terms of ovarian reserve, as measured by ultrasound examination and hormonal status. By stratifying patients for gonadotoxicity of the therapy received and time elapsed from the end of the therapy, we report that patients treated with low gonadotoxic therapies, while being similar to age-matched controls in their ovarian reserve when evaluated within a few years from the end of the therapy, show a clear impairment over longer times. We also report that anti-Müllerian hormone is the most sensitive hormonal parameter in detecting changes in ovarian reserve when compared with follicle-stimulating hormone or inhibin-B. This study stresses the importance of accurate counseling at the time of diagnosis of cancer and emphasizes the risks of infertility with low gonadotoxic therapies that may reduce the reproductive window of survivors.

  7. Repeatability of antral follicle counts and anti-Müllerian hormone and their associations determined at an unknown stage of follicular growth and an expected day of follicular wave emergence in dairy cows.

    PubMed

    Gobikrushanth, M; Dutra, P A; Bruinjé, T C; Colazo, M G; Butler, S T; Ambrose, D J

    2017-04-01

    The objectives were to determine (1) the repeatability of antral follicle counts (AFC) and anti-Müllerian hormone (AMH) concentrations at an unknown stage of follicular growth (FG) and at an expected day of follicular wave emergence (FWE) in dairy cows, and (2) the association between AFC and AMH at the above two stages. Transrectal ultrasound imaging of the ovaries and blood sample collection were performed on 91 lactating Holstein cows (33 primiparous, 58 multiparous) to determine AFC and serum concentrations of AMH at an unknown stage of FG (mean ± SEM; 14 ± 0.5 d postpartum) and an expected day of FWE (mean ± SEM; 73 ± 0.5 d postpartum, approximately 36 h after the 2nd GnRH of Ovsynch protocol). The AFC ranged from 10 to 53 (mean ± SEM; 26.1 ± 1.0) and 6 to 45 (mean ± SEM; 23.4 ± 1.0) at an unknown stage of FG and expected day of FWE, respectively (P = 0.02). Serum concentrations (pg/mL) of AMH ranged from 13.9 to 528.8 (mean ± SEM; 187.3 ± 13.1) and 38.2 to 774.1 (mean ± SEM; 218.7 ± 14.5) at an unknown stage of FG and expected day of FWE, respectively (P < 0.01). The repeatability estimates for AFC and serum AMH concentrations between an unknown stage of FG and the expected day of FWE were 0.37 and 0.73 (P < 0.01), respectively. The correlation between AFC and AMH was moderate at an unknown stage of FG (r = 0.54; P < 0.01) and at an expected day of FWE (r = 0.59; P < 0.01). In summary, both AFC and AMH were repeatable when measured at an unknown stage of FG and an expected day of FWE, but the repeatability was greater for AMH. Thus, if cows are tested at an unknown stage of FG, serum AMH may be a more reliable phenotype than AFC to test association with reproductive outcomes. Furthermore, AFC and AMH were moderately correlated at the two stages examined.

  8. Monohaloacetic acid drinking water disinfection by-products inhibit follicle growth and steroidogenesis in mouse ovarian antral follicles in vitro

    PubMed Central

    Jeong, Clara H.; Gao, Liying; Dettro, Tyler; Wagner, Elizabeth D.; Ricke, William A.; Plewa, Michael J.; Flaws, Jodi A.

    2016-01-01

    Water disinfection greatly reduced the incidence of waterborne diseases, but the reaction between disinfectants and natural organic matter in water leads to the formation of drinking water disinfection by-products (DBPs). DBPs have been shown to be toxic, but their effects on the ovary are not well defined. This study tested the hypothesis that monohalogenated DBPs (chloroacetic acid, CAA; bromoacetic acid, BAA; iodoacetic acid, IAA) inhibit antral follicle growth and steroidogenesis in mouse ovarian follicles. Antral follicles were isolated and cultured with either vehicle or DBPs (0.25–1.00 mM of CAA; 2–15 µM of BAA or IAA) for 48 and 96 h. Follicle growth was measured every 24 h and the media were analyzed for estradiol levels at 96 h. Exposure to DBPs significantly inhibited antral follicle growth and reduced estradiol levels compared to controls. These data demonstrate that DBP exposure caused ovarian toxicity in vitro. PMID:27151372

  9. Monohaloacetic acid drinking water disinfection by-products inhibit follicle growth and steroidogenesis in mouse ovarian antral follicles in vitro.

    PubMed

    Jeong, Clara H; Gao, Liying; Dettro, Tyler; Wagner, Elizabeth D; Ricke, William A; Plewa, Michael J; Flaws, Jodi A

    2016-07-01

    Water disinfection greatly reduced the incidence of waterborne diseases, but the reaction between disinfectants and natural organic matter in water leads to the formation of drinking water disinfection by-products (DBPs). DBPs have been shown to be toxic, but their effects on the ovary are not well defined. This study tested the hypothesis that monohalogenated DBPs (chloroacetic acid, CAA; bromoacetic acid, BAA; iodoacetic acid, IAA) inhibit antral follicle growth and steroidogenesis in mouse ovarian follicles. Antral follicles were isolated and cultured with either vehicle or DBPs (0.25-1.00mM of CAA; 2-15μM of BAA or IAA) for 48 and 96h. Follicle growth was measured every 24h and the media were analyzed for estradiol levels at 96h. Exposure to DBPs significantly inhibited antral follicle growth and reduced estradiol levels compared to controls. These data demonstrate that DBP exposure caused ovarian toxicity in vitro.

  10. Sphingosine-1-phosphate and ceramide are associated with health and atresia of bovine ovarian antral follicles.

    PubMed

    Hernández-Coronado, C G; Guzmán, A; Espinosa-Cervantes, R; Romano, M C; Verde-Calvo, J R; Rosales-Torres, A M

    2015-02-01

    The follicle destiny towards ovulation or atresia is multi-factorial in nature and involves outcries, paracrine and endocrine factors that promote cell proliferation and survival (development) or unchain apoptosis as part of the atresia process. In several types of cells, sphingosine-1-phospate (S1P) promotes cellular proliferation and survival, whereas ceramide (CER) triggers cell death, and the S1P/CER ratio may determine the fate of the cell. The aim of present study was to quantify S1P and CER concentrations and their ratio in bovine antral follicles of 8 to 17 mm classified as healthy and atretic antral follicles. Follicles were dissected from cow ovaries collected from a local abattoir. The theca cell layer, the granulosa cells and follicular fluid were separated, and 17β-estradiol (E2) and progesterone (P4) concentrations were measured in the follicular fluid by radioimmunoassay. Based on the E2/P4 ratio, the follicles were classified as healthy (2.2±0.3) or atretic (0.2±0.3). In both follicular compartments (granulosa and theca cell layer), sphingolipids were extracted and S1P and CER concentrations were quantified by HPLC (XTerra RP18; 5 µm, 3.0×150 mm column). Results showed that in both follicular compartments, S1P concentrations were higher in healthy antral follicles than in atretic antral follicles (P<0.05). The concentration of CER in the granulosa cells was higher in atretic antral follicles than in healthy antral follicles, but no differences were observed in the theca cell layer. The S1P/CER ratio in both follicular compartments was also higher in healthy antral follicles. Interestingly, in these follicles, there was a 45-fold greater concentration of S1P than CER in the granulosa cells (P<0.05), whereas in the theca cell layer, S1P had only a 14-fold greater concentration than CER when compared with atretic antral follicles. These results suggest that S1P plays a role in follicle health, increasing cellular proliferation and survival. In

  11. Genistein exposure inhibits growth and alters steroidogenesis in adult mouse antral follicles.

    PubMed

    Patel, Shreya; Peretz, Jackye; Pan, Yuan-Xiang; Helferich, William G; Flaws, Jodi A

    2016-02-15

    Genistein is a naturally occurring isoflavone phytoestrogen commonly found in plant products such as soybeans, lentils, and chickpeas. Genistein, like other phytoestrogens, has the potential to mimic, enhance, or impair the estradiol biosynthesis pathway, thereby potentially altering ovarian follicle growth. Previous studies have inconsistently indicated that genistein exposure may alter granulosa cell proliferation and hormone production, but no studies have examined the effects of genistein on intact antral follicles. Thus, this study was designed to test the hypothesis that genistein exposure inhibits follicle growth and steroidogenesis in intact antral follicles. To test this hypothesis, antral follicles isolated from CD-1 mice were cultured with vehicle (dimethyl sulfoxide; DMSO) or genistein (6.0 and 36μM) for 18-96h. Every 24h, follicle diameters were measured to assess growth. At the end of each culture period, the media were pooled to measure hormone levels, and the cultured follicles were collected to measure expression of cell cycle regulators and steroidogenic enzymes. The results indicate that genistein (36μM) inhibits growth of mouse antral follicles. Additionally, genistein (6.0 and 36μM) increases progesterone, testosterone, and dehydroepiandrosterone (DHEA) levels, but decreases estrone and estradiol levels. The results also indicate that genistein alters the expression of steroidogenic enzymes at 24, 72 and 96h, and the expression of cell cycle regulators at 18h. These data indicate that genistein exposure inhibits antral follicle growth by inhibiting the cell cycle, alters sex steroid hormone levels, and dysregulates steroidogenic enzymes in cultured mouse antral follicles. Copyright © 2016 Elsevier Inc. All rights reserved.

  12. Genistein Exposure Inhibits Growth and Alters Steroidogenesis in Adult Mouse Antral Follicles

    PubMed Central

    Patel, Shreya; Peretz, Jackye; Pan, Yuan-Xiang; Helferich, William G.; Flaws, Jodi A.

    2016-01-01

    Genistein is a naturally occurring isoflavone phytoestrogen commonly found in plant products such as soybeans, lentils, and chickpeas. Genistein, like other phytoestrogens, has the potential to mimic, enhance, or impair the estradiol biosynthesis pathway, thereby potentially altering ovarian follicle growth. Previous studies have inconsistently indicated that genistein exposure may alter granulosa cell proliferation and hormone production, but no studies have examined the effects of genistein on intact antral follicles. Thus, this study was designed to test the hypothesis that genistein exposure inhibits follicle growth and steroidogenesis in intact antral follicles. To test this hypothesis, antral follicles isolated from CD-1 mice were cultured with vehicle (dimethyl sulfoxide; DMSO) or genistein (6.0 and 36 μM) for 18 – 96 hours (h). Every 24 h, follicle diameters were measured to assess growth. At the end of each culture period, the media were pooled to measure hormone levels, and the cultured follicles were collected to measure expression of cell cycle regulators and steroidogenic enzymes. The results indicate that genistein (36 μM) inhibits growth of mouse antral follicles. Additionally, genistein (6.0 and 36 μM) increases progesterone, testosterone, and dehydroepiandrosterone (DHEA) levels, but decreases estrone and estradiol levels. The results also indicate that genistein alters the expression of steroidogenic enzymes at 24, 72 and 96 h, and the expression of cell cycle regulators at 18 h. These data indicate that genistein exposure inhibits antral follicle growth by inhibiting the cell cycle, alters sex steroid hormone levels, and dysregulates steroidogenic enzymes in cultured mouse antral follicles. PMID:26792615

  13. Quantitative bioluminescence imaging of transgene expression in intact porcine antral follicles in vitro

    USDA-ARS?s Scientific Manuscript database

    The porcine oocyte maturation in vivo occurs within the ovarian follicle and is regulated by the interactions between oocytes and surrounding follicular components, including theca, granulosa, and cumulus cells, and follicular fluid. Therefore, the antral follicle is an essential microenvironment fo...

  14. Localization and expression of follicle-stimulating hormone receptor gene in buffalo (Bubalus bubalis) pre-antral follicles.

    PubMed

    Sharma, G Taru; Dubey, P K; Kumar, G Sai

    2011-02-01

    Follicle-stimulating hormone (FSH) stimulates antral follicles to grow, but its role in earlier stages (pre-antral) of follicle development, if any, is obscure. Aim of this study was to study the expression of follicle-stimulating hormone receptor (FSHR) gene in different sizes of pre-antral follicles (PFs) (<150, 200, 250, 300, 350, 400 μm) and to find out an optimum dose of FSH for better growth, development and steroidogenesis of PFs in vitro. Buffalo ovaries were collected from a local abattoir, and PFs were isolated by mechanical method. A semi-quantitative RT-PCR amplification strategy was used for mRNA expression, while FSHR protein was localized by immunohistochemistry. Isolated pre-antral follicles (80-85 μm) were cultured in TCM-199 supplemented with 10% foetal bovine serum, 1% ITS and 30 ng/ml EGF served as control medium. Addition of three different doses of FSH (0.5, 1.0, 2.0 μg/ml) in control medium was considered as treatment groups. A single 2.184-kb receptor mRNA transcript was present in all sizes (<150-400 μm) of follicles. Follicle-stimulating hormone receptor was also localized immunohistochemically in granulosa cells of all sizes of follicles. Survival and growth rate of follicles significantly (p<0.05) increased following supplementation of FSH at a concentration of 1.0 μg/ml and the culture medium also showed a significantly (p<0.05) greater accumulation of oestradiol and progesterone. In conclusion, FSHR is expressed in all sizes of PFs and in vitro survival, growth and steroidogenesis of follicles are optimally stimulated by 1.0 μg/ml FSH. These findings demonstrate that FSH has an important role during the recruitment, growth and development of buffalo ovarian PFs.

  15. Beef heifers with diminished numbers of antral follicles have decreased uterine protein concentrations.

    PubMed

    McNeel, Anthony K; Soares, Émerson M; Patterson, Allyson L; Vallet, Jeffrey L; Wright, Elane C; Larimore, Erin L; Amundson, Olivia L; Miles, Jeremy R; Chase, Chadwick C; Lents, Clay A; Wood, Jennifer R; Cupp, Andrea S; Perry, George A; Cushman, Robert A

    2017-04-01

    Previous research demonstrated a favorable relationship between the number of follicles detectable in the bovine ovary by ultrasonography and fertility, and bovine females with diminished numbers of antral follicles had smaller reproductive tracts. Therefore, we hypothesized that uterine function would be compromised in beef heifers with diminished numbers of antral follilcles. Angus heifers (n=480) were submitted for ultrasonographic evaluation of antral follicle number at 325 and 355d of age. After the second ultrasonographic examination, 40 pubertal heifers with the greatest average number of antral follicles (30.9±0.7) and 40 pubertal heifers with the lowest average number of antral follicles (14.2±0.7) were synchronized with two i.m. injections of prostaglandin F2α (25mg) administered 11d apart, and heifers were slaughtered on d6 (n=26 heifers/group) or d16 (n=14 heifers/group) of the resultant estrous cycle. The uterus was weighed, flushed for determination of protein content, and representative samples were fixed for determination of endometrial gland morphometry. Heifers in the Low group had fewer surface antral follicles and smaller reproductive tracts than heifers in the High group (P<0.01). Protein content of the uterine flushes was decreased in heifers in the Low group (P<0.01); however, there was no difference in the percent area of the endometrium occupied by endometrial glands. From these results, we conclude that the uterine environment of beef heifers with diminished numbers of antral follicles is less conducive to supporting early embryonic survival. Published by Elsevier B.V.

  16. Infertile women below the age of 40 have similar anti-Müllerian hormone levels and antral follicle count compared with women of the same age with no history of infertility.

    PubMed

    Hvidman, H W; Bentzen, J G; Thuesen, L L; Lauritsen, M P; Forman, J L; Loft, A; Pinborg, A; Nyboe Andersen, A

    2016-05-01

    Do infertile patients below the age of 40 years have a lower ovarian reserve, estimated by anti-Müllerian hormone (AMH) and total antral follicle count (AFC), than women of the same age with no history of infertility? Serum AMH and AFC were not lower in infertile patients aged 20-39 years compared with a control group of the same age with no history of infertility. WHAT IS KNOWN ALREADY?: The management of patients with a low ovarian reserve and a poor response to controlled ovarian stimulation (COS) remains a challenge in assisted reproductive technologies (ART). Both AMH levels and AFC reflect the ovarian reserve and are valuable predictors of the ovarian response to exogenous gonadotrophins. However, there is a large inter-individual variation in the age-related depletion of the ovarian reserve and a broad variability in the levels of AMH and AFC compatible with conception. Women with an early depletion of the ovarian reserve may experience infertility as a consequence of postponement of childbearing. Thus, low ovarian reserve is considered to be overrepresented among infertile patients. A prospective cohort study including 382 women with a male partner referred to fertility treatment at Rigshospitalet, Copenhagen, Denmark during 2011-2013 compared with a control group of 350 non-users of hormonal contraception with no history of infertility recruited during 2008-2010. Included patients and controls were aged 20-39 years. Women with polycystic ovary syndrome were excluded. On Cycle Days 2-5, AFC and ovarian volume were measured by transvaginal sonography, and serum levels of AMH, FSH and LH were assessed. Infertile patients had similar AMH levels (11%, 95% confidence interval (CI): -1;24%) and AFC (1%, 95% CI: -7;8%) compared with controls with no history of infertility in an age-adjusted linear regression analysis. The prevalence of very low AMH levels (<5 pmol/l) was similar in the two cohorts (age-adjusted odds ratio: 0.9, 95% CI: 0.5;1.7). The findings

  17. Equol inhibits growth, induces atresia, and inhibits steroidogenesis of mouse antral follicles in vitro

    PubMed Central

    Mahalingam, Sharada; Gao, Liying; Gonnering, Marni; Helferich, William; Flaws, Jodi A.

    2016-01-01

    Equol is a non-steroidal estrogen metabolite produced by microbial conversion of daidzein, a major soy isoflavone, in the gut of some humans and many animal species. Isoflavones and their metabolites can affect endogenous estradiol production, action, and metabolism, potentially influencing ovarian follicle function. However, no studies have examined the effects of equol on intact ovarian antral follicles, which are responsible for sex steroid synthesis and further development into ovulatory follicles. Thus, the present study tested the hypothesis that equol inhibits antral follicle growth, increases follicle atresia, and inhibits steroidogenesis in the adult mouse ovary. To test this hypothesis, antral follicles isolated from adult CD-1 mice were cultured with vehicle control (dimethyl sulfoxide; DMSO) or equol (600 nM, 6 μM, 36 μM, 100 μM) for 48 and 96 h. Every 24 h, follicle diameters were measured to monitor growth. At 48 and 96 h, the culture medium was subjected to measurement of hormone levels, and the cultured follicles were subjected to gene expression analysis. Additionally, follicles were histologically evaluated for signs of atresia after 96 h of culture. The results indicate that equol (100 μM) inhibited follicle growth, altered the mRNA levels of bcl2-associated X protein and B cell leukemia/lymphoma 2, and induced follicle atresia. Further, equol decreased the levels of estradiol, testosterone, androstenedione, and progesterone, and it decreased mRNA levels of cholesterol side-chain cleavage, steroid 17-α-hydroxalase, and aromatase. Collectively, these data indicate that equol inhibits growth, increases atresia, and inhibits steroidogenesis of cultured mouse antral follicles. PMID:26876617

  18. Quantitative bioluminescence imaging of transgene expression in intact porcine antral follicles in vitro

    PubMed Central

    2014-01-01

    Background The porcine oocyte maturation in vivo occurs within the ovarian follicle and is regulated by the interactions between oocytes and surrounding follicular components, including theca, granulosa, and cumulus cells, and follicular fluid. Therefore, the antral follicle is an essential microenvironment for efficient oocyte maturation and its developmental competence. Quantitative bioluminescence imaging of firefly luciferase reporter genes in an intact antral follicle would allow investigation of changes in cellular and molecular events and in the context of the whole follicles. In this study, we investigate factors influencing bioluminescence measurements as a first step towards developing a new bioluminescence imaging system for intact antral follicles. Methods We analyzed the time course of bioluminescence emitted from transfected living intact follicles using a cationic lipid mediated gene transfer method with increasing doses (1-3 μg) of firefly luciferase reporter gene (pGL4). In addition, a standard luciferase assay was used to confirm the luciferase expression in granulosa cells in the transfected intact antral follicles. Finally, the dose effects of substrate, D-luciferin, were determined for optimal quantitative bioluminescence imaging of intact porcine antral follicles in vitro. Results The level of luciferase activity of follicles with 3 μg pGL4 was significantly (P < 0.05) greater than the 1 μg and 2 μg groups at 1 min after D-luciferin injection. The bioluminescence intensity of transfected follicles reached a peak at 1 min, and then it was significantly (P < 0.05) reduced within 2 min after injection of D-luciferin; with the level of bioluminescence emission remained constant from 2.5 to 10 min. The bioluminescence emission was maximal with 300 μg of D-luciferin. Conclusions The results of this study suggested that the investigation of factors influencing bioluminescence measurements is a critical step toward developing a

  19. Quantitative bioluminescence imaging of transgene expression in intact porcine antral follicles in vitro.

    PubMed

    Jung, Song-yi; Willard, Scott T

    2014-01-30

    The porcine oocyte maturation in vivo occurs within the ovarian follicle and is regulated by the interactions between oocytes and surrounding follicular components, including theca, granulosa, and cumulus cells, and follicular fluid. Therefore, the antral follicle is an essential microenvironment for efficient oocyte maturation and its developmental competence. Quantitative bioluminescence imaging of firefly luciferase reporter genes in an intact antral follicle would allow investigation of changes in cellular and molecular events and in the context of the whole follicles. In this study, we investigate factors influencing bioluminescence measurements as a first step towards developing a new bioluminescence imaging system for intact antral follicles. We analyzed the time course of bioluminescence emitted from transfected living intact follicles using a cationic lipid mediated gene transfer method with increasing doses (1-3 μg) of firefly luciferase reporter gene (pGL4). In addition, a standard luciferase assay was used to confirm the luciferase expression in granulosa cells in the transfected intact antral follicles. Finally, the dose effects of substrate, D-luciferin, were determined for optimal quantitative bioluminescence imaging of intact porcine antral follicles in vitro. The level of luciferase activity of follicles with 3 μg pGL4 was significantly (P < 0.05) greater than the 1 μg and 2 μg groups at 1 min after D-luciferin injection. The bioluminescence intensity of transfected follicles reached a peak at 1 min, and then it was significantly (P < 0.05) reduced within 2 min after injection of D-luciferin; with the level of bioluminescence emission remained constant from 2.5 to 10 min. The bioluminescence emission was maximal with 300 μg of D-luciferin. The results of this study suggested that the investigation of factors influencing bioluminescence measurements is a critical step toward developing a new bioluminescence imaging model. This

  20. Global changes in uterine protein secretion are associated with differences in the number of antral follicles in heifers

    USDA-ARS?s Scientific Manuscript database

    The numbers of antral follicles on the ovary have been reported to be associated with differences in fertility in beef cattle. To further explore potential mechanisms for these differences, an experiment was undertaken to determine the association between numbers of antral follicles and differences ...

  1. Oocyte ultrastructural characteristics in camel (Camelus dromedarius) primordial to large antral follicles.

    PubMed

    Davoodian, N; Mesbah, F; Kafi, M

    2011-04-01

    This study was conducted to describe in detail the ultrastructural features and morphological characteristics of camel oocytes from preantral follicles in relation to the sequential stages of follicular development and also for oocytes from antral follicles in relation to their diameter. Camel oocytes from primordial, primary, secondary and also early to late antral follicles were processed and examined by light and transmission electron microscopy. Primordial follicular oocytes were characterized by a layer of flattened granulosa cells around and also eccentric nucleus and few cytoplasmic organelles in the peripheral region. Up to the secondary follicle stage, flat cells were replaced by cuboidal granulosa cells and their number increased and also an increase in the number of organelles such as vesicles, mitochondria and endoplasmic reticulum was observed. In the early antral stage, the formation of zona pellucida, appearance of microvilli and pleomorphic mitochondria was seen and the nucleus was dislocated to the peripheral region. During final growth phase, the extent of endoplasmic reticulum, vesicles and mitochondria increased, the number of lipid droplets decreased and cumulus cell process endings (CCPE) were observed. In conclusion, the growth of camel oocyte is associated with progressive increase in the number of mitochondria, endoplasmic reticulum, Golgi complexes and cytoplasmic vesicles as well as decrease in the number of lipid droplets and the nucleus migration from an eccentric in preantral to a peripheral location in antral follicles. © 2010 Blackwell Verlag GmbH.

  2. Methoxychlor inhibits growth of antral follicles by altering cell cycle regulators

    SciTech Connect

    Gupta, Rupesh K. Meachum, Sharon Hernandez-Ochoa, Isabel Peretz, Jackye Yao, Humphrey H. Flaws, Jodi A.

    2009-10-01

    Methoxychlor (MXC) reduces fertility in female rodents, decreases antral follicle numbers, and increases atresia through oxidative stress pathways. MXC also inhibits antral follicle growth in vitro. The mechanism by which MXC inhibits growth of follicles is unknown. The growth of follicles is controlled, in part, by cell cycle regulators. Thus, we tested the hypothesis that MXC inhibits follicle growth by reducing the levels of selected cell cycle regulators. Further, we tested whether co-treatment with an antioxidant, N-acetyl cysteine (NAC), prevents the MXC-induced reduction in cell cycle regulators. For in vivo studies, adult cycling CD-1 mice were dosed with MXC or vehicle for 20 days. Treated ovaries were subjected to immunohistochemistry for proliferating cell nuclear antigen (PCNA) staining. For in vitro studies, antral follicles isolated from adult cycling CD-1 mouse ovaries were cultured with vehicle, MXC, and/or NAC for 48, 72 and 96 h. Levels of cyclin D2 (Ccnd2) and cyclin dependent kinase 4 (Cdk4) were measured using in vivo and in vitro samples. The results indicate that MXC decreased PCNA staining, and Ccnd2 and Cdk4 levels compared to controls. NAC co-treatment restored follicle growth and expression of Ccnd2 and Cdk4. Collectively, these data indicate that MXC exposure reduces the levels of Ccnd2 and Cdk4 in follicles, and that protection from oxidative stress restores Ccnd2 and Cdk4 levels. Therefore, MXC-induced oxidative stress may decrease the levels of cell cycle regulators, which in turn, results in inhibition of the growth of antral follicles.

  3. Di (2-ethylhexyl) phthalate inhibits growth of mouse ovarian antral follicles through an oxidative stress pathway

    SciTech Connect

    Wang, Wei Craig, Zelieann R. Basavarajappa, Mallikarjuna S. Gupta, Rupesh K. Flaws, Jodi A.

    2012-01-15

    Di (2-ethylhexyl) phthalate (DEHP) is a plasticizer that has been shown to inhibit growth of mouse antral follicles, however, little is known about the mechanisms by which DEHP does so. Oxidative stress has been linked to follicle growth inhibition as well as phthalate-induced toxicity in non-ovarian tissues. Thus, we hypothesized that DEHP causes oxidative stress and that this leads to inhibition of the growth of antral follicles. To test this hypothesis, antral follicles isolated from CD-1 mice (age 31–35 days) were cultured with vehicle control (dimethylsulfoxide [DMSO]) or DEHP (1–100 μg/ml) ± N-acetyl cysteine (NAC, an antioxidant at 0.25–1 mM). During culture, follicles were measured daily. At the end of culture, follicles were collected and processed for in vitro reactive oxygen species (ROS) assays to measure the presence of free radicals or for measurement of the expression and activity of various key antioxidant enzymes: Cu/Zn superoxide dismutase (SOD1), glutathione peroxidase (GPX) and catalase (CAT). The results indicate that DEHP inhibits the growth of follicles compared to DMSO control and that NAC (0.25–1 mM) blocks the ability of DEHP to inhibit follicle growth. Furthermore, DEHP (10 μg/ml) significantly increases ROS levels and reduces the expression and activity of SOD1 compared to DMSO controls, whereas NAC (0.5 mM) rescues the effects of DEHP on ROS levels and SOD1. However, the expression and activity of GPX and CAT were not affected by DEHP treatment. Collectively, these data suggest that DEHP inhibits follicle growth by inducing production of ROS and by decreasing the expression and activity of SOD1. -- Highlights: ► DEHP inhibits growth and increases reactive oxygen species in ovarian antral follicles in vitro. ► NAC rescues the effects of DEHP on the growth and reactive oxygen species levels in follicles. ► DEHP decreases the expression and activity of Cu/Zn superoxide dismutase, which can be rescued by NAC, in antral

  4. Effects of an Environmentally Relevant Phthalate Mixture on Cultured Mouse Antral Follicles.

    PubMed

    Zhou, Changqing; Flaws, Jodi A

    2017-03-01

    Phthalates are used in building materials, medical devices, and personal care products. Most studies on phthalates have focused on single phthalates, but it is important to study mixtures of phthalates because humans are exposed to such mixtures daily. We tested the hypothesis that phthalate mixture exposure decreases antral follicle growth, compromises steroidogenic capacity, and induces atresia. Antral follicles from adult CD-1 mice were cultured with vehicle control or phthalate mixture (1-500 µg/ml) for 96 h. The mixture was made of 35% diethyl phthalate, 21% di(2-ethylhexyl) phthalate, 15% dibutyl phthalate, 15% diisononyl phthalate, 8% diisobutyl phthalate, and 5% benzylbutyl phthalate. During culture, antral follicle diameters were measured every 24 h to monitor growth. After culture, media were subjected to measurements of sex steroid hormones and follicles were subjected to evaluation of gene expression and atresia. The phthalate mixture (100 and 500 µg/ml) decreased antral follicle growth starting at 24 h compared to controls. The mixture at 10, 100, and 500 µg/ml also decreased androstenedione, testosterone, estrone, and estradiol levels compared to control. The mixture (10, 100, and 500 µg/ml) reduced atresia rating, but it induced more oocyte fragmentation compared to control. The phthalate mixture at different doses adversely affected cell cycle regulators, antioxidant enzymes, apoptotic factors, steroidogenic enzymes, and receptors. Collectively, these data indicate that exposure to an environmentally relevant phthalate mixture reduces antral follicle growth, induces oocyte fragmentation, and decreases hormone production by adversely affecting the expression of cell cycle regulators, apoptotic factors, steroidogenic enzymes, and receptors. © The Author 2016. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  5. Di (2-ethylhexyl) phthalate inhibits growth of ovarian antral follicles through an oxidative stress pathway

    PubMed Central

    Wang, Wei; Craig, Zelieann R.; Basavarajappa, Mallikarjuna S.; Gupta, Rupesh; Flaws, Jodi A.

    2011-01-01

    Di (2-ethylhexyl) phthalate (DEHP) is a plasticizer that has been shown to inhibit growth of mouse antral follicles, however, little is known about the mechanisms by which DEHP does so. Oxidative stress has been linked to follicle growth inhibition as well as phthalate-induced toxicity in non-ovarian tissues. Thus, we hypothesized that DEHP causes oxidative stress and that this leads to inhibition of the growth of antral follicles. To test this hypothesis, antral follicles isolated from CD-1 mice (age 32–35 days) were cultured with vehicle control (dimethylsulfoxide [DMSO]) or DEHP (1–100μg/ml) ± N-acetyl cysteine (NAC, an antioxidant at 0.25–1mM). During culture, follicles were measured daily. At the end of culture, follicles were collected and processed for in vitro reactive oxygen species (ROS) assays to measure the presence of free radicals or for measurement of the expression and activity of various key antioxidant enzymes: Cu/Zn superoxide dismutase (SOD1), glutathione peroxidase (GPX) and catalase (CAT). The results indicate that DEHP inhibits the growth of follicles compared to DMSO control and that NAC (0.25–1mM) blocks the ability of DEHP to inhibit follicle growth. Furthermore, DEHP (10μg/ml) significantly increases ROS levels and reduces the expression and activity of SOD1 compared to DMSO controls, whereas NAC (0.5mM) rescues the effects of DEHP on ROS levels and SOD1. However, the expression and activity of GPX and CAT were not affected by DEHP treatment. Collectively, these data suggest that DEHP inhibits follicle growth by inducing production of ROS and by decreasing the expression and activity of SOD1. PMID:22155089

  6. Antral follicle populations and embryo production--in vitro and in vivo--of Bos indicus-taurus donors from weaning to yearling ages.

    PubMed

    Silva-Santos, K C; Santos, G M G; Koetz Júnior, C; Morotti, F; Siloto, L S; Marcantonio, T N; Urbano, M R; Oliveira, R L; Lima, D C M; Seneda, M M

    2014-04-01

    Interest in indicus-taurus cattle has been increasing, as these animals are likely to present the best characteristics of Zebu and European bovine breeds. The aim of this study was to compare the embryo production of indicus-taurus donors with high vs low antral follicle counts obtained by ovum pickup/in vitro production (OPU/IVP) and superovulation (SOV)/embryo collection. Braford females at weaning age (3/8 Nelore × 5/8 Hereford, n = 137, 9 ± 1 month old) were subjected to six serial ovarian ultrasonographs and were assigned to two groups according to the number of antral follicles ≥ 3 mm as follows: G-High antral follicular count (AFC, n = 20, mean ≥ 40 follicles) and G-Low AFC (n = 20, mean ≤ 10 follicles). When the females (n = 40) reached 24 months of age, they were subjected to both OPU/IVP and SOV/embryo collection. The average number of follicles remained highly stable throughout all of the ultrasound evaluations (range 0.90-0.92). The mean number of COCs recovered (36.90 ± 13.68 vs 5.80 ± 3.40) was higher (p < 0.05) for females with high AFC, resulting in higher (p < 0.05) numbers of total embryos among females with high vs low AFC (6.10 ± 4.51 vs 0.55 ± 0.83). The mean number of embryos per collection was also higher (p < 0.05) for G-High vs G-Low (6.95 ± 5.34 vs 1.9 ± 2.13). We conclude that a single ultrasound performed at pre-pubertal ages to count antral follicles can be used as a predictor of embryo production following IVP and SOV/embryo collection in indicus-taurus females. © 2014 Blackwell Verlag GmbH.

  7. Methoxychlor reduces estradiol levels by altering steroidogenesis and metabolism in mouse antral follicles in vitro.

    PubMed

    Basavarajappa, Mallikarjuna S; Craig, Zelieann R; Hernández-Ochoa, Isabel; Paulose, Tessie; Leslie, Traci C; Flaws, Jodi A

    2011-06-15

    The organochlorine pesticide methoxychlor (MXC) is a known endocrine disruptor that affects adult rodent females by causing reduced fertility, persistent estrus, and ovarian atrophy. Since MXC is also known to target antral follicles, the major producer of sex steroids in the ovary, the present study was designed to test the hypothesis that MXC decreases estradiol (E₂) levels by altering steroidogenic and metabolic enzymes in the antral follicles. To test this hypothesis, antral follicles were isolated from CD-1 mouse ovaries and cultured with either dimethylsulfoxide (DMSO) or MXC. Follicle growth was measured every 24 h for 96 h. In addition, sex steroid hormone levels were measured using enzyme-linked immunosorbent assays (ELISA) and mRNA expression levels of steroidogenic enzymes as well as the E₂ metabolic enzyme Cyp1b1 were measured using qPCR. The results indicate that MXC decreased E₂, testosterone, androstenedione, and progesterone (P₄) levels compared to DMSO. In addition, MXC decreased expression of aromatase (Cyp19a1), 17β-hydroxysteroid dehydrogenase 1 (Hsd17b1), 17α-hydroxylase/17,20-lyase (Cyp17a1), 3β hydroxysteroid dehydrogenase 1 (Hsd3b1), cholesterol side-chain cleavage (Cyp11a1), steroid acute regulatory protein (Star), and increased expression of Cyp1b1 enzyme levels. Thus, these data suggest that MXC decreases steroidogenic enzyme levels, increases metabolic enzyme expression and this in turn leads to decreased sex steroid hormone levels.

  8. Di (2-ethylhexyl) Phthalate Exposure Impairs Growth of Antral Follicle in Mice

    PubMed Central

    Li, Lan; Liu, Jing-Cai; Lai, Fang-Nong; Liu, Huan-Qi; Zhang, Xi-Feng; Dyce, Paul W.; Shen, Wei; Chen, Hong

    2016-01-01

    Di (2-ethylhexyl) phthalate (DEHP) is a widely used plastic additive. As an environmental endocrine disruptor, it has been shown to be harmful to the mammalian reproductive system. Previous studies indicated that DEHP inhibited the development of mouse ovarian follicles. However, the mechanisms by which DEHP affects ovarian antral follicle development during the pre-puberty stage are poorly understand. Thus, we investigated the effects of direct DEHP exposure on antral follicle growth in pre-pubescent mice by use of intraperitoneal injection. Our results demonstrated that the percentage of large antral follicles was significantly reduced when mice were exposed to 20 or 40 μg/kg DEHP every 5 days from postnatal day 0 (0 dpp) to 15 dpp. In 20 dpp, we performed microarray of these ovaries. The microarray results indicated that mRNA levels of apoptosis related genes were increased. The mRNA levels of the apoptosis and cell proliferation (negative) related genes Apoe, Agt, Glo1 and Grina were increased after DEHP exposure. DEHP induced the differential gene expression of Hsp90ab1, Rhoa, Grina and Xdh which may play an important role in this process. In addition, TUNEL staining and immunofluorescence showed that DEHP exposure significantly increased the number of TUNEL, Caspase3 and γH2AX positive ovarian somatic cells within the mouse ovaries. Flow cytometer analyses of redox-sensitive probes showed that DEHP caused the accumulation of reactive oxygen species. Moreover, the mRNA expression of ovarian somatic cell antioxidative enzymes was down-regulated both in vivo and in vitro. In conclusion, our data here demonstrated that DEHP exposure induced oxidative stress and ovarian somatic cell apoptosis, and thus may impact antral follicle enlargement during the pre-pubertal stage in mice. PMID:26845775

  9. Methoxychlor reduces estradiol levels by altering steroidogenesis and metabolism in mouse antral follicles in vitro

    SciTech Connect

    Basavarajappa, Mallikarjuna S. Craig, Zelieann R. Hernandez-Ochoa, Isabel Paulose, Tessie Leslie, Traci C. Flaws, Jodi A.

    2011-06-15

    The organochlorine pesticide methoxychlor (MXC) is a known endocrine disruptor that affects adult rodent females by causing reduced fertility, persistent estrus, and ovarian atrophy. Since MXC is also known to target antral follicles, the major producer of sex steroids in the ovary, the present study was designed to test the hypothesis that MXC decreases estradiol (E{sub 2}) levels by altering steroidogenic and metabolic enzymes in the antral follicles. To test this hypothesis, antral follicles were isolated from CD-1 mouse ovaries and cultured with either dimethylsulfoxide (DMSO) or MXC. Follicle growth was measured every 24 h for 96 h. In addition, sex steroid hormone levels were measured using enzyme-linked immunosorbent assays (ELISA) and mRNA expression levels of steroidogenic enzymes as well as the E{sub 2} metabolic enzyme Cyp1b1 were measured using qPCR. The results indicate that MXC decreased E{sub 2}, testosterone, androstenedione, and progesterone (P{sub 4}) levels compared to DMSO. In addition, MXC decreased expression of aromatase (Cyp19a1), 17{beta}-hydroxysteroid dehydrogenase 1 (Hsd17b1), 17{alpha}-hydroxylase/17,20-lyase (Cyp17a1), 3{beta} hydroxysteroid dehydrogenase 1 (Hsd3b1), cholesterol side-chain cleavage (Cyp11a1), steroid acute regulatory protein (Star), and increased expression of Cyp1b1 enzyme levels. Thus, these data suggest that MXC decreases steroidogenic enzyme levels, increases metabolic enzyme expression and this in turn leads to decreased sex steroid hormone levels. - Highlights: > MXC inhibits steroidogenesis > MXC inhibits steroidogenic enzymes > MXC induces metabolic enzymes

  10. Scrambled and fried: Cigarette smoke exposure causes antral follicle destruction and oocyte dysfunction through oxidative stress

    SciTech Connect

    Sobinoff, A.P.; Beckett, E.L.; Jarnicki, A.G.; Sutherland, J.M.; McCluskey, A.; Hansbro, P.M.; McLaughlin, E.A.

    2013-09-01

    Cigarette smoke is a reproductive hazard associated with pre-mature reproductive senescence and reduced clinical pregnancy rates in female smokers. Despite an increased awareness of the adverse effects of cigarette smoke exposure on systemic health, many women remain unaware of the adverse effects of cigarette smoke on female fertility. This issue is compounded by our limited understanding of the molecular mechanisms behind cigarette smoke induced infertility. In this study we used a direct nasal exposure mouse model of cigarette smoke-induced chronic obstructive pulmonary disease to characterise mechanisms of cigarette-smoke induced ovotoxicity. Cigarette smoke exposure caused increased levels of primordial follicle depletion, antral follicle oocyte apoptosis and oxidative stress in exposed ovaries, resulting in fewer follicles available for ovulation. Evidence of oxidative stress also persisted in ovulated oocytes which escaped destruction, with increased levels of mitochondrial ROS and lipid peroxidation resulting in reduced fertilisation potential. Microarray analysis of ovarian tissue correlated these insults with a complex mechanism of ovotoxicity involving genes associated with detoxification, inflammation, follicular activation, immune cell mediated apoptosis and membrane organisation. In particular, the phase I detoxifying enzyme cyp2e1 was found to be significantly up-regulated in developing oocytes; an enzyme known to cause molecular bioactivation resulting in oxidative stress. Our results provide a preliminary model of cigarette smoke induced sub-fertility through cyp2e1 bioactivation and oxidative stress, resulting in developing follicle depletion and oocyte dysfunction. - Highlights: • Cigarette smoke exposure targets developing follicle oocytes. • The antral follicle oocyte is a primary site of ovarian cigarette smoke metabolism. • Cyp2e1 is a major enzyme involved in ameliorating smoke-induced ovotoxicity. • Cigarette smoke causes oocyte

  11. Di-(2-ethylhexyl) phthalate and mono-(2-ethylhexyl) phthalate inhibit growth and reduce estradiol levels of antral follicles in vitro

    SciTech Connect

    Gupta, Rupesh K.; Singh, Jeffery M.; Leslie, Tracie C.; Meachum, Sharon; Flaws, Jodi A.; Yao, Humphrey H-C

    2010-01-15

    Any insult that affects survival of ovarian antral follicles can cause abnormal estradiol production and fertility problems. Phthalate esters (PEs) are plasticizers used in a wide range of consumer and industrial products. Exposure to these chemicals has been linked to reduced fertility in humans and animal models. Di-(2-ethylhexyl) phthalate (DEHP) and mono-(2-ethylhexyl) phthalate (MEHP) decrease serum estradiol levels and aromatase (Arom) expression, prolong estrous cycles, and cause anovulation in animal and culture models. These observations suggest PEs directly target antral follicles. We therefore tested the hypothesis that DEHP (1-100 mug/ml) and MEHP (0.1-10 mug/ml) directly inhibit antral follicular growth and estradiol production. Antral follicles from adult mice were cultured with DEHP or MEHP, and/or estradiol for 96 h. During culture, follicle size was measured every 24 h as a measurement of follicle growth. After culture, media were collected for measurement of estradiol levels and follicles were subjected to measurement of cylin-D-2 (Ccnd2), cyclin-dependant-kinase-4 (Cdk4), and Arom. We found that DEHP and MEHP inhibited growth of follicles and decreased estradiol production compared to controls at the highest doses. DEHP and MEHP also decreased mRNA expression of Ccnd2, Cdk4, and Arom at the highest dose. Addition of estradiol to the culture medium prevented the follicles from DEHP- and MEHP-induced inhibition of growth, reduction in estradiol levels, and decreased Ccnd2 and Cdk4 expression. Collectively, our results indicate that DEHP and MEHP may directly inhibit antral follicle growth via a mechanism that partially includes reduction in levels of estradiol production and decreased expression of cell cycle regulators.

  12. Scrambled and fried: cigarette smoke exposure causes antral follicle destruction and oocyte dysfunction through oxidative stress.

    PubMed

    Sobinoff, A P; Beckett, E L; Jarnicki, A G; Sutherland, J M; McCluskey, A; Hansbro, P M; McLaughlin, E A

    2013-09-01

    Cigarette smoke is a reproductive hazard associated with pre-mature reproductive senescence and reduced clinical pregnancy rates in female smokers. Despite an increased awareness of the adverse effects of cigarette smoke exposure on systemic health, many women remain unaware of the adverse effects of cigarette smoke on female fertility. This issue is compounded by our limited understanding of the molecular mechanisms behind cigarette smoke induced infertility. In this study we used a direct nasal exposure mouse model of cigarette smoke-induced chronic obstructive pulmonary disease to characterise mechanisms of cigarette-smoke induced ovotoxicity. Cigarette smoke exposure caused increased levels of primordial follicle depletion, antral follicle oocyte apoptosis and oxidative stress in exposed ovaries, resulting in fewer follicles available for ovulation. Evidence of oxidative stress also persisted in ovulated oocytes which escaped destruction, with increased levels of mitochondrial ROS and lipid peroxidation resulting in reduced fertilisation potential. Microarray analysis of ovarian tissue correlated these insults with a complex mechanism of ovotoxicity involving genes associated with detoxification, inflammation, follicular activation, immune cell mediated apoptosis and membrane organisation. In particular, the phase I detoxifying enzyme cyp2e1 was found to be significantly up-regulated in developing oocytes; an enzyme known to cause molecular bioactivation resulting in oxidative stress. Our results provide a preliminary model of cigarette smoke induced sub-fertility through cyp2e1 bioactivation and oxidative stress, resulting in developing follicle depletion and oocyte dysfunction. Copyright © 2013 Elsevier Inc. All rights reserved.

  13. Oocyte maturation and expression pattern of follicular genes during in-vitro culture of vitrified mouse pre-antral follicles.

    PubMed

    Jamalzaei, Parisa; Valojerdi, Mojtaba Rezazadeh; Ebrahimi, Bita; Farrokhi, Ali

    2016-01-01

    Our aim was to evaluate the oocyte maturation rate and follicular genes expression pattern during in-vitro culture of vitrified mouse pre-antral follicles. Middle sized pre-antral follicles were isolated mechanically from the ovaries of pre-pubertal mice and distributed in vitrification and control groups. In the vitrification group, follicles were washed in equilibration and vitrification solutions and then were immersed in liquid nitrogen after loading on cryotop tips. After warming in descending concentrations of sucrose solutions, fresh and vitrified-warmed follicles were cultured for 13 days. Follicles survival rate and follicular genes expression were assessed during in vitro culture. Finally, at the end of the culture period oocytes maturation rate were compared in both groups. In the vitrification group, follicles survival rate was lower significantly comparing to the control group (P < 0.05), whereas oocytes maturation rate were similar. Although at the beginning of the culture period, expression of some genes such as Gdf9, Bmp15, Tgfβ1 and BmprII were higher in the vitrification group (P < 0.05), during the rest of the culture period expression pattern of all follicular genes were similar in both groups. In conclusion, survival rate of cryotop vitrified pre-antral follicles reduced during culture period while oocytes maturation and follicular genes expression did not show any noticeable alteration.

  14. Total Oxidative status of Mouse Vitrified Pre-Antral Follicles with Pre-Treatment of Alpha Lipoic Acid

    PubMed Central

    Hatami, Sahar; Zavareh, Saeed; Salehnia, Mojdeh; Lashkarbolouki, Taghi; Ghorbanian, Mohammad Taghi; Karimi, Isaac

    2014-01-01

    Background: Cryopreservation of pre-antral follicles is a hopeful technique to preserve female fertility. The aim of the present study was to evaluate reactive oxygen species (ROS) and total antioxidant capacity (TAC) levels of mouse vitrified pre-antral follicles in the presence of alpha lipoic acid (ALA). Methods: Isolated pre-antral follicles (140–150 µm in diameter) were divided into vitrified–warmed and fresh groups. Each group was subjected to in vitro maturation with or without ALA for 12 days, followed by adding human chronic gonadotropin to induce ovulation. In vitro fertilization was performed to evaluate their developmental competence. In parallel, the amount of ROS and TAC were assessed after 0, 24, 48, 72, and 96 h of culture by 2',7'-dichlorofluorescin assay and ferric reducing/antioxidant power assay, respectively. Results: The respective rates of survival, antrum formation, and metaphase II oocytes were significantly higher in ALA-supplemented groups compared to the groups not treated with ALA. TAC and ROS levels were significantly decreased and increased, respectively during the culture period up to 96 h in the absence of ALA in both vitrified and non-vitrified samples. However, with pretreatment of ALA, TAC levels were increased significantly and remained constant up to 96 h in vitrified-warmed pre-antral follicles, while ROS levels completely returned to the level of starting point after 96 h of culture in the presence of ALA. Conclusion: Pretreatment of ALA positively influences development of pre-antral follicles in vitrified and non-vitrified samples through increasing follicular TAC level and decreasing ROS levels. PMID:24842145

  15. Ovarian Primordial and Non-Growing Follicle Counts According to the Stages of Reproductive Aging Workshop (STRAW) Staging System

    PubMed Central

    Hansen, Karl R.; Craig, LaTasha B.; Zavy, Michael T.; Klein, Nancy A.; Soules, Michael R.

    2011-01-01

    Objective To characterize the ovarian primordial and non-growing follicle number according to the Stages of Reproductive Aging Workshop (STRAW) staging system as defined by menstrual cycle characteristics. Methods Normal ovaries were collected from 63 women (age 26-52 years) undergoing oophorectomy for benign indications. Prior to surgery, each participant completed a detailed questionnaire collecting information regarding menstrual cycle characteristics and were classified by bleeding patterns into STRAW stages -4, -3, -2, and -1. A single ovary was selected for determination of the ovarian primordial and total non-growing follicle number utilizing a validated fractionator/optical disector method. A subset of the participants (n = 43) underwent transvaginal ultrasound examination for the determination of the ovarian antral follicle count and serum measurements of FSH, estradiol, anti-müllerian hormone and inhibin B. All measurements were obtained within two weeks of surgery, irrespective of cycle day. Results Significant differences were identified in ovarian primordial (p <0.0001) and non-growing follicle (p <0.0001) counts across the STRAW stages. In post-hoc testing, the differences in primordial follicle counts were significant between each of the STRAW stages. Significant differences were also identified in serum levels of anti-müllerian hormone, FSH and the ovarian antral follicle count across the STRAW stages. Conclusions Progression through the STRAW stages as defined by menstrual cycle characteristics is associated with progressive and significant decreases in the ovarian primordial follicle number. PMID:22189385

  16. Involvement of miRNAs and Cell-Secreted Vesicles in Mammalian Ovarian Antral Follicle Development.

    PubMed

    da Silveira, Juliano C; de Andrade, Gabriella M; Nogueira, Marcelo F G; Meirelles, Flávio V; Perecin, Felipe

    2015-12-01

    Ovarian follicular development is a controlled series of events culminating with an ovulatory or atretic follicle. MicroRNAs (miRNAs) are small noncoding RNAs involved in translational regulation of genes in different developmental processes. Deletion of Dicer in mice ovaries demonstrated the importance of miRNAs in reproduction, which led to infertility. The miRNAs were thought to act only within host cells; however, these molecules are also present in cell-secreted vesicles. These vesicles are present in body fluids such as milk, serum, and ovarian follicular fluid. Vesicles are secreted in extracellular fluids and travel from donor to target cells, mediating transfer of bioactive material. Herein we discuss the role of hormonal-regulated miRNAs within different ovarian follicular cells as well as cell-secreted vesicles participation in mammalian ovarian follicular fluid. Furthermore, we discuss the possibility of miRNAs transference mediated by cell-secreted vesicles present in ovarian follicular fluid, increasing the versatility of miRNA functions during antral follicle development. © The Author(s) 2015.

  17. Co-culture of buffalo (Bubalus bubalis) preantral follicles with antral follicles: a comparative study of developmental competence of oocytes derived from in vivo developed and in vitro cultured antral follicles.

    PubMed

    Sharma, G Taru; Dubey, Pawan K; Nath, Amar; Saikumar, G

    2013-08-01

    The present study was undertaken to examine whether the presence of antral follicles (AFs) affects the survival, growth and steroidogenesis of preantral follicles (PFs) and compare the maturation and developmental competence of buffalo oocytes derived from in vivo developed and in vitro cultured AFs. Two experiments were carried out. In experiment I, PFs (200-250 μm) were isolated and cultured with or without AFs (3-5 mm) in TCM-199 medium that contained 10% fetal bovine serum (FBS), 1% insulin transferin selenium (ITS), 20 ng/ml epidermal growth factor (EGF), 0.5 μg/ml follicle-stimulating hormone (FSH) and 100 ng/ml insulin-like growth factor (IGF)-I. In experiment II, in vitro developmental competence was compared for the cumulus-oocyte complexes (COCs) recovered from in vivo developed and in vitro cultured AFs. Survival, growth, development of antrum, accumulation of estradiol and progesterone was (P < 0.05) higher when PFs were co-cultured with AFs. Developmental competence of both types of follicular oocytes did not differ significantly in terms of maturation and cleavage rate, but morula and blastocyst production rate were (P < 0.05) higher with in vivo developed AFs as compared with the in vitro cultured antral follicular oocytes. In conclusion, co-culture of PFs with AFs supports long-term survival and growth of buffalo PFs and this co-culture system plays a dual role for in vitro production of embryos as well as understanding the relationship between developing PFs and AFs.

  18. Effects of cyclic increase in gonadotropins on the in vitro development of primordial follicles to antral stage.

    PubMed

    Fabbri, Raffaella; Pasquinelli, Gianandrea; Parazza, Isabella; Macciocca, Maria; Magnani, Valentina; Battaglia, Cesare; Paradisi, Roberto; Venturoli, Stefano

    2012-10-01

    The aim of this study was to evaluate the effects of FSH and LH on follicle development during a long-term culture of cryopreserved human ovarian tissue, using morphological and ultrastructural examinations. Thawed ovarian tissue slices from a 4-year-old child with Wilms tumor were cultured for 32 weeks in two different culture conditions, without (medium A) and with (medium B) a monthly peaked increase in FSH and LH. At week 32, in the medium B cultured tissue, a cluster of preantral follicles associated with two oocytes prematurely ovulated was observed, suggesting that the cyclic increase of gonadotropins promoted thawed follicles to grow up to the antral stage. However, the integrity and coordinated follicle development were not maintained. Indeed, ultrastructural analysis showed a well-preserved "naked" oocyte with concomitant features of immaturity and maturity, as if this culture condition had led to an asynchronous maturation of oocyte cytoplasmic components.

  19. High numbers of antral follicles are positively associated with in vitro embryo production but not the conception rate for FTAI in Nelore cattle.

    PubMed

    Santos, Gustavo Martins Gomes Dos; Silva-Santos, Katia Cristina; Barreiros, Thales Ricardo Rigo; Morotti, Fábio; Sanches, Bruno Valente; de Moraes, Fábio Lucas Zito; Blaschi, Wanessa; Seneda, Marcelo Marcondes

    2016-02-01

    The objective was to compare the conception rates for FTAI and in vitro embryo production between Nelore cows with different antral follicle counts (AFC=number of follicles ≤3mm in diameter in the ovaries). Nelore cows (n=547) were subjected to ovulation synchronization. Randomly during the estrous cycle (D0), cows received an intravaginal device containing 1.9g P4 (CIDR(®)) and 2mg BE (Estrogin(®)), IM. When the device was removed (D8), the cows received 500μg PGF2α (Ciosin(®)), 300IU eCG (Novormon(®)) and 1mg EC (ECP(®)), IM. All cows were inseminated 48h after P4 device removal. Antral follicles ≥3mm were counted using an intravaginal microconvex transducer (D0), and the cows were assigned to high (G-High, ≥25 follicles, n=183), intermediate (G-Intermediate, 16-20 follicles, n=183) or low AFC groups (G-Low, ≤10 follicles, n=181). In another experiment, COCs were retrieved by OPU from Nelore cows (n=66), which were assigned to groups according to oocyte production: G-High (n=22, ≥40 oocytes), G-Intermediate (n=25, 18-25 oocytes) or G-Low (n=19, ≤7 oocytes). All COCs from the same cow were cultured individually (maximum of 25 COCs per drop) and then in vitro fertilized using thawed frozen sperm (2×10(8)/dose) from a Nelore sire of known fertility. The data were analyzed using a Kruskal-Wallis and a Chi-square test (P≤0.05). There was no difference in the conception rates after FTAI between Nelore cows with high, intermediate or low AFC (51.9 vs. 48.6 vs. 58.6%). The number of viable embryos was 18.4±6.7 (G-High), 6.1±3.6 (G-Intermediate) and 0.6±0.7 (G-Low; P<0.05). Therefore, AFC had no influence on the conception rates for FTAI; however, Nelore cows with high oocyte production exhibited better in vitro embryo production. Copyright © 2015 Elsevier B.V. All rights reserved.

  20. Regulation of Pcsk6 expression during the preantral to antral follicle transition in mice: opposing roles of FSH and oocytes.

    PubMed

    Diaz, Francisco J; Sugiura, Koji; Eppig, John J

    2008-01-01

    Several secreted products of the TGFbeta superfamily have important roles during follicular development and are produced by both oocytes and somatic cells (granulosa and theca) in the follicle. The proprotein convertases are a family of seven known proteins that process TGFbeta ligands and other secreted products to their mature active form. The present study examined the regulation of steady-state levels of Pcsk6 mRNA, which encodes a convertase protein known to process members of the TGFbeta superfamily, during mouse follicular development. Pcsk6 mRNA and protein were expressed in preantral but not cumulus or mural granulosa cells. Pcsk6 mRNA levels in preantral granulosa cells were not regulated by growing oocytes of preantral follicles, but were elevated by FSH. Furthermore, Pcsk6 mRNA in preantral granulosa cells was potently suppressed by factor(s) secreted by fully grown oocytes from antral follicles, in part through SMAD2/3-mediated pathways. Oocytes acquired the ability to suppress the steady-state levels of Pcsk6 mRNA in granulosa cells during the preantral to antral follicle transition. Suppression of Pcsk6 mRNA by oocytes could reflect a change in the mechanism(s) regulating the activity of members of the TGFbeta superfamily.

  1. Isolation and culture of ovine and bubaline small and large pre-antral follicles: effect of cyclicity and presence of a dominant follicle.

    PubMed

    Nandi, S; Girish Kumar, V; Ramesh, H S; Manjunatha, B M; Gupta, P S P

    2009-02-01

    Studies were conducted to examine the effects of the cyclicity and the presence of a dominant follicle (DF) in ovary on the recovery and in vitro growth of pre-antral follicles (PFs) in sheep and buffalo. Small pre-antral follicles (SPFs, 100-250 microm) and large pre-antral follicles (LPFs, 250-450 microm) were isolated from slaughterhouse ovaries in the breeding seasons by a mechanical and enzymatic method. The sheep and buffalo PFs were cultured in vitro for 6 and 15 days, respectively, and examined for their growth, survival and antrum formation rates and growth rates of oocytes in cultured pre-antral follicles. The follicles of the sheep and buffalo were recovered and cultured simultaneously within replicates. The recovery rates (number per ovary) of both SPFs and LPFs were significantly (p < 0.05) higher in cyclic ewes (SPFs: 22.0 +/- 3.3 vs 12.1 +/- 2.6 and LPFs: 16.0 +/- 3.6 vs 9.2 +/- 1.8) and buffaloes (SPFs: 9.2 +/- 1.3 vs 4.1 +/- 1.0 and LPFs: 10.3 +/- 2.7 vs 5.4 +/- 0.7) compared with those recovered from acyclic ones. Presence of a DF in ovary significantly (p < 0.05) reduced the recovery rates of LPFs in ewes (9.06 +/- 2.7 vs 16.4 +/- 3.8) but had no effect in buffalo. Cyclicity of animals or follicular dominance had no effects on in vitro growth, survival and antrum formation rates and growth rates of oocytes in cultured PFs of SPFs and LPFs in both sheep and buffalo. The in vitro growth, survival and antrum formation rates of LPFs and growth rates of oocytes in cultured LPFs were significantly (p < 0.05) higher than those observed in SPFs in both sheep and buffalo. The overall recovery and growth rates of the PFs were lower in buffaloes compared with ewes.

  2. A light- and electron-microscope study of the oocyte nucleus during development of the antral follicle in the prepubertal mouse.

    PubMed

    Chouinard, L A

    1975-05-01

    The ordered changes which occur in the structural organization of the mouse oocyte nucelus during the preparatory, the maturative and the preovulatory stages of antral follicle development, have been studied under both light and electron microscopy. All observations have been made on those antral follicles whose development is initiated on postnatal day 14 and completed by postnatal day 28 in prepubertal animals of the ICR albino mouse strain. The formed entities that can be recognized within the oocyte nucleus during that period are the condensing bivalents, the heterochromatic knobs, the nucleolus and the extranucleolar bodies. At the onset of antral follicle development, the highly unravelled dictyate bivalents are seen to take on a lampbrush-type configuration. Subsequent condensation of these lampbursh bivalents appears to be a very gradual and lengthy process that extends over almost the entire period of antral follicle development. The shortening and thickening of the lampbrush bivalents are best interpreted as resulting from the withdrawal of their lateral loop-like projections into the chromosome axes and from the focal aggregation of these axes into compact chromatin masses. Electron-opaque granules, which appear within the oocyte nucleus during the preparatory and maturative follicle stages, are seen to be intimately associated with these condensing bivalents. A number of Feulgen-positive heterochromatic knobs make their appearance in contact with certain bivalents during the preparatory follicle stage. These knobs are not reincorporated as such into the condensing chromatin masses and undergo disintegration and dissolution during the preovulatory follicle stage. The size, shape and ultrastructural features of the nucleolus remain unchanged thoughout the period of antral follicle development. Breakdown and dissolution of the nucleolar mass is a swift process that takes place only in the fully mature preovulatory follicle and more or less concomitantly

  3. Effect of insulin supplementation on in vitro maturation of pre-antral follicles from adult and pre-pubertal mice.

    PubMed

    Nath, Amar; Hakim, Bilal Ahmad; Rajender, Singh; Singh, Kavita; Sachdev, Monika; Konwar, Rituraj

    2016-05-01

    This study was aimed to determine the impact of insulin concentrations on in vitro pre-antral follicle growth, survival, antrum formation rate, and retrieval of mature oocytes in mice. Mice pre-antral follicle growth were recorded on days 2, 4, 6, 8, 10, and 12 in α-modified essential media (α-MEM) supplemented with insulin concentrations of 6, 8, and 10 μg/ml along with 10% FBS, 100 mIU/ml follicle stimulating hormone, 10 mIU/ml luteinizing hormone, 100 μg/ml penicillin, and 50 μg/ml streptomycin. After 12 d of growth in vitro, follicles were allowed to mature for 16-18 h in α-MEM supplemented with 1.5 IU/ml human chorionic gonadotrophin (hCG) and 5 ng/ml epidermal growth factor (EGF). The initial diameter (54.86 ± 2.5 μm) of mice oocyte progressively increased in all the three insulin concentration groups and attained a maximum size on day 12 (71.90 ± 2.8 μm). Supplementation with higher concentrations of insulin (both 8 and 10 μg/ml) significantly enhanced antrum formation without effecting the oocyte diameter and percent retrieval of mature oocyte in all the three concentration groups. Both in vitro cultured as well as in vivo collected follicles and oocytes showed similar localization and expression of oocyte maturation markers SAS1B and GDF9. Insulin concentration of 8 μg/ml was found to be optimal for in vitro follicle culture of adult mice (42-49 d). Optimized follicle culture conditions were also assessed successfully with pre-pubertal mice (12-14 d); however, adult mice showed higher follicle survival, antrum formation, and more mature oocytes production in comparison to pre-pubertal mice.

  4. Transcriptome profiling of granulosa cells of bovine ovarian follicles during growth from small to large antral sizes

    PubMed Central

    2014-01-01

    Background At later stages of folliculogenesis, the mammalian ovarian follicle contains layers of epithelial granulosa cells surrounding an antral cavity. During follicle development granulosa cells replicate, secrete hormones and support the growth of the oocyte. In cattle, the follicle needs to grow > 10 mm in diameter to allow an oocyte to ovulate, following which the granulosa cells cease dividing and differentiate into the specialised cells of the corpus luteum. To better understand the molecular basis of follicular growth and granulosa cell maturation, we undertook transcriptome profiling of granulosa cells from small (< 5 mm; n = 10) and large (> 10 mm, n = 4) healthy bovine follicles using Affymetrix microarrays (24,128 probe sets). Results Principal component analysis for the first two components and hierarchical clustering showed clustering into two groups, small and large, with the former being more heterogeneous. Size-frequency distributions of the coefficient of variation of the signal intensities of each probe set also revealed that small follicles were more heterogeneous than the large. IPA and GO enrichment analyses revealed that processes of axonal guidance, immune signalling and cell rearrangement were most affected in large follicles. The most important networks were associated with: (A) Notch, SLIT/ROBO and PI3K signalling, and (B) ITGB5 and extracellular matrix signalling through extracellular signal related kinases (ERKs). Upstream regulator genes which were predicted to be active in large follicles included STAT and XBP1. By comparison, developmental processes such as those stimulated by KIT, IHH and MEST were most active in small follicles. MGEA5 was identified as an upstream regulator in small follicles. It encodes an enzyme that modifies the activity of many target proteins, including those involved in energy sensing, by removal of N-acetylglucosamine from serine and threonine residues. Conclusions Our data suggest that as

  5. Dioxin exposure reduces the steroidogenic capacity of mouse antral follicles mainly at the level of HSD17B1 without altering atresia

    SciTech Connect

    Karman, Bethany N. Basavarajappa, Mallikarjuna S. Hannon, Patrick Flaws, Jodi A.

    2012-10-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a potent ovarian toxicant. Previously, we demonstrated that in vitro TCDD (1 nM) exposure decreases production/secretion of the sex steroid hormones progesterone (P4), androstenedione (A4), testosterone (T), and 17β-estradiol (E2) in mouse antral follicles. The purpose of this study was to determine the mechanism by which TCDD inhibits steroidogenesis. Specifically, we examined the effects of TCDD on the steroidogenic enzymes, atresia, and the aryl hydrocarbon receptor (AHR) protein. TCDD exposure for 48 h increased levels of A4, without changing HSD3B1 protein, HSD17B1 protein, estrone (E1), T or E2 levels. Further, TCDD did not alter atresia ratings compared to vehicle at 48 h. TCDD, however, did down regulate the AHR protein at 48 h. TCDD exposure for 96 h decreased transcript levels for Cyp11a1, Cyp17a1, Hsd17b1, and Cyp19a1, but increased Hsd3b1 transcript. TCDD exposure particularly lowered both Hsd17b1 transcript and HSD17B1 protein. However, TCDD exposure did not affect levels of E1 in the media nor atresia ratings at 96 h. TCDD, however, decreased levels of the proapoptotic factor Bax. Collectively, these data suggest that TCDD exposure causes a major block in the steroidogenic enzyme conversion of A4 to T and E1 to E2 and that it regulates apoptotic pathways, favoring survival over death in antral follicles. Finally, the down‐regulation of the AHR protein in TCDD exposed follicles persisted at 96 h, indicating that the activation and proteasomal degradation of this receptor likely plays a central role in the impaired steroidogenic capacity and altered apoptotic pathway of exposed antral follicles. -- Highlights: ► TCDD disrupts steroidogenic enzymes in mouse antral follicles. ► TCDD particularly affects the HSD17B1 enzyme in mouse antral follicles. ► TCDD does not affect atresia ratings in mouse antral follicles. ► TCDD decreases levels of the proapoptitic factor Bax in mouse antral follicles.

  6. Improvement of the developmental competence of porcine oocytes collected from early antral follicles by cytoplast fusion

    PubMed Central

    DANG-NGUYEN, Thanh Quang; APPELTANT, Ruth; SOMFAI, Tamas; ISHIHARA, Shinya; MEN, Nguyen Thi; SANTOS, Elisa Caroline da Silva; NOGUCHI, Junko; KANEKO, Hiroyuki; KIKUCHI, Kazuhiro

    2016-01-01

    In the present study, we propose an alternative technique called cytoplast fusion to improve the maturation rate and developmental competence of growing oocytes collected from early antral follicles in pigs. We examined whether the fusion of a growing oocyte with the cytoplast from a fully-grown oocyte (CFR group) could better promote maturation and developmental competence of the growing oocyte compared to germinal vesicle (GV) transfer (GVTR group). After 44 h of in vitro maturation (IVM), most growing oocytes (GR group) were still arrested at the GV stage (64.0 ± 5.1%); this number was significantly higher (P < 0.01) than that of the other groups. No matured oocyte was observed in the GR group. The maturation rate of GVTR oocytes was significantly improved (18.8 ± 3.5%) compared with that of growing oocytes. The proportion of oocytes that reached the metaphase-II (M-II) stage in the CFR group (37.8 ± 2.0%) was significantly higher (P < 0.05) than that in the GVTR group, although still lower than that in the control group (75.2 ± 4.4%). No blastocyst was derived from growing oocytes. Among in vitro fertilized GVTR oocytes, 3.0 ± 1.9% developed into blastocysts; however, this percentage showed an insignificant increase compared with the GR group. On the other hand, the percentage of CFR embryos that developed into blastocysts (12.0 ± 4.3%) was significantly higher than that of GR embryos (0.0%), although still lower than that of control embryos (27.0 ± 5.5%). Total cell number in blastocysts in the GVTR group (23.3 ± 6.9) was significantly lower (P < 0.05) than that in the control group (50.4 ± 5.0). Meanwhile, the total cell number in blastocysts derived from CFR oocytes (36.3 ± 4.8) was comparable to that of the control group. In summary, cytoplast fusion significantly improves maturation rate and developmental competence of growing oocytes compared with GV transfer. PMID:27795465

  7. Mono(2-Ethylhexyl) Phthalate Accelerates Early Folliculogenesis and Inhibits Steroidogenesis in Cultured Mouse Whole Ovaries and Antral Follicles1

    PubMed Central

    Hannon, Patrick R.; Brannick, Katherine E.; Wang, Wei; Flaws, Jodi A.

    2015-01-01

    Humans are ubiquitously exposed to di(2-ethylhexyl) phthalate (DEHP), which is an environmental toxicant present in common consumer products. DEHP potentially targets the ovary through its metabolite mono(2-ethylhexyl) phthalate (MEHP). However, the direct effects of MEHP on ovarian folliculogenesis and steroidogenesis, two processes essential for reproductive and nonreproductive health, are unknown. The present study tested the hypotheses that MEHP directly accelerates early folliculogenesis via overactivation of phosphatidylinositol 3-kinase (PI3K) signaling, a pathway that regulates primordial follicle quiescence and activation, and inhibits the synthesis of steroid hormones by decreasing steroidogenic enzyme levels. Neonatal ovaries from CD-1 mice were cultured for 6 days with vehicle control, DEHP, or MEHP (0.2–20 μg/ml) to assess the direct effects on folliculogenesis and PI3K signaling. Further, antral follicles from adult CD-1 mice were cultured with vehicle control or MEHP (0.1–10 μg/ml) for 24–96 h to establish the temporal effects of MEHP on steroid hormones and steroidogenic enzymes. In the neonatal ovaries, MEHP, but not DEHP, decreased phosphatase and tensin homolog levels and increased phosphorylated protein kinase B levels, leading to a decrease in the percentage of germ cells and an increase in the percentage of primary follicles. In the antral follicles, MEHP decreased the mRNA levels of 17alpha-hydroxylase-17,20-desmolase, 17beta-hydroxysteroid dehydrogenase, and aromatase leading to a decrease in testosterone, estrone, and estradiol levels. Collectively, MEHP mediates the effect of DEHP on accelerated folliculogenesis via overactivating PI3K signaling and inhibits steroidogenesis by decreasing steroidogenic enzyme levels. PMID:25810477

  8. Di-n-Butyl Phthalate Disrupts the Expression of Genes Involved in Cell Cycle and Apoptotic Pathways in Mouse Ovarian Antral Follicles1

    PubMed Central

    Craig, Zelieann R.; Hannon, Patrick R.; Wang, Wei; Ziv-Gal, Ayelet; Flaws, Jodi A.

    2012-01-01

    ABSTRACT Di-n-butyl phthalate (DBP) is present in many consumer products, such as infant, beauty, and medical products. Several studies have shown that DBP causes reproductive toxicity in rodents, but no studies have evaluated its effects on ovarian follicles. Therefore, we used a follicle culture system to evaluate the effects of DBP on antral follicle growth, cell cycle and apoptosis gene expression, cell cycle staging, atresia, and 17β-estradiol (E2) production. Antral follicles were isolated from adult CD-1 mice and exposed to DBP at 1, 10, 100, and 1000 μg/ml for 24 or 168 h. Follicles treated with vehicle or DBP at 1–100 μg/ml grew over time, but DBP at 1000 μg/ml significantly suppressed follicle growth. Regardless of effect on follicle growth, DBP-treated follicles had decreased mRNA for cyclins D2, E1, A2, and B1 and increased p21. Levels of the proapoptotic genes Bax, Bad, and Bok were not altered by DBP treatment, but DBP 1000 μg/ml increased levels of Bid and decreased levels of the antiapoptotic gene Bcl2. DBP-treated follicles contained significantly more cells in G1 phase, significantly less cells in S, and exhibited a trend for fewer cells in G2. Although DBP did not affect E2 production and atresia at 24 h, follicles treated with DBP had reduced levels of E2 at 96 h and underwent atresia at 168 h. These data suggest that DBP targets antral follicles and alters the expression of cell cycle and apoptosis factors, causes cell cycle arrest, decreases E2, and triggers atresia, depending on dose. PMID:23242528

  9. Pregnenolone co-treatment partially restores steroidogenesis, but does not prevent growth inhibition and increased atresia in mouse ovarian antral follicles treated with mono-hydroxy methoxychlor

    SciTech Connect

    Craig, Zelieann R. Hannon, Patrick R. Flaws, Jodi A.

    2013-11-01

    Mono-hydroxy methoxychlor (mono-OH MXC) is a metabolite of the pesticide, methoxychlor (MXC). Although MXC is known to decrease antral follicle numbers, and increase follicle death in rodents, not much is known about the ovarian effects of mono-OH MXC. Previous studies indicate that mono-OH MXC inhibits mouse antral follicle growth, increases follicle death, and inhibits steroidogenesis in vitro. Further, previous studies indicate that CYP11A1 expression and production of progesterone (P{sub 4}) may be the early targets of mono-OH MXC in the steroidogenic pathway. Thus, this study tested whether supplementing pregnenolone, the precursor of progesterone and the substrate for HSD3B, would prevent decreased steroidogenesis, inhibited follicle growth, and increased follicle atresia in mono-OH MXC-treated follicles. Mouse antral follicles were exposed to vehicle (dimethylsulfoxide), mono-OH MXC (10 μg/mL), pregnenolone (1 μg/mL), or mono-OH MXC and pregnenolone together for 96 h. Levels of P{sub 4}, androstenedione (A), testosterone (T), estrone (E{sub 1}), and 17β-estradiol (E{sub 2}) in media were determined, and follicles were processed for histological evaluation of atresia. Pregnenolone treatment alone stimulated production of all steroid hormones except E{sub 2}. Mono-OH MXC-treated follicles had decreased sex steroids, but when given pregnenolone, produced levels of P{sub 4}, A, T, and E{sub 1} that were comparable to those in vehicle-treated follicles. Pregnenolone treatment did not prevent growth inhibition and increased atresia in mono-OH MXC-treated follicles. Collectively, these data support the idea that the most upstream effect of mono-OH MXC on steroidogenesis is by reducing the availability of pregnenolone, and that adding pregnenolone may not be sufficient to prevent inhibited follicle growth and survival. - Highlights: • Mono-OH MXC inhibited antral follicle steroidogenesis, growth, and survival. • Pregnenolone partially restored steroidogenesis

  10. Survival and growth of isolated pre-antral follicles from human ovarian medulla tissue during long-term 3D culture.

    PubMed

    Yin, H; Kristensen, S G; Jiang, H; Rasmussen, A; Andersen, C Yding

    2016-07-01

    Can human pre-antral follicles isolated enzymatically from surplus medulla tissue survive and grow in vitro during long-term 3D culture? Secondary human follicles can develop to small antral follicles and remain hormonally active in an alginate-encapsulation culture system for more than 30 days. Ovarian tissue cryopreservation followed by transplantation is a promising fertility preservation approach for cancer patients. However, transplantation of cryopreserved tissue to patients may carry the risk of re-implanting malignant cells. Grafting of follicles enzymatically isolated from ovarian tissue or developing a method for follicular culture and maturation in vitro may provide fertility to such patients without the risk of reintroducing the malignancy. However, the growth of pre-antral follicles isolated by enzymatic digestion from medulla tissue during long-term culture has received only little attention. Two to ten human pre-antral follicles were encapsulated together within an alginate bead and cultured with or without ovarian interstitial tissue for either 7 days or >30 days. Follicles were cultured in either 20% oxygen or 5% oxygen or encapsulated in a lower concentration of alginate together with a lower concentration of FSH in high oxygen. A total of 395 pre-antral follicles from 16 cancer patients, aged 9-37 years, were co-cultured for either 7 days or >30 days. A proportion of follicle (64) were removed from culture on Day 7 and assessed for viability using confocal fluorescence microscopy following calcein-AM and ethidium homodimer-1 staining or histology. The remaining follicles (331) were continued in culture for >30 days then assessed for survival and growth. Anti-Müllerian hormone (AMH) and estradiol levels were quantified in the medium. An optimized protocol for isolation of intact healthy pre-antral follicles from ovarian medulla was developed. After 7 days of culture, secondary follicles had a significantly higher survival rates compared with

  11. Morphological and functional preservation of pre-antral follicles after vitrification of macaque ovarian tissue in a closed system

    PubMed Central

    Ting, A. Y.; Yeoman, R. R.; Campos, J. R.; Lawson, M. S.; Mullen, S. F.; Fahy, G. M.; Zelinski, M. B.

    2013-01-01

    STUDY QUESTION What are the appropriate conditions to vitrify the macaque ovarian cortex in a large-volume, closed system that will preserve functional pre-antral follicles? SUMMARY ANSWER The combination of glycerol, ethylene glycol (EG) and polymers with cooling in liquid nitrogen (LN2) vapor and a two-step warming procedure was able to preserve tissue and follicle morphology as well as function of a small population of secondary follicles in the macaque ovarian cortex following vitrification in a closed system. WHAT IS KNOWN ALREADY For prepubertal cancer patients or those who require immediate cancer therapy, ovarian tissue cryopreservation offers the only hope for future fertility. However, the efficacy of live birth from the transplantation of cryopreserved ovarian tissue is still unclear. In addition, live birth from cryopreserved ovarian tissue has only been demonstrated after tissue autotransplantation, which poses the risk of transmitting metastatic cancer cells back to the cancer survivor in certain cancers. STUDY DESIGN, SIZE, DURATION Non-human primate model, n = 4, randomized, control versus treatment. End-points were collected from tissue histology, tissue culture (48 h) and isolated secondary follicle culture (6 weeks). PARTICIPANTS/MATERIALS, SETTING, METHODS Two vitrification solutions (VSs) containing EG + glycerol (VEG) and EG + dimethylsulfoxide (VED) were examined for vitrification, devitrification and thermodynamic properties. Once the optimal VS was determined, macaque ovarian cortical pieces (3 × 3 × 0.5 mm3) were divided into fresh and two vitrified groups (VEG and VED). For the vitrification groups, tissues were exposed to 1/4, 1/2 and 1× VS for 5 min/step as well as 1× VS + polymers for 1 min at 37°C, loaded into high-security straws with 1 ml of VS + polymers, heat sealed and cooled in LN2 vapor. Samples were warmed in a 40°C water bath and cryoprotective agents were diluted with 1, 0.5, 0.25 and 0 M sucrose. Tissues were fixed

  12. Does AMH Reflect Follicle Number Similarly in Women with and without PCOS?

    PubMed Central

    Christiansen, Sverre C.; Eilertsen, Tina B.; Vanky, Eszter; Carlsen, Sven M.

    2016-01-01

    Context Increased Anti-Mullerian Hormone in polycystic ovary syndrome, may be due to overactive follicles rather than reflect antral follicle count. Objective Does Anti-Mullerian Hormone reflect antral follicle count similarly in women with or without polycystic ovary syndrome or polycystic ovarian morphology? Design Cross-sectional, case-control. Setting Women who delivered preterm in 1999–2006. For each index woman, a woman with a term delivery was identified. Patients Participation rate was 69%. Between 2006–2008, 262 women were included, and diagnosed to have polycystic ovary syndrome, polycystic ovarian morphology or to be normal controls. Intervention(s) Blood tests, a clinical examination and vaginal ultrasound. Main Outcome Measure(s) Anti-Mullerian Hormone / antral follicle count -ratio, SHBG, androstenedione and insulin, to test potential influence on the Anti-Mullerian Hormone / antral follicle count -ratio. Results Mean Anti-Mullerian Hormone / antral follicle count ratio in women with polycystic ovary syndrome or polycystic ovarian morphology was similar to that of the controls (polycystic ovary syndrome: 1,2 p = 0,10 polycystic ovarian morphology: 1,2, p = 0,27 Controls 1,3). Anti-Mullerian Hormone showed a positive linear correlation to antral follicle count in all groups. Multivariate analysis did not change the results. Conclusions We confirmed the positive correlation between AMH and follicle count. Anti-Mullerian Hormone seems to be a reliable predictor of antral follicle count, independent of polycystic ovary syndrome diagnosis or ovarian morphology. PMID:26799212

  13. Mono-hydroxy methoxychlor alters levels of key sex steroids and steroidogenic enzymes in cultured mouse antral follicles

    SciTech Connect

    Craig, Zelieann R.; Leslie, Traci C.; Hatfield, Kimberly P.; Gupta, Rupesh K.; Flaws, Jodi A.

    2010-12-01

    Methoxychlor (MXC) is an organochlorine pesticide that reduces fertility in female rodents by decreasing antral follicle numbers and increasing follicular death. MXC is metabolized in the body to mono-hydroxy MXC (mono-OH). Little is known about the effects of mono-OH on the ovary. Thus, this work tested the hypothesis that mono-OH exposure decreases production of 17{beta}-estradiol (E{sub 2}) by cultured mouse antral follicles. Antral follicles were isolated from CD-1 mice (age 35-39 days) and exposed to dimethylsulfoxide (DMSO), or mono-OH (0.1-10 {mu}g/mL) for 96 h. Media and follicles were collected for analysis of sex steroid levels and mRNA expression, respectively. Mono-OH treatment (10 {mu}g/mL) decreased E{sub 2} (DMSO: 3009.72 {+-} 744.99 ng/mL; mono-OH 0.1 {mu}g/mL: 1679.66 {+-} 461.99 ng/mL; 1 {mu}g/mL: 1752.72 {+-} 532.41 ng/mL; 10 {mu}g/mL: 45.89 {+-} 33.83 ng/mL), testosterone (DMSO: 15.43 {+-} 2.86 ng/mL; mono-OH 0.1 {mu}g/mL: 17.17 {+-} 4.71 ng/mL; 1 {mu}g/mL: 13.64 {+-} 3.53 ng/mL; 10 {mu}g/mL: 1.29 {+-} 0.23 ng/mL), androstenedione (DMSO: 1.92 {+-} 0.34 ng/mL; mono-OH 0.1 {mu}g/mL: 1.49 {+-} 0.43 ng/mL; 1 {mu}g/mL: 0.64 {+-} 0.31 ng/mL; 10 {mu}g/mL: 0.12 {+-} 0.06 ng/mL) and progesterone (DMSO: 24.11 {+-} 4.21 ng/mL; mono-OH 0.1 {mu}g/mL: 26.77 {+-} 4.41 ng/mL; 1 {mu}g/mL: 20.90 {+-} 3.75 ng/mL; 10 {mu}g/mL: 9.44 {+-} 2.97 ng/mL) levels. Mono-OH did not alter expression of Star, Hsd3b1, Hsd17b1 and Cyp1b1, but it did reduce levels of Cyp11a1, Cyp17a1 and Cyp19a1 mRNA. Collectively, these data suggest that mono-OH significantly decreases levels of key sex steroid hormones and the expression of enzymes required for steroidogenesis.

  14. 2,3,7,8-Tetrachlorodibenzo-p-dioxin activates the aryl hydrocarbon receptor and alters sex steroid hormone secretion without affecting growth of mouse antral follicles in vitro

    SciTech Connect

    Karman, Bethany N. Basavarajappa, Mallikarjuna S. Craig, Zelieann R. Flaws, Jodi A.

    2012-05-15

    The persistent environmental contaminant, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) is an ovarian toxicant. These studies were designed to characterize the actions of TCDD on steroidogenesis and growth of intact mouse antral follicles in vitro. Specifically, these studies tested the hypothesis that TCDD exposure leads to decreased sex hormone production/secretion by antral follicles as well as decreased growth of antral follicles in vitro. Since TCDD acts through binding to the aryl hydrocarbon receptor (AHR), and the AHR has been identified as an important factor in ovarian function, we also conducted experiments to confirm the presence and activation of the AHR in our tissue culture system. To do so, we exposed mouse antral follicles for 96 h to a series of TCDD doses previously shown to have effects on ovarian tissues and cells in culture, which also encompass environmentally relevant and pharmacological exposures (0.1–100 nM), to determine a dose response for TCDD in our culture system for growth, hormone production, and expression of the Ahr and Cyp1b1. The results indicate that TCDD decreases progesterone, androstenedione, testosterone, and estradiol levels in a non-monotonic dose response manner without altering growth of antral follicles. The addition of pregnenolone substrate (10 μM) restores hormone levels to control levels. Additionally, Cyp1b1 levels were increased by 3–4 fold regardless of the dose of TCDD exposure, evidence of AHR activation. Overall, these data indicate that TCDD may act prior to pregnenolone formation and through AHR transcriptional control of Cyp1b1, leading to decreased hormone levels without affecting growth of antral follicles. -- Highlights: ►TCDD disrupts sex steroid hormone levels, but not growth of antral follicles. ►Pregnenolone co-treatment by-passes TCDD-induced steroid hormone disruption. ►TCDD affects steroid hormone levels through an AHR pathway in antral follicles.

  15. Development of the membrana granulosa of bovine antral follicles: structure, location of mitosis and pyknosis, and immunolocalization of involucrin and vimentin.

    PubMed

    van Wezel, I L; Krupa, M; Rodgers, R J

    1999-01-01

    The membrana granulosa of the ovarian follicle is termed the 'follicular epithelium', yet there have been no studies considering its epithelial nature and how it changes during follicular development. Therefore, these issues were investigated using histology (n = 45 ovaries), considering its structure and the location of proliferating and dying cells, and drawing analogies with other epithelia. Additionally, differences between the layers of granulosa cells were demonstrated by immunohistochemistry (n = 7 ovaries). The structure of the membrana granulosa differed between follicles. Six arbitrary classifications were designed based on these structures, 80 follicles were allocated (n = 13 ovaries) to these classes and the follicular diameters were then measured. For the first time, differences in membrana granulosa structure were shown to correspond to follicle size. Follicles in classes 1-3, where basal granulosa cells were columnar with nuclei positioned basally in the cell, were all < or = 3 mm in diameter. All follicles larger than 3 mm had either columnar basal cells with nuclei positioned centrally (class 4), or had rounded basal cells (class 5), and all follicles > 5 mm had only rounded basal cells. In all these classes, cells in the middle zone were rounded; cells aligning the antrum were often flattened. Irrespective of follicle class, cell proliferation and cell death were shown to be predominantly in the middle portions, rather than the most antral or most basal portions, of the membrana granulosa of healthy and atretic follicles. Involucrin, a marker of keratinocyte differentiation, was localized to the suprabasal region of the membrana granulosa of healthy follicles, particularly in the second and third cellular layers in from the follicular basal lamina. Conversely, the staining intensity for the intermediate filament protein vimentin was lowest in this region, and greatest in the more antral and basal regions. In atretic follicles, there was widespread

  16. Computer-generated ovaries to assist follicle counting experiments.

    PubMed

    Skodras, Angelos; Marcelli, Gianluca

    2015-01-01

    Precise estimation of the number of follicles in ovaries is of key importance in the field of reproductive biology, both from a developmental point of view, where follicle numbers are determined at specific time points, as well as from a therapeutic perspective, determining the adverse effects of environmental toxins and cancer chemotherapeutics on the reproductive system. The two main factors affecting follicle number estimates are the sampling method and the variation in follicle numbers within animals of the same strain, due to biological variability. This study aims at assessing the effect of these two factors, when estimating ovarian follicle numbers of neonatal mice. We developed computer algorithms, which generate models of neonatal mouse ovaries (simulated ovaries), with characteristics derived from experimental measurements already available in the published literature. The simulated ovaries are used to reproduce in-silico counting experiments based on unbiased stereological techniques; the proposed approach provides the necessary number of ovaries and sampling frequency to be used in the experiments given a specific biological variability and a desirable degree of accuracy. The simulated ovary is a novel, versatile tool which can be used in the planning phase of experiments to estimate the expected number of animals and workload, ensuring appropriate statistical power of the resulting measurements. Moreover, the idea of the simulated ovary can be applied to other organs made up of large numbers of individual functional units.

  17. Expression of androgen-producing enzyme genes and testosterone concentration in Angus and Nellore heifers with high and low ovarian follicle count.

    PubMed

    Loureiro, Bárbara; Ereno, Ronaldo L; Favoreto, Mauricio G; Barros, Ciro M

    2016-07-15

    Follicle population is important when animals are used in assisted reproductive programs. Bos indicus animals have more follicles per follicular wave than Bos taurus animals. On the other hand, B taurus animals present better fertility when compared with B indicus animals. Androgens are positively related with the number of antral follicles; moreover, they increase growth factor expression in granulose cells and oocytes. Experimentation was designed to compare testosterone concentration in plasma, and follicular fluid and androgen enzymes mRNA expression (CYP11A1, CYP17A1, 3BHSD, and 17BHSD) in follicles from Angus and Nellore heifers. Heifers were assigned into two groups according to the number of follicles: low and high follicle count groups. Increased testosterone concentration was measured in both plasma and follicular fluid of Angus heifers. However, there was no difference within groups. Expression of CYP11A1 gene was higher in follicles from Angus heifers; however, there was no difference within groups. Expression of CYP17A1, 3BHSD, and 17BHSD genes was higher in follicles from Nellore heifers, and expression of CYP17A1 and 3BHSD genes was also higher in HFC groups from both breeds. It was found that Nellore heifers have more antral follicles than Angus heifers. Testosterone concentration was higher in Angus heifers; this increase could be associated with the increased mRNA expression of CYP11A1. Increased expression of androgen-producing enzyme genes (CYP17A1, 3BHSD, and 17BHSD) was detected in Nellore heifers. It can be suggested that testosterone is acting through different mechanisms to increase follicle development in Nellore and improve fertility in Angus heifers. Copyright © 2016 Elsevier Inc. All rights reserved.

  18. Caprine ovarian follicle requirements differ between preantral and early antral stages after IVC in medium supplemented with GH and VEGF alone or in combination.

    PubMed

    Cadenas, J; Leiva-Revilla, J; Vieira, L A; Apolloni, L B; Aguiar, F L N; Alves, B G; Lobo, C H; Rodrigues, A P R; Apgar, G A; Smitz, J; Figueiredo, J R; Maside, C

    2017-01-01

    The aim of the present study was to evaluate the effect of growth hormone (GH) and vascular endothelial growth factor (VEGF) added alone, sequentially or in combination, in the presence of insulin at physiological concentration (10 ng/mL) on the IVC of two different follicular categories: preantral (experiment 1; Exp.1) and early antral (experiment 2; Exp.2). Isolated follicles were individually cultured for 24 (Exp.1) and 18 days (Exp.2) in the following treatments: αMEM(+) (Control), or Control medium supplemented with 50 ng/mL GH (GH), 100 ng/mL VEGF (VEGF), the combination of both (GH + VEGF), GH during the first 12 days and VEGF from Day 12 until the end of the culture (GH/VEGF) and vice versa (VEGF/GH). At the end of the culture, cumulus-oocyte complexes from in vitro-grown follicles were recovered and subjected to IVM. The following end points were evaluated: Follicle morphology, growth rates and antrum formation, production of estradiol, progesterone and testosterone, oocyte viability and meiotic stage, as well as relative expression of LHR, Amh, HAS2, PTGS2, CYP17, CYP19A1, and 3βHSD. A considerable amount of viable fully grown oocytes were recovered after the IVC of early antral follicles in all treatments. Nevertheless, the GH treatment presented the highest percentage of fully grown oocytes (60%), mean oocyte diameter (117.74 ± 2.61 μm), and meiotic resumption (50%). Furthermore, GH treatment produced higher (P < 0.05) rates of metaphase II oocytes than all the other treatments, and similar LHR, Amh, and PTGS2 transcript levels to in vivo. Contrary to early antral follicles, preantral follicles were not affected by medium supplementation. In conclusion, the addition of GH to a culture medium containing physiological concentrations of insulin improves oocyte growth and maturation after the IVC of goat early antral follicles.

  19. Dioxin exposure reduces the steroidogenic capacity of mouse antral follicles mainly at the level of HSD17B1 without altering atresia.

    PubMed

    Karman, Bethany N; Basavarajappa, Mallikarjuna S; Hannon, Patrick; Flaws, Jodi A

    2012-10-01

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a potent ovarian toxicant. Previously, we demonstrated that in vitro TCDD (1nM) exposure decreases production/secretion of the sex steroid hormones progesterone (P4), androstenedione (A4), testosterone (T), and 17β-estradiol (E2) in mouse antral follicles. The purpose of this study was to determine the mechanism by which TCDD inhibits steroidogenesis. Specifically, we examined the effects of TCDD on the steroidogenic enzymes, atresia, and the aryl hydrocarbon receptor (AHR) protein. TCDD exposure for 48h increased levels of A4, without changing HSD3B1 protein, HSD17B1 protein, estrone (E1), T or E2 levels. Further, TCDD did not alter atresia ratings compared to vehicle at 48h. TCDD, however, did down regulate the AHR protein at 48h. TCDD exposure for 96h decreased transcript levels for Cyp11a1, Cyp17a1, Hsd17b1, and Cyp19a1, but increased Hsd3b1 transcript. TCDD exposure particularly lowered both Hsd17b1 transcript and HSD17B1 protein. However, TCDD exposure did not affect levels of E1 in the media nor atresia ratings at 96h. TCDD, however, decreased levels of the proapoptotic factor Bax. Collectively, these data suggest that TCDD exposure causes a major block in the steroidogenic enzyme conversion of A4 to T and E1 to E2 and that it regulates apoptotic pathways, favoring survival over death in antral follicles. Finally, the down-regulation of the AHR protein in TCDD exposed follicles persisted at 96h, indicating that the activation and proteasomal degradation of this receptor likely plays a central role in the impaired steroidogenic capacity and altered apoptotic pathway of exposed antral follicles.

  20. Quantification of ZP1, ZP2 and ZP3 mRNA of marmoset monkey (Callithrix jacchus) oocytes from periantral and antral follicles.

    PubMed

    Konrad, L; Kuhnert, S; Nayudu, P L; Einspanier, R; Hinsch, K D; Hinsch, E

    2012-05-01

    In mammals, the oocyte and preimplantation embryo are protected by the zona pellucida (ZP) consisting mainly of ZP glycoproteins, which are responsible for sperm binding, induction of the acrosome reaction and zona pellucida hardening to prevent polyspermia. The ZP proteins become increasingly important as possible predictors for in vitro cultured oocytes competence. As little is known about the stage-dependent expression of ZP1, ZP2 and ZP3 in marmoset monkey (Callithrix jacchus) oocytes, mRNA expression was investigated with real-time RT-PCR. Total-RNA was isolated from three different classes of marmoset oocytes; Class 1 oocytes from periantral follicles (<600 μm, n = 10), Class 2 oocytes from small antral follicles (600-1000 μm, n = 10) and Class 3 oocytes from large antral follicles (>1000 μm, n = 9). Compared with Class 1 oocytes mRNA expression of ZP1, ZP2 and ZP3 in Class 2 oocytes was significantly decreased. In Class 3 oocytes, the transcription of ZP1, ZP2 and ZP3 genes showed also a significant decrease compared with Class 1 oocytes. In this study a differently regulated expression of the ZP genes during late folliculogenesis with an obvious downregulation of ZP1, ZP2 and ZP3 could be demonstrated for the first time in the marmoset monkey.

  1. A western-style diet, with and without chronic androgen treatment, alters the number, structure and function of small antral follicles in ovaries of young adult monkeys

    PubMed Central

    Bishop, Cecily V.; Xu, Fuhua; Xu, Jing; Ting, Alison Y.; Galbreath, Etienne; McGee, Whitney K.; Zelinski, Mary B.; Hennebold, Jon D.; Cameron, Judy L.; Stouffer, Richard L.

    2015-01-01

    Objective To examine the small antral follicle (SAF) cohort in ovaries of adult rhesus monkeys following consumption of a western-style diet (WSD), with or without chronically elevated androgen levels since before puberty. Design Cholesterol or testosterone (T; n=6/group) implants were placed subcutaneously in female rhesus macaques beginning at 1 yr of age (pre-pubertal), with addition of a WSD (high fat/fructose) at 5.5 yrs (menarche ~2.6 yrs). Ovaries were collected at 7 yrs of age. One ovary/female was embedded in paraffin for morphological and immunohistochemical analyses. The SAFs (<2.5mm) were dissected from the other ovary obtained at/near menses in a subgroup of females (n=3/group), and processed for microarray analyses of the SAF transcriptome. Ovaries of adult monkeys consuming a standard macaque diet (low in fats and sugars) were obtained at similar stages of the menstrual cycle and used as controls for all analyses. Setting National primate research center Animals Adult, female rhesus monkeys (Macaca mulatta) Interventions None Main outcome measures Histological analyses, SAF counts and morphology, protein localization and abundance in SAFs, transcriptome in SAFs (mRNAs) Results Compared to controls, consumption of a WSD, with and without T treatment, increased the numbers of SAFs per ovary, due to the presence of more atretic follicles. Numbers of granulosa cells expressing cellular proliferation markers (pRb and pH3) was greater in healthy SAFs, while numbers of cells expressing the cell cycle inhibitor (p21) was higher in atretic SAFs. Intense CYP17A1 staining was observed in the theca cells of SAFs from WSD+/− T groups, compared to controls. Microarray analyses of the transcriptome in SAFs isolated from WSD and WSD+T treated females and controls consuming a standard diet, identified 1944 genes whose mRNA levels changed ≥2-fold among the three groups. Further analyses identified several gene pathways altered by WSD and/or WSD+T associated with

  2. Bisphenol A down-regulates rate-limiting Cyp11a1 to acutely inhibit steroidogenesis in cultured mouse antral follicles

    SciTech Connect

    Peretz, Jackye; Flaws, Jodi A.

    2013-09-01

    Bisphenol A (BPA) is the backbone of polycarbonate plastic products and the epoxy resin lining of aluminum cans. Previous studies have shown that exposure to BPA decreases sex steroid hormone production in mouse antral follicles. The current study tests the hypothesis that BPA first decreases the expression levels of the steroidogenic enzyme cytochrome P450 side-chain cleavage (Cyp11a1) and steroidogenic acute regulatory protein (StAR) in mouse antral follicles, leading to a decrease in sex steroid hormone production in vitro. Further, the current study tests the hypothesis that these effects are acute and reversible after removal of BPA. Exposure to BPA (10 μg/mL and 100 μg/mL) significantly decreased expression of Cyp11a1 and StAR beginning at 18 h and 72 h, respectively, compared to controls. Exposure to BPA (10 μg/mL and 100 μg/mL) significantly decreased progesterone levels beginning at 24 h and decreased androstenedione, testosterone, and estradiol levels at 72 h and 96 h compared to controls. Further, after removing BPA from the culture media at 20 h, expression of Cyp11a1 and progesterone levels were restored to control levels by 48 h and 72 h, respectively. Additionally, expression of StAR and levels of androstenedione, testosterone, and estradiol never decreased compared to controls. These data suggest that BPA acutely decreases expression of Cyp11a1 as early as 18 h and this reduction in Cyp11a1 may lead to a decrease in progesterone production by 24 h, followed by a decrease in androstenedione, testosterone, and estradiol production and expression of StAR at 72 h. Therefore, BPA exposure likely targets Cyp11a1 and steroidogenesis, but these effects are reversible with removal of BPA exposure. - Highlights: • BPA may target Cyp11a1 to inhibit steroidogenesis in antral follicles. • BPA may decrease the expression of Cyp11a1 prior to inhibiting steroidogenesis. • The adverse effects of BPA on steroidogenesis in antral follicles are reversible.

  3. Grid analysis improves reliability in follicle counts made by ultrasonography in women with polycystic ovary syndrome.

    PubMed

    Lujan, Marla E; Brooks, Eric D; Kepley, Anna L; Chizen, Donna R; Pierson, Roger A; Peppin, Andrew K

    2010-05-01

    Poor reliability has been reported when counting the total number of follicles in polycystic ovaries using conventional two-dimensional (2-D) ultrasound viewing methods. In the current study, we report good reliability in follicle counts when observers imposed a programmable grid system over the viewing window. Four observers estimated total follicle counts in 45 ovarian ultrasound scans by compartmentalizing the ovary into 9 to 12 grid sections and performing focused follicle counts per section. The mean number of follicles counted per ovary was 44.6 +/- 2.3. The level of inter-observer agreement when making follicle counts was 0.82 and total follicle counts did not differ among observers. The level of intra-observer agreement was 0.93 which further corroborated the utility of this method for making dependable follicle counts. In summary, the ability to obtain reproducible follicle counts will help to establish reliable diagnostic criteria for polycystic ovarian morphology. Copyright 2010 World Federation for Ultrasound in Medicine & Biology. Published by Elsevier Inc. All rights reserved.

  4. Lifestyle factors associated with histologically derived human ovarian non-growing follicle count in reproductive age women†

    PubMed Central

    Peck, Jennifer D.; Quaas, Alexander M.; Craig, LaTasha B.; Soules, Michael R.; Klein, Nancy A.; Hansen, Karl R.

    2016-01-01

    -users. Neither heavier alcohol use (>3 drink-years), BMI, OCP use, nor tobacco use were significantly associated with the ovarian NGF count. Similar patterns of association with moderate cumulative alcohol use were observed when evaluating associations with pre-antral follicles and total follicle counts. LIMITATIONS, REASONS FOR CAUTION All participants in this convenience sample had a benign indication for hysterectomy, and therefore may not be broadly representative of the population without such an indication. Additionally, lifestyle factors were self-reported, and the sample size of the present investigation limits our ability to detect associations of smaller magnitude. WIDER IMPLICATIONS OF THE FINDINGS While our findings are in disagreement with a single investigation that utilized human follicle density as the outcome of interest, they are consistent with many studies investigating the relationship between lifestyle factors and the age of spontaneous menopause. Furthermore, they suggest a mechanism that does not involve accelerated follicular atresia to explain the association between smoking and an earlier age of menopause. STUDY FUNDING/COMPETING INTEREST(S) This investigation was funded by NIA R29-HD37360-04 (N.A.K.) and OCAST HR04-115 (K.R.H.) and by the National Institute of General Medical Sciences, Grant 1 U54GM104938 (J.D.P.). There is no conflict of interest. PMID:26497957

  5. Effects of in vitro growth culture duration and prematuration culture on maturational and developmental competences of bovine oocytes derived from early antral follicles.

    PubMed

    Huang, Weiping; Nagano, Masashi; Kang, Sung-Sik; Yanagawa, Yojiro; Takahashi, Yoshiyuki

    2013-10-15

    Bovine ovaries offer a large pool of oocytes that could be used for in vitro production of embryos of genetically valuable animals. The effects of in vitro growth (IVG) culture duration (10, 12, and 14 days) on the viability and growth of bovine oocytes derived from early antral follicles (0.5-1 mm diameter) in this study. In addition, the effect of pre-IVM culture with phosphodiesterase inhibitor (3-isobutyl-1-methylxanthine) on nuclear maturation of IVG oocytes was also evaluated. In experiment 1, oocyte viability observed after 10 or 12 days of IVG culture was greater (P < 0.05) than that observed after 14 days of culture. Oocyte diameters and proportions of oocytes at metaphase II stage were greater (P < 0.05) when 12 or 14 days of IVG culture where used when compared with 10 days culture. In addition, the proportion of oocytes at metaphase II stage was greater (P < 0.05) when pre-IVM culture was performed for oocytes derived from 12 and 14 days of IVG culture. When 12 and 14 days of IVG culture followed by pre-IVM culture were compared in experiment 2, cumulus cell membrane integrity was greater (P < 0.05) after 12 days. Blastocyst production rate for oocytes obtained after 12 days of IVG culture (24.5%) was greater (P < 0.05) than for oocytes obtained after 14 days (9.9%). In conclusion, 12 days IVG followed by pre-IVM culture was considered the optimal processing system for bovine oocytes derived from early antral follicles when oocyte viability, diameter, maturation, and development competences were considered.

  6. Influence of puberty and antral follicle count on calving day in crossbred beef heifers

    USDA-ARS?s Scientific Manuscript database

    The ability of a cow to produce a calf every 12 months beginning at 24 months of age is one of the primary factors contributing to the long-term profitability of a cow-calf operation. Cows that calve early in the calving season have the greatest productive lifespan, but which factors contribute the...

  7. Impact of oxygen concentration on adult murine pre-antral follicle development in vitro and the corresponding metabolic profile.

    PubMed

    Gook, Debra A; Edgar, D H; Lewis, K; Sheedy, J R; Gardner, D K

    2014-01-01

    Oxygen concentration during in vitro culture has a significant effect on the physiology of embryos, altering metabolic profile and developmental outcome. Although atmospheric oxygen has been used routinely for the culture of ovarian follicles, oxygen concentration may also be critical for follicle growth but the optimal concentration has not been determined. In this study, mechanically isolated primary and secondary follicles (80-140 µm diameter) from adult mouse ovaries were cultured in serum-free conditions for 8 days in either 5 or 20% oxygen to determine growth (follicular diameter), morphology and viability. For each oxygen concentration, half of the medium was replaced on Days 2, 4 and 6 or on Day 4 only. In the latter group, metabolic analysis of spent follicular culture media was performed by (1)H-NMR. The proportion of viable, growing follicles was significantly (P < 0.0001) higher in 5% than in 20% oxygen (59% versus 8%). Reducing the frequency of medium replacement during culture in 5% oxygen resulted in significantly (P < 0.001) more viable follicles (79 versus 46%). In 20% oxygen, poor follicular viability was observed irrespective of the frequency of medium replacement (8 and 10% respectively). Metabolic profiles showed marked differences in amino acid and carbohydrate utilization with respect to both oxygen concentration and between Days 4 and 8 of development. Metabolites which significantly discriminated between oxygen concentration at both time points were glucose consumption, lactate utilization, alanine, alanyl-glutamine, leucine and proline. In conclusion, the poor in vitro follicular development previously observed in minimal culture conditions may reflect the use of 20% oxygen. Frequent medium replenishment is not necessary and does not overcome the detrimental effect of high oxygen on follicle viability. Further optimization of culture conditions would benefit from metabolic analyses and the use of 5% oxygen should be tested further for

  8. Prophase I arrest of mouse oocytes mediated by natriuretic peptide precursor C requires GJA1 (connexin-43) and GJA4 (connexin-37) gap junctions in the antral follicle and cumulus-oocyte complex.

    PubMed

    Richard, Samantha; Baltz, Jay M

    2014-06-01

    Fully grown germinal vesicle stage mouse oocytes remain arrested in meiotic prophase I until ovulation. This arrest is maintained by cGMP produced in cumulus granulosa cells surrounding the oocyte. Recently, it was found that cGMP production in cumulus cells depends on NPR2 guanylate cyclase activated by its ligand natriuretic peptide precursor C (NPPC). It is assumed that cGMP reaches the oocyte through gap junctions that couple cumulus granulosa cells to each other and to the oocyte. Previous work identified two main types of gap junctions in the follicle, connexin-43 gap junctions (GJA1 protein) between granulosa cells and connexin-37 gap junctions (GJA4) between cumulus cells and the oocyte. However, it had not been established that both types are required for meiotic arrest mediated by NPPC/NPR2 signaling. To investigate this, we used connexin mimetic peptides (CMPs) that specifically disrupt gap junction isoforms within cumulus-oocyte complexes (COCs) and isolated antral follicles in culture. We furthermore developed a punctured antral follicle preparation to permit CMP access to the antral cavity in an otherwise intact follicle. CMP directed against connexin-43 (Cx43 CMP) overcame NPPC-mediated meiotic arrest in both isolated COCs and antral follicles. Cx37 CMP, in contrast, had no effect when present in the medium, but released oocyte arrest in the presence of NPPC when microinjected into the perivitelline space near the oocyte surface in COCs. This is consistent with both connexin isoforms being required for meiotic arrest and with the reported localization of connexin-43 throughout the cumulus cells and connexin-37 at the oocyte surface. © 2014 by the Society for the Study of Reproduction, Inc.

  9. Multiple ovarian antral follicles in a preterm infant with neonatal intrahepatic cholestasis caused by citrin deficiency: a clinical, genetic and transcriptional analysis.

    PubMed

    Lin, Wei-Xia; Zhang, Zhan-Hui; Deng, Mei; Cai, Xiang-Ran; Song, Yuan-Zong

    2012-09-01

    Neonatal intrahepatic cholestasis caused by citrin deficiency (NICCD) is an autosomal recessive disease caused by the dysfunction of citrin, an aspartate/glutamate carrier encoded by the SLC25A13 gene. Considerable progress has been made on the diagnosis and treatment of NICCD, but its clinical and molecular features still remain far from being completely elucidated and generally understood. The infant, a preterm female delivered at a gestational age of 31 weeks, was referred to our hospital at the age of 8 months because of jaundice lasting for 4.5 months and ovarian masses uncovered for 3 months. Besides serum indices indicating cholestasis, elevated serum levels of luteinizing hormone, follicle stimulating hormone and estradiol were also detected. Abdominal magnetic resonance imaging demonstrated bilateral multi-cystic ovarian masses, with the largest size being 7.4 × 6.2 × 9.6 mm(3). SLC25A13 gene analysis revealed that the patient was a compound heterozygote of c.1177+1G>A and c.754G>A. The paternally-inherited mutation c.754G>A was a novel one with a carrier rate of less than 1%. SLC25A13 transcriptional study in peripheral blood lymphocytes (PBLs) documented a novel splice variant r.616_848del which resulted from c.754G>A, with another variant r.1019_1177del from the maternally-inherited c.1177+1G>A mutation. The diagnoses were NICCD and multiple ovarian antral follicles (minipuberty), and the patient responded well to a galactose-free and medium chain triglyceride (MCT)-enriched formula. The findings in this paper expanded the clinical and molecular spectrum of the SLC25A13 gene, and lent support to the concept that PBLs could be taken as a feasible specimen source for SLC25A13 transcriptional analysis. Copyright © 2012 Elsevier B.V. All rights reserved.

  10. Infertility and ovarian follicle reserve depletion are associated with dysregulation of the FSH and LH receptor density in human antral follicles.

    PubMed

    Regan, Sheena L P; Knight, Phil G; Yovich, John L; Stanger, James D; Leung, Yee; Arfuso, Frank; Dharmarajan, Arun; Almahbobi, Ghanim

    2017-02-07

    The low take-home baby rate in older women in Australia (5.8%) undergoing IVF (5.8%) is linked to the depletion of the ovarian reserve of primordial follicles. Oocyte depletion causes an irreversible change to ovarian function. We found that the young patient FSH receptor and LH receptor expression profile on the granulosa cells collected from different size follicles were similar to the expression profile reported in natural cycles in women and sheep. This was reversed in the older patients with poor ovarian reserve. The strong correlation of BMPR1B and FSH receptor density in the young was not present in the older women; whereas, the LH receptor and BMPR1B correlation was weak in the young but was strongly correlated in the older women. The reduced fertilisation and pregnancy rate was associated with a lower LH receptor density and a lack of essential down-regulation of the FSH and LH receptor. The mechanism regulating FSH and LH receptor expression appears to function independently, in vivo, from the dose of FSH gonadotrophin, rather than in response to it. Restoring an optimum receptor density may improve oocyte quality and the pregnancy rate in older women.

  11. Expression of TGF-beta superfamily growth factors, their receptors, the associated SMADs and antagonists in five isolated size-matched populations of pre-antral follicles from normal human ovaries.

    PubMed

    Kristensen, Stine Gry; Andersen, Kasper; Clement, Christian Alexandro; Franks, Stephen; Hardy, Kate; Andersen, Claus Yding

    2014-04-01

    In mammals, members of the transforming growth factor-beta (TGF-β) superfamily are known to have key roles in the regulation of follicular growth and development. The aim of the study was to evaluate the expression of TGF-β superfamily growth factors, their receptors, downstream SMAD signalling molecules and TGF-β/bone morphogenetic protein (BMP) antagonists during early human folliculogenesis. Human pre-antral follicles were enzymatically isolated from surplus ovarian tissue obtained from women having ovarian cortical tissue frozen for fertility preservation. A total of 348 human pre-antral follicles, ranging from 40 to 200 µm in diameter, were isolated from ovarian tissue obtained from 15 women, aged 24-34 years. Isolated pre-antral follicles were grouped according to diameter in five size-matched populations spanning the primordial, primary and secondary stage follicles and analysed by whole-genome microarray analysis. Selected proteins/genes were analysed by immunocytochemistry and quantitative RT-PCR. TGF-β superfamily genes with overall highest mRNA expressions levels included growth differentiation factors 9 (GDF9), BMP15, BMP6, BMP-receptor-2 (BMPR2), anti-Müllerian hormone receptor 2 (AMHR2), TGFβR3, inhibin-α (INHA) and intracellular SMAD3 and SMAD4. Moreover, genes which were differentially expressed from the primordial to the late secondary stage follicles included GDF9, BMP15, AMH, INHBB, TGFβR3, SMAD4 and antagonists Follistatin (FST) and GREM1. Collectively, these data indicate that the active TGF-β superfamily pathways in early human folliculogenesis consist of primarily GDF9 combined with possible synergistic effects of BMP15 through the BMPR2 and intracellular activation of SMAD3 and SMAD4, and that AMH and INHBB are engaged in intrafollicular events from the onset of follicular growth. Moreover, the presence of multiple TGF-β/BMP antagonists imply that certain growth factors are subjected to local regulation on different levels that

  12. Co-treatment of mouse antral follicles with 17β-estradiol interferes with mono-2-ethylhexyl phthalate (MEHP)-induced atresia and altered apoptosis gene expression

    PubMed Central

    Craig, Zelieann R.; Singh, Jeffrey; Gupta, Rupesh K.; Flaws, Jodi A.

    2014-01-01

    Mono-2-ethyhexyl phthalate (MEHP) is a metabolite of a plasticizer found in many consumer products. MEHP inhibits mouse ovarian follicle growth by reducing 17β-estradiol (E2) production. Yet, whether MEHP causes follicle death (atresia) is unclear. We hypothesized that MEHP causes atresia by altering apoptosis gene expression, and that E2 co-treatment blocks these effects. Follicles were exposed to MEHP (0.36–36 µM) ± E2 for 48–96h to determine the effect of MEHP ± E2 on atresia and gene expression. MEHP increased atresia, but this effect was blocked by co-treatment with E2. MEHP increased the expression of the pro-apoptotic gene Aifm1, but decreased that of the pro-apoptotic gene Bok and the anti-apoptotic gene Bcl2l10. E2 interfered with MEHP-induced changes in Aifm1 and Bcl2l10. Our findings suggest that decreased E2 levels are required for MEHP-induced follicle atresia and that Aifm1, Bok, and Bcl2l10 are involved in this process. PMID:24412242

  13. Co-treatment of mouse antral follicles with 17β-estradiol interferes with mono-2-ethylhexyl phthalate (MEHP)-induced atresia and altered apoptosis gene expression.

    PubMed

    Craig, Zelieann R; Singh, Jeffrey; Gupta, Rupesh K; Flaws, Jodi A

    2014-06-01

    Mono-2-ethyhexyl phthalate (MEHP) is a metabolite of a plasticizer found in many consumer products. MEHP inhibits mouse ovarian follicle growth by reducing 17β-estradiol (E2) production. Yet, whether MEHP causes follicle death (atresia) is unclear. We hypothesized that MEHP causes atresia by altering apoptosis gene expression, and that E2 co-treatment blocks these effects. Follicles were exposed to MEHP (0.36-36μM)±E2 for 48-96h to determine the effect of MEHP±E2 on atresia and gene expression. MEHP increased atresia, but this effect was blocked by co-treatment with E2. MEHP increased the expression of the pro-apoptotic gene Aifm1, but decreased that of the pro-apoptotic gene Bok and the anti-apoptotic gene Bcl2l10. E2 interfered with MEHP-induced changes in Aifm1 and Bcl2l10. Our findings suggest that decreased E2 levels are required for MEHP-induced follicle atresia and that Aifm1, Bok, and Bcl2l10 are involved in this process.

  14. Understanding follicle growth in vivo.

    PubMed

    Oktem, Ozgur; Urman, Bulent

    2010-12-01

    Ovarian reserve is determined by the number of primordial follicles in the ovary. Quiescent primordial follicles are activated for growth and pass through stages of development before they reach the antral stage. Then a cohort of antral follicles is recruited for further growth, dominance and ovulation under the cyclic stimulation of gonadotrophins. What triggers the initiation of growth in primordial follicles has remained a mystery for decades. However, recent studies on mutant mouse models have shown that primordial follicles are maintained in a dormant state by the actions of various inhibitory molecules to preserve the follicle pool, such as the transcription factor Foxo3a, PTEN (phosphotase and tensin homolog deleted on chromosome 10) and Tsc-1 (tumour suppressor tuberous schlerosis complex). Mice with deletions of these oocyte-specific genes exhibit premature activation of dormant primordial follicles, and all primordial follicles become depleted in early adulthood, causing premature ovarian failure. Other oocyte and somatic cell-derived growth factors are also involved in the early, gonadotrophin-independent phase of follicle growth via autocrine and paracrine interactions. Interestingly, some of these factors also play critical roles at later stages of follicle growth, such as the process of selecting the dominant follicle, by modifying the response of the follicles to gonadotrophins and inhibiting premature luteinization. Therefore, a thorough understanding of the molecular aspects of folliculogenesis is of paramount importance in the context of translational medicine and future clinical applications in human reproduction.

  15. Polycystic ovarian morphology and the diagnosis of polycystic ovary syndrome: redefining threshold levels for follicle count and serum anti-Müllerian hormone using cluster analysis.

    PubMed

    Lie Fong, S; Laven, J S E; Duhamel, A; Dewailly, D

    2017-08-01

    ' clusters had their similarly aged counterpart of 'non-PCOM' clusters. Likewise, two clusters comprised women younger than 30 years, with (n = 28, 'PCOM regularly cycling women') or without (n = 118, 'normal regularly cycling women') features of PCOM (increased FNPO and/or serum AMH). The two other clusters in older women could be labelled 'normal regularly cycling women' or 'PCOM regularly cycling women' (n = 100 and 51, respectively). The prevalence of PCOM was significantly greater in old than in young regularly cycling women controls. In the young population, after exclusion of the 'PCOM regularly cycling women', the diagnostic performance of AMH, expressed by area under the curve (AUC) (AUC = 0.903; CI (0.876-0.930)) to differentiate PCOS women from normal regularly cycling women was similar to that using the FNPO (AUC = 0.915, CI (0.891-0.940)) (P = 0.25), confirming results from earlier studies. In the old population, the diagnostic performance of AMH was greater than that of FNPO (AUCs = 0.948 (0.927-0.970) vs 0.874 (0.836-0.912), respectively, P = 0.00035). Cut-off levels of AMH and antral follicle count distinguishing regularly cycling non-PCOM women from PCOS women were higher in young women than in older women. Data of normal women were obtained from earlier studies, aiming to measure normal endocrine values. Apparently, the strong effect of age in cluster analysis revealed a dichotomy in the age distribution among the cohort of regularly cycling women included. This was involuntary since in none of the original studies, eligibility was limited by age and there was considerable overlap in age ranges of the cohorts. Transvaginal ultrasound was performed using a 6.5-8 mHz probe and our data confirm that this threshold level for FNPO is still valid if using such probe frequencies, although the use of devices with a maximum frequency lower than 8 mHz has become obsolete. Obviously, newer ultrasound scanner using higher transducer frequency will facilitate the

  16. Attaining dominance in vivo: distinguishing dominant from challenger follicles in humans.

    PubMed

    Gore, M A; Nayudu, P L; Vlaisavljevic, V

    1997-12-01

    This study provides the first report of local suppression of follicle growth in vivo by the dominant follicle in humans. Analyses were focused on antral follicles > or =6 mm in diameter, classified as dominant (ovulatory) and challenger (large, non-ovulatory) follicles. We examined antral follicle characteristics, interactions and location in the ovary in the follicular phase of natural, ovulatory cycles in humans. Individual antral follicles (> or =2 mm) were identified and monitored using a system, developed by the authors, incorporating mapping and computer modelling from ultrasound imaging of the ovary. Results showed that antral follicle development was not restricted to a particular location in the ovary. The apparent randomization of location may provide a developmental advantage for follicles. The population of challenger follicles peaked at mid follicular phase, suggesting a period when the ovarian environment was supportive of larger antral follicle development. As ovulation date approached, however, challenger follicles did not attain the visible characteristics of dominance: mid-level echogenicity, large size, smooth antral wall, round shape. Furthermore, dominant follicles subsequently appeared to reduce the number of neighbouring challenger follicles. The results strongly suggest that a paracrine secretion of the dominant follicle was the source of this localized effect. This process may provide increased nutrient level and space at a critical growth stage for dominant follicles.

  17. Investigation of a functional role for Titin in the bovine ovary based on the results of an initial whole genome scan for antral follicle count

    USDA-ARS?s Scientific Manuscript database

    A world-wide food shortage is predicted by the year 2050, and biotechnologies are needed to improve production efficiency in agriculture. Biotechnologies that improve reproductive efficiency in domestic farm species will improve the availability and price of food for the growing world population. ...

  18. The progression of small-follicle reserves in the ovaries of wild African elephants (Loxodonta africana) from puberty to reproductive senescence.

    PubMed

    Stansfield, F J; Nöthling, J O; Allen, W R

    2013-01-01

    This study aimed to determine whether the follicle reserve in the ovary of the African elephant declines progressively after puberty and whether its depletion constrains the fertility of older females. Elephant ovaries were fixed in 4% neutral buffered formalin and small-follicle counts made using stereological protocols. Excepting a slight rise in small-follicle numbers between 16 and 25 years of age, there was a trend for follicle numbers to fall from puberty to 70 years. Reproductive status did not impact significantly on small-follicle numbers (P=0.31). The number of early primary follicles, initially higher in number than true primary follicles, fell from post-puberty to nil at 45 years of age. Six of the seven oldest animals in the study showed signs of recent ovarian activity in the form of antral follicles, corpora lutea or large corpora nigra. The four oldest elephants (mean age 69 years) had a median small-follicle count of 11,113. In summary, it appears that the elephant ovary is capable of supplying oocytes for ovulation right up to the time of death at the age of maximum life expectancy, although the follicle reserve becomes depleted in some older elephants.

  19. Expression pattern of vascular endothelial growth factor in canine folliculogenesis and its effect on the growth and development of follicles after ovarian organ culture.

    PubMed

    Abdel-Ghani, M A; Shimizu, T; Suzuki, H

    2014-10-01

    In this study, the expressions of VEGF in dog follicles were detected by immunohistochemistry and the effects of VEGF treatment on the primordial to primary follicle transition and on subsequent follicle progression were examined using a dog ovary organ culture system. The frozen-thawed canine ovarian follicles within slices of ovarian cortical tissue were cultured for 7 and 14 days in presence or absence of VEGF. After culture, the ovaries were fixed, sectioned, stained and counted for morphologic analysis. The results showed that VEGF was expressed in the theca cells of antral follicles and in the granulosa cells nearest the oocyte in preantral follicle but not in granulosa cells of primordial and primary follicles; however, the VEGF protein was expressed in CL. After in vitro culture, VEGF caused a decrease in the number of primordial follicles and concomitant increase in the number of primary follicles that showed growth initiation and reached the secondary and preantral stages of development after 7 and 14 days. Follicular viability was also improved in the presence of VEGF after 7 and 14 days in culture. In conclusion, treatment with VEGF was found to promote the activation of primordial follicle development that could provide an alternative approach to stimulate early follicle development in dogs. © 2014 Blackwell Verlag GmbH.

  20. Dysregulation of ovarian follicular development in female rat: LH decreases FSH sensitivity during preantral-early antral transition.

    PubMed

    Orisaka, Makoto; Hattori, Katsushige; Fukuda, Shin; Mizutani, Tetsuya; Miyamoto, Kaoru; Sato, Takashi; Tsang, Benjamin K; Kotsuji, Fumikazu; Yoshida, Yoshio

    2013-08-01

    Several clinical studies have shown a correlation of hypersecretion of LH and polycystic ovary syndrome (PCOS), infertility, and miscarriage in women, suggesting that chronically elevated LH impairs fertility. Growth arrest of small antral follicles in PCOS is also assumed to be associated with an abnormal endocrine environment involving increased LH stimulation, a hyperandrogenic milieu, and subsequent dysregulated FSH action in the ovarian follicles. In this study, we examined whether and how LH modulates follicular development and steroid production during preantral-early antral follicle transition by using a rat preantral follicle culture system. LH augments testosterone and estradiol production in preantral follicles via up-regulating mRNA abundance of CYP17A1 and CYP19A1. LH promotes rat preantral follicle growth, and the follicular size reaches that of early antral follicles in vitro, a response attenuated by the specific androgen receptor antagonist and a targeted disruption of androgen receptor gene. Sustained follicle stimulation by LH, but not by androgen, decreases FSH receptor mRNA levels and FSH receptor signaling and inhibits FSH-induced follicular growth. The data suggest that LH promotes preantral-early antral transition via the increased synthesis and growth-promoting action of androgen. However, chronic LH stimulation impairs FSH-dependent antral follicle growth by suppressing granulosa cell FSHR expression via the modulation of intraovarian regulators, including LH-induced thecal factors.

  1. Impact of age and sex of Rocky Mountain elk (Cervus elaphus nelsoni) on follicle counts from rectal mucosal biopsies for preclinical detection of chronic wasting disease.

    PubMed

    Spraker, Terry R; VerCauteren, Kurt C; Gidlewski, Tom L; Munger, Randy D; Walter, W David; Balachandran, Aru

    2009-11-01

    To determine if the number of rectal lymphoid follicles decreases with respect to age and sex relative to diagnosis of chronic wasting disease (CWD), rectal biopsies (n = 1,361) were taken from captive Rocky Mountain elk (Cervus elaphus nelsoni) at 4 ranches in the western United States between 2005 and 2008. Rectal tissues were stained with a monoclonal antibody (F99/97.6.1), which selectively stains the abnormal isoform of the prion protein associated with CWD of elk. The number of lymphoid follicles obtained from typical biopsy tissues decreased with the age of the animal. The acceptable number of lymphoid follicles for detection of CWD was not considered to be a problem in elk up to 8.5 years of age, but in elk over 8.5 years of age, the follicle count was considered to be low. Sex of the animal had no effect on the number of lymphoid follicles observed in each age group. Rectal biopsies were an accurate test to diagnose preclinical stages of CWD in elk but may be best suited to elk that are less then 8.5 years of age.

  2. Follicular interactions affect the in vitro development of isolated goat preantral follicles.

    PubMed

    Duarte, Ana Beatriz Graça; Chaves, Roberta Nogueira; Araújo, Valdevane Rocha; Celestino, Juliana Jales; Silva, Gerlane Modesto; Lopes, Cláudio Afonso Pinho; Tavares, Líliam Mara Trevisan; Campelo, Cláudio Cabral; de Figueiredo, José Ricardo

    2011-08-01

    The aim of this study was to evaluate the influence of the number of follicles per drop (one or three) and antral follicles on in vitro development of isolated goat preantral follicles. Preantral follicles were isolated through microdissection and distributed individually (control) or in groups of three follicles (treatment) in microdroplets of α-MEM with or without 1000 ng/ml follicle stimulating hormone (FSH) for Experiments 1 and 2, respectively. Experiment 3 was divided into four treatments according to the presence of one or three preantral follicles, associated or not with antral follicles. After culture, oocytes were retrieved from morphologically normal follicles and submitted to in vitro maturation (IVM) and live/dead fluorescent labelling. Results of Experiment 1 (basic medium without FSH) showed that culture of preantral follicles in groups enhances viability, growth and antrum formation after 12 days. However, in the presence of FSH (Experiment 2), only the recovery rate of fully grown oocytes for IVM was significantly affected by grouping of follicles. In Experiment 3, in general, co-culture of preantral follicles with an early antral follicle had a detrimental effect on viability, antrum formation and production of oocytes for IVM. In conclusion, the performance of in vitro culture of goat preantral follicles is affected by the number of follicles per drop, the presence of an antral follicle and FSH.

  3. Sperm counts and serum follicle-stimulating hormone levels before and after radiotherapy and chemotherapy in men with testicular germ cell cancer

    SciTech Connect

    Berthelsen, J.G.

    1984-02-01

    Sperm counts were low (median, 15 X 10(6) per ejaculate) and serum follicle-stimulating hormone (FSH) levels were moderately elevated (median, 31 IU/l) after unilateral orchiectomy and immediately before radiotherapy and chemotherapy in 34 patients with seminomas and 20 patients with nonseminomatous germ cell tumors. The scattered radiation (0.2 to 1.3 Gray (Gy)) reaching the remaining testicle during radiotherapy caused azoospermia in more than two thirds of the patients. A median of 540 days elapsed after the end of treatment before spermatozoa were again found in semen samples, while a median of 1250 days passed before the pretreatment sperm count was reached. One to 5 years after treatment, sperm counts were still low (median, 6 X 10(6) per ejaculate) and serum FSH was elevated (median, 61 IU/l). The adjuvant chemotherapy given to the 20 patients with nonseminomatous tumors did not appear to affect restitution appreciably.

  4. Plasma antimullerian hormone as a predictor of ovarian antral follicular population in Bos indicus (Nelore) and Bos taurus (Holstein) heifers.

    PubMed

    Batista, E O S; Macedo, G G; Sala, R V; Ortolan, M D D V; Sá Filho, M F; Del Valle, T A; Jesus, E F; Lopes, R N V R; Rennó, F P; Baruselli, P S

    2014-06-01

    In Bos taurus cattle, antimullerian hormone (AMH) has been demonstrated to have a high degree of correlation with ovarian antral follicle count and the number of healthy follicles and oocytes. To document the correlation between the plasma concentration of AMH and follicular number in Bos indicus and Bos taurus heifers, Nelore (Bos indicus, n = 16) and Holstein heifers (Bos taurus, n = 16) had their ovarian follicular waves synchronized. After synchronization, ovarian antral follicular population (AFP) was evaluated three times at 60-day (d) intervals (T-120 d, 120 days before plasma AMH determination; T-60 d, 60 days before; and T0, at the time of plasma AMH determination). The plasma AMH concentration was positively correlated with the number of ovarian follicles on the day of the follicular wave emergence in Bos indicus (Nelore) and Bos taurus (Holstein) heifers at each evaluation time (p < 0.05). The AFP was higher in Bos indicus (Nelore) than in Bos taurus (Holstein) heifers (p < 0.05). Similarly, the AMH concentration was higher in Bos indicus (Nelore) than in Bos taurus (Holstein) heifers (p < 0.0001). When heifers were classified as to present high or low AFP according to the mean of the AFP within each genetic group, high-AFP heifers presented a greater (p < 0.0001) AMH concentration than low-AFP heifers, regardless of the genetic group. In conclusion, the AFP is positively correlated with plasma AMH concentration in both Bos indicus (Nelore) and Bos taurus (Holstein) heifers. Furthermore, Bos indicus (Nelore) heifers presented both greater plasma AMH concentrations and AFP than Bos taurus (Holstein) heifers. © 2014 Blackwell Verlag GmbH.

  5. Follicle dynamics and anovulation in polycystic ovary syndrome.

    PubMed

    Franks, Stephen; Stark, Jaroslav; Hardy, Kate

    2008-01-01

    Polycystic ovary syndrome (PCOS) is the commonest cause of anovulatory infertility and menstrual cycle abnormalities, but the factors responsible for failure to select a dominant follicle remain unclear. Source is authors' own studies and search of the relevant literature. Arrest of antral follicle growth is associated with an abnormal endocrine environment involving hypersecretion of luteinizing hormone and insulin (and perhaps hyperandrogenism). The net effect is secondary suppression of FSH, which leads to inhibition of maturation of otherwise healthy follicles in the cohort. There is, however, emerging evidence for an intrinsic abnormality of folliculogenesis in PCOS that affects the very earliest, gonadotrophin independent, stages of follicle development. There is an increased density of small pre-antral follicles and an increased proportion of early growing follicles. These abnormalities in anovulatory PCOS are further defined by abnormal granulosa cell proliferation and disparate growth of oocyte and surrounding granulosa cells. This suggests that the normal 'dialogue' between oocyte and granulosa cells in these early growing follicles is altered. There is evidence that abnormal, local (follicle-to-follicle) signalling of anti-Müllerian hormone may play a part in disordered folliculogenesis, but it is plausible that other local regulators that have been implicated in normal and abnormal pre-antral follicle development-such as insulin-like growth factors and sex steroids-have a role in aberrant folliculogenesis in PCOS. Significant abnormalities in the very earliest stages of folliculogenesis may be the root cause of anovulation in PCOS.

  6. Multiple follicle culture supports primary follicle growth through paracrine-acting signals.

    PubMed

    Hornick, J E; Duncan, F E; Shea, L D; Woodruff, T K

    2013-01-01

    In vitro follicle growth in alginate hydrogels is a unique and versatile method for studying ovarian and follicle biology that may also have implications for fertility preservation. Current culture systems support the development of isolated mouse follicles from the secondary stage onward. However, it has been a challenge to grow smaller follicles in vitro due to the dissociation of the oocyte from companion somatic cells. Recent work has demonstrated that coculturing primary follicles with mouse embryonic fibroblasts or ovarian stromal cells supports follicle survival and growth. In this study, we demonstrate that follicles themselves can exert a beneficial coculture effect. When primary follicles were cultured in groups of five or ten (multiple follicle culture), there was increased growth and survival. The multiple follicle culture approach maintained follicle integrity and resulted in the formation of antral stage follicles containing meiotically competent gametes. The growth and survival of primary follicles were highly number dependent, with the most significant enhancement observed when the largest number of follicles was grown together. Our data suggest that the follicle unit is necessary to produce the secreted factors responsible for the supportive effects of multiple follicle culture, as neither denuded oocytes, oocyte-secreted factors, nor granulosa cells alone were sufficient to support early follicle growth in vitro. Therefore, there may be signaling from both the oocyte and the follicle that enhances growth but requires both components in a feedback mechanism. This work is consistent with current in vivo models for follicle growth and thus advances the movement to recapitulate the ovarian environment in vitro.

  7. Ultrastructure of the basal lamina of bovine ovarian follicles and its relationship to the membrana granulosa.

    PubMed

    Irving-Rodgers, H F; Rodgers, R J

    2000-03-01

    Different morphological phenotypes of follicular basal lamina and of membrana granulosa have been observed. Ten preantral follicles (< 0. 1 mm), and 17 healthy and six atretic antral follicles (0.5-12 mm in diameter) were processed for light and electron microscopy to investigate the relationship the between follicular basal lamina and membrana granulosa. Within each antral follicle, the shape of the basal cells of the membrana granulosa was uniform, and either rounded or columnar. There were equal proportions of follicles follicles had only rounded basal cells. Conventional basal laminae of a single layer adjacent to the basal granulosa cells were observed in healthy follicles at the preantral and antral stages. However, at the preantral stage, the conventional types of basal lamina were enlarged or even partially laminated. A second type of basal lamina, described as 'loopy', occurred in about half the preantral follicles and in half the antral follicles follicles. 'Loopy' basal laminae were composed of basal laminae aligning the basal surface of basal granulosa cells, but with additional layers or loops often branching from the innermost layer. Each loop was usually < 1 microm long and had vesicles (20-30 nm) attached to the inner aspect. Basal cellular processes were also common, and vesicles could be seen budding off from these processes. In antral follicles, conventional basal laminae occurred in follicles with rounded basal granulosa cells. Other follicles with columnar cells, and atretic follicles, had the 'loopy' basal lamina phenotype. Thus, follicles have different basal laminae that relate to the morphology of the membrana granulosa.

  8. Anti-angiogenic VEGFA164B isoform mRNA is more abundant in E2-inactive, atretic follicles while expression of angiogenic VEGFA isoforms is greater in granulosa cells from developing bovine follicles prior to the LH surge

    USDA-ARS?s Scientific Manuscript database

    Vascular endothelial growth factor A (VEGFA) is expressed by granulosa cells of the follicle and if its actions are blocked, ovulation and antral follicle development is inhibited. However, the role of anti-angiogenic VEGFA isoforms in bovine dominant follicle development, especially prior to and a...

  9. Regulators of ovarian preantral follicle development.

    PubMed

    McGee, Elizabeth A; Raj, Renju S

    2015-05-01

    Preantral follicle development has become an increasingly recognized area of study in the last two decades. Factors that regulate the growth survival and differentiation of these small, yet complex structures have been identified. The field of fertility preservation and a need for increased numbers of mature oocytes for stem cell research revealed how little we knew of how follicles got from the primordial stage to the antral stage with a healthy and competent oocyte inside. This work discusses the role of gonadotropins in regulating preantral follicles and also the role of the TGF-β family members and their associated Smad signaling molecules in preantral follicle development. Preantral follicle development is a necessary step to fertility in females and further understanding of this process is essential for progress in both infertility care and the enlarging field of in vitro folliculogenesis.

  10. Assessment of ovarian reserve: Anti-Mullerian hormone versus follicle stimulating hormone

    PubMed Central

    Jamil, Zehra; Fatima, Syeda Sadia; Cheema, Zahra; Baig, Safia; Choudhary, Roha Ahmed

    2016-01-01

    Background: This study aimed to evaluate the strength of anti-Mullerian hormone (AMH) and follicle stimulating hormone (FSH) in reflecting the antral follicle count (AFC) in infertile females. Materials and Methods: This cross-sectional study was conducted on 160 females, visiting infertility clinic for assisted reproduction. Serum samples collected on the 3rd day of the cycle were assayed for FSH, luteinizing hormone, and AMH while AFC was assessed via transvaginal ultrasound. The study cohort was segregated into three groups based on AFC. Results: Chronological age and FSH was significantly high in females with very low AFC (P < 0.01 and 0.009, respectively), yet they failed to discriminate patients with normal and higher follicle count (P = 0.65 and 0.84). Conversely, AMH reported highly significant difference between very low AFC and with those having either normal AFC (P = 0.002) or higher AFC (P = 0.001). Moreover, a significant difference in AMH was observed between normal and higher AFC group (P = 0.04). Conclusion: Compared to female’s age and FSH, AMH is superior in clustering study cohort on the bases of antral follicular pool, especially in setups with nonavailability of technological expertise to assess AFC. Incorporation of AMH along with other biomarkers improves estimation of baseline ovarian reserve, required to standardize dose for optimum response; avoiding the risk of failure to retrieve oocyte or inappropriate stimulation leading to ovarian hyperstimulation syndrome. Further prospective studies are required to ascertain its role in predicting the outcomes of ART in such patients. PMID:28163746

  11. Use of an electronic activity detection system to characterize estrus activity in crossbred beef heifers differing in follicle number

    USDA-ARS?s Scientific Manuscript database

    Increased numbers of antral follicles have been associated with decreased calving day, increased fertility, increased serum estradiol concentrations, increased serum progesterone concentrations, and increased estrus behavior in cattle. In addition, cows with increased fertility have been shown to h...

  12. The magnitude of gonadotoxicity of chemotherapy drugs on ovarian follicles and granulosa cells varies depending upon the category of the drugs and the type of granulosa cells.

    PubMed

    Yuksel, Aytac; Bildik, Gamze; Senbabaoglu, Filiz; Akin, Nazli; Arvas, Macit; Unal, Fehmi; Kilic, Yagmur; Karanfil, Isil; Eryılmaz, Baldan; Yilmaz, Pelin; Ozkanbaş, Can; Taskiran, Cagatay; Aksoy, Senai; Guzel, Yılmaz; Balaban, Basak; Ince, Umit; Iwase, Akira; Urman, Bulent; Oktem, Ozgur

    2015-12-01

    groups of 13 to receive a single IP injection of: saline (control), gemcitabine (200 mg/kg), cisplatin (50 mg/kg) or cyclophosphamide (200 mg/kg). The animals were euthanized 72 h later. Follicle counts and serum AMH levels were compared between the groups. In vitro cytotoxicity studies were performed using mitotic non-luteinized rat (SIGC) and human (COV434, HGrC1) granulosa cells, and non-mitotic luteinized human (HLGC) granulosa cells. The cells were plated at a density of 5000 cells/well using DMEM-F12 culture media supplemented with 10% FBS. Chemotherapy agents were used at their therapeutic blood concentrations. The growth of mitotic granulosa cells was monitored real-time using xCelligence system. Live/dead cell and apoptosis assays were also carried out using intravital Yo-Pro-1 staining and cleaved caspase-3 expression, respectively. Estradiol (E2), progesterone (P) and anti-Mullerian hormone (AMH) levels were assayed with ELISA. Cyclophosphamide and cisplatin caused massive atresia of both primordials and growing follicles in the rat ovary whereas gemcitabine impacted pre-antral/antral follicles only. Cyclophosphamide and cisplatin induced apoptosis of both mitotic non-luteinized and non-mitotic luteinized granulosa cells in vitro. By contrast, cytotoxicity of gemcitabine was confined to mitotic non-luteinized granulosa cells. This study tested only three chemotherapeutic agents. The experimental methodology described here could be applied to other drugs for detailed analysis of their ovarian cytotoxicity. These findings indicate that in vivo and in vitro cytotoxic actions of chemotherapy drugs on the ovarian follicles and granulosa cells vary depending upon the their mechanism of action and the nature of the granulosa cells. © The Author 2015. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  13. Presence of growth hormone receptor (GH-R) mRNA and protein in goat ovarian follicles and improvement of in vitro preantral follicle survival and development with GH.

    PubMed

    Martins, F S; Saraiva, M V A; Magalhães-Padilha, D M; Almeida, A P; Celestino, J J H; Padilha, R T; Cunha, R M S; Silva, J R V; Campello, C C; Figueiredo, J R

    2014-07-01

    This study aimed to demonstrate the expression of growth hormone receptor (GH-R) mRNA and protein in goat ovarian follicles in order to investigate the effects of GH on the survival and development of preantral follicles. The ovaries were processed for the isolation of follicles to study GH-R mRNA expression or to localization of GH-R by immunohistochemical analysis. Pieces of ovarian cortex were cultured for 7 days in minimum essential medium(+) (MEM(+)) in the presence or absence of GH at different concentrations (1, 10, 50, 100, and 200 ng/mL). High expression levels of GH-R mRNA were observed in granulosa/theca cells from large antral follicles. However, preantral follicles do not express mRNA for GH-R. Immunohistochemistry demonstrated that the GH-R protein was expressed in the oocytes/granulosa cells of antral follicles, but any protein expression was observed in preantral follicles. The highest (P < 0.05) rate of normal follicles and intermediate follicles was observed after 7 days in MEM(+) plus 10 ng/mL GH (70%). In conclusion, GH-R mRNA and protein are expressed in caprine antral follicles, but not in preantral follicles. Moreover, GH maintains the survival of goat preantral follicles and promotes the development of primordial follicles. Copyright © 2014 Elsevier Inc. All rights reserved.

  14. Hormonal regulation of early follicle development in the rat ovary.

    PubMed

    Hsueh, A J; McGee, E A; Hayashi, M; Hsu, S Y

    2000-05-25

    Although earlier studies focused on the hormonal regulation of antral and preovulatory follicles, recent studies indicate the importance of the hormonal control mechanism for preantral follicles. The endocrine hormone FSH is not only a survival factor for early antral follicles but also a potent growth and differentiation factor for preantral follicles. In addition, KGF secreted by theca cells and c-kit ligand secreted by granulosa cells play paracrine roles in the regulation of preantral follicle growth and development. Furthermore oocyte-derived GDF-9 promotes the growth and differentiation of early follicles by acting on somatic cells in the follicle. It is likely that the genetic makeup of an oocyte could determine the secretion of oocyte hormones which would, in turn, regulate the growth and differentiation of the surrounding somatic cells of that follicle. A better understanding of the hormonal mechanisms underlying early follicle development could provide a refined culture system for the in vitro maturation of fertilizable oocytes and future design of fertility and contraceptive agents.

  15. The presence of VEGF and Notch2 during preantral-antral follicular transition in infantile rats: Anatomical evidence and its implications.

    PubMed

    Torres-Ortiz, Minerva Carolina; Gutiérrez-Ospina, Gabriel; Gómez-Chavarín, Margarita; Murcia, Clara; Alonso-Morales, Rogelio A; Perera-Marín, Gerardo

    2017-08-01

    Folliculogenesis is a process that depends on angiogenesis, in which VEGF and Notch signaling pathway members are involved. Although this pathway is present in preantral and antral follicular structures during the second stage of folliculogenesis, this association has not been described. Therefore, this study aimed to identify VEGF and Notch2 in ovary structures of infantile rats after induction of follicular development with a gonadotropin stimulus. In order to explore this possibility we analyzed rat ovary morphology from days 10-25 after birth; subsequently, the transition from preantral follicle to an antral stage was analyzed by the induction of follicular development with equine chorionic gonadotropin (eCG) and VEGF and Notch were identified in the rat ovary by fluorescence. The histological analysis revealed that the ovary of a 10-day-old rat has the highest percentage of preantral follicles and based on this a 10IU eCG dose promoted an increase in the number of antral follicles, as well as a decrease in the number of preantral follicles, related to which there was an increase in ovary weight and size. In addition, a higher concentration of circulating estradiol was observed, proliferation of granulosa cells in both follicle groups was stimulated, and the accumulation of VEGF in granulosa and theca cells and in the antral follicle oocyte was increased (p<0.05), whereas the presence of Notch2 was limited to mural granulosa cells, in granulosa cells that formed the cumulus oophorus and in the oocyte of both groups of follicles. The multiple correspondence analysis allowed us to support an association between VEGF and Notch2 during the transition from preantral to antral follicles in the ovary of an infantile rat. Copyright © 2017 Elsevier Inc. All rights reserved.

  16. Ovarian follicle vascularization in fasted pig.

    PubMed

    Barboni, Barbara; Barbara, Barboni; Martelli, Alessandra; Alessandra, Martelli; Berardinelli, Paolo; Paolo, Berardinelli; Russo, Valentina; Valentina, Russo; Turriani, Maura; Maura, Turriani; Bernabò, Nicola; Nicola, Bernabò; Lucidi, Pia; Pia, Lucidi; Mattioli, Mauro; Mauro, Mattioli

    2004-09-01

    The authors have investigated in the different classes of ovarian follicles the vascular area, the blood vessel distribution, the vascular endothelial growth factor (VEGF) mRNA expression and the VEGF secretion during equine chorionic gonadotropin (eCG) induced follicle growth in prepubertal gilts fed ad libitum or fasted. Immunohistochemistry staining of Von Willebrand factor showed that fasting caused a dramatic increase in the vascular area of medium-large tertiary follicles. The increase involved the two concentric vessel networks and the area between them that, becoming crossed by several anastomosis, modified the whole vessel architecture. Both in situ hybridization and in vitro culture experiments demonstrate that granulosa cells from medium-large follicles are engaged in a copious VEGF production upon eCG stimulation both in gilts fed ad libitum or fasted. More surprisingly, the production of VEGF becomes diffuse amongst theca cells of fasted animals thus recruiting a compartment that in condition of normal feeding regimen appears nearly quiescent. In conclusion, the data presented describe a local angiogenic process that develops in the follicle wall of growing antral follicle in case of acute severe food restriction. The mechanism, essentially confined to follicles that potentially approach ovulation, appears to assume the meaning of a local compensatory mechanism that may help maintaining adequate nutrient delivery to follicles that undergo ovulation.

  17. Ultrastructural localisation of calcium deposits in pig ovarian follicles.

    PubMed

    Rozinek, J; Rajmon, R; Petr, J; Rohlík, J; Jeseta, M; Sedmíková, M; Rehák, D; Jílek, F

    2006-01-01

    Calcium intracellular signaling regulates many intracellular events including oocyte maturation. This signaling is strongly dependent on the influx of calcium ions from extracellular spaces and on the state of intracellular calcium stores. In this study, intracellular calcium deposits were detected in follicle-enclosed pig oocytes using the combined oxalate-pyroantimonate method. These deposits were observed in the nucleus, the mitochondria, the cytoplasm, and on the surface of lipid droplets. The amount of calcium deposits was expressed as a percentage of the area of the respective cellular compartment, which is covered with calcium deposits on ultrathin sections. The distribution of calcium deposits in oocytes changed during folliculogenesis. The amount of calcium deposits in nuclei (1.11% of the area of oocyte nuclei) and cytoplasm (1.02%) in oocytes from secondary and early antral follicles (0.90% nuclei; 0.99% cytoplasm) is significantly lower (P < 0.05) than the amount of calcium deposits in these compartments in oocytes from primary follicles (2.51% nuclei; 2.34% cytoplasm) or antral follicles with growing oocyte (2.91% nuclei; 2.21% cytoplasm). The amount of calcium deposits in mitochondria of oocytes from primary follicles (1.27%) or antral follicles with growing oocyte (1.14%) is significantly lower (P < 0.05) than in the nucleus (2.51% in oocytes from primary follicles; 2.91% in growing oocytes from antral follicles) or cytoplasm (2.34% in oocytes from primary follicles; 2.21% in growing oocytes from antral follicles). The amount of calcium deposits in the cytoplasm of fully-grown oocytes (1.46%) dropped to levels significantly lower (P < 0.05) than those observed in the oocyte nucleus (2.29%). On the basis of these data, we can conclude that the population of follicles on pig ovaries differs in the distribution and concentration of calcium deposits in oocytes, and these changes may be involved in the regulation of the meiotic competence of oocytes.

  18. Inhibin in individual buffalo ovarian follicles in relation to size.

    PubMed

    Palta, P; Prakash, B S; Manik, R S; Madan, M L

    1996-06-01

    A sensitive and specific radioimmunoassay was validated and applied for measurement of inhibin in follicular fluid (bFF) obtained from individual buffalo ovarian follicles. Follicular size was measured with an ultrasound machine and follicles were categorized as small, medium and large. Presence of inhibin was detected in all the antral follicles above 3 mm diameter. Inhibin concentration showed a positive relationship (R = 0.27, P < 0.01) with follicular diameter and was significantly higher (P < 0.05) in bFF from medium and large follicles (8.44 +/- 0.54 and 7.70 +/- 0.45 micrograms/ml, respectively) in comparison to that from small follicles (5.74 +/- 0.80 micrograms/ml). Total inhibin content was highly correlated (R = 0.92, P < 0.001) with follicular diameter and the inhibin content was higher (P < 0.001) in large > medium > small follicles.

  19. The prevalence of lymphoid follicles in Helicobacter pylori associated gastritis in patients with ulcers and non-ulcer dyspepsia.

    PubMed Central

    Zaitoun, A M

    1995-01-01

    AIMS--To determine the prevalence of lymphoid follicles in Helicobacter pylori positive and negative gastritis in antral and body type gastric mucosa in patients with non-ulcer dyspepsia (NUD), duodenal ulcer, or gastric ulcer; to correlate follicle presence with patient age; to evaluate the correlation between the prevalence of lymphoid follicles and active and inactive gastritis and its severity; and to assess the positive predictive value of lymphoid follicle prevalence with respect to H pylori infection. METHODS--Gastric biopsy specimens, graded according to the Sydney system, from 337 patients were studied. RESULTS--Lymphoid follicles occurred more often in antral mucosa (78%) than in body type mucosa (41%) and were observed in 85% of patients with H pylori positive gastritis. There was no significant difference between NUD and gastric and duodenal ulcer disease with regard to the presence of lymphoid follicles. The positive predictive value of the presence of lymphoid follicles in H pylori infection was 96%. Lymphoid follicles were more commonly observed in patients aged between 10 and 29 years. Lymphoid follicles were more frequently found in pangastritis of all subtypes than in antral gastritis and also in active gastritis than in inactive gastritis. The presence of lymphoid follicles correlated strongly with the degree and severity of gastritis. CONCLUSION--Lymphoid follicles are a constant morphological feature of H pylori associated gastritis. Images PMID:7615851

  20. Biology and Biotechnology of Follicle Development

    PubMed Central

    Palma, Gustavo Adolfo; Argañaraz, Martin Eduardo; Barrera, Antonio Daniel; Rodler, Daniela; Mutto, Adrian Ángel; Sinowatz, Fred

    2012-01-01

    Growth and development of ovarian follicles require a series of coordinated events that induce morphological and functional changes within the follicle, leading to cell differentiation and oocyte development. The preantral early antral follicle transition is the stage of follicular development during which gonadotropin dependence is obtained and the progression into growing or atresia of the follicle is made. Follicular growth during this period is tightly regulated by oocyte-granulosatheca cell interactions. A cluster of early expressed genes is required for normal folliculogenesis. Granulosa cell factors stimulate the recruitment of theca cells from cortical stromal cells. Thecal factors promote granulosa cell proliferation and suppress granulosa cell apoptosis. Cell-cell and cell-extracellular matrix interactions influence the production of growth factors in the different follicular compartments (oocyte, granulosa, and theca cells). Several autocrine and paracrine factors are involved in follicular growth and differentiation; their activity is present even at the time of ovulation, decreasing the gap junction communication, and stimulating the theca cell proliferation. In addition, the identification of the factors that promote follicular growth from the preantral stage to the small antral stage may provide important information for the identification for assisted reproduction techniques. PMID:22666170

  1. C-type natriuretic peptide stimulates ovarian follicle development.

    PubMed

    Sato, Yorino; Cheng, Yuan; Kawamura, Kazuhiro; Takae, Seido; Hsueh, Aaron J W

    2012-07-01

    C-type natriuretic peptide (CNP) encoded by the NPPC (Natriuretic Peptide Precursor C) gene expressed in ovarian granulosa cells inhibits oocyte maturation by activating the natriuretic peptide receptor (NPR)B (NPRB) in cumulus cells. RT-PCR analyses indicated increased NPPC and NPRB expression during ovarian development and follicle growth, associated with increases in ovarian CNP peptides in mice. In cultured somatic cells from infantile ovaries and granulosa cells from prepubertal animals, treatment with CNP stimulated cGMP production. Also, treatment of cultured preantral follicles with CNP stimulated follicle growth whereas treatment of cultured ovarian explants from infantile mice with CNP, similar to FSH, increased ovarian weight gain that was associated with the development of primary and early secondary follicles to the late secondary stage. Of interest, treatment with FSH increased levels of NPPC, but not NPRB, transcripts in ovarian explants. In vivo studies further indicated that daily injections of infantile mice with CNP for 4 d promoted ovarian growth, allowing successful ovulation induction by gonadotropins. In prepubertal mice, CNP treatment alone also promoted early antral follicle growth to the preovulatory stage, leading to efficient ovulation induction by LH/human chorionic gonadotropin. Mature oocytes retrieved after CNP treatment could be fertilized in vitro and developed into blastocysts, allowing the delivery of viable offspring. Thus, CNP secreted by growing follicles is capable of stimulating preantral and antral follicle growth. In place of FSH, CNP treatment could provide an alternative therapy for female infertility.

  2. The Value of Automated Follicle Volume Measurements in IVF/ICSI

    PubMed Central

    Vandekerckhove, Frank; Bracke, Victoria; De Sutter, Petra

    2014-01-01

    Background/Aims: The objective of this literature study is to investigate the place of recent software technology sonography-based automated volume count (SonoAVC) for the automatic measurement of follicular volumes in IVF/ICSI. Its advantages and disadvantages and potential future developments are evaluated. Methods: A total of 74 articles were read via a PubMed literature study. The literature study included 53 articles, 32 of which for the systematic review. Results: The SonoAVC software shows excellent accuracy. Comparing the technology with the “golden standard” two-dimensional (2D) manual follicle measurements, SonoAVC leads to a significantly lower intra- and inter-observer variability. However, there is no significant difference in clinical outcome (pregnancy rate). We noted a significant advantage in the time gained, both for doctor and patient. By storing the images, the technology offers the possibility of including a quality control and continuous training and further standardization of follicular monitoring can be expected. Ovarian reserve testing by measuring the antral follicle count with SonoAVC is highly reliable. Conclusion: This overview of previously published literature shows how SonoAVC offers advantages for clinical practice, without losing any accuracy or reliability. Doctors should be motivated to the general use of follicular volumes instead of follicular diameters. PMID:25593942

  3. Expression of follicle-stimulating hormone receptor (FSHR) in goat ovarian follicles and the impact of sequential culture medium on in vitro development of caprine preantral follicles.

    PubMed

    Saraiva, M V A; Celestino, J J H; Araújo, V R; Chaves, R N; Almeida, A P; Lima-Verde, I B; Duarte, A B G; Silva, G M; Martins, F S; Bruno, J B; Matos, M H T; Campello, C C; Silva, J R V; Figueiredo, J R

    2011-08-01

    This study evaluated the expression of FSH receptors (FSHR) in the different stages of goat follicle development and investigated whether the addition of increasing concentrations of FSH throughout the culture period influences the survival, growth and antral formation of in vitro-cultured caprine preantral follicles. The expression of FSHR was analysed before and after culturing follicles using real-time RT-PCR. For the culture, preantral follicles (≥150 μm) were isolated from ovarian fragments and cultured for 18 days in α-MEM+ alone or associated with recombinant FSH (rFSH: 100 or 1000 ng/ml), or in α-MEM+ supplemented with increasing concentrations of FSH throughout culture periods as follows: (a) sequential medium 1: FSH 100 ng/ml (from day 0 to 6), FSH 500 ng/ml (from day 6 to 12) and FSH 1000 ng/ml (from day 12 to 18); and (b) sequential medium 2: FSH 500 ng/ml (from day 0 to 9) and 1000 ng/ml (from day 9 to 18). Follicle development was evaluated on the basis of antral cavity formation, follicular and oocyte growth, and cumulus-oocyte complex health. The expression of FSHR in isolated caprine follicles increased from the preantral to antral phase. Regarding the culture, after 18 days, sequential medium 1 promoted follicular survival, antrum formation and a reduction in oocyte extrusion. Both sequential media promoted a higher rate of meiotic resumption compared with the other treatments. In conclusion, the addition of increased concentrations of FSH (sequential medium) has a significant impact on the in vitro development of caprine preantral follicles.

  4. Analysis of LH receptor in canine ovarian follicles throughout the estrous cycle.

    PubMed

    De Los Reyes, Monica; Palomino, Jaime; Parraguez, Victor H; Ramirez, Fernando

    2017-04-15

    The aim of this study was to determine the mRNA LHR and LHR protein expression pattern in the canine ovarian follicles at different stage of development throughout the estrous cycle. Dog ovaries were obtained from 1-6y bitches at proestrus/estrus, anestrus and diestrus stages following ovariohysterectomy. Follicular cells were mechanically recovered from follicles distributed into four types (preantral, small antral, medium antral and large antral). Total RNA extraction was performed and the evaluation of gene expression levels was achieved by relative quantification q-PCR analysis. Intrafollicular amounts of LHR were assessed by western blot method. All results were evaluated by ANOVA. The expression levels of mRNA LHR in follicular cells were observed in every stage of development, however this gene expression varied over the estrous cycle. LHR transcripts increased (P < 0.05) from preantral to antral stage. There were not differences in LHR gene expression among follicles at preantral stages; however, at antral stages the lowest (P < 0.05) LHR mRNA expression was found at anestrus and the highest (P < 0.05) at proestrus/estrus. The LHR protein was also detected in dog follicles in all reproductive phases with patterns varying with stage of follicular development over the reproductive cycle. The antibody against human LHR revealed two bands at ∼90 and ∼67 kDa, probably representing the matured protein and its precursor respectively. Both bands LHR appeared already at preantral follicles increasing (P < 0.05) with growth. A high proportion of LHR was presented as immature forms in all follicles stages during different phases of the estrous cycle. In conclusion, the gene and protein of LHR are differentially expressed in dog follicles over the estrous cycle, increasing with growth and the precursor protein is the most predominant LHR form present in canine follicles. Copyright © 2017 Elsevier Inc. All rights reserved.

  5. Antral function in duodenal ulcer

    PubMed Central

    Tovey, F. I.; Parker, K.; Swaminathan, M.; Daniell, A.

    1969-01-01

    A dye-dilution technique is used to compare the concentration and output of acid, chloride and pepsin in duodenal ulcer patients and controls following stimulation of the antrum with alcohol, sodium bicarbonate and peptone with the response to insulin hypoglycaemia and maximal histamine stimulation. The mean secretory rate was higher in response to all the stimuli in duodenal ulcer patients except to sodium bicarbonate. Following antral stimulation by peptone and sodium bicarbonate the acid and chloride concentrations rose to very high levels. The mean outputs were the same as those following maximal histamine stimulation although there were wide individual variations. There was no difference between duodenal ulcer patients and controls. The ‘neutral chloride’ concentration was much less than in the other phases. The basal secretion and the secretions in response to insulin and histamine stimulation showed higher concentrations, as well as higher outputs of acid and chloride in duodenal ulcer patients. Pepsin concentration in response to the various stimuli showed no difference between duodenal ulcer patients and controls and the pepsin output reflected the changes in secretory volume. PMID:4892935

  6. Felis catus ovary as a model to study follicle biology in vitro.

    PubMed

    Rojo, Julieta L; Linari, Martina; Musse, Mariana P; Peluffo, Marina C

    2015-07-01

    The current study was designed to evaluate the response of individual intact antral follicles from adult female domestic cats to a luteinizing hormone (LH) stimulus in vitro by assessing cumulus-oocyte expansion (C-OE) and steroid production. C-OE and steroid levels (estradiol [E2] and progesterone [P4]) obtained from individual antral feline follicles (n = 366 follicles; n = 56 cats) were analyzed after 12 or 24 h of culture in the presence or absence of LH (low [3.4 ng/ml] or high [100 ng/ml]). At the end of the culture, the highest percentage of expanded cumulus-oocyte complexes (COCs) was observed in the LH groups at 12 or 24 h in comparison to their controls (p < 0.001). There was a significant increase in expanded COCs when comparing LH concentrations (high vs. low) at 12 or 24 h. Higher levels of both E2 and P4 were observed in the media from antral follicles after 12 and 24 h of culture in the presence of LH (both concentration, p < 0.05). There was no association between hormone levels and follicle diameter; high variability was observed in the steroid levels produced by antral follicles within all treatment groups. These data indicate, for the first time, that feline antral follicles (0.5-2 mm) from different stages of the natural estrous cycle can be cultured and will respond to an LH stimulus, based on an increase in steroid levels as well as C-OE after 12 or 24 h in culture.

  7. The Relationship Between Variation in Size of the Primordial Follicle Pool and Age at Natural Menopause.

    PubMed

    Depmann, M; Faddy, M J; van der Schouw, Y T; Peeters, P H M; Broer, S L; Kelsey, T W; Nelson, S M; Broekmans, F J M

    2015-06-01

    Menopause has been hypothesized to occur when the nongrowing follicle (NGF) number falls below a critical threshold. Age at natural menopause can be predicted using NGF numbers and this threshold. These predictions support the use of ovarian reserve tests, reflective of the ovarian follicle pool, in menopause forecasting. The objective of the study was to investigate the hypothesis that age-specific NGF numbers reflect age at natural menopause. Histologically derived NGF numbers obtained from published literature (n = 218) and distribution of menopausal ages derived from the population-based Prospect-European Prospective Investigation into Cancer and Nutrition (Prospect-EPIC) cohort (n = 4037) were combined. NGF data were from single ovaries that had been obtained postnatally for various reasons, such as elective surgery or autopsy. From the Prospect-EPIC cohort, women aged 58 years and older with a known age at natural menopause were selected. There were no interventions. Conformity between observed age at menopause in the Prospect-EPIC cohort and NGF-predicted age at menopause from a model for age-related NGF decline constructed using a robust regression analysis. A critical threshold for NGF number was estimated by comparing the probability distribution of the age at which the NGF numbers fall below this threshold with the observed distribution of age at natural menopause from the Prospect-EPIC cohort. The distributions of observed age at natural menopause and predicted age at natural menopause showed close conformity. The close conformity observed between NGF-predicted and actual age at natural menopause supports the hypothesis that that the size of the primordial follicle pool is an important determinant for the length of the individual ovarian life span and supports the concept of menopause prediction using ovarian reserve tests, such as anti-Müllerian hormone and antral follicle count, as derivatives of the true ovarian reserve.

  8. The Mare Model to Study the Effects of Ovarian Dynamics on Preantral Follicle Features

    PubMed Central

    Alves, Kele A.; Alves, Benner G.; Gastal, Gustavo D. A.; de Tarso, Saulo G. S.; Gastal, Melba O.; Figueiredo, José R.; Gambarini, Maria L.; Gastal, Eduardo L.

    2016-01-01

    Ovarian tissue collected by biopsy procedures allows the performance of many studies with clinical applications in the field of female fertility preservation. The aim of the present study was to investigate the influence of reproductive phase (anestrous vs. diestrous) and ovarian structures (antral follicles and corpus luteum) on the quality, class distribution, number, and density of preantral follicles, and stromal cell density. Ovarian fragments were harvested by biopsy pick-up procedures from mares and submitted to histological analysis. The mean preantral follicle and ovarian stromal cell densities were greater in the diestrous phase and a positive correlation of stromal cell density with the number and density of preantral follicles was observed. The mean area (mm2) of ovarian structures increased in the diestrous phase and had positive correlations with number of preantral follicles, follicle density, and stromal cell density. Biopsy fragments collected from ovaries containing an active corpus luteum had a higher follicle density, stromal cell density, and proportion of normal preantral follicles. In conclusion, our results showed: (1) the diestrous phase influenced positively the preantral follicle quality, class distribution, and follicle and stromal cell densities; (2) the area of ovarian structures was positively correlated with the follicle and stromal cell densities; and (3) the presence of an active corpus luteum had a positive effect on the quality of preantral follicles, and follicle and stromal densities. Therefore, herein we demonstrate that the presence of key ovarian structures favors the harvest of ovarian fragments containing an appropriate number of healthy preantral follicles. PMID:26900687

  9. Changes in gonadotrophin secretion and ovarian antral follicular activity in seasonally breeding sheep throughout the year.

    PubMed

    McNatty, K P; Hudson, N L; Henderson, K M; Lun, S; Heath, D A; Gibb, M; Ball, K; McDiarmid, J M; Thurley, D C

    1984-01-01

    Overall, significantly more antral follicles greater than or equal to 1 mm diameter were present in Romney ewes during anoestrus than in the breeding season (anoestrus, 35 +/- 3 (mean +/- s.e.m.) follicles per ewe, 23 sheep; Day 9-10 of oestrous cycle, 24 +/- 1 follicles per ewe, 22 sheep; P less than 0.01), although the mean numbers of preovulatory-sized follicles (greater than or equal to 5 mm diam.) were similar (anoestrus, 1.3 +/- 0.2 per ewe; oestrous cycle, 1.0 +/- 0.1 per ewe). The ability of ovarian follicles to synthesize oestradiol did not differ between anoestrus and the breeding season as assessed from the levels of extant aromatase enzyme activity in granulosa cells and steroid concentrations in follicular fluid. Although the mean plasma concentration of LH did not differ between anoestrus and the luteal phase of the breeding season, the pattern of LH secretion differed markedly; on Day 9-10 of the oestrous cycle there were significantly more (P less than 0.001) high-amplitude LH peaks (i.e. greater than or equal to 1 ng/ml) in plasma and significantly fewer (P less than 0.001) low amplitude peaks (less than 1 ng/ml) than in anoestrous ewes. Moreover, the mean concentrations of FSH and prolactin were significantly lower during the luteal phase of the cycle than during anoestrus (FSH, P less than 0.05, prolactin, P less than 0.001). It is concluded that, in Romney ewes, the levels of antral follicular activity change throughout the year in synchrony with the circannual patterns of prolactin and day-length. Also, these data support the notion that anovulation during seasonal anoestrus is due to a reduced frequency of high-amplitude LH discharges from the pituitary gland.

  10. Gonadotropin-binding sites in the rhesus monkey ovary: role of the vasculature in the selective distribution of human chorionic gonadotropin to the preovulatory follicle

    SciTech Connect

    Zeleznik, A.J.; Schuler, H.M.; Reichert, L.E. Jr.

    1981-08-01

    These experiments were initiated to determine if differences exist in the vasculature of individual follicles in the rhesus monkey ovary during the late follicular phase of the menstrual cycle and to determine whether differences in vascularity result in differential exposure of certain follicles to gonadotropic hormones. The density of blood vessels within the thecal layer of the dominant follicle and other antral follicles was determined in ovaries from four animals removed on day 9 or 10 of the menstrual cycle. Blood vessels were identified using a histochemical stain for hemoglobin. Morphometric analysis indicated that the percentage of the thecal layer occupied by blood vessels in the dominant follicles (48%) was significantly greater (P less than 0.005) than that of other smaller antral follicles either within the same ovary as the dominant follicle (24%) or in the contralateral ovary (26%). To determine if differences in vascularity result in a differential supply of gonadotropins to the dominant follicle, we studied, by autoradiography, the in vivo and in vitro binding of (125I)hCG in four rhesus monkeys on day 9 of the menstrual cycle. Results of in vitro binding studies indicated that the thecal layer of virtually every antral follicle possessed hCG-binding sites. However, when (125I)hcg was injected iv into animals and allowed to distribute via the vasculature, the dominant follicle was heavily labeled while other smaller antral follicles accumulated little, if any, radioiodinated hCG. These observations indicate that increased vascularization of individual follicles results in preferential delivery of gonadotropins, and suggest that blood flow to individual follicles may play an instrumental role in the selective maturation of the preovulatory follicle in the rhesus monkey.

  11. Diagnosis of polycystic ovary syndrome (PCOS): revisiting the threshold values of follicle count on ultrasound and of the serum AMH level for the definition of polycystic ovaries.

    PubMed

    Dewailly, D; Gronier, H; Poncelet, E; Robin, G; Leroy, M; Pigny, P; Duhamel, A; Catteau-Jonard, S

    2011-11-01

    Polycystic ovarian morphology (PCOM) at ultrasound is currently used in the diagnosis of polycystic ovary syndrome (PCOS). We hypothesized that the previously proposed threshold value of 12 as an excessive number of follicles per ovary (FN) is no longer appropriate because of current technological developments. In this study, we have revisited the thresholds for FN and for the serum Anti-Müllerian hormone (AMH) level (a possible surrogate for FN) for the definition of PCOM. Clinical, hormonal and ultrasound data were consecutively recorded in 240 patients referred to our department between 2008 and 2010 for exploration of hyperandrogenism (HA), menstrual disorders and/or infertility. According to only their symptoms, patients were grouped as: non-PCOS without HA and with ovulatory cycles (group 1, n = 105), presumption of PCOS with only HA or only oligo-anovulation (group 2, n = 73) and PCOS with HA and oligo-anovulation (group 3, n = 62). By cluster analysis using androgens, LH, FSH, AMH, FN and ovarian volume, group 1 appeared to be constituted of two homogeneous clusters, most likely a non-PCOM non-PCOS subgroup (n = 66) and a PCOM, non-PCOS (i.e. asymptomatic) subgroup (n = 39). Receiver operating characteristic curve analysis was applied to distinguish the non-PCOM non-PCO members of group 1 and to group 3. For FN and serum AMH respectively, the areas under the curve were 0.949 and 0.973 and the best compromise between sensitivity (81 and 92%) and specificity (92 and 97%) was obtained with a threshold values of 19 follicles and 35 pmol/l (5 ng/ml). For the definition of PCOM, the former threshold of >12 for FN is no longer valid. A serum AMH >35 pmol/l (or >5 ng/ml) appears to be more sensitive and specific than a FN >19 and should be therefore included in the current diagnostic classifications for PCOS.

  12. Levels of BMP-6 mRNA in goat ovarian follicles and in vitro effects of BMP-6 on secondary follicle development.

    PubMed

    Frota, Isana M A; Leitão, Cintia C F; Costa, José J N; van den Hurk, Robert; Saraiva, Márcia V A; Figueiredo, José R; Silva, José R V

    2013-08-01

    Expression of BMP-6 mRNA was quantified by real-time polymerase chain reaction (PCR) and the BMP-6 protein was demonstrated by immunohistochemistry in the primordial, primary, secondary, small and large antral follicles of goat. Furthermore, the influence of BMP-6 on increase in diameter, antrum formation and expression of BMP-6 and FSH-R in in vitro cultured secondary follicles was studied. Therefore, goat primordial, primary and secondary follicles, as well as small and large antral follicles were obtained and the mRNA levels of BMP-6 were quantified by PCR in real time. Expression of BMP-6 protein in goat follicles was demonstrated by immunohistochemistry. The influence of BMP-6 in the presence or absence of follicle-stimulating hormone (FSH) on both the development of secondary follicles and the expression of mRNA for BMP-6 and FSH-R was evaluated after 6 days of culture. Furthermore, the follicular diameter and the formation of the antrum were evaluated before and after 6 days of culture and compared by Kruskal-Wallis and chi-squared tests (P < 0.05), respectively. The results show that the level of mRNA for BMP-6 in primary and secondary follicles was significantly higher than in the primordial follicles (P < 0.05). Similar levels of BMP-6 mRNA were observed in cumulus-oocyte complexes and mural granulosa/theca cells from small and large antral follicles, respectively. BMP-6 protein was expressed in oocytes of all categories of follicles and in granulosa cells from secondary follicles onwards. Addition of BMP-6 to the culture medium increased the diameter of secondary follicles mainly by antrum formation after 6 days' culture, in the presence or absence of FSH (P < 0.05). Furthermore, addition of FSH resulted in increased levels of BMP-6 mRNA in these follicles (P < 0.05). Simultaneous administration of FSH and BMP-6 enhanced the levels of FSH receptor (FSH-R) mRNA (P < 0.05). It is concluded that BMP-6 mRNA is increased during transition from primordial to

  13. Fibrin promotes development and function of macaque primary follicles during encapsulated three-dimensional culture.

    PubMed

    Xu, J; Lawson, M S; Yeoman, R R; Molskness, T A; Ting, A Y; Stouffer, R L; Zelinski, M B

    2013-08-01

    Does fibrin introduced into the extracellular matrix affect the growth and maturation of individual primate follicles during encapsulated three-dimensional (3D) culture? While not altering follicle survival, fibrin-alginate (FIBRIN) improves macaque primary, but not secondary, follicle development during encapsulated 3D culture in terms of growth, steroidogenesis, anti-Müllerian hormone (AMH)/vascular endothelial growth factor (VEGF) production and oocyte maturation. Efforts to grow non-human primate ovarian follicles from the secondary to the antral stage during encapsulated 3D culture have been successful. However, the growth and maturation of primary follicles in vitro has not been reported in primates, especially in chemically defined conditions. In vitro follicle maturation was investigated using the rhesus macaque (Macaca mulatta). Ovaries (n = 7 pairs) were obtained during the early follicular phase of the menstrual cycle (cycle day 1-4). Primary (80-120 µm diameter) and secondary (125-225 µm diameter) follicles were isolated mechanically, randomly assigned to experimental groups, encapsulated into alginate (0.25% w/v) or FIBRIN (25 mg/ml fibrinogen-0.25% alginate) and cultured for 13 and 5 weeks, respectively. Individual follicles were cultured in alpha minimum essential medium supplemented with FSH. Follicle survival and growth were assessed by microscopy. Follicles that reached the antral stage were treated with recombinant hCG. Metaphase II (MII) oocytes were inseminated via ICSI. Follicle morphology was evaluated by hematoxylin and eosin (H&E) staining. Immunohistochemistry was performed for cytochrome P450 family 17 subfamily A polypeptide 1 (CYP17A1) and 19 subfamily A polypeptide 1 (CYP19A1). Culture medium was analyzed for estradiol (E2) and progesterone by chemiluminescence, androstenedione (A4) by radioimmunoassay, as well as anti-Müllerian hormone (AMH) and vascular endothelial growth factor (VEGF) by enzyme-linked immunosorbent assay. A

  14. Quantitative expression of antiapoptotic and proapoptotic genes in sheep ovarian follicles grown in vivo or cultured in vitro.

    PubMed

    Praveen Chakravarthi, V; Kona, S S R; Siva Kumar, A V N; Bhaskar, M; Rao, V H

    2015-03-01

    To test the hypothesis that the poor development of the oocytes in cultured ovarian follicles of mammals is due to aberrant expression of developmentally important genes, quantitative expression patterns of Bcl2 (B-cell leukemia/lymphoma 2; antiapoptotic) and Bax (Bcl2-associated X protein; proapoptotic) genes in preantral, early antral, antral, large antral follicles, and cumulus-oocyte complexes (COCs) grown in vivo or cultured in vitro were studied. The level and pattern of expression of Bcl2 in the cumulus cells isolated from different development stages of in vivo- and in vitro-grown ovarian follicles were similar suggesting that in vitro culture did not alter the expression of this antiapoptotic gene in the cumulus cells. However, between the in vivo- and in vitro-grown ovarian follicles (1) Bcl2 expression levels in the oocytes from antral follicles (2.21 ± 0.14 vs. 0.87 ± 0.19), large antral follicles (0 ± 0.35 vs. 1.56 ± 0.13), and COCs (0.45 ± 0.31 vs. 2.69 ± 0.15), Bax expression levels in the (2) cumulus cells from early antral (2.09 ± 0.11 vs. 0.98 ± 0.13) and large antral follicle (0.63 ± 0.44 vs. 0 ± 0.21), and (3) oocytes from antral follicles (1.65 ± 0.20 vs. 0.97 ± 0.15), large antral follicles (0.93 ± 0.18 vs. 2.08 ± 0.11), and COCs (1.03 ± 0.17 vs. 2.09 ± 0.11) were significantly different (P ≤ 0.05). Similarly, Bcl2 to Bax ratios were also significantly different between some but not all stages of in vivo and in vitro development. From the present results, it is concluded that imbalance in the expression of proapoptotic and antiapoptotic genes may be an important cause for the compromised development potential of the oocytes in cultured ovarian follicles of sheep. Copyright © 2015 Elsevier Inc. All rights reserved.

  15. [Dynamics of the growth and maturation of oocytes and follicles in the ovaries of the cow].

    PubMed

    Kurilo, L F; Zelenina, I A; Skhirtladze, Z Sh; Tepliakova, N P

    1986-01-01

    Chromosome despiralization and nucleolus vacuolization have been studied during the oocyte intensive growth. Oocyte and nucleolus growth has been found to stop at the secondary antral follicles with the diameter more than 1000 mkm. Chromosomal and nucleolar activity decreases at this stage. Chromosomes condense and concentrate around the nucleolus and chromatine mass (karyosphere) forms.

  16. Secondary follicle growth and oocyte maturation during encapsulated three-dimensional culture in rhesus monkeys: effects of gonadotrophins, oxygen and fetuin.

    PubMed

    Xu, J; Lawson, M S; Yeoman, R R; Pau, K Y; Barrett, S L; Zelinski, M B; Stouffer, R L

    2011-05-01

    An alginate-based matrix supports the three-dimensional (3D) architecture of non-human primate follicles and, in the presence of FSH, permits the in vitro development of pre-antral follicles to the small antral stage, including the production of ovarian steroids and paracrine factors. The current study investigated the ability of gonadotrophins, fetuin and oxygen (O₂) to improve primate follicle growth and oocyte maturation in vitro. Macaque secondary follicles were isolated from the early follicular phase ovaries, encapsulated in a sodium alginate matrix and cultured individually for 40 days in supplemented medium. The effects of recombinant human (rh) FSH (15, 3 and 0.3 ng/ml for high, medium and low FSH, respectively), bovine fetuin (1 or 0 mg/ml) and O₂ (5 or 20% v/v) were examined. Half of the follicles in each culture condition received rhLH on Day 30-40. Follicles that reached antral stage were treated with rh chorionic gonadotrophin for 34 h to initiate oocyte meiotic maturation. Media were analyzed for ovarian steroids and anti-müllerian hormone (AMH). Improved culture conditions supported non-human primate, secondary follicle growth to the antral stage and, for the first time, promoted oocyte maturation to the MII stage. In the presence of fetuin at 5% O₂, follicles had the highest survival rate if cultured with high or medium FSH, whereas follicles grew to larger diameters at Week 5 in low FSH. Oocyte health and maturation were promoted under 5% O₂. High FSH stimulated steroid production by growing follicles, and steroidogenesis by follicles cultured with low FSH was promoted by LH. AMH biosynthesis was elevated with high compared with low FSH and for longer under 5% O₂ than under 20% O₂. This encapsulated 3D culture model permits further studies on the endocrine and local factors that influence primate follicle growth and oocyte maturation, with relevance to enhancing fertility preservation options in women.

  17. Morphometry and immunohistochemistry of follicles growth and steroidogenesis in saharian wild sand rat, Psammomys obesus, ovary.

    PubMed

    Boubekri, Amina; Gernigon-Spychalowicz, Thérese; Khammar, Farida; Exbrayat, Jean-Marie

    2009-01-01

    The sand rat (Psammomys obesus) constitutes a model to study seasonal changes and several metabolic disorders. In order to perform breeding laboratory conditions, the reproductive function of this species living in North Occidental Algerian Sahara was studied. The aim of this work was to investigate the follicular growth changes and the steroidogenic associated aspects. The study was performed using morphometrical and immunohistochemical methods. From primordial to preantral states, the follicle diameter increased progressively from 17-20 mum to 192-225 mum. The preovulatory follicles reached about 500 mum in diameter. Immunoreactivity to progesterone, androstenediol and estradiol, varied in the different parts of the ovary and follicular cells. The progesterone antibody appeared clearly labelled in the theca interna of the growing follicle and increased in the granulosa; the androgen antibody was continuously weak and diffuses in all follicles; the estradiol labelling appeared weak and diffuse in preantral follicles then increased in antral follicles in both theca and granulosa or only in granulosa. In antral follicles, estradiol label was clearly localized in granulosa cells and totally devoid in theca cells. In Psammomys ovary, labels of hormone were diffuse or localized, weak or intense in the theca and or in the granulosa according to the follicle size.

  18. Correlations among antral follicular echotexture, apoptosis and expression of key steroidogenic enzymes in sheep

    PubMed Central

    VANDUZER, Taylor; DUGGAVATHI, Raj; MURAWSKI, Maciej; ZIEBA, Dorota A.; SROKA, Patrycja; BARTLEWSKI, Pawel M.

    2014-01-01

    Nineteen cycling ewes underwent transrectal ultrasonography of ovaries followed by ovariectomies during the growth phase of the first follicular wave of the interovulatory interval or the proestrus/estrus phase of the cycle. Quantitative ultrasonographic characteristics of the antrum and follicular wall in a total of forty-three ovine antral follicles were examined for correlations with the protein expression of three steroidogenic enzymes (cytochrome P450 17α-hydroxylase, CYP17; cytochrome P450 aromatase, CYP19; and 3β-hydroxysteroid dehydrogenase, 3β-HSD) determined by densitometric analysis of immunohistochemical slides, follicular dimensions, granulosa layer thickness and the percentage of apoptotic granulosa cells. Significant correlations were found between echotextural attributes of ovine antral follicles and the percentage of apoptotic granulosa cells, CYP17 expression (theca), CYP19 expression (granulosa) and 3β-HSD expression (theca cells). Computer-aided analyses of ultrasonographic images can be beneficial to the development of assisted reproductive technologies and diagnosis of hormonal imbalances without the need for ovarian biopsies or hormone assays. PMID:25109269

  19. Notch Signaling Regulates Ovarian Follicle Formation and Coordinates Follicular Growth

    PubMed Central

    Vanorny, Dallas A.; Prasasya, Rexxi D.; Chalpe, Abha J.; Kilen, Signe M.

    2014-01-01

    Ovarian follicles form through a process in which somatic pregranulosa cells encapsulate individual germ cells from germ cell syncytia. Complementary expression of the Notch ligand, Jagged1, in germ cells and the Notch receptor, Notch2, in pregranulosa cells suggests a role for Notch signaling in mediating cellular interactions during follicle assembly. Using a Notch reporter mouse, we demonstrate that Notch signaling is active within somatic cells of the embryonic ovary, and these cells undergo dramatic reorganization during follicle histogenesis. This coincides with a significant increase in the expression of the ligands, Jagged1 and Jagged2; the receptor, Notch2; and the target genes, Hes1 and Hey2. Histological examination of ovaries from mice with conditional deletion of Jagged1 within germ cells (J1 knockout [J1KO]) or Notch2 within granulosa cells (N2 knockout [N2KO]) reveals changes in follicle dynamics, including perturbations in the primordial follicle pool and antral follicle development. J1KO and N2KO ovaries also contain multi-oocytic follicles, which represent a failure to resolve germ cell syncytia, and follicles with enlarged oocytes but lacking somatic cell growth, signifying a potential role of Notch signaling in follicle activation and the coordination of follicle development. We also observed decreased cell proliferation and increased apoptosis in the somatic cells of both conditional knockout lines. As a consequence of these defects, J1KO female mice are subfertile; however, N2KO female mice remain fertile. This study demonstrates important functions for Jagged1 and Notch2 in the resolution of germ cell syncytia and the coordination of somatic and germ cell growth within follicles of the mouse ovary. PMID:24552588

  20. Glycomic analyses of ovarian follicles during development and atresia

    PubMed Central

    Hatzirodos, Nicholas; Nigro, Julie; Irving-Rodgers, Helen F.; Vashi, Aditya V.; Hummitzsch, Katja; Caterson, Bruce; Sullivan, Thomas R.; Rodgers, Raymond J.

    2012-01-01

    To examine the detailed composition of glycosaminoglycans during bovine ovarian follicular development and atresia, the specialized stromal theca layers were separated from the stratified epithelial granulosa cells of healthy (n = 6) and atretic (n = 6) follicles in each of three size ranges: small (3–5 mm), medium (6-9 mm) and large (10 mm or more) (n = 29 animals). Fluorophore-assisted carbohydrate electrophoresis analyses (on a per cell basis) and immunohistochemistry (n = 14) were undertaken. We identified the major disaccharides in thecal layers and the membrana granulosa as chondroitin sulfate-derived ∆uronic acid with 4-sulfated N-acetylgalactosamine and ∆uronic acid with 6-sulfated N-acetylgalactosamine and the heparan sulfate-derived Δuronic acid with N-acetlyglucosamine, with elevated levels in the thecal layers. Increasing follicle size and atresia was associated with increased levels of some disaccharides. We concluded that versican contains 4-sulfated N-acetylgalactosamine and it is the predominant 4-sulfated N-acetylgalactosamine proteoglycan in antral follicles. At least one other non- or 6-sulfated N-acetylgalactosamine proteoglycan(s), which is not decorin or an inter-α-trypsin inhibitor family member, is present in bovine antral follicles and associated with hitherto unknown groups of cells around some larger blood vessels. These areas stained positively for chondroitin/dermatan sulfate epitopes [antibodies 7D4, 3C5, and 4C3], similar to stem cell niches observed in other tissues. The sulfation pattern of heparan sulfate glycosaminoglycans appears uniform across follicles of different sizes and in healthy and atretic follicles. The heparan sulfate products detected in the follicles are likely to be associated with perlecan, collagen XVIII or betaglycan. PMID:22057033

  1. The phenotypic diversity in per-follicle anti-Müllerian hormone production in polycystic ovary syndrome.

    PubMed

    Alebić, M Š; Stojanović, N; Duhamel, A; Dewailly, D

    2015-08-01

    Is intrinsic dysregulation of granulosa cells (GC) and consequent increases in the per-follicle production of anti-Müllerian hormone (AMH), correlated with the phenotypic presentation of women with polycystic ovaries? Involvement of intrinsic GC dysregulation in oligo-anovulation associated with polycystic ovary syndrome (PCOS) is likely because among women with PCOS, those with oligo-amenorrhea have higher per-follicle AMH production than those who ovulate normally, irrespective of their androgen and/or metabolic status. Women with PCOS have higher serum AMH level than non-PCOS women due to an increased follicle number and excessive AMH production per follicle, the latter reflecting a putative GC dysfunction that may vary between PCOS phenotypes. This is a retrospective analysis of data collected from 1021 women undergoing infertility evaluation from March 2011 to October 2013. The study included women with polycystic ovarian morphology (PCOM) who met the Rotterdam criteria for PCOS (n = 272), women with PCOM only (n = 168) and controls (n = 581). We used serum AMH to antral follicle count (AFC) ratio (AMH/AFC) as a marker of per-follicle AMH production and checked whether this ratio was associated with the PCOS phenotype and to the menstrual, androgen and metabolic status in women with PCOS, women with PCOM only and in controls. AMH/AFC was significantly higher in oligo-amenorrheic women with PCOS than in eumenorrheic women with PCOS or PCOM (P < 0.001) but also in the latter group compared with controls (P < 0.001) regardless of androgen status. Stepwise discriminant analysis yielded a significant score for the menstrual status with a discriminant power of 26.5% (P < 0.001). This score included AFC, AMH/AFC, waist circumference and LH with partial R(2) of 0.172, 0.042, 0.024 and 0.023, respectively. The AMH to AFC ratio as a surrogate marker for average AMH may be subject to error because follicles below the sensitivity limit of the ultrasonography used may

  2. Adrenomedullin in rat follicles and corpora lutea: expression, functions and interaction with endothelin-1

    PubMed Central

    2011-01-01

    Background Adrenomedullin (ADM), a novel vasorelaxant peptide, was found in human/rat ovaries. The present study investigated the interaction of ADM and endothelin-1 (ET-1) in follicles and newly formed corpora lutea (CL) and the actions of ADM on progesterone production in CL during pregnancy. Methods The peptide and gene expression level of adrenomedullin in small antral follicles, large antral follicles and CL was studied by real-time RT-PCR and EIA. The effect of ADM treatment on oestradiol production in 5-day follicular culture and on progesterone production from CL of different pregnant stages was measured by EIA. The interaction of ADM and ET-1 in follicles and CL at their gene expression level was studied by real-time RT-PCR. Results In the rat ovary, the gene expression of Adm increased during development from small antral follicles to large antral follicles and CL. In vitro treatment of preantral follicular culture for 5 days with ADM increased oestradiol production but did not affect follicular growth or ovulation rate. The regulation of progesterone production by ADM in CL in culture was pregnancy-stage dependent, inhibitory at early and late pregnancy but stimulatory at mid-pregnancy, which might contribute to the high progesterone production rate of the CL at mid-pregnancy. Moreover, the interaction between ADM and ET-1 at both the production and functional levels indicates that these two vasoactive peptides may form an important local, fine-tuning regulatory system together with LH and prolactin for progesterone production in rat CL. Conclusions As the CL is the major source of progesterone production even after the formation of placenta in rats, ADM may be an important regulator in progesterone production to meet the requirement of pregnancy. PMID:21824440

  3. Lipopolysaccharide reduces the primordial follicle pool in the bovine ovarian cortex ex vivo and in the murine ovary in vivo.

    PubMed

    Bromfield, John J; Sheldon, I Martin

    2013-04-01

    Infections of the uterus or mammary gland with Gram-negative bacteria cause infertility in cattle, not only during disease but also for some time afterward. Even though these infections are in organs distant from the ovary, metritis and mastitis perturb antral follicle development and function in vivo. Although granulosa cells from antral follicles express toll-like receptor 4 (TLR4), and detect and mount an inflammatory response to lipopolysaccharide (LPS) from Gram-negative bacteria, it is not known whether LPS impacts preantral follicle development. The present study tested the hypothesis that LPS perturbs the development of primordial ovarian follicles. Exposure of bovine ovarian cortex ex vivo to LPS reduced the primordial follicle pool associated with increased primordial follicle activation. Ovarian cortex culture supernatants accumulated the inflammatory mediators IL-1beta, IL-6, and IL-8 in an LPS concentration-dependent manner. In addition, LPS exposure modulated key intracellular regulators of follicle activation with loss of the primordial follicle PTEN and cytoplasmic translocation of FOXO3. Acute exposure of mice in vivo to LPS also reduced the primordial follicle pool associated with increased follicle atresia. The increased follicle atresia was TLR4-dependent as Tlr4-deficient mice were insensitive to LPS-mediated follicle atresia. However, LPS did not affect the diameter of individually cultured bovine secondary follicles or their enclosed oocytes. In conclusion, LPS reduced the primordial ovarian follicle pool in the bovine ovarian cortex ex vivo and in the murine ovary in vivo. These observations provide an insight into how bacterial infections distant from the ovary have long term effects on fertility.

  4. Effects of gonadotrophins, growth hormone, and activin A on enzymatically isolated follicle growth, oocyte chromatin organization, and steroid secretion.

    PubMed

    Ola, Safiriyu Idowu; Ai, Jun-Shu; Liu, Jing-He; Wang, Qiang; Wang, Zhen-Bo; Chen, Da-Yuan; Sun, Qing-Yuan

    2008-01-01

    So far, standard follicle culture systems can produce blastocyst from less than 40% of the in vitro matured oocytes compared to over 70% in the in vivo counterpart. Because the capacity for embryonic development is strictly associated with the terminal stage of oocyte growth, the nuclear maturity status of the in vitro grown oocyte was the subject of this study. Mouse early preantral follicles (100-130 microm) and early antral follicles (170-200 microm) isolated enzymatically were cultured for 12 and 4 days, respectively, in a collagen-free dish. The serum-based media were supplemented with either 100 mIU/ml FSH (FSH only); 100 mIU/ml FSH + 10 mIU/ml LH (FSH-LH); 100 mIU/ml FSH + 1 mIU/ml GH (FSH-GH) or 100 mIU/ml FSH + 100 ng/ml activin A (FSH-AA). Follicle survival was highest in follicle stimulating hormone (FSH)-AA group in both cultured preantral (91.8%) and antral follicles (82.7%). Survival rates in the other groups ranged between 48% (FSH only, preantral follicle culture) and 78.7% (FSH only, antral follicle culture). Estradiol and progesterone were undetectable in medium lacking gonadotrophins while AA supplementation in synergy with FSH caused increased estradiol secretion and a simultaneously lowered progesterone secretion. Chromatin configuration of oocytes from surviving follicles at the end of culture revealed that there were twice more developmentally incompetent non-surrounded nucleolus (NSN) oocytes (>65%) than the competent surrounded nucleolus (SN) oocytes (<34%). We conclude that the present standard follicle culture system does not produce optimum proportion of developmentally competent oocytes.

  5. A novel method for toxicology: in vitro culture system of a rat preantral follicle.

    PubMed

    Wan Xuying; Zhu Jiangbo; Zhu Yuping; Xili, Ma; Liu Zhen; Wang Fei; Xu Guifeng; Zhang Tianbao

    2011-08-01

    Preantral follicle in vitro culture systems have been successfully or nearly successfully established for sheep, pig and mouse, and applied on follicle development and regulation research on reproductive biology and physiology. However, there have been few studies concerning rat preantral follicle in vitro development. The objective is to establish an in vitro culture system for rat preantral follicles which can be used for reproductive biology and toxicology research. Rat preantral follicles are mechanically separated, cultured in vitro in single follicle mode for continuous 12 days using 96-well plates, and then administrated ovulation induction. The observation on follicle development, hormone level, and ovum formation are recorded and assessed. Taking in vivo growth and in vitro maturation of oocytes group as control group, in vitro growth and maturation of oocytes group is assessed to see whether this in vitro culture method is successful. The conditions for rat follicle culture are determined based on the mouse pre-antral follicle culture. The in vitro culture system for rat preantral follicles established in this study is feasible and successful, and can serve as model for reproductive biology and toxicology research.

  6. Bioengineering the Hair Follicle

    PubMed Central

    Parimoo, S; Zheng, Y; Barrows, T; Boucher, M; Washenik, K

    2007-01-01

    The hair follicle develops from the primitive embryonic epidermis as a result of complex epithelial-mesenchymal interactions. The full follicle, consisting of epithelial cylinders under control of a proximal lying mesenchymal papilla, grows in cycles giving rise to a new hair shaft during each cycle. The ability to cycle endows the follicle with regenerative properties. The evolution of hair follicle engineering began with the recognition in the early 1960's that hair follicles could be transplanted clinically into a foreign site and still grow a shaft typical of the donor site. Since that time, it has been found that the follicular papilla has hair follicle inducing properties and that the hair follicle houses within it epithelial stem cells that can respond to hair inductive signals. These findings have laid the foundation for isolating hair-forming cells, for expanding the cells in culture, and for forming new follicles in vivo. PMID:19279694

  7. Evaluation of antral motility in humans using manometry and scintigraphy.

    PubMed Central

    Jones, K; Edelbroek, M; Horowitz, M; Sun, W M; Dent, J; Roelofs, J; Muecke, T; Akkermans, L

    1995-01-01

    Recent studies suggest that scintigraphy can be used to evaluate non-invasively antral motility in humans, although scintigraphic techniques have not yet been compared with more conventional measurements of intraluminal pressures by manometry. Simultaneous scintigraphic and manometric measurements of antral motility were performed in nine healthy volunteers. After intubation with a sleeve/sidehole catheter which incorporated five pressure sideholes located at 1.5 cm intervals spanning the antrum, each subject ingested 100 g minced beef labelled with 100 MBq 99mTc-chicken liver and 150 ml water. Between 40-43, 60-63, 80-83, and 100-103 minutes after meal ingestion, radioisotopic data were acquired in two second frames. Time-activity curves showing antral 'contractions' resulting from wall motion were derived by drawing small regions of interest over the antrum to coincide with the position of the antral manometric sideholes. Scintigraphic contraction rates approximated 3/minute, whereas antral pressure waves that occluded the lumen were less frequent (p < 0.01 for all), particularly in the proximal antrum. The amplitude of wall motion, evaluated scintigraphically, and the amplitude of pressure waves were both inversely related to the distance from the pylorus (r > -0.32, p < 0.05) and antral volume r > -0.29 (p < 0.05). There were significant relationships between the amplitude of contractions assessed scintigraphically and the number of lumen-occlusive antral pressure waves in the distal antrum (r -0.48, p < 0.05) but not in the more proximal antral regions. It is concluded that scintigraphy can detect antral wall motion with greater sensitivity than manometry, particularly in the proximal antrum. As manometry gives information on the amplitude as well as the temporal and spatial organisation of those contractions which result in lumen occlusion, the combination of scintigraphic and manometric techniques in the evaluation of antral motility shows considerable

  8. Alginate encapsulation supports the growth and differentiation of human primordial follicles within ovarian cortical tissue.

    PubMed

    Laronda, Monica M; Duncan, Francesca E; Hornick, Jessica E; Xu, Min; Pahnke, Jennifer E; Whelan, Kelly A; Shea, Lonnie D; Woodruff, Teresa K

    2014-08-01

    In vitro follicle growth (IVFG) is an investigational fertility preservation technique in which immature follicles are grown in culture to produce mature eggs that can ultimately be fertilized. Although progress has been made in growing primate primary and secondary follicles in vitro, it has been a relatively greater challenge to isolate and culture primordial follicles. The purpose of this study was to develop methods to grow human primordial follicles in vitro using alginate hydrogels. We obtained human ovarian tissue for research purposes through the National Physicians Cooperative from nationwide sites and used it to test two methods for culturing primordial follicles. First, primordial follicles were isolated from the ovarian cortex and encapsulated in alginate hydrogels. Second, 1 mm × 1 mm pieces of 500 μm-thick human ovarian cortex containing primordial follicles were encapsulated in alginate hydrogels, and survival and follicle development within the tissue was assessed for up to 6 weeks. We found that human ovarian tissue could be kept at 4 °C for up to 24 h while still maintaining follicle viability. Primordial follicles isolated from ovarian tissue did not survive culture. However, encapsulation and culture of ovarian cortical pieces supported the survival, differentiation, and growth of primordial and primary follicles. Within several weeks of culture, many of the ovarian tissue pieces had formed a defined surface epithelium and contained growing preantral and antral follicles. The early stages of in vitro human follicle development require the support of the native ovarian cortex.

  9. The domestic dog and cat as models for understanding the regulation of ovarian follicle development in vitro.

    PubMed

    Songsasen, N; Comizzoli, P; Nagashima, J; Fujihara, M; Wildt, D E

    2012-12-01

    The culture of ovarian follicles is an important tool for understanding the mechanisms controlling follicle development and differentiation of the oocyte. The benefit of recovering meiotically and developmentally competent oocytes from early stage follicles (primordial, primary, pre-antral and early antral) also would be significant, ranging from rescue of genomes from endangered species to preserving fertility in women facing cancer treatments. This research field is at an early stage of scientific discovery. To-date, live offspring from cultured primordial follicles that produced fertilizable oocytes has occurred only in the mouse. Progress in other more complex species has been limited because larger animals have longer durations of natural folliculogenesis, thereby requiring more culture time to generate fully grown follicles and oocytes. We believe the dog and cat are excellent models for understanding more about folliculogenesis in vitro. This review highlights what is known about this topic for these two species as well as future priorities. We have discovered that it is more challenging to maintain viability of primordial follicles within ovarian tissues in vitro in the dog than the cat. Nonetheless, it is possible to grow both isolated cat and dog pre-antral follicles in culture. Although the follicles of both species have the capacity to increase in size and produce steroids, only cat oocytes appear morphologically normal. The reason for this striking difference between these two species is an area of high research priority. While much more fundamental data are required, we envision advanced technology that will allow harvesting oocytes from the vast, unused follicle stores sequestered within carnivore ovaries. These gametes have utility for reproducing genetically valuable dogs and cats that are 'companions' or biomedical models for investigating human disorders as well as for salvaging the genomes of rare canid and felid species that die before

  10. Gene expression patterns in granulosa cells and oocytes at various stages of follicle development as well as in in vitro grown oocyte-and-granulosa cell complexes.

    PubMed

    Munakata, Yasuhisa; Kawahara-Miki, Ryoka; Shiratsuki, Shogo; Tasaki, Hidetaka; Itami, Nobuhiko; Shirasuna, Koumei; Kuwayama, Takehito; Iwata, Hisataka

    2016-08-25

    Follicle development is accompanied by proliferation of granulosa cells and increasing oocyte size. To obtain high-quality oocytes in vitro, it is important to understand the processes that occur in oocytes and granulosa cells during follicle development and the differences between in vivo and in vitro follicle development. In the present study, oocytes and granulosa cells were collected from early antral follicles (EAFs, 0.5-0.7 mm in diameter), small antral follicles (SAFs, 1-3 mm in diameter), large antral follicles (LAFs, 3-7 mm in diameter), and in vitro grown oocyte-and-granulosa cell complexes (OGCs), which were cultured for 14 days after collection from EAFs. Gene expression was analyzed comprehensively using the next-generation sequencing technology. We found top upstream regulators during the in vivo follicle development and compared them with those in in vitro developed OGCs. The comparison revealed that HIF1 is among the top regulators during both in vivo and in vitro development of OGCs. In addition, we found that HIF1-mediated upregulation of glycolysis in granulosa cells is important for the growth of OGCs, but the cellular metabolism differs between in vitro and in vivo grown OGCs. Furthermore, on the basis of comparison of upstream regulators between in vivo and in vitro development of OGCs, we believe that low expression levels of FLT1 (VEGFA receptor), SPP1, and PCSK6 can be considered causal factors of the suboptimal development under in vitro culture conditions.

  11. Gene expression patterns in granulosa cells and oocytes at various stages of follicle development as well as in in vitro grown oocyte-and-granulosa cell complexes

    PubMed Central

    MUNAKATA, Yasuhisa; KAWAHARA-MIKI, Ryoka; SHIRATSUKI, Shogo; TASAKI, Hidetaka; ITAMI, Nobuhiko; SHIRASUNA, Koumei; KUWAYAMA, Takehito; IWATA, Hisataka

    2016-01-01

    Follicle development is accompanied by proliferation of granulosa cells and increasing oocyte size. To obtain high-quality oocytes in vitro, it is important to understand the processes that occur in oocytes and granulosa cells during follicle development and the differences between in vivo and in vitro follicle development. In the present study, oocytes and granulosa cells were collected from early antral follicles (EAFs, 0.5–0.7 mm in diameter), small antral follicles (SAFs, 1–3 mm in diameter), large antral follicles (LAFs, 3–7 mm in diameter), and in vitro grown oocyte-and-granulosa cell complexes (OGCs), which were cultured for 14 days after collection from EAFs. Gene expression was analyzed comprehensively using the next-generation sequencing technology. We found top upstream regulators during the in vivo follicle development and compared them with those in in vitro developed OGCs. The comparison revealed that HIF1 is among the top regulators during both in vivo and in vitro development of OGCs. In addition, we found that HIF1-mediated upregulation of glycolysis in granulosa cells is important for the growth of OGCs, but the cellular metabolism differs between in vitro and in vivo grown OGCs. Furthermore, on the basis of comparison of upstream regulators between in vivo and in vitro development of OGCs, we believe that low expression levels of FLT1 (VEGFA receptor), SPP1, and PCSK6 can be considered causal factors of the suboptimal development under in vitro culture conditions. PMID:27108636

  12. Vascular endothelial growth factor and angiopoietin production by primate follicles during culture is a function of growth rate, gonadotrophin exposure and oxygen milieu.

    PubMed

    Fisher, T E; Molskness, T A; Villeda, A; Zelinski, M B; Stouffer, R L; Xu, J

    2013-12-01

    What is the time course of production of vascular endothelial growth factor-A (VEGF-A), angiopoietin (ANGPT)-1 and ANGPT-2 by primate follicles during encapsulated three-dimensional culture, and what conditions affect their production? Primate follicles produce VEGF-A and ANGPT-2 in vitro, particularly after developing to the antral stage, with VEGF production influenced by FSH concentration and O(2) tension. Folliculogenesis, i.e. the development of primordial follicles into mature, antral follicles, requires the creation of a vascular network in the follicle wall via a process called angiogenesis. Angiogenic factors including VEGFs and ANGPTs have documented roles in angiogenesis. However, direct studies on the production and regulation of angiogenic factors by individual, growing follicles are limited. Ovaries (n = 9 pairs) were obtained from rhesus macaques during the early follicular phase of the menstrual cycle (cycle days 1-4). Secondary (125-225 µm) follicles were isolated mechanically, encapsulated into alginate (0.25% w/v) and cultured for 40 days. Individual follicles were cultured in a 5 or 20% O(2) environment in alpha minimum essential medium supplemented with recombinant human (h) FSH. Half of the follicles had recombinant hLH added to the media from Days 30 to 40. Follicle diameters were measured weekly. Follicles were categorized at Week 5 as no-grow (NG; <250 μm in diameter), slow-grow (SG; 251-499 μm) and fast-grow (FG; >500 μm). VEGF-A, ANGPT-1 and -2 concentrations in media were measured by ELISA. VEGF concentrations were low throughout the culture for NG follicles. SG and FG follicles had detectable VEGF concentrations at Week 2, which continued to rise throughout culture. VEGF concentrations were distinct (P < 0.05) among all three follicle categories during Weeks 4 and 5. VEGF concentrations were higher (P < 0.05) in SG follicles in the presence of high/mid-dose FSH at 5% O(2). In contrast, there were no dose-dependent differences in

  13. Growth and development of the ovary and small follicle pool from mid fetal life to pre-puberty in the African elephant (Loxodonta africana).

    PubMed

    Stansfield, Fiona J; Nöthling, Johan O; Allen, William R

    2012-07-23

    Follicle numbers and developing ovarian morphology, particularly with reference to the presence of interstitial tissue, are intimately linked within the ovary of the African elephant during the period spanning mid-gestation to puberty. These have not been previously quantified in any studies. The collection of 7 sets of elephant fetal ovaries between 11.2 and 20.2 months of gestation, and 29 pairs of prepubertal calf ovaries between 2 months and 9 years of age during routine management off-takes of complete family groups in private conservancies in Zimbabwe provided an opportunity for a detailed study of this period. The changing morphology of the ovary is described as the presumptive cortex and medulla components of the fetal ovary settled into their adult form. Interstitial tissue dominated the ovary in late fetal life and these cells stained strongly for 3β-hydroxysteroid dehydrogenase. This staining continued postnatally through to 4.5 years of age suggesting continued secretion of progestagens by the ovary during this period. The considerable growth of antral follicles peaked at 28% of ovarian volume at around 16.7 months of fetal age. The numbers of small follicles (primordial, early primary and true primary), counted in the cortex using stereological protocols, revealed fewer small follicles in the ovaries of animals aged 0 to 4.5 years of age than during either late fetal life or prepubertal life. The small follicle populations of the late-fetal and prepubertal ovaries of the African elephant were described along with the changing morphology of these organs. The changes noted represent a series of events that have been recorded only in the elephant and the giraffe species to date. The expansion of the interstitial tissue of the fetal ovary and its continued presence in early post natal life may well contribute to the control of follicle development in these early years. Further research is required to determine the reasons behind the variation

  14. Protein and messenger RNA expression of interleukin 1 system members in bovine ovarian follicles and effects of interleukin 1β on primordial follicle activation and survival in vitro.

    PubMed

    Passos, J R S; Costa, J J N; da Cunha, E V; Silva, A W B; Ribeiro, R P; de Souza, G B; Barroso, P A A; Dau, A M P; Saraiva, M V A; Gonçalves, P B D; van den Hurk, R; Silva, J R V

    2016-01-01

    This study aimed to investigate the expression of interleukin 1 (IL-1) system members (proteins and messenger RNA of ligands and receptors) and its distribution in ovarian follicles of cyclic cows and to evaluate the effects of IL-1β on the survival and activation of primordial follicles in vitro. The ovaries were processed for localization of IL-1 system in preantral and antral follicles by immunohistochemical, real-time polymerase chain reaction, and Western blot analysis. For in vitro studies, ovarian fragments were cultured in α-MEM(+) supplemented with IL-1β (0, 1, 10, 50, or 100 ng/mL), and after 6 d, the cultured tissues were processed for histologic analysis. Immunohistochemical results showed that the IL-1 system proteins IL-1β, IL-1RA, IL-1RI, and IL-1RII were detected in the cytoplasm of oocytes and granulosa cells from all follicular categories and theca cells of antral follicles. Variable levels of messenger RNA for the IL-1 system members were observed at different stages of development. After 6 d of culture, the presence of IL-1β (10 or 50 ng/mL) was effective in maintaining the percentage of normal follicles and in promoting primordial follicle activation. In conclusion, IL-1 system members are differentially expressed in ovarian follicles according to their stage of development. Moreover, IL-1β promotes the development of primordial follicles. These results indicate an important role of the IL-1 system in the regulation of bovine folliculogenesis. Copyright © 2016 Elsevier Inc. All rights reserved.

  15. Effect of gonadotropin-releasing hormone antagonist on primordial follicle survival in the primate ovary.

    PubMed

    Attaman, Jill; Arbogast, Laura K; Friedman, Chad I; Danforth, Douglas R

    2014-01-01

    To examine the effects of gonadotropin-releasing hormone (GnRH) antagonist on primordial follicle reserve in the primate ovary. A prospective basic research study in which 10 juvenile cynomolgus monkeys (Macaca fascicularis) had 1 ovary surgically removed. Six animals were then treated with the GnRH antagonist antide (1.0 mg/kg/day) for 14 days, and 4 animals were treated with vehicle. After treatment the contralateral ovary was removed and both ovaries were prepared for assessment of primordial, primary, and secondary follicle numbers. Antide treatment resulted in a modest (13%) but significant decrease in primordial follicle number in juvenile macaques (p = 0.048, n = 6). Three animals demonstrated a marked reduction in primordial follicles (19%, 25%, 36%) and 3 animals had no (< 5%) change in primordial follicles after antide treatment. Control animals demonstrated no change in primordial follicle number following vehicle treatment. Antide had no effect on primary, secondary, or early antral follicle numbers and did not affect circulating estradiol concentrations. In contrast to mice, in which GnRH antagonist treatment markedly reduces primordial follicle reserve, the effect of antide in nonhuman primates was less dramatic and somewhat variable. These data suggest there may be a subset of animals susceptible to the adverse effects of GnRH antagonist on primordial follicle survival.

  16. In vitro culture of early secondary preantral follicles in hanging drop of ovarian cell-conditioned medium to obtain MII oocytes from outbred deer mice.

    PubMed

    Choi, Jung Kyu; Agarwal, Pranay; He, Xiaoming

    2013-12-01

    The ovarian follicle (each contains a single oocyte) is the fundamental functional tissue unit of mammalian ovaries. In humans, it has been long held true that females are born with a maximum number of follicles (or oocytes) that are not only nonrenewable, but also undergoing degeneration with time with a sharply decreased oocyte quality after the age of ∼35. Therefore, it is of importance to isolate and bank ovarian follicles for in vitro culture to obtain fertilizable oocytes later, to preserve the fertility of professional women who may want to delay childbearing, young and unmarried women who may lose gonadal function because of exposure to environmental/occupational hazards or aggressive medical treatments, such as radiation and chemotherapy, and even endangered species and breeds. Although they contributed significantly to the understanding of follicle science and biology, most studies reported to date on this topic were done using the man-made, unnatural inbred animal species. It was found in this study that the conventional two-dimensional microliter drop and three-dimensional hanging drop (HD) methods, reported to be effective for in vitro culture of preantral follicles from inbred mice, are not directly transferrable to outbred deer mice. Therefore, a modified HD method was developed in this study to achieve a much higher (>5 times compared to the best conventional methods) percentage of developing early secondary preantral follicles from the outbred mice to the antral stage, for which, the use of an ovarian cell-conditioned medium and multiple follicles per HD were identified to be crucial. It was further found that the method for in vitro maturation of oocytes in antral follicles obtained by in vitro culture of preantral follicles could be very different from that for oocytes in antral follicles obtained by hormone stimulation in vivo. Therefore, this study should provide important guidance for establishing effective protocols of in vitro follicle

  17. Relationship Between 17-Hydroxyprogesterone Responses to Human Chorionic Gonadotropin and Markers of Ovarian Follicle Morphology in Women With Polycystic Ovary Syndrome

    PubMed Central

    Maas, Kevin H.; Chuan, Sandy S.; Cook-Andersen, Heidi; Su, H. Irene; Duleba, A.

    2015-01-01

    Context: Women with polycystic ovary syndrome (PCOS) have increased 17-hydroxyprogesterone (17-OHP) responses to gonadotropin stimulation although individual variability is substantial, as reflected by exaggerated as well as normal responses. The relationship between 17-OHP responses to gonadotropin stimulation and markers of ovarian function has not been assessed. Objective: To determine whether 17-OHP responses are associated with antral follicle count (AFC), anti-Mullerian hormone (AMH), or inhibin B (Inh B) levels in PCOS and normal women. Design: Prospective study. Setting: Research center at an academic medical center. Participants: Women with PCOS (n = 18) and normal controls (n = 18). Interventions: Blood samples were obtained before and 24 hours after administration of 25 μg recombinant-human chorionic gonadotropin. Ovarian imaging was conducted with three-dimensional pelvic ultrasound. Main Outcome Measures: Basal and stimulated levels of 17-OHP, androgens, estrogen, AMH, Inh B, and AFC. Results: In women with PCOS, 17-OHP responses were heterogeneous and inversely correlated with AMH and Inh B levels, but not AFC. In a subgroup of PCOS women with exaggerated 17-OHP responses, AMH levels were equivalent to that of normal women. In PCOS women with normal 17-OHP responses, AMH levels were markedly elevated. Conclusion: Based on heterogeneous 17-OHP responses to human chorionic gonadotropin in women with PCOS, AMH levels are inversely linked to ovarian androgen production while positively correlated with AFC. These findings suggest that in PCOS, AMH production may reflect redistribution of the follicle population or regulation by intraovarian mechanisms. PMID:25313914

  18. A non-neuronal cholinergic system of the ovarian follicle.

    PubMed

    Mayerhofer, Artur; Kunz, Lars

    2005-11-01

    We have recently provided evidence that acetylcholine (ACh) is a non-neuronal intraovarian signalling molecule, produced by granulosa cells (GCs) and which appears to act as signalling factor in the growing follicle. The ACh biosynthesis enzyme, choline-acetyltransferase (ChAT), is expressed only in growing, antral follicles in rodent and primate species. This restriction to follicle stages, which depend on the activity of follicle-stimulating hormone (FSH), may suggest that ACh could be an as yet unknown local mediator of FSH actions. In respect of ACh actions, our ongoing studies indicate that they may be exerted via different muscarinic ACh-receptors (MR) in GCs, but also in oocytes in an overlapping fashion. To elucidate functional details we have studied cultured human GCs isolated from preovulatory follicles. Activation of MRs increases intracellular calcium and, e.g., induces the master transcription factor egr-1, implying involvement in cell differentiation events. ACh agonists also activate a calcium-activated potassium channel (BK(Ca)) resulting in membrane hyperpolarization, which allows activation of other voltage-dependent ion channels. Experimental modulation of the chain of these events causes altered steroidogenesis, implying a crucial role of ACh in endocrine functions. Further ACh actions include phosphorylation of the gap junction molecule connexin 43 and disruption of intercellular communication between GCs. This may allow strongly coupled GCs to escape from the functional syncytium of the follicle in order to initiate proliferation. Proliferation is indeed strongly increased in cultured human GCs when treated with cholinergic agents. The repertoire of ACh/MR actions is far from being fully appreciated and may include epigenetic regulation in healthy growing follicles. Although many aspects of the ovarian cholinergic system, including, for instance, influence of follicular ACh on the MR-bearing oocyte, remain to be examined. The present data

  19. Promoting extracellular matrix remodeling via ascorbic acid enhances the survival of primary ovarian follicles encapsulated in alginate hydrogels.

    PubMed

    Tagler, David; Makanji, Yogeshwar; Tu, Tao; Bernabé, Beatriz Peñalver; Lee, Raymond; Zhu, Jie; Kniazeva, Ekaterina; Hornick, Jessica E; Woodruff, Teresa K; Shea, Lonnie D

    2014-07-01

    The in vitro growth of ovarian follicles is an emerging technology for fertility preservation. Various strategies support the culture of secondary and multilayer follicles from various species including mice, non-human primate, and human; however, the culture of early stage (primary and primordial) follicles, which are more abundant in the ovary and survive cryopreservation, has been limited. Hydrogel-encapsulating follicle culture systems that employed feeder cells, such as mouse embryonic fibroblasts (MEFs), stimulated the growth of primary follicles (70-80 µm); yet, survival was low and smaller follicles (<70 µm) rapidly lost structure and degenerated. These morphologic changes were associated with a breakdown of the follicular basement membrane; hence, this study investigated ascorbic acid based on its role in extracellular matrix (ECM) deposition/remodeling for other applications. The selection of ascorbic acid was further supported by a microarray analysis that suggested a decrease in mRNA levels of enzymes within the ascorbate pathway between primordial, primary, and secondary follicles. The supplementation of ascorbic acid (50 µg/mL) significantly enhanced the survival of primary follicles (<80 µm) cultured in alginate hydrogels, which coincided with improved structural integrity. Follicles developed antral cavities and increased to diameters exceeding 250 µm. Consistent with improved structural integrity, the gene/protein expression of ECM and cell adhesion molecules was significantly changed. This research supports the notion that modifying the culture environment (medium components) can substantially enhance the survival and growth of early stage follicles. © 2013 Wiley Periodicals, Inc.

  20. Transcriptome Profiling of the Theca Interna from Bovine Ovarian Follicles during Atresia

    PubMed Central

    Hatzirodos, Nicholas; Irving-Rodgers, Helen F.; Hummitzsch, Katja; Rodgers, Raymond J.

    2014-01-01

    The theca interna is a specialized stromal layer that envelops each growing ovarian follicle. It contains capillaries, fibroblasts, immune cells and the steroidogenic cells that synthesize androgens for conversion to estradiol by the neighboring granulosa cells. During reproductive life only a small number of follicles will grow to a sufficient size to ovulate, whereas the majority of follicles will undergo regression/atresia and phagocytosis by macrophages. To identify genes which are differentially regulated in the theca interna during follicular atresia, we undertook transcriptome profiling of the theca interna from healthy (n = 10) and antral atretic (n = 5) bovine follicles at early antral stages (<5 mm). Principal Component Analyses and hierarchical classification of the signal intensity plots for the arrays showed primary clustering into two groups, healthy and atretic. A total of 543 probe sets were differentially expressed between the atretic and healthy theca interna. Further analyses of these genes by Ingenuity Pathway Analysis and Gene Ontology Enrichment Analysis Toolkit software found most of the genes being expressed were related to cytokines, hormones and receptors as well as the cell cycle and DNA replication. Cell cycle genes which encode components of the replicating chromosome complex and mitotic spindle were down-regulated in atretic theca interna, whereas stress response and inflammation-related genes such as TP53, IKBKB and TGFB1 were up-regulated. In addition to cell cycle regulators, upstream regulators that were predicted to be inhibited included Retinoblastoma 1, E2 transcription factor 1, and hepatocyte growth factor. Our study suggests that during antral atresia of small follicles in the theca interna, arrest of cell cycle and DNA replication occurs rather than up- regulation of apoptosis-associated genes as occurs in granulosa cells. PMID:24956388

  1. The Hair Follicle Enigma.

    PubMed

    Bernard, Bruno A

    2017-03-07

    The hair follicle is a mini organ endowed with a unique structure and cyclic behavior. Despite the intense research efforts which have been devoted at deciphering the hair follicle biology over the past 70 years, one must admit that hair follicle remains an enigma. In this brief review, various aspects of hair follicle biology will be addressed, and more importantly, unsolved questions and new possible research tracks will be highlighted, including hair follicle glycobiology and exosome mediated cell-cell interactions. Even though bricks of knowledge are solidly being acquired, an integrative picture remains to emerge. One can predict that computer science, algorithms and bioinformatics will assist in fostering our understanding hair biology. This article is protected by copyright. All rights reserved.

  2. Phosphoinositide 3-kinase p110δ mediates estrogen- and FSH-stimulated ovarian follicle growth.

    PubMed

    Li, Qian; He, Hui; Zhang, Yin-Li; Li, Xiao-Meng; Guo, Xuejiang; Huo, Ran; Bi, Ye; Li, Jing; Fan, Heng-Yu; Sha, Jiahao

    2013-09-01

    In the mammalian ovary, primordial follicles are generated early in life and remain dormant for prolonged periods. Their growth resumes via primordial follicle activation, and they continue to grow until the preovulatory stage under the regulation of hormones and growth factors, such as estrogen, FSH, and IGF-1. Both FSH and IGF-1 activate the phosphatidylinositol-3 kinase (PI3K)/Akt (acute transforming retrovirus thymoma protein kinase) signaling pathway in granulosa cells (GCs), yet it remains inconclusive whether the PI3K pathway is crucial for follicle growth. In this study, we investigated the p110δ isoform (encoded by the Pik3cd gene) of PI3K catalytic subunit expression in the mouse ovary and its function in fertility. Pik3cd-null females were subfertile, exhibited fewer growing follicles and more atretic antral follicles in the ovary, and responded poorly to exogenous gonadotropins compared with controls. Ovary transplantation showed that Pik3cd-null ovaries responded poorly to FSH stimulation in vitro; this confirmed that the follicle growth defect was intrinsically ovarian. In addition, estradiol (E2)-stimulated follicle growth and GC proliferation in preantral follicles was impaired in Pik3cd-null ovaries. FSH and E2 substantially activated the PI3K/Akt pathway in GCs of control mice but not in those of Pik3cd-null mice. However, primordial follicle activation and oocyte meiotic maturation were not affected by Pik3cd knockout. Taken together, our findings indicate that the p110δ isoform of the PI3K catalytic subunit is a key component of the PI3K pathway for both FSH and E2-stimulated follicle growth in ovarian GCs; however, it is not required for primordial follicle activation and oocyte development.

  3. The effects of insulin and follicle-simulating hormone (FSH) during in vitro development of ovarian goat preantral follicles and the relative mRNA expression for insulin and FSH receptors and cytochrome P450 aromatase in cultured follicles.

    PubMed

    Chaves, Roberta N; Duarte, Ana Beatriz G; Rodrigues, Giovanna Q; Celestino, Juliana J H; Silva, Gerlane M; Lopes, Claudio Afonso P; Almeida, Anderson P; Donato, Mariana A M; Peixoto, C A; Moura, Arlindo A A; Lobo, Carlos H; Locatelli, Yann; Mermillod, Pascalle; Campello, Claudio C; Figueiredo, Jose Ricardo

    2012-09-01

    The actions of different concentrations of insulin alone or in combination with follicle-stimulating hormone (FSH) were evaluated by in vitro follicular development and mRNA expression of cytochrome P450 aromatase (CYP19A1) and as receptors for insulin (INSR) and FSH (FSHR) from isolated, cultured goat preantral follicles. Goat preantral follicles were microdissected and cultured for 18 days in the absence or presence of insulin (5 and 10 ng/ml or 10 μg/ml) alone or in combination with FSH. After 18 days, the addition of the maximum concentration of insulin to the culture medium reduced follicular survival and antrum formation rates significantly compared to the other treatments. However, when FSH was added to the culture medium, no differences between these two parameters were observed. Preantral and antral follicles from the fresh control as well as from all cultured follicles still presented a normal ultrastructural pattern. In medium supplemented with FSH, only insulin at 10 ng/ml presented oocytes with higher rates of meiosis resumption compared to control, as well as oocytes in metaphase II. Treatment with insulin (10 ng/ml) plus FSH resulted in significantly increased levels of INSR and CYP19A1 mRNA compared to that with other treatments. In conclusion, 10 ng/ml insulin associated with FSH was more efficient in promoting resumption of oocyte meiosis, maintaining survival, stimulating follicular development, and increasing expression of the INSR and CYP19A1 genes in goat preantral follicles.

  4. Induction of multiple ovulation via modulation of angiotensin II receptors in in vitro ovarian follicle culture models.

    PubMed

    Kim, Yong Jin; Kim, Yoon Young; Kang, Byeong-Cheol; Kim, Moon Suk; Ko, In Kap; Liu, Hung Ching; Rosenwaks, Zev; Ku, Seung-Yup

    2016-09-15

    In vitro culture of ovarian follicles is a promising bioengineering technique for retrieving fertilizable oocytes from preserved ovarian tissues of cancer survivors. However, current in vitro follicle culture techniques are labour-intensive and of low efficiency, as only single follicle culture (SFC) has been possible to date. The present study investigated the feasibility of multifollicular cluster culture (MFCC) system using angiotensin II receptor (ATII-Rc) analogues. Ovarian pre-antral follicles isolated from 2-week-old C57BL6 mice were cultured with ATII-Rc agonist or antagonist and their maturation outcomes were compared with control group. When single follicles were cultured, the ovulation and maturation rates were similar in all three groups. When three-follicle clusters were cultured, up to three follicles were ovulated in the ATII-Rc agonist group while none or one follicle ovulated in control or antagonist groups (p < 0.0001). Significantly higher numbers of mature oocytes were obtained in the agonist group (three-follicle 28.2 ± 4.9 vs. SFC 11.0 ± 1.3, per 25 cultured droplets) (p < 0.0001), and the development of each fertilized oocytes was comparable to those from SFC. It is therefore concluded that this novel MFCC system can significantly improve the efficiency of in vitro mature oocyte retrieval via ATII-Rc modulation. Copyright © 2016 John Wiley & Sons, Ltd.

  5. The crestal approach: antral membrane elevation via a post graft.

    PubMed

    Soltan, Muna; Smiler, Dennis; Ghostine, Michelle; Prasad, Hari S; Rohrer, Michael D

    2011-06-01

    The crestal approach to elevating the antral membrane by a resorbable StemVie Post is a modification of the sinus lift technique. This technique can add 4 to 10 mm bone height for severely atrophic ridges in areas that are difficult to access through a lateral window. The procedure is minimally invasive, simple, predictable, and has less postoperative morbidity because of smaller flap design and minimal osteotomy. If sufficient alveolar bone is present for stabilization, an implant can be placed simultaneous with antral elevation and graft. The StemVie Post completely resorbs and is replaced by the patient's own bone.

  6. Abnormal in vitro development of ovarian follicles explanted from mice exposed to tetrachlorvinphos.

    PubMed

    Nayudu, P L; Kiesel, P S; Nowshari, M A; Hodges, J K

    1994-01-01

    A system of mouse ovarian follicle culture in which follicles can be grown from a preantral stage of development through antral formation has been developed and modified recently by Nayudu and colleagues. Follicles have been shown to grow in this culture system at a relatively constant rate and show responsiveness to LH at the end of the culture by ovulation of mature oocytes. Reported here are the distinctly different in vitro growth patterns of follicles explanted from 22- to 24-day-old mice during a period when the colony was being treated for skin parasites with tetrachlorvinphos (TCVP) (Rabond). There is to date no information on the effects of this compound on the mammalian female reproductive system. For follicles from the TCVP treated group, the duration of growth as intact follicles was markedly reduced in comparison to mice of the same strain and source not treated with TCVP. In the treated group, premature termination of follicular growth was also associated with the spontaneous expulsion of oocytes with immature nuclei and without cumulus cells. For those follicles from treated mice that did remain in culture until the day luteinizing hormone was given, the ovulatory response was poor and the maturation response of the oocytes was low in comparison with the follicles from untreated mice. The effect of the treatment on the follicles was further characterized by obvious differences in the patterns of growth. Follicles in the untreated group grew in a linear pattern at around 25 microns/day; a single phase, fast pattern for the whole culture period.(ABSTRACT TRUNCATED AT 250 WORDS)

  7. Is assessment of anti-Müllerian hormone and/or antral follicle count useful in the prediction of ovarian response in expected normal responders treated with a fixed dose of recombinant FSH and GnRH antagonists? A prospective observational study.

    PubMed

    Ganidou, Maria A; Kolibianakis, Efstratios M; Venetis, Christos A; Gerou, Spiros; Makedos, Georgios A; Klearchou, Nikolaos; Tarlatzis, Basil C

    2014-11-01

    The purpose of this prospective observational study was to evaluate whether the assessment of AMH and AFC is useful in the prediction of ovarian response in expected normal responders treated with a fixed dose of recombinant FSH (rec-FSH) and GnRH antagonists. A base model including age and basal FSH as independent predictors of COCs could explain 15% of the variance observed in the number of COCs retrieved (p = 0.002). The addition of AFC did not increase significantly the predictive ability of the above model, whereas the addition of AMH increased the performance of the base model by 13% (p < 0.001). Logistic regression analysis showed that only when AMH was added to the base model, including age and FSH, its predictive capacity for high ovarian response was statistically significant (F-test: p = 0.001; c-statistic: 0.80, 95% CI: 0.70-0.88), but this was not the case for poor ovarian response. In conclusion, the addition of AMH, but not of AFC, to a model including female age and basal FSH, is useful in the prediction of ovarian response in expected normal responders treated with a fixed dose of recombinant FSH and GnRH antagonists.

  8. Evaluation of gastric antral motility in four dimensions

    NASA Astrophysics Data System (ADS)

    Bergmann, Helmar; Minear, Greg; Kugi, Andrea; Stacher, Georg

    1994-09-01

    Gastric motion after the ingestion of a radioactively labeled standard meal was visualized using a triple headed gamma camera and dynamic SPECT acquisitions consisting of 30 scans of 6 s duration each. After the ingestion of a radiolabeled standard meal tomographic reconstruction produced, after prefiltering with a Metz filter, images of reasonable quality, in spite of the short acquisition time per view. Oblique slices rectangular to the longitudinal axis of the antrum were positioned employing 3D rendering techniques. These slices were extracted from the sequential volumes to produce time activity curves (TACs) of antral contractions. From the TACs the amplitudes and the frequencies of the antral contraction curves obtained from planar acquisitions, were markedly higher for the curves from the tomographic slices. This was due to the removal of oblique components of motion in the oblique slices. The effect of the long sampling interval of 6 seconds was checked on simulations using data from planar acquisitions and found to permit sampling of the antral waves with acceptable accuracy. 3D display of the stomach contributed to the anatomical knowledge since it showed clearly differences to the positions that would have been expected from conventional x-ray views. Antral contraction strength was not necessarily related with the rate of gastric emptying, which emphasizes the role of other factors, mainly the tone of the fundus, for the emptying process.

  9. Comparisons of mRNA expression for aromatase, FSH receptor, and IGF-I in the granulosa of small ovarian follicles between cattle selected and unselected for twin ovulations

    USDA-ARS?s Scientific Manuscript database

    Long-term selection of cattle for the production of twin ovulations and births has enhanced the development of preantral and antral ovarian follicles and increased the frequency of twin or triplet ovulations to greater than 60%. However, these differences have not been linked to differences in FSH s...

  10. Comparisons of mRNA expression for insulin-like growth factor (IGF) type 2 receptor (IGF2R) and IGF-1 in small ovarian follicles between cattle selected and not selected for twin ovulations

    USDA-ARS?s Scientific Manuscript database

    Both IGF-1 and -2 stimulate ovarian follicular cell proliferation and antral follicle development. Actions of IGF-1 and -2 are mediated through the IGF type 1 receptor, whereas binding of IGF-2 to the IGF2R results in its degradation. Information on the role of IGF2R in regulating bovine follicula...

  11. Inhibitors of c-Jun phosphorylation impede ovine primordial follicle activation.

    PubMed

    Bertoldo, Michael J; Bernard, Jérémy; Duffard, Nicolas; Tsikis, Guillaume; Alves, Sabine; Calais, Laure; Uzbekova, Svetlana; Monniaux, Danielle; Mermillod, Pascal; Locatelli, Yann

    2016-05-01

    Is the c-Jun-N-terminal kinase (JNK) pathway implicated in primordial follicle activation? Culture of ovine ovarian cortex in the presence of two different c-Jun phosphorylation inhibitors impeded pre-antral follicle activation. Despite its importance for fertility preservation therapies, the mechanisms of primordial follicle activation are poorly understood. Amongst different signalling pathways potentially involved, the JNK pathway has been previously shown to be essential for cell cycle progression and pre-antral follicle development in mice. Ovine ovarian cortex pieces were cultured with varying concentrations of SP600125, JNK inhibitor VIII or anti-Mullerian hormone (AMH) in the presence of FSH for 9 days. Follicular morphometry and immunohistochemistry for proliferating cell nuclear antigen (PCNA), apoptosis and follicle activation (Foxo3a) were assessed. Inhibition of primordial follicle activation occurred in the presence of SP600125, JNK inhibitor VIII and AMH when compared with controls (all P < 0.05) after 2 days of culture. However, only in the highest concentrations used was the inhibition of activation associated with induction of follicular apoptosis (P < 0.05). In growing follicles, PCNA antigen expression was reduced when the JNK inhibitors or AMH were used (P < 0.05 versus control), indicating reduced proliferation of the somatic compartment. Although we evaluated the effects of inhibition of c-Jun phosphorylation on primordial follicle development, we did not determine the cellular targets and mechanism of action of the inhibitors. These results are the first to implicate the JNK pathway in primordial follicle activation and could have significant consequences for the successful development of fertility preservation strategies and our understanding of primordial follicle activation. n/a. Dr Michael J. Bertoldo and the laboratories involved in the present study were supported by a grant from 'Région Centre' (CRYOVAIRE, Grant number #320000268

  12. Phenotypic variation in anti-Mullerian hormone (AMH) production per follicle in women with polycystic ovary syndrome (PCOS) and isolated polycystic ovarian morphology (PCOM): an observational cross-sectional study.

    PubMed

    Bhide, Priya; Kulkarni, Abhijit; Dilgil, Merve; Dhir, Puja; Shah, Amit; Gudi, Anil; Homburg, Roy

    2017-10-01

    This observational study compares the ratio of serum anti-Mullerian hormone (AMH) to the total antral follicle count (AFC) (as a marker of AMH production per follicle) in the various phenotypes of women with polycystic ovary syndrome (PCOS) and isolated polycystic ovarian morphology (PCOM). Two hundred and sixty-two women were recruited. Women with PCOS were divided into four phenotypes based on the diagnostic inclusion criteria of oligo-anovulation (OA), hyperandrogenism (HA) and polycystic ovarian morphology (PCOM). These included Group A (OA + HA + PCOM), Group B (OA + HA), Group C (HA + PCOM) and Group D (OA + PCOM). A ratio of serum AMH to total AFC was calculated and expressed as the AMH/AFC ratio which was compared in the phenotypes of PCOS and isolated PCOM. The median AMH/AFC ratios in PCOS-A, PCOS-D, PCOS-C and PCOM were 1.5, 1.6, 1.2 and 1.1, respectively. There were significant differences in the groups compared [F(3, 238) = 6.14, p = 0.000)]. The ratios were significantly higher in the oligo-anovulatory phenotypes PCOS-A and PCOS-D than the PCOM (p = 0.004 and 0.002, respectively). There was no significant difference in the ratio between ovulatory phenotype PCOS-C and PCOM (p = 0.59). The role of androgens and LH in per-follicle AMH production remains limited. The findings support the hypothesis of a key role for AMH in the mechanism of anovulation in PCOS.

  13. Cell Autonomous Phosphoinositide 3-Kinase Activation in Oocytes Disrupts Normal Ovarian Function Through Promoting Survival and Overgrowth of Ovarian Follicles

    PubMed Central

    Ebbert, Katherine; Cordeiro, Marilia H.; Romero, Megan; Zhu, Jie; Serna, Vanida Ann; Whelan, Kelly A.; Woodruff, Teresa K.

    2015-01-01

    In this study, we explored the effects of oocytic phosphoinositide 3-kinase (PI3K) activation on folliculogensis by generating transgenic mice, in which the oocyte-specific Cre-recombinase induces the expression of constitutively active mutant PI3K during the formation of primordial follicles. The ovaries of neonatal transgenic (Cre+) mice showed significantly reduced apoptosis in follicles, which resulted in an excess number of follicles per ovary. Thus, the elevation of phosphatidylinositol (3,4,5)-trisphosphate levels within oocytes promotes the survival of follicles during neonatal development. Despite the increase in AKT phosphorylation, primordial follicles in neonatal Cre+ mice remained dormant demonstrating a nuclear accumulation of phosphatase and tensin homolog deleted on chromosome 10 (PTEN). These primordial follicles containing a high level of nuclear PTEN persisted in postpubertal females, suggesting that PTEN is the dominant factor in the maintenance of female reproductive lifespan through the regulation of primordial follicle recruitment. Although the oocytic PI3K activity and PTEN levels were elevated, the activation of primordial follicles and the subsequent accumulation of antral follicles with developmentally competent oocytes progressed normally in prepubertal Cre+ mice. However, mature Cre+ female mice were anovulatory. Because postnatal day 50 Cre+ mice released cumulus-oocyte complexes with developmentally competent oocytes in response to super-ovulation treatment, the anovulatory phenotype was not due to follicular defects but rather endocrine abnormalities, which were likely caused by the excess number of overgrown follicles. Our current study has elucidated the critical role of oocytic PI3K activity in follicular function, as well as the presence of a PTEN-mediated mechanism in the prevention of immature follicle activation. PMID:25594701

  14. Oocyte cryopreservation for fertility preservation in postpubertal female children at risk for premature ovarian failure due to accelerated follicle loss in Turner syndrome or cancer treatments.

    PubMed

    Oktay, K; Bedoschi, G

    2014-12-01

    To preliminarily study the feasibility of oocyte cryopreservation in postpubertal girls aged between 13 and 15 years who were at risk for premature ovarian failure due to the accelerated follicle loss associated with Turner syndrome or cancer treatments. Retrospective cohort and review of literature. Academic fertility preservation unit. Three girls diagnosed with Turner syndrome, 1 girl diagnosed with germ-cell tumor. and 1 girl diagnosed with lymphoblastic leukemia. Assessment of ovarian reserve, ovarian stimulation, oocyte retrieval, in vitro maturation, and mature oocyte cryopreservation. Response to ovarian stimulation, number of mature oocytes cryopreserved and complications, if any. Mean anti-müllerian hormone, baseline follical stimulating hormone, estradiol, and antral follicle counts were 1.30 ± 0.39, 6.08 ± 2.63, 41.39 ± 24.68, 8.0 ± 3.2; respectively. In Turner girls the ovarian reserve assessment indicated already diminished ovarian reserve. Ovarian stimulation and oocyte cryopreservation was successfully performed in all female children referred for fertility preservation. A range of 4-11 mature oocytes (mean 8.1 ± 3.4) was cryopreserved without any complications. All girls tolerated the procedure well. Oocyte cryopreservation is a feasible technique in selected female children at risk for premature ovarian failure. Further studies would be beneficial to test the success of oocyte cryopreservation in young girls. Copyright © 2014 North American Society for Pediatric and Adolescent Gynecology. Published by Elsevier Inc. All rights reserved.

  15. Follicles of various maturation stages react differently to enzymatic isolation: a comparison of different isolation protocols.

    PubMed

    Lierman, Sylvie; Tilleman, Kelly; Cornelissen, Maria; De Vos, Winnok H; Weyers, Steven; T'Sjoen, Guy; Cuvelier, Claude A; De Sutter, Petra

    2015-02-01

    Isolation of human follicles is based on digestion of the tissue by combinations of enzymes. Follicle vitality and morphology are often based on the analysis of pooled follicles of different maturation stages. Information is therefore lacking on the effect of the isolation protocol to individual follicles of different maturation stages. A study was conducted using five protocols combining different enzymes and varying concentrations. Isolated follicles were classified according to their maturation stages, counted and characterized for vitality, morphology, early apoptosis and organization of transzonal projections. No statistical differences were found between the protocols when outcome parameters were analysed on a pool of follicles regardless of their maturation status. Differences were observed in quality when the follicles were analysed separately according to their maturation status. Combining morphologic characteristics and vitality, both Liberase DH and Liberase TM combined with collagenase IV were better at isolating high-quality primordial follicles, compared with collagenase IV. No statistical difference between the isolation protocols was found for primary follicles. If only high-quality isolated secondary follicles are needed, collagenase IV is found to be most advantageous. Follicles of different maturation stages react differently when enzymatic isolation protocols are compared. Copyright © 2014 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

  16. Maxillary antral bone grafts for repair of orbital fractures.

    PubMed

    Copeland, M; Meisner, J

    1991-04-01

    Use of bone from the maxillary antrum to repair defects in the orbital floor was described more than 20 years ago but has not been reported for correction of orbital rim fractures. The method is appealing because the source is contiguous with the recipient site; enhanced exposure might allow better fracture reduction and evacuation of debris and hematoma from the maxillary sinus. The intraoral approach also avoids an external incision and scar, prevents such complications as pneumothorax or dural perforation, and reduces postoperative pain. In 60 cases of orbital and zygomatic complex fractures seen between 1985 and 1990, less than 8% required more extensive graft material than the maxillary antra could provide. To assess the potential advantages of local over extraanatomical bone grafts, we evaluated maxillary antral bone grafts obtained through buccal sulcus incisions in 14 patients for restoration following fractures of the orbit. Several of these patients are described. Bone union was complete in all patients and there was no morbidity related to infection, oroantral fistula formation, dehiscence, or disfigurement. Sufficient bone was available from the uninvolved contralateral side to repair even severely comminuted fractures. In zygomatic complex fractures, maxillary antral grafts appeared to provide additional strength in the region of the fractured maxillary buttress. The success of the procedure in our experience, coupled with the safety of bone harvesting from this source, and the avoidance of an external scar make maxillary antral bone well suited to reconstruction of all areas of the orbit.

  17. Somatostatin modulates cholinergic neurotransmission in canine antral muscle

    SciTech Connect

    Koelbel, C.B.; van Deventer, G.; Khawaja, S.; Mogard, M.; Walsh, J.H.; Mayer, E.A. UCLA Medical Center, Torrance, CA )

    1988-02-01

    Somatostatin has been shown to inhibit antral motility in vivo. To examine the effect of somatostatin on cholinergic neurotransmission in the canine antrum, we studied the mechanical response of and the release of ({sup 3}H)acetylcholine from canine longitudinal antral muscle in response to substance P, gastrin 17, and electrical stimulation. In unstimulated tissues, somatostatin had a positive inotropic effect on spontaneous phasic contractions. In tissues stimulated with substance P and gastrin 17, but not with electrical stimulation, somatostatin inhibited the phasic inotropic response dose dependently. This inhibitory effect was abolished by indomethacin. Somatostatin stimulated the release of prostaglandin E{sub 2} radioimmunoreactivity, and prostaglandin E{sub 2} inhibited the release of ({sup 3}H)acetylcholine induced by substance P and electrical stimulation. Somatostatin increased the release of ({sup 3}H)acetylcholine from unstimulated tissues by a tetrodotoxin-sensitive mechanism but inhibited the release induced by substance P and electrical stimulation. These results suggest that somatostatin has a dual modulatory effect on cholinergic neutrotransmission in canine longitudinal antral muscle. This effect is excitatory in unstimulated tissues and inhibitory in stimulated tissues. The inhibitory effect is partially mediated by prostaglandins.

  18. Context based algorithmic framework for identifying and classifying embedded images of follicle units

    NASA Astrophysics Data System (ADS)

    Rahman, Md. Mahbubur; Iyengar, S. S.; Zeng, Wei; Hernandez, Frank; Nusbaum, Bernard P.; Rose, Paul

    2014-03-01

    Medical image processing has been very emerging research areas in recent days. These types of images are naturally so noisy. To count the target objects is never easy. But the proper treatment depends on the accuracy of the successful locating and counting of the desired objects in an image. Some research work can do this type of segmentation of images, but they include so many constraints on the input images that these solutions cannot be applied in a generalized way to most of the images. Even a slight variation in nature of an input image can lead to a major incorrectness of the result. So we developed a generalized way to count a very noisy part of human body, the hair follicle on the scalp. The objective of this research is to count the number of hair follicle groups and the number of follicles into each group in a microscopic image of human scalp. The follicles are nonstandard in shape i.e. they do not maintain any standard shape like rectangle, oval, circle etc. Moreover the follicles are overlapping with one another in many cases. So it is hard to separate them. Here we will present a technique to count the number of follicle group as well as number of follicles in each group. We also applied well-known techniques to cluster the objects detected and a method to generate a neighboring connected graph in order to calculate the inter follicular distances.

  19. Knockout of RNA Binding Protein MSI2 Impairs Follicle Development in the Mouse Ovary: Characterization of MSI1 and MSI2 during Folliculogenesis

    PubMed Central

    Sutherland, Jessie M.; Sobinoff, Alexander P.; Gunter, Kara M.; Fraser, Barbara A.; Pye, Victoria; Bernstein, Ilana R.; Boon, Evan; Siddall, Nicole A.; De Andres, Luisa I.; Hime, Gary R.; Holt, Janet E.; Graf, Thomas; McLaughlin, Eileen A.

    2015-01-01

    Characterizing the mechanisms underlying follicle development in the ovary is crucial to understanding female fertility and is an area of increasing research interest. The RNA binding protein Musashi is essential for post-transcriptional regulation of oocyte maturation in Xenopus and is expressed during ovarian development in Drosophila. In mammals Musashi is important for spermatogenesis and male fertility, but its role in the ovary has yet to be characterized. In this study we determined the expression of mammalian Musashi proteins MSI1 and MSI2 during mouse folliculogenesis, and through the use of a MSI2-specific knockout mouse model we identified that MSI2 is essential for normal follicle development. Time-course characterization of MSI1 and MSI2 revealed distinct differences in steady-state mRNA levels and protein expression/localization at important developmental time-points during folliculogenesis. Using a gene-trap mouse model that inactivates Msi2, we observed a significant decrease in ovarian mass, and change in follicle-stage composition due to developmental blocking of antral stage follicles and pre-antral follicle loss through atresia. We also confirmed that hormonally stimulated Msi2-deficient mice produce significantly fewer MII oocytes (60.9% less than controls, p < 0.05). Furthermore, the majority of these oocytes are of poor viability (62.2% non-viable/apoptotic, p < 0.05), which causes a reduction in female fertility evidenced by decreased litter size in Msi2-deficient animals (33.1% reduction to controls, p < 0.05). Our findings indicate that MSI1 and MSI2 display distinct expression profiles during mammalian folliculogenesis and that MSI2 is required for pre-antral follicle development. PMID:26131972

  20. Serum luteinizing hormone level and luteinizing hormone/follicle-stimulating hormone ratio but not serum anti-Müllerian hormone level is related to ovarian volume in Korean women with polycystic ovary syndrome.

    PubMed

    Chun, Sungwook

    2014-06-01

    The aim of the present study was to investigate the relationship between ovarian follicle count and volume on ultrasonography and serum hormone levels including the levels of the anti-Müllerian hormone (AMH) and gonadotropin in women with the polycystic ovary syndrome (PCOS). A total of 118 Korean women aged 18-35 years who were newly diagnosed with PCOS at a university hospital were included in this study. Serum LH, FSH, and AMH levels were measured in the early follicular phase, and the total antral follicle count (TFC) and the total ovarian volume (TOV) were assessed by ultrasonography. The correlations between serum hormonal parameters and ultrasonography characteristics in women with PCOS were evaluated using Pearson's correlation coefficients and a linear regression analysis. Serum AMH levels were significantly correlated with serum LH levels and LH/FSH ratios, and TFC and TOV were significantly correlated with each other on ultrasonography. Serum AMH and LH levels and the LH/FSH ratio were significantly correlated with TFC. Statistically significant correlations between TOV and the LH level (r=0.208, p=0.024) and the LH/FSH ratio (r=0.237, p=0.010) were observed. However, the serum AMH level was not significantly correlated with the ovarian volume, and this result did not change after adjusting for age and body mass index. Serum AMH is not related to the ovarian volume in women with PCOS. My results suggest that serum LH level and the LH/FSH ratio may be more useful than the serum AMH level for representing the status of the ovarian volume in women with PCOS.

  1. Transcriptome profiling of granulosa cells from bovine ovarian follicles during atresia

    PubMed Central

    2014-01-01

    Background The major function of the ovary is to produce oocytes for fertilisation. Oocytes mature in follicles surrounded by nurturing granulosa cells and all are enclosed by a basal lamina. During growth, granulosa cells replicate and a large fluid-filled cavity (the antrum) develops in the centre. Only follicles that have enlarged to over 10 mm can ovulate in cows. In mammals, the number of primordial follicles far exceeds the numbers that ever ovulate and atresia or regression of follicles is a mechanism to regulate the number of oocytes ovulated and to contribute to the timing of ovulation. To better understand the molecular basis of follicular atresia, we undertook transcriptome profiling of granulosa cells from healthy (n = 10) and atretic (n = 5) bovine follicles at early antral stages (< 5 mm). Results Principal Component Analysis (PCA) and hierarchical classification of the signal intensity plots for the arrays showed primary clustering into two groups, healthy and atretic. These analyses and size-frequency plots of coefficients of variation of signal intensities revealed that the healthy follicles were more heterogeneous. Examining the differentially-expressed genes the most significantly affected functions in atretic follicles were cell death, organ development, tissue development and embryonic development. The overall processes influenced by transcription factor gene TP53 were predicted to be activated, whereas those of MYC were inhibited on the basis of known interactions with the genes in our dataset. The top ranked canonical pathway contained signalling molecules common to various inflammatory/fibrotic pathways such as the transforming growth factor-β and tumour necrosis factor-α pathways. The two most significant networks also reflect this pattern of tissue remodelling/fibrosis gene expression. These networks also contain molecules which are present in the canonical pathways of hepatic fibrosis/hepatic stellate cell activation and transforming

  2. A Morphometric Study of Antral G-Cell Density in a Sample of Adult General Population: Comparison of Three Different Methods and Correlation with Patient Demography, Helicobacter pylori Infection, Histomorphology and Circulating Gastrin Levels

    PubMed Central

    Petersson, Fredrik; Borch, Kurt; Rehfeld, Jens F; Franzén, Lennart E

    2009-01-01

    Helicobacter pylori infection has been linked to hypergastrinemia and either decreased or normal G-cell content in the antral mucosa. To clarify this controversial issue, we quantitatively determined antral G-cell content on the same biopsy specimens with three different methods and examined whether these methods are intercorrelated and the relation of these methods to plasma gastrin concentrations, demography, the occurrence of H. pylori infection and chronic gastritis. Gastric antral mucosal biopsy sections from 273 adults (188 with and 85 without H pylori infection) from a general population sample were examined immunohistochemically for G-cells using cell counting, stereology (point counting) and computerized image analysis. Gastritis was scored according to the updated Sydney system. Basal plasma gastrin concentrations were measured by radioimmunoassay. The three methods for G-cell quantification were poorly correlated and the results showed no correlation with basal plasma gastrin concentrations. The antral G-cell density and scores for H. pylori colonization were positively related to age. Neither the scores for chronic inflammation, nor the scores for inflammatory activity, atrophy or intestinal metaplasia were consistently related to the antral G-cell content. In conclusion, the results of three techniques for G-cell quantification in the gastric antral mucosa were poorly intercorrelated and none of the methods correlated with plasma gastrin concentrations. Age and scores for H pylori colonization seem to be determinants of the G-cell density. That common morphometric techniques correlate poorly is of utmost importance to bear in mind when quantitative morphological studies are planned, compared or interpreted. PMID:19079618

  3. Hair Follicle Pigmentation

    PubMed Central

    Slominski, Andrzej; Wortsman, Jacobo; Plonka, Przemyslaw M.; Schallreuter, Karin U.; Paus, Ralf; Tobin, Desmond J.

    2005-01-01

    Hair shaft melanin components (eu- or/and pheomelanin) are a long-lived record of precise interactions in the hair follicle pigmentary unit, e.g., between follicular melanocytes, keratinocytes, and dermal papilla fibroblasts. Follicular melanogenesis (FM) involves sequentially the melanogenic activity of follicular melanocytes, the transfer of melanin granules into cortical and medulla keratinocytes, and the formation of pigmented hair shafts. This activity is in turn regulated by an array of enzymes, structural and regulatory proteins, transporters, and receptors and their ligands, acting on the developmental stages, cellular, and hair follicle levels. FM is stringently coupled to the anagen stage of the hair cycle, being switched-off in catagen to remain absent through telogen. At the organ level FM is precisely coupled to the life cycle of melanocytes with changes in their compartmental distribution and accelerated melanoblast/melanocyte differentiation with enhanced secretory activity. The melanocyte compartments in the upper hair follicle also provides a reservoir for the repigmentation of epidermis and, for the cyclic formation of new anagen hair bulbs. Melanin synthesis and pigment transfer to bulb keratinocytes are dependent on the availability of melanin precursors, and regulation by signal transduction pathways intrinsic to skin and hair follicle, which are both receptor dependent and independent, act through auto-, para- or intracrine mechanisms and can be modified by hormonal signals. The important regulators are MC1 receptor its and adrenocorticotropic hormone, melanocyte stimulating hormone, agouti protein ligands (in rodents), c-Kit, and the endothelin receptors with their ligands. Melanin itself has a wide range of bioactivities that extend far beyond its determination of hair color. PMID:15654948

  4. Follicle Loss and Apoptosis in Cyclophosphamide-Treated Mice: What’s the Matter?

    PubMed Central

    Chen, Xiu-Ying; Xia, He-Xia; Guan, Hai-Yun; Li, Bin; Zhang, Wei

    2016-01-01

    With increasing numbers of young female cancer survivors following chemotherapy, chemotherapy-induced fertility loss must be considered. Menstrual disorder and infertility are of particular concern in female cancer patients. We showed that treatment with the alkylating agent cyclophosphamide (CTX) could cause severe primordial follicle loss and growing follicle apoptosis, resulting in loss of ovarian reserve. SPF C57BL/6 female mice were treated with a single dose of 120 mg/kg of CTX or saline as a control, and both sides of ovaries were collected three or seven days after injection. Following CTX treatment, the ovaries were mostly composed of collapsed oocytes and presented marked cortical fibrosis and a reduced number of follicles, especially primordial follicles. The loss of primordial follicles was confirmed by primordial follicle counting, immunohistochemistry and Western blot detection of DDx4/MVH. Follicle apoptosis was tested by a TUNEL assay and the number of TUNEL-positive follicle cells increased, as expected, in CTX-treated mice. Furthermore, expression of APAF-1 and cleaved caspase-3 was also increased after CTX treatment. Analysis of the PI3K/Akt/mTOR signaling pathway showed that CTX increased phosphorylation of Akt, mTOR and downstream proteins without affecting total levels. These results demonstrated that the CTX treatment led to the hyperactivation of the PI3K/Akt/mTOR signaling pathway in ovaries which may be related to primordial follicle loss and growing follicle apoptosis. PMID:27248997

  5. Transcriptomic diversification of developing cumulus and mural granulosa cells in mouse ovarian follicles.

    PubMed

    Wigglesworth, Karen; Lee, Kyung-Bon; Emori, Chihiro; Sugiura, Koji; Eppig, John J

    2015-01-01

    Cumulus cells and mural granulosa cells (MGCs) have functionally distinct roles in antral follicles, and comparison of their transcriptomes at a global and systems level can propel future studies on mechanisms underlying their functional diversity. These cells were isolated from small and large antral follicles before and after stimulation of immature mice with gonadotropins, respectively. Both cell types underwent dramatic transcriptomic changes, and differences between them increased with follicular growth. Although cumulus cells of both stages of follicular development are competent to undergo expansion in vitro, they were otherwise remarkably dissimilar with transcriptomic changes quantitatively equivalent to those of MGCs. Gene ontology analysis revealed that cumulus cells of small follicles were enriched in transcripts generally associated with catalytic components of metabolic processes, while those from large follicles were involved in regulation of metabolism, cell differentiation, and adhesion. Contrast of cumulus cells versus MGCs revealed that cumulus cells were enriched in transcripts associated with metabolism and cell proliferation while MGCs were enriched for transcripts involved in cell signaling and differentiation. In vitro and in vivo models were used to test the hypothesis that higher levels of transcripts in cumulus cells versus MGCs is the result of stimulation by oocyte-derived paracrine factors (ODPFs). Surprisingly ∼48% of transcripts higher in cumulus cells than MGCs were not stimulated by ODPFs. Those stimulated by ODPFs were mainly associated with cell division, mRNA processing, or the catalytic pathways of metabolism, while those not stimulated by ODPFs were associated with regulatory processes such as signaling, transcription, phosphorylation, or the regulation of metabolism.

  6. The domestic dog and cat as models for understanding the regulation of ovarian follicle development in vitro

    PubMed Central

    Songsasen, N.; Comizzoli, P.; Nagashima, J.; Fujihara, M.; Wildt, D.E

    2012-01-01

    The culture of ovarian follicles is an important tool for understanding of the mechanisms controlling follicle development and differentiation of its oocyte. The benefit of recovering meiotically and developmentally competent oocytes from early stage follicles (primordial, primary, preantral and early antral) also would be significant, ranging from rescue of genomes from endangered species to preserving fertility in women facing cancer treatments. This field of research is at an early stage of scientific discovery. To-date, live offspring from cultured primordial follicles that produced fertilizable oocytes has occurred only in the mouse. Progress in other more complex species has been limited because larger animals have longer durations of natural folliculogenesis, thereby requiring more culture time to generate fully grown follicles and oocytes. We believe the dog and cat are excellent models for understanding more about folliculogenesis in vitro. This review highlights what is known about this topic for these two species as well as future priorities. In brief, it is more challenging to maintain viability of primordial follicles within ovarian tissues in vitro in the dog than the cat. Nonetheless, it is possible to grow both isolated cat and dog preantral follicles in culture. Although the follicles of both species have the capacity to increase in size and produce steroids, only cat oocytes are morphologically normal. This striking difference between the dog and cat is an area of high research priority. While much more fundamental data are required, we envision advanced technology that will allow harvesting oocytes from the vast, unused follicle stores sequestered within carnivore ovaries. These gametes have utility for reproducing genetically valuable dogs and cats that are ‘companions’ or biomedical models for investigating human disorders or for salvaging the genomes of rare canid and felid species that die before contributing to genetic management

  7. Endoscopic features of lymphoid follicles in Helicobacter pylori-associated chronic gastritis.

    PubMed

    Hayashi, Seishu; Imamura, Jun; Kimura, Kiminori; Saeki, Shunichi; Hishima, Tsunekazu

    2015-01-01

    Small, round, yellowish-white nodules (YWN) are frequently observed in Helicobacter pylori-associated gastritis. The aim of the present study was to investigate the clinical significance of these YWN. Participants comprised 211 patients with H. pylori-associated gastritis, ranging in age from 23 to 86 years. YWN were detected in 23% of participants, more frequently in women (33%) than in men (12%; P < 0.01). YWN were observed on the antral mucosa in 4.7% of cases, lesser curvature of the corpus mucosa in 20%, greater curvature of the corpus mucosa in 0.9%, and fundic mucosa in 12%. Most YWN located on the antral mucosa showed nodular type, and most YWN located on the corpus mucosa and fundic mucosa showed flat type. On magnifying endoscopy with narrow-band imaging, YWN appeared as round whitish lesions with radial or branching microvessels on the surface and hypovascular globe structures just beneath the surface of the mucosa. Targeted biopsies of YWN revealed lymphoid follicles with lymphocyte infiltration or intense inflammatory cell infiltration. The endoscopic finding of YWN could be observed at any site of the gastric mucosa in H. pylori-associated gastritis, and represented histological lymphoid follicles. © 2014 The Authors. Digestive Endoscopy © 2014 Japan Gastroenterological Endoscopy Society.

  8. Differences between rats and mice in the involvement of the aryl hydrocarbon receptor in 4-vinylcyclohexene diepoxide-induced ovarian follicle loss

    SciTech Connect

    Thompson, Kary E.; Bourguet, Shannon M.; Christian, Patricia J.; Benedict, Jamie C.; Sipes, I. Glenn; Flaws, Jodi A.; Hoyer, Patricia B. . E-mail: hoyer@u.arizona.edu

    2005-03-01

    Repeated dosing with the occupational chemical 4-vinylcyclohexene diepoxide (VCD) selectively depletes small pre-antral follicles in the ovaries of rats and mice via apoptosis. The aryl hydrocarbon receptor (AhR) plays a role in mediating the effects of several xenobiotics. Therefore, this study was designed to investigate a potential role of the AhR in VCD-induced ovotoxicity. Female F344 rats, C57BL/6 mice, or AhR-deficient (-/-, AhRKO) mice were dosed daily (15 days) with vehicle, VCD (80 mg/kg, i.p.) and/or the AhR antagonist, alpha-naphthoflavone (ANF; 80 mg/kg, i.p.). Compared with controls, VCD caused a 60% reduction (P < 0.05) in primordial and primary follicles in mice and rats. Concurrent dosing with ANF protected against the VCD-induced follicle loss in rats, but not in mice. As with AhR-intact mice and rats, VCD induced a 70% loss (P < 0.05) of small pre-antral follicles in AhRKO mice. AhR mRNA expression was increased (P < 0.05) by VCD dosing in small pre-antral follicles isolated from ovaries of rats but not mice. AhR protein in rats was increased by VCD dosing in oocyte nuclei in primordial and primary follicles when measured by immunofluorescence and confocal microscopy. In rat small pre-antral follicles, apoptosis-associated caspase-3-like activity was increased (P < 0.05) by VCD treatment, decreased (P < 0.05) by ANF treatment, and unaffected by VCD plus ANF treatment. VCD had no effect on expression of GST Ya1 or GST Ya2 mRNA or CYP 1A1 protein in rats. Taken together, these findings demonstrate a difference between rats and mice in the potential involvement of AhR as related to VCD-induced ovotoxicity. Whereas, AhR appears to be involved in rats, no evidence for a similar role in mice was obtained. Overall, these findings point out that there can be mechanistic species differences in ovarian responses to xenobiotic chemicals.

  9. Notch signaling regulates gastric antral LGR5 stem cell function.

    PubMed

    Demitrack, Elise S; Gifford, Gail B; Keeley, Theresa M; Carulli, Alexis J; VanDussen, Kelli L; Thomas, Dafydd; Giordano, Thomas J; Liu, Zhenyi; Kopan, Raphael; Samuelson, Linda C

    2015-10-14

    The major signaling pathways regulating gastric stem cells are unknown. Here we report that Notch signaling is essential for homeostasis of LGR5(+) antral stem cells. Pathway inhibition reduced proliferation of gastric stem and progenitor cells, while activation increased proliferation. Notch dysregulation also altered differentiation, with inhibition inducing mucous and endocrine cell differentiation while activation reduced differentiation. Analysis of gastric organoids demonstrated that Notch signaling was intrinsic to the epithelium and regulated growth. Furthermore, in vivo Notch manipulation affected the efficiency of organoid initiation from glands and single Lgr5-GFP stem cells, suggesting regulation of stem cell function. Strikingly, constitutive Notch activation in LGR5(+) stem cells induced tissue expansion via antral gland fission. Lineage tracing using a multi-colored reporter demonstrated that Notch-activated stem cells rapidly generate monoclonal glands, suggesting a competitive advantage over unmanipulated stem cells. Notch activation was associated with increased mTOR signaling, and mTORC1 inhibition normalized NICD-induced increases in proliferation and gland fission. Chronic Notch activation induced undifferentiated, hyper-proliferative polyps, suggesting that aberrant activation of Notch in gastric stem cells may contribute to gastric tumorigenesis. © 2015 The Authors.

  10. Characterization of the effects of neurokinins on canine antral muscle

    SciTech Connect

    Koelbel, C.B.; Mayer, E.A.; Van Deventer, G.; Snape, W.J. Jr.; Patel, A. Wadsworth Hospital, Los Angeles, CA )

    1988-12-01

    The excitation of longitudinal antral muscle by substance P (SP) involves both a myogenic and a cholinergic effect. To examine if these responses are mediated by different neurokinin receptors, the authors studied the mechanical response and the release of ({sup 3}H)acetylcholine from antral muscle strips in response to SP, substance P methylester (SPME), neurokinin A (NKA), neurokinin B (NKB), and several nonmammalian tachykinins. All peptides studied showed a dose-dependent inotropic and chronotropic effect on spontaneous phasic contractions. This ionotropic effect in longitudinal muscle was partially atropine sensitive for SPME, SP, and NKB but not for NKA, whereas neither atropine nor tetrodotoxin had an effect in circular muscle. In longitudinal muscle, all three neurokinins were equipotent. In longitudinal muscle treated with atropine and in circular muscle, the rank order of potency for the inotropic response was NKA > NKB > SP > SPME. For the chronotropic response the rank order was SPME, SP > NKA > NKB. NKA, NKB, and SP caused a dose-dependent, tetrodotoxin-sensitive increase in ({sup 3}H)acetylcholine release from strips preincubated with ({sup 3}H)choline. NKA was significantly more potent to release ({sup 3}H)acetylcholine than either NKB or SP. The stimulated release was inhibited by (D-Ala{sup 2},D-Met{sup 5})methionine enkephalinamide and the SP antagonist, spantide. These results are consistent with the hypothesis that NKA is the natural ligand mediating the myogenic inotropic response in both muscle layers and the cholinergic response in longitudinal muscle.

  11. Novel sonographic clues for diagnosis of antral gastritis and Helicobacter pylori infection: a clinical study.

    PubMed

    Cakmakci, Emin; Ucan, Berna; Colak, Bayram; Cinar, Hasibe Gokçe

    2014-09-01

    The purpose of this study was to find out whether transabdominal sonography may have a predictive role for detection of antral gastritis and Helicobacter pylori infection in the antrum. A total of 108 patients and 54 control participants were allocated into 3 groups: group 1, controls without any symptoms or findings of antral gastritis and H pylori infection; group 2, patients with symptoms and endoscopic findings consistent with gastritis in the absence of documented H pylori infection; and group 3, patients with symptoms and endoscopic findings consistent with gastritis and documented H pylori infection. These groups were compared in terms of demographics, antral wall thickness, mucosal layer (together with muscularis mucosa) thickness, and mucosal layer-to-antral wall thickness ratio. The groups had no statistically significant differences with respect to age, sex, body mass index, and smoking habits. However, it turned out that both antral walls and muscularis mucosa layers were thicker and the mucosal layer-to-antral wall thickness ratio was higher in groups 2 and 3 compared to group 1 (P > .001). In addition, group 3 had statistically significantly thicker antral walls and muscularis mucosa layers and a significantly increased mucosal layer-to-antral wall thickness ratio than group 2 (P < .001). Our results suggest that antral gastritis caused by H pylori infection is associated with characteristic features such as thickening of antral walls and mucosal layers on sonography. These novel clues may be useful in the diagnosis of gastritis, and unnecessary interventions and measures can be avoided in some cases. © 2014 by the American Institute of Ultrasound in Medicine.

  12. Basic fibroblast growth factor promotes the development of human ovarian early follicles during growth in vitro.

    PubMed

    Wang, Tian-ren; Yan, Li-ying; Yan, Jie; Lu, Cui-ling; Xia, Xi; Yin, Tai-lang; Zhu, Xiao-hui; Gao, Jiang-man; Ding, Ting; Hu, Wei-hong; Guo, Hong-yan; Li, Rong; Qiao, Jie

    2014-03-01

    What is the effect of basic fibroblast growth factor (bFGF) on the growth of individual early human follicles in a three-dimensional (3D) culture system in vitro? The addition of 200 ng bFGF/ml improves human early follicle growth, survival and viability during growth in vitro. It has been demonstrated that bFGF enhances primordial follicle development in human ovarian tissue culture. However, the growth and survival of individual early follicles in encapsulated 3D culture have not been reported. The maturation in vitro of human ovarian follicles was investigated. Ovarian tissue (n= 11) was obtained from 11 women during laparoscopic surgery for gynecological disease, after obtaining written informed consent. One hundred and fifty-four early follicles were isolated by enzymic digestion and mechanical disruption. They were individually encapsulated into alginate (1% w/v) and randomly assigned to be cultured with 0, 100, 200 or 300 ng bFGF/ml for 8 days. Individual follicles were cultured in minimum essential medium α (αMEM) supplemented with bFGF. Follicle survival and growth were assessed by microscopy. Follicle viability was evaluated under confocal laser scanning microscope following Calcein-AM and Ethidium homodimer-I (Ca-AM/EthD-I) staining. After 8 days in culture, all 154 follicles had increased in size. The diameter and survival rate of the follicles and the percentage with good viability were significantly higher in the group cultured with 200 ng bFGF/ml than in the group without bFGF (P < 0.05). The percentage of follicles in the pre-antral stage was significantly higher in the 200 ng bFGF/ml group than in the group without bFGF (P < 0.05), while the percentages of primordial and primary follicles were significantly lower (P < 0.05). The study focuses on the effect of bFGF on the development of individual human early follicles in 3D culture in vitro and has limited ability to reveal the specific effect of bFGF at each different stage. The findings

  13. The hedgehog system in ovarian follicles of cattle selected for twin ovulations and births: evidence of a link between the IGF and hedgehog systems.

    PubMed

    Aad, Pauline Y; Echternkamp, Sherrill E; Sypherd, David D; Schreiber, Nicole B; Spicer, Leon J

    2012-10-01

    Hedgehog signaling is involved in regulation of ovarian function in Drosophila, but its role in regulating mammalian ovarian folliculogenesis is less clear. Therefore, gene expression of Indian hedgehog (IHH) and its type 1 receptor, patched 1 (PTCH1), were quantified in bovine granulosa (GC) or theca (TC) cells of small (1-5 mm) antral follicles by in situ hybridization and of larger (5-17 mm) antral follicles by real-time RT-PCR from ovaries of cyclic cows genetically selected (Twinner) or not selected (control) for twin ovulations. Expression of IHH mRNA was localized to GC and cumulus cells, whereas PTCH1 mRNA was greater in TC than in GC. Estrogen-active (E-A; follicular fluid concentration of estradiol > progesterone) versus estrogen-inactive follicles had a greater abundance of mRNA for IHH in GC and PTCH1 in TC. Abundance of IHH mRNA in GC was not affected by cow genotype, whereas TC PTCH1 mRNA was less in large E-A follicles of Twinners than in controls. In vitro, estradiol and wingless-type (WNT) 3A increased IHH mRNA in IGF1-treated GC. IGF1 and BMP4 treatments decreased PTCH1 mRNA in small TC. Estradiol and LH increased PTCH1 mRNA in IGF1-treated TC from large and small follicles, respectively. In summary, functional status of ovarian follicles was associated with differences in hedgehog signaling in GC and TC. We hypothesize that as follicles grow and develop, increased free IGF1 may suppress expression of IHH mRNA by GC and PTCH1 mRNA by TC, and these effects are regulated in a paracrine way by estradiol and other intra- and extragonadal factors.

  14. Mechanisms mediating cholinergic antral circular smooth muscle contraction in rats

    PubMed Central

    Wrzos, Helena F; Tandon, Tarun; Ouyang, Ann

    2004-01-01

    AIM: To investigate the pathway (s) mediating rat antral circular smooth muscle contractile responses to the cholinomimetic agent, bethanechol and the subtypes of muscarinic receptors mediating the cholinergic contraction. METHODS: Circular smooth muscle strips from the antrum of Sprague-Dawley rats were mounted in muscle baths in Krebs buffer. Isometric tension was recorded. Cumulative concentration-response curves were obtained for (+)-cis-dioxolane (cD), a nonspecific muscarinic agonist, at 10-8-10-4 mol/L, in the presence of tetrodotoxin (TTX, 10-7 mol/L). Results were normalized to cross sectional area. A repeat concentration-response curve was obtained after incubation of the muscle for 90 min with antagonists for M1 (pirenzepine), M2 (methoctramine) and M3 (darifenacin) muscarinic receptor subtypes. The sensitivity to PTX was tested by the ip injection of 100 mg/kg of PTX 5 d before the experiment. The antral circular smooth muscles were removed from PTX-treated and non-treated rats as strips and dispersed smooth muscle cells to identify whether PTX-linked pathway mediated the contractility to bethanechol. RESULTS: A dose-dependent contractile response observed with bethanechol, was not affected by TTX. The pretreatment of rats with pertussis toxin decreased the contraction induced by bethanechol. Lack of calcium as well as the presence of the L-type calcium channel blocker, nifedipine, also inhibited the cholinergic contraction, with a reduction in response from 2.5 ± 0.4 g/mm2 to 1.2 ± 0.4 g/mm2 (P < 0.05). The dose-response curves were shifted to the right by muscarinic antagonists in the following order of affinity: darifenacin (M3) > methocramine (M2) > pirenzepine (M1). CONCLUSION: The muscarinic receptors-dependent contraction of rat antral circular smooth muscles was linked to the signal transduction pathway(s) involving pertussis-toxin sensitive GTP-binding proteins and to extracellular calcium via L-type voltage gated calcium channels. The

  15. Growth and antrum formation of bovine primary follicles in long-term culture in vitro.

    PubMed

    Sun, Jing; Li, Xiangdong

    2013-09-01

    Successful antral formation in vitro from bovine preantral follicles (145-170 μm) has been described previously, but antrum formation from the primary follicle (50-70 μm) has not yet been achieved in vitro. The aim of the study was to establish an optimal culture system supporting the growth and maturation of bovine primary follicles (50-70 μm) in vitro. Bovine primary follicles were cultured in a three-dimensional culture system for 13 or 21 days in alpha-minimum essential medium. Various treatments including follicle stimulating hormone (FSH), luteinizing hormone (LH), 17β-estradiol (E2), basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF) were tested. The follicular diameter and antrum formation rate were recorded, and follicular maturation markers (P450 aromatase, CYP19A1; anti-Mullerian hormone, AMH; growth differentiation factor-9, GDF9; bone morphogenetic protein-15, BMP15; and type III transforming growth factor β receptor, TGFβR3) were analyzed by real-time RT-PCR. After 21 days of culture under each treatment condition, the follicular diameter was significantly enlarged in the presence of FSH + LH + E2 + bFGF or FSH + LH + E2 + bFGF + EGF (p<0.05). An addition of 50 ng/ml bFGF or bFGF +25 ng/ml EGF initiated antrum formation by day 19 and day 17 of culture, and the antral cavity formation rate was 16.7% and 33.3% by 21 days of culture, respectively. The expression of follicular maturation markers (CYP19A1, AMH, GDF9, BMP15 and TGFβR3) was significantly altered. We conclude that addition of 50 ng/ml bFGF +25 ng/ml EGF to media containing FSH + LH + E2 turned out to be the most effective optimized culture conditions to support the growth and maturation of bovine primary follicles in vitro.

  16. Resveratrol has dose-dependent effects on DNA fragmentation and mitochondrial activity of ovine secondary follicles cultured in vitro.

    PubMed

    Macedo, T J S; Barros, V R P; Monte, A P O; Gouveia, B B; Bezerra, M É S; Cavalcante, A Y P; Barberino, R S; Menezes, V G; Matos, M H T

    2017-08-01

    The worldwide consumption of red wine, nuts and grapes has resulted in increased human exposure to resveratrol, which could affect reproductive function. However, the effect of resveratrol on in vitro culture of early-stage ovarian follicles has never been investigated. The aims of the present study were to evaluate the effect of resveratrol on sheep secondary follicle morphology, growth, DNA fragmentation, intracellular levels of glutathione (GSH) and active mitochondria. Secondary follicles were isolated from the ovaries and cultured for 18 days in supplemented α-MEM+ (control medium) or in control medium containing resveratrol (2, 10 or 30 µM). The parameters analyzed were morphology, antrum formation, follicle diameter, DNA fragmentation, GSH levels and mitochondrial activity. After 18 days, all resveratrol groups significantly decreased the percentages of morphologically normal follicles compared with the control group (α-MEM+). Antrum formation was higher in both α-MEM+ and 2 µM resveratrol groups than in the 10 µM resveratrol group. In addition, 30 µM resveratrol increased the percentage of oocytes with DNA damage compared with the control. Oocytes from follicles treated with 10 or 30 µM resveratrol significantly decreased intracellular GSH levels compared with the 2 µM resveratrol group. Moreover, follicles in α-MEM+ (control) showed more active mitochondria than those in 10 or 30 µM resveratrol. In conclusion, ovine isolated secondary follicles are able to grow to the antral stage after in vitro culture in medium containing 2 µM resveratrol, maintaining the same rates of DNA damage, GSH levels and mitochondrial function as the control medium. However, the addition of 30 µM resveratrol increased DNA fragmentation and oxidative stress through decreasing mitochondrial activity.

  17. Hair follicle differentiation and regulation.

    PubMed

    Rogers, George E

    2004-01-01

    Ten years ago, Hardy (1992) wrote a timely review on the major features of hair follicle development and hair growth which she referred to as a secret life. Many of these secrets are now being revealed. The information discussed in this brief review comprises the structure of the hair and hair follicle, the continuing characterisation of the genes for keratin and keratin associated proteins, the determination of the location of their expression in the different cell layers of the hair follicle, molecular signals which control keratin gene expression and post-translational events in the terminal stages of hair formation.

  18. LAPS-FSH: a new and effective long-acting follicle-stimulating hormone analogue for the treatment of infertility.

    PubMed

    Jung, Sunyoung; Park, Youngjin; Kim, YoungHoon; Kim, Yu Yon; Choi, Hyun-Ji; Son, Woo-Chan; Kwon, SeChang

    2014-10-01

    Although several long-acting follicle-stimulating hormone (FSH) therapies have been developed to enhance the ovarian response, a disadvantage of FSH therapy is its relatively short half-life, which requires women to receive one to two injections per day for almost 2 weeks. In the present study, we developed a novel FSH analogue by conjugating recombinant human FSH (rhFSH) and the constant region of the human immunoglobulin G4 fragment via non-peptidyl linkers. The efficacy of the FSH analogue was evaluated in vitro by cAMP level assessments, pharmacokinetic studies and a determination of ovarian weight and by comparing these findings with the results from other FSH analogues. In addition, the total number of antral and Graafian follicles was determined after 7 days of treatment with control, 6µgkg(-1) follitropin β, 6, 12 or 42µgkg(-1) corifollitropin α or 3, 6 or 12µgkg(-1) long acting protein/peptide discovery-follicle-stimulating hormone (LAPS-FSH). As a result, the animals treated with 12µgkg(-1) LAPS-FSH produced additional and larger healthy follicles. These data demonstrate that LAPS-FSH promotes growth and inhibits atresia of the ovarian follicle compared with other available drugs, suggesting that our new drug enhances the efficacy and duration of treatment. It is expected that our new FSH analogue will result in a higher chance of pregnancy in patients who are unresponsive to other drugs.

  19. A study of operative treatment and bacteriological examination of persistent oro-antral fistulas.

    PubMed

    Sindet-Pedersen, S; Skoglund, L A; Hvidegaard, T; Holst, E

    1983-10-01

    A retrospective study of 35 patients with oro-antral fistulas of more than 2 weeks duration is presented. Treatment results by using the Rehrmann plastic procedure, as well as complications and sequelae are described. The results of bacteriological examination in 15 patients with maxillary sinusitis in association with oro-antral fistulas are presented. The bacteriological findings suggest that broad spectrum antibiotics in certain conditions should be administered in cases of persisting oro-antral fistulas associated with maxillary sinusitis. The results of the present study show that the Rehrmann-operation proves satisfactory even in treatment-resistent cases.

  20. A Case of Oro-antral Communication Closed by Autotransplantation of Third Molar.

    PubMed

    Nagori, Shakil Ahmed; Jose, Anson; Bhutia, Ongkila; Roychoudhury, Ajoy

    2015-03-01

    A number of surgical treatment modalities have been described in literature for closure of oro-antral communications. None of the methods however provide for immediate prosthetic rehabilitation of the communication site. We describe a case of oro-antral communication treated using autogenous third molar transplantation. At 18 months of follow-up, the transplanted tooth was functioning well with radiographic evidence of lamina dura and periodontal ligament. Its root-tip was still seen projecting into the maxillary sinus. This report suggests that when possible, autotransplantation of third molar can be a simple and excellent choice for closure of oro-antral communications.

  1. A case of gastric polyposis in antral area of stomach following prolonged proton-pump therapy

    PubMed Central

    Alqutub, Adel Nazmi; Masoodi, Ibrahim

    2010-01-01

    We describe the clinical scenario of a young male with history of non ulcer dyspepsia who had endoscopic evidence of gastric polyposis in antral area. The polyps disappeared four months after proton pump inhibitors were stopped. Proton pump inhibitors have been linked to gastric fundal polyposis and not antral gland polyposis. This is the first report originating from an Asian country describing antral gland polyposis (AGPs) in a patient on long-term PPI therapy with no evidence of Helicobacter pylori. A case report with brief review is presented. PMID:20930929

  2. Chocolate cysts from ovarian follicles.

    PubMed

    Jain, S; Dalton, M E

    1999-11-01

    To study the development of chocolate cysts by serial transvaginal ultrasonographic tracking of ovarian follicles. Retrospective study. Department of Obstetrics and Gynaecology, Sunderland Royal Hospital, Sunderland, United Kingdom. We reviewed case notes of all patients who underwent laparoscopy for diathermy to endometriosis/ovarian diathermy/aspiration of ovarian cysts from 1989 to 1998. Twelve women with histories of infertility and proven chocolate cysts with documented ultrasonographic findings were included in the study. Serial ultrasonographic tracking of ovarian follicles in the cycle leading to the development of the chocolate cysts, followed by ultrasonographic tracking of cysts for 3 months and laparoscopy. Development and formation of chocolate cysts. The diagnosis of chocolate cysts was confirmed laparoscopically in all patients and histopathologically in four. Ultrasound confirmed that they had all developed from follicles. Chocolate cysts can develop from ovarian follicles.

  3. Melatonin prevents cisplatin-induced primordial follicle loss via suppression of PTEN/AKT/FOXO3a pathway activation in the mouse ovary.

    PubMed

    Jang, Hoon; Lee, Ok-Hee; Lee, Youngeun; Yoon, Hyemin; Chang, Eun Mi; Park, Miseon; Lee, Jeong-Woong; Hong, Kwonho; Kim, Jung Oh; Kim, Nam Keun; Ko, Jung Jae; Lee, Dong Ryul; Yoon, Tae Ki; Lee, Woo Sik; Choi, Youngsok

    2016-04-01

    Premature ovarian failure (POF) is a major side effect of chemotherapy in young cancer patients. To develop pharmaceutical agents for preserving fertility, it is necessary to understand the mechanisms responsible for chemotherapy-induced follicle loss. Here, we show that treatment with cisplatin, a widely used anticancer drug, depleted the dormant follicle pool in mouse ovaries by excessive activation of the primordial follicles, without inducing follicular apoptosis. Moreover, we show that co-treatment with the antioxidant melatonin prevented cisplatin-induced disruption of the follicle reserve. We quantified the various stages of growing follicles, including primordial, primary, secondary, and antral, to demonstrate that cisplatin treatment alone significantly decreased, whereas melatonin co-treatment preserved, the number of primordial follicles in the ovary. Importantly, analysis of the PTEN/AKT/FOXO3a pathway demonstrated that melatonin significantly decreased the cisplatin-mediated inhibitory phosphorylation of PTEN, a key negative regulator of dormant follicle activation. Moreover, melatonin prevented the cisplatin-induced activating phosphorylation of AKT, GSK3β, and FOXO3a, all of which trigger follicle activation. Additionally, we show that melatonin inhibited the cisplatin-induced inhibitory phosphorylation and nuclear export of FOXO3a, which is required in the nucleus to maintain dormancy of the primordial follicles. These findings demonstrate that melatonin attenuates cisplatin-induced follicle loss by preventing the phosphorylation of PTEN/AKT/FOXO3a pathway members; thus, melatonin is a potential therapeutic agent for ovarian protection and fertility preservation during chemotherapy in female cancer patients. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  4. Differences in abundance of GnRH-I and GnRH-II among bovine antral follicles

    USDA-ARS?s Scientific Manuscript database

    Estradiol plays a critical role in fertility among cattle, specifically it has been reported that preovulatory estradiol regulates uterine pH, sperm transport, and uterine secretions that impact embryo development and survival. In addition, GnRH and GnRH receptors have been reported in granulosa cel...

  5. Genomic study and MeSH enrichment analysis of early pregnancy rate and antral follicle numbers in Nelore heifers

    USDA-ARS?s Scientific Manuscript database

    Zebu animals (Bos indicus) are known to take longer to reach puberty when compared to taurine animals (Bos taurus), limiting the supply of animals for harvest or breeding and impacting profitability. Genomic information can be a helpful tool to better understand complex traits, and improve genetic g...

  6. Impact of environmental tobacco smoke exposure in women on oxidative stress in the antral follicle and assisted reproduction outcomes

    PubMed Central

    Kazemi, Ashraf; Ramezanzadeh, Fatemeh; Esfahani, Mohammad Hosein Nasr; Saboor-Yaraghi, Ali Akbar; Nejat, Saharnaz; Rahimi-Foroshani, Abbas

    2013-01-01

    Background: Cigarette smoke contains many oxidants and may alter the human reproduction by inducing oxidative stress (OS) in both active and passive smokers. This study was designed to evaluate the effect of environmental tobacco smoke (ETS) exposure on oxidative stress in the follicular fluid and the assisted reproduction outcomes. Materials and Methods: An observational prospective study was carried out on 236 infertile women, who underwent assisted reproduction cycles. The ETS exposure was assessed using self-reported ETS exposure and the cotinine level in follicular fluid. To evaluate the OS in follicular fluid (FF) malon-di-aldehyde (MDA) and total antioxidant capacity (TAC) were measured. The number of retrieved oocytes, rate of metaphase II stage oocytes, fertilization rate, good cleavage rate, and no-fragmented embryo rate were considered as the assisted reproduction outcomes. The results were adjusted for age, body mass index, duration, and etiology of infertility; P-values less than 0.05 were considered significant. Results: The MDA and TAC levels in FF were not related to the self-report number of the weekly ETS exposure and cotinine levels in FF. Also, the number of retrieved oocytes, MII stage oocytes, fertilization rate, good cleavage rate, and no-fragmented embryo rate were not related to the cotinine level and weekly ETS exposure. However, in women whose cotinine levels in FF were lower and equal/above 3.5 ng/ml, the number of retrieved oocytes was higher (12.63 ± .71 vs. 9.28 ± 1.11, P = 0.01). The relationship between the MDA level and cleavage rate (Beta = −18.5, confidence interval-34.9 and-2.1, P < 0.05) was negatively significant and the relationship between the MII stage rate with TAC (Beta = 0.02, confidence interval 0.01 and 0.04, P < 0.05) was positively significant. Conclusion: The ETS exposure may alter the assisted reproduction success by influencing the number of available oocytes. Although, the OS in a follicular environment affect the ability of oocytes to reach the specific cleavage stages at appropriate time intervals, it does not mediate poor-assisted reproduction outcomes due to ETS exposure. PMID:24379845

  7. WBC count

    MedlinePlus

    Leukocyte count; White blood cell count ... blood is 4,500 to 11,000 white blood cells per microliter (mcL) or 4.5 to 11. ... LOW WHITE BLOOD CELL (WBC) COUNT A low number of WBCs is called leukopenia. A WBC count below 4500 is below normal ...

  8. Sympathetic nerve activity in normal and cystic follicles from isolated bovine ovary: local effect of beta-adrenergic stimulation on steroid secretion.

    PubMed

    Paredes, Alfonso H; Salvetti, Natalia R; Diaz, Ariel E; Dallard, Bibiana E; Ortega, Hugo H; Lara, Hernan E

    2011-05-16

    Cystic ovarian disease (COD) is an important cause of abnormal estrous behavior and infertility in dairy cows. COD is mainly observed in high-yielding dairy cows during the first months post-partum, a period of high stress. We have previously reported that, in lower mammals, stress induces a cystic condition similar to the polycystic ovary syndrome in humans and that stress is a definitive component in the human pathology. To know if COD in cows is also associated with high sympathetic activity, we studied isolated small antral (5 mm), preovulatory (10 mm) and cystic follicles (25 mm). Cystic follicles which present an area 600 fold greater compared with preovulatory follicles has only 10 times less concentration of NE as compared with small antral and preovulatory follicles but they had 10 times more NE in follicular fluid, suggesting a high efflux of neurotransmitter from the cyst wall. This suggestion was reinforced by the high basal release of recently taken-up 3H-NE found in cystic follicles. While lower levels of beta-adrenergic receptor were found in cystic follicles, there was a heightened response to the beta-adrenergic agonist isoproterenol and to hCG, as measured by testosterone secretion. There was however an unexpected capacity of the ovary in vitro to produce cortisol and to secrete it in response to hCG but not to isoproterenol. These data suggest that, during COD, the bovine ovary is under high sympathetic nerve activity that in addition to an increased response to hCG in cortisol secretion could participate in COD development.

  9. Genetic variations altering FSH action affect circulating hormone levels as well as follicle growth in healthy peripubertal girls.

    PubMed

    Busch, Alexander S; Hagen, Casper P; Almstrup, Kristian; Main, Katharina M; Juul, Anders

    2016-04-01

    Do variants of the genes encoding follicle stimulating hormone (FSH) beta subunit (B) and FSH receptor (R) impact circulating reproductive hormone levels and ovarian follicle maturation in healthy peripubertal girls? FSHB and FSHR genetic variants exert, alone or their combination, distinct effects on reproductive hormone levels as well as ovarian follicle maturation in healthy peripubertal girls. FSHB and FSHR genetic variants impact reproductive hormone levels as well as associated pathologies in women. While FSHR c. 2039A>G is known to alter gonadotrophin levels in women, FSHR c.-29G>A has not yet been shown to exert effect and there are conflicting results concerning FSHB c.-211G>T. This population-based study included 633 girls recruited as part of two cohorts, the COPENHAGEN Puberty Study (2006-2014, a cross-sectional and ongoing longitudinal study) and the Copenhagen Mother-Child Cohort (1997-2002, including transabdominal ultrasound (TAUS) of the ovaries in a subset of 91 peripubertal girls). Clinical examinations, including pubertal breast stage (Tanner's classification B1-B5) were performed. Circulating levels of FSH, luteinizing hormone (LH), estradiol, anti-Mullerian hormone (AMH) and inhibin-B were assessed by immunoassays. In a subset of the girls (n = 91), ovarian volume and the number/size of antral follicles were assessed by TAUS. Genotypes were determined by competitive PCR. FSHR c.2039A>G minor alleles were positively associated with serum FSH (β = 0.08, P = 0.004), LH (β = 0.06, P = 0.012) and estradiol (β = 0.06, P = 0.017) (adjusted for Tanner stages). In a combined model, FSHR c.-29G>A and FSHR c.2039A>G alleles were positively associated with FSH levels in early-pubertal girls (B2 + B3, n = 327, r = 0.1, P = 0.02) and in young adolescents (B4 + B5, n = 149, r = 0.2, P = 0.01). Serum AMH and inhibin B levels were not significantly influenced by the single nucleotide polymorphisms (SNPs). Single SNPs were not associated with follicles

  10. Endoplasmic reticulum-derived multilamellar bodies in oocytes of mouse follicle cultures under oxidized low-density lipoprotein treatment.

    PubMed

    Spanel-Borowski, Katharina; Nowicki, Marcin; Borlak, Juergen; Trapphoff, Tom; Eichenlaub-Ritter, Ursula

    2013-01-01

    Multilamellar bodies associated with an organized endoplasmic reticulum (ER) arise in various somatic cell types, and a subtype called multivesicular bodies is described in oocytes. Both entities, so far undetermined in significance, may occur in oocytes of follicles under oxidative stress. In preovulatory follicles, oxidative stress appears to be caused by oxidized low-density lipoprotein (ox-LDL). Cultures of preantral mouse follicles were treated with 100 µg/ml ox-LDL or normal LDL (n-LDL) for 12-48 h or for 12 days during antral follicle growth followed by in vitro ovulation and harvest of cumulus oophorus complexes (COCs) with metaphase II (MII) oocytes on day 13. Preantral follicles, COCs, or MII oocytes were immunostained with anti-tubulin antibody or stained with actin-binding phalloidin for confocal microscopy. Ultrathin sections were prepared for electron microscopy. Preantral follicles exposed to n-LDL or ox-LDL developed normally, and MII oocytes in COCs possessed normal spindles with well-aligned chromosomes. In contrast, treated cumulus cells underwent apoptosis. Only the ox-LDL-treated preantral follicle oocytes showed ER-derived multilamellar bodies (EMBs) of type I, consisting of rough ER membranes for the envelope. The MII oocytes of COCs showed type II EMBs consisting of smooth/vesicular ER and were more prominent after ox-LDL than after n-LDL exposure. Degenerating mitochondria were prominent in oocytes of the ox-LDL group and judged as a sign of oxidative stress. Oxidative stress presumably induces damage of proteins and organelles in the oocytes. The EMBs might sequester the damaged structures for oocyte survival. Thus, EMBs could represent a novel form of autophagy. Copyright © 2012 S. Karger AG, Basel.

  11. [Forskolin inhibits spontaneous contraction of gastric antral smooth muscle in rats].

    PubMed

    Jiang, Jing-Zhi; Sun, Qian; Xu, Dong-Yuan; Zhang, Mo-Han; Piao, Li-Hua; Cai, Ying-Lan; Jin, Zheng

    2013-04-25

    The aim of the present study was to investigate the effects of cyclic adenosine monophosphate (cAMP) on rat gastric antral circular smooth muscle function. Forskolin, a direct activator of adenylyl cyclase (AC), was used to observe the influences of cAMP. Multi-channel physiological recorder was used to record spontaneous contraction activity of gastric antral circular muscle from Wistar rats. And ELISA method was used to detect the change of cAMP production in perfusate. The results showed that forskolin concentration-dependently suppressed the amplitude and frequency of the spontaneous contraction of the gastric antral muscle, and lowered the baseline of contraction movement significantly. Forskolin concentration-dependently increased the production of cAMP in the perfusate, which showed a significant negative correlation with the contraction amplitude of gastric antral ring muscle. The inhibitory effect of forskolin on spontaneous contraction activity of rat gastric antral circular muscle could be blocked by cAMP-dependent protein kinase (PKA) inhibitor H-89. These results suggest forskolin increases cAMP production and then activates PKA pathway, resulting in the inhibition of the spontaneous contraction activity of rat gastric antral circular smooth muscle.

  12. Expression of connexin 43 mRNA and protein in developing follicles of prepubertal porcine ovaries

    USGS Publications Warehouse

    Melton, C.M.; Zaunbrecher, G.M.; Yoshizaki, G.; Patio, R.; Whisnant, S.; Rendon, A.; Lee, V.H.

    2001-01-01

    A major form of cell-cell communication is mediated by gap junctions, aggregations of intercellular channels composed of connexins (Cxs), which are responsible for exchange of low molecular weight (< 1200 Da) cytosolic materials. These channels are a growing family of related proteins. This study was designed to determine the ontogeny of connexin 43 (Cx43) during early stages of follicular development in prepubertal porcine ovaries. A partial-length (412 base) cDNA clone was obtained from mature porcine ovaries and determined to have 98% identity with published porcine Cx43. Northern blot analysis demonstrated a 4.3-kb mRNA in total RNA isolated from prepubertal and adult porcine ovaries. In-situ hybridization revealed that Cx43 mRNA was detectable in granulosa cells of primary follicles but undetectable in dormant primordial follicles. The intensity of the signal increased with follicular growth and was greatest in the large antral follicles. Immunohistochemical evaluation indicated that Cx43 protein expression correlated with the presence of Cx43 mRNA. These results indicate that substantial amounts of Cx43 are first expressed in granulosa cells following activation of follicular development and that this expression increases throughout follicular growth and maturation. These findings suggest an association between the enhancement of intercellular gap-junctional communication and onset of follicular growth. ?? 2001 Elsevier Science Inc. All rights reserved.

  13. Loss of Gremlin Delays Primordial Follicle Assembly but Does Not Affect Female Fertility in Mice1

    PubMed Central

    Myers, Michelle; Tripurani, Swamy K.; Middlebrook, Brooke; Economides, Aris N.; Canalis, Ernesto; Pangas, Stephanie A.

    2011-01-01

    ABSTRACT The transforming growth factor beta (TGFB) protein family is renowned for its diverse roles in developmental biology including reproduction. Gremlin is a member of the differential screening-selected gene aberrative in neuroblastoma (DAN)/cerberus family of bone morphogenetic protein (BMP) antagonists. Recent studies on gremlin focus on its involvement in embryonic skeletal, lung, and kidney development. To define the role of gremlin (Grem1) in female reproduction, we analyzed postnatal folliculogenesis using global and conditional knockout (cKO) mice for gremlin. Grem1−/− mice die within 48 h after birth, and ovaries collected from neonatal Grem1−/− mice demonstrated reduced oocyte numbers and delayed primordial follicle development. Transplanting Grem1−/− neonatal ovaries showed that folliculogenesis proceeded to large antral follicle stage, but Grem1−/− ovaries contained corpora lutea-like structures not found in control-transplanted ovaries. However, Grem1 cKO mice had comparable fertility to control mice. These data suggest that gremlin plays a previously uncharacterized role in the regulation of oocyte numbers and the timing of primordial follicle development, but either it is not required for later folliculogenesis or its loss is possibly compensated by other BMP antagonists. PMID:21832168

  14. The Molecular Basis of Impaired Follicle-Stimulating Hormone Action

    PubMed Central

    Siegel, Eric T.; Kim, Hyung-Goo; Nishimoto, Hiromi Koso

    2013-01-01

    The pituitary gonadotropin follicle-stimulating hormone (FSH) interacts with its membrane-bound receptor to produce biologic effects. Traditional functions of FSH include follicular development and estradiol production in females, and the regulation of Sertoli cell action and spermatogenesis in males. Knockout mice for both the ligand (Fshb) and the receptor (Fshr) serve as models for FSH deficiency, while Fshb and Fshr transgenic mice manifest FSH excess. In addition, inactivating mutations of both human orthologs (FSHB and FSHR) have been characterized in a small number of patients, with phenotypic effects of the ligand disruption being more profound than those of its receptor. Activating human FSHR mutants have also been described in both sexes, leading to a phenotype of normal testis function (male) or spontaneous ovarian hyperstimulation syndrome (females). As determined from human and mouse models, FSH is essential for normal puberty and fertility in females, particularly for ovarian follicular development beyond the antral stage. In males, FSH is necessary for normal spermatogenesis, but there are differences in human and mouse models. The FSHB mutations in humans result in azoospermia; while FSHR mutations in humans and knockouts of both the ligand and the receptor in mice affect testicular function but do not result in absolute infertility. Available evidence also indicates that FSH may also be necessary for normal androgen synthesis in males and females. PMID:23184658

  15. [Gastric vascular lesions in cirrhosis: gastropathy and antral vascular ectasia].

    PubMed

    Casas, Meritxell; Calvet, Xavier; Vergara, Mercedes; Bella, Maria Rosa; Junquera, Félix; Martinez-Bauer, Eva; Campo, Rafael

    2015-02-01

    Portal hypertensive gastropathy (GHP) is a complication of portal hypertension usually associated with liver cirrhosis. The pathogenesis is unclear but the presence of portal hypertension is an essential factor for its development. GHP may be asymptomatic or present as gastrointestinal bleeding or iron deficiency anemia. Endoscopic lesions vary from a mosaic pattern to diffuse red spots; the most common location is the fundus. Treatment is indicated when there is acute or chronic bleeding, as secondary prophylaxis. There is insufficient evidence to recommend primary prophylaxis in patients who have never bled. Drugs that decrease portal pressure, such as non-cardioselective beta-blockers, and/or endoscopic ablative treatments, such as argon-beam coagulation, may be used. The role of transarterial intrahepatic portosystemic shunt) or bypass surgery has been insufficiently analyzed. Antral vascular ectasia (EVA) is a rare entity in liver cirrhosis, whose pathophysiology is still unknown. Clinical presentation is similar to that of GHP and endoscopy usually shows red spots in the antrum. Biopsy is often required to differentiate EVA from GHP. There is no effective medical therapy, so endoscopic ablative therapy and, in severe cases, antrectomy are recommended.

  16. Idiopathic bilateral antral exostoses: A rare case in maxillary sinus

    PubMed Central

    Borie, Eduardo; Watanabe, Plauto C.A.; Orsi, Iara A.; Fuentes, Ramón

    2014-01-01

    INTRODUCTION Exostoses in paranasal sinuses have been reported in the otolaryngology literature, but they have not been described in the dental literature to our knowledge. The aim of this article is to describe an idiopathic and rare case of bilateral exostosis obtained by cone-beam computed tomography. PRESENTATION OF CASE The case shows a healthy and asymptomatic patient with a different size and form of exostoses in both maxillary sinuses. DISCUSSION It is difficult to clinically diagnose the antral exostosis due the asymptomatic nature of this condition, unless the approach would be through endoscope. Sometimes this condition is related with nasal irrigants, however in this case the patient asserted not having used nasal irrigation ever; thus, it is impossible to relate this kind of treatment as a principal cause. CONCLUSION The published data of exostoses in maxillary sinus seem to be limited in the dental literature, and this condition is important to consider in an implant treatment planning. Also, it is important to perform a follow-up of the cases in trying to find the possible causes of exostosis. PMID:25128728

  17. Characterization and quantification of wound-induced hair follicle neogenesis using in vivo confocal scanning laser microscopy

    PubMed Central

    Fan, Chengxiang; Luedtke, Michael A.; Prouty, Stephen M.; Burrows, Michelle; Kollias, Nikiforos

    2011-01-01

    Background In vivo confocal scanning laser microscopy (CSLM) is a recently-developed non-invasive technique for visualizing microscopic structures with the skin. CSLM has been used to characterize proliferative and inflammatory skin diseases, neoplastic skin lesions and pigmented lesions. Objective Here, we assessed the ability of CSLM to evaluate the formation of neogenic hair follicles after a full thickness wound in mice. Methods Full-thickness wounds were made on the dorsal skin of 3-week old mice. After scab detachment (SD), the number, width, length, space and volume of neogenic hair follicles were analyzed using CSLM. The results were compared with those from conventional methods, including staining for alkaline phosphatase (AP) and keratin 17 (K17) as well as histology. Results Quantification of neogenic hair follicles using CSLM compared favorably with results from direct measurements on isolated epidermal tissue after immunostaining for K17, a marker for the epithelial portion of new hair follicles. CSLM detected 89% of K17-stained follicles. CSLM more accurately quantitated the number of new follicles compared to AP staining, which detects the dermal portion of the new follicle. The width and length measurement from CSLM and histology were very close and correlated with each other. The minimum length of a neogenic hair follicle that could be detected by CSLM was 21 μm. The space between neogenic hair follicles was decreased in histological sections compared to CSLM. Conclusions CSLM is an accurate and valuable method for counting and measuring neogenic hair follicles non-invasively. CSLM produces images similar to histology in mice. Measurements of microstructures using CSLM more accurately reflect actual sizes since this technique avoids fixation artifact. In vivo visualization of developing follicles with CSLM permits detection of serial changes in hair follicle formation, thus conserving numbers of mice required for studies and improving detection of

  18. Culture of bovine ovarian follicle wall sections maintained the highly estrogenic profile under basal and chemically defined conditions.

    PubMed

    Vasconcelos, R B; Salles, L P; Oliveira e Silva, I; Gulart, L V M; Souza, D K; Torres, F A G; Bocca, A L; Rosa e Silva, A A M

    2013-08-01

    Follicle cultures reproduce in vitro the functional features observed in vivo. In a search for an ideal model, we cultured bovine antral follicle wall sections (FWS) in a serum-free defined medium (DM) known to induce 17β-estradiol (E2) production, and in a nondefined medium (NDM) containing serum. Follicles were sectioned and cultured in NDM or DM for 24 or 48 h. Morphological features were determined by light microscopy. Gene expression of steroidogenic enzymes and follicle-stimulating hormone (FSH) receptor were determined by RT-PCR; progesterone (P4) and E2 concentrations in the media were measured by radioimmunoassay. DM, but not NDM, maintained an FWS morphology in vitro that was similar to fresh tissue. DM also induced an increase in the expression of all steroidogenic enzymes, except FSH receptor, but NDM did not. In both DM and NDM, there was a gradual increase in P4 throughout the culture period; however, P4 concentration was significantly higher in NDM. In both media, E2 concentration was increased at 24 h, followed by a decrease at 48 h. The E2:P4 ratio was higher in DM than in NDM. These results suggest that DM maintains morphological structure, upregulates the expression of steroidogenic enzyme genes, and maintains steroid production with a high E2:P4 ratio in FWS cultures.

  19. Culture of bovine ovarian follicle wall sections maintained the highly estrogenic profile under basal and chemically defined conditions

    PubMed Central

    Vasconcelos, R.B.; Salles, L.P.; Silva, I. Oliveira e; Gulart, L.V.M.; Souza, D.K.; Torres, F.A.G.; Bocca, A.L.; Silva, A.A.M. Rosa e

    2013-01-01

    Follicle cultures reproduce in vitro the functional features observed in vivo. In a search for an ideal model, we cultured bovine antral follicle wall sections (FWS) in a serum-free defined medium (DM) known to induce 17β-estradiol (E2) production, and in a nondefined medium (NDM) containing serum. Follicles were sectioned and cultured in NDM or DM for 24 or 48 h. Morphological features were determined by light microscopy. Gene expression of steroidogenic enzymes and follicle-stimulating hormone (FSH) receptor were determined by RT-PCR; progesterone (P4) and E2 concentrations in the media were measured by radioimmunoassay. DM, but not NDM, maintained an FWS morphology in vitro that was similar to fresh tissue. DM also induced an increase in the expression of all steroidogenic enzymes, except FSH receptor, but NDM did not. In both DM and NDM, there was a gradual increase in P4 throughout the culture period; however, P4 concentration was significantly higher in NDM. In both media, E2 concentration was increased at 24 h, followed by a decrease at 48 h. The E2:P4 ratio was higher in DM than in NDM. These results suggest that DM maintains morphological structure, upregulates the expression of steroidogenic enzyme genes, and maintains steroid production with a high E2:P4 ratio in FWS cultures. PMID:23969977

  20. Hair follicle changes following intense pulsed light axillary hair reduction: histometrical, histological and immunohistochemical evaluation.

    PubMed

    El-Domyati, Moetaz; Hosam, Wael; Moftah, Noha H; Abdel Raouf, Hamza; Saad, Selwet M

    2017-04-01

    Intense pulsed light (IPL) has been used for years in hair reduction; however, no previous studies discussed quantitative histological and immunohistochemical changes of hair follicles after IPL. Accordingly, this study aims to objectively quantify histological and immunohistochemical changes of hair follicles after IPL hair reduction. Right axillae of 21 volunteers were subjected to 6 IPL sessions using Quanta system IPL and evaluated at 1 week and 1 month after last session (i.e., 3 and 4 months from the start of treatment, respectively) in comparison to baseline and left control axillae. Using hair count, histological and immunohistochemical assessment of vertical and serial transverse sections coupled with computerized morphometric analysis, determination of hair reduction percentage, measurement of hair shaft (HS) diameter, calculation of percentage of hair follicle types and quantitative evaluation of PCNA, Ki67 and P53 markers were performed. After IPL, there was significant decrease of hair count, HS diameter, percentage of terminal anagen follicles, terminal/vellus (T/V) ratio, anagen/telogen (A/T) ratio and expression of PCNA and Ki67; however, significant increase of percentage of terminal telogen and total vellus follicles with vellus-like type and P53 expression was identified. So, reduction of hair number and thickness occurred after IPL by induction of telogenesis and miniaturization through decreased hair follicle proliferation and increase in DNA damage that could favor apoptosis.

  1. Ultrasonographic gastric antral area and gastric contents volume in children.

    PubMed

    Schmitz, Achim; Thomas, Schraner; Melanie, Fruehauf; Rabia, Liamlahi; Klaghofer, Richard; Weiss, Markus; Kellenberger, Christian

    2012-02-01

    Cross-sectional gastric antral area (GAA) measurements by ultrasonography (US) have been proposed for preoperative assessment of gastric volume in adults but not been validated in children. This study investigates whether in children gastric volumes can be predicted by US performed in different patient positions. Gastric fluid and air volumes were examined by magnetic resonance imaging before or up to 120 min after ingestion of 7 ml·kg(-1) diluted raspberry syrup in healthy volunteers who had fasted overnight. GAA was measured with US three times each in supine (SUP), elevated 45° degree supine (E45) and right decubital (RDC) position using imaging planes defined by vascular landmarks. Correlation coefficients (Pearson) between GAA and gastric volumes were calculated and Bland-Altman analysis performed. Sixteen children aged from 6.4 to 12.8 (9.2) years were included in 23 examinations: 6 after overnight fasting, 3 directly after, and 14 with a delay of 74 ± 35 min after fluid intake. GAA was 221 ± 116, 218 ± 112, and 347 ± 188 mm(2) for SUP, E45, and RDC position, respectively. The best correlation between body weight corrected total gastric/gastric fluid volume (TGV(w)/GFV(w)) with GAA was found for RDC position (R = 0.79; P < 0.01/R = 0.78; P < 0.01). Bias and precision of calculated and measured GFV(w) was 0 ± 2.8 ml·kg(-1). Correlations between GAA and TGV(w) or GFV(w) in children are best in the RDC position, but not sufficient to predict GFV(w) with a given GAA. Interpretation of isolated GAA values may be misleading. © 2011 Blackwell Publishing Ltd.

  2. Expression and localization of members of the thrombospondin family during final follicle maturation and corpus luteum formation and function in the bovine ovary

    PubMed Central

    BERISHA, Bajram; SCHAMS, Dieter; RODLER, Daniela; SINOWATZ, Fred; PFAFFL, Michael W.

    2016-01-01

    The aim of this study was to characterize the expression patterns and localization of the thrombospondin family members (THBS1, THBS2) and their receptors (CD36 and CD47) in bovine ovaries. First, the antral follicles were classified into 5 groups based on the follicle size and estradiol-17beta (E2) concentration in the follicular fluid (< 0.5, 0.5–5, 5–40, 40–180 and >180 E2 ng/ml). Second, the corpus luteum (CL) was assigned to the following stages: days 1–2, 3–4, 5–7, 8–12, 13–16 and >18 of the estrous cycle and of pregnancy (month 1–2, 3–4, 6–7 and > 8). Third, the corpora lutea were collected by transvaginal ovariectomy before and 0.5, 2, 4, 12, 24, 48 and 64 h after inducing luteolysis by injecting a prostaglandin F2alpha analog. The mRNA expression of examined factors was measured by RT-qPCR, steroid hormone concentration by EIA, and localization by immunohistochemistry. The mRNA expression of THBS1, THBS2, CD36, and CD47 in the granulosa cells and theca interna was high in the small follicles and reduced in the preovulatory follicles. The mRNA expression of THBS1, THBS2, and CD47 in the CL during the estrous cycle was high, but decreased significantly during pregnancy. After induced luteolysis, thrombospondins increased significantly to reach the maximum level at 12 h for THBS1, 24 h for THBS2, and 48 h for CD36. The temporal expression and localization pattern of the thrombospondins and their specific receptors in the antral follicles and corpora lutea during the different physiological phases of the estrous cycle and induced luteolysis appear to be compatible with their inhibitory role in the control of ovarian angiogenesis. PMID:27396384

  3. Single-chain bifunctional vascular endothelial growth factor (VEGF)-follicle-stimulating hormone (FSH)-C-terminal peptide (CTP) is superior to the combination therapy of recombinant VEGF plus FSH-CTP in stimulating angiogenesis during ovarian folliculogenesis.

    PubMed

    Trousdale, Rhonda K; Pollak, Susan V; Klein, Jeffrey; Lobel, Leslie; Funahashi, Yasuhiro; Feirt, Nikki; Lustbader, Joyce W

    2007-03-01

    Infertility technologies often employ exogenous gonadotropin therapy to increase antral follicle production. In an effort to enhance ovarian response, several long-acting FSH therapies have been developed including an FSH-C-terminal peptide (CTP), where the FSH subunits are linked by the CTP moiety from human chorionic gonadotropin, which is responsible for the increased half-life of human chorionic gonadotropin. We found that administration of FSH-CTP for ovarian hyperstimulation in rats blunted ovarian follicle vascular development. In women, reduced ovarian vasculature has been associated with lower pregnancy rates. We were interested in determining whether vascular endothelial growth factor (VEGF) therapy could enhance ovarian angiogenesis in FSH-CTP-treated rats. Coadministration of systemic FSH-CTP plus recombinant VEGF was compared with treatment with a novel, single-chain bifunctional VEGF-FSH-CTP (VFC) analog. For VFC, the FSH portion targets the protein to the ovary and stimulates follicle growth, whereas VEGF enhances local vascular development. Both in vitro and in vivo studies confirm the dual FSH and VEGF action of the VFC protein. Evaluation of ovarian follicle development demonstrates that administration of combination therapy using VEGF and FSH-CTP failed to increase follicle vasculature above levels seen with FSH-CTP monotherapy. However, treatment with VFC significantly increased follicle vascular development while concurrently increasing the number of large antral follicles produced. In conclusion, we report the production and characterization of a long-acting, bifunctional VEGF-FSH-CTP protein that is superior to combination therapy for enhancing VEGF activity in the ovary and stimulating follicular angiogenesis in rats.

  4. Notch1 and Notch2 receptors regulate mouse and human gastric antral epithelial cell homoeostasis

    PubMed Central

    Gifford, Gail B; Demitrack, Elise S; Keeley, Theresa M; Tam, Andrew; La Cunza, Nilsa; Dedhia, Priya H; Spence, Jason R; Simeone, Diane M; Saotome, Ichiko; Louvi, Angeliki; Siebel, Christian W; Samuelson, Linda C

    2016-01-01

    Objective We tested the ability of Notch pathway receptors Notch1 and Notch2 to regulate stem and epithelial cell homoeostasis in mouse and human gastric antral tissue. Design Mice were treated with the pan-Notch inhibitor dibenzazepine (DBZ) or inhibitory antibodies targeting Notch1 and/or Notch2. Epithelial proliferation, apoptosis and cellular differentiation were measured by histological and molecular approaches. Organoids were established from mouse and human antral glands; growth and differentiation were measured after treatment with Notch inhibitors. Results Notch1 and Notch2 are the predominant Notch receptors expressed in mouse and human antral tissue and organoid cultures. Combined inhibition of Notch1 and Notch2 in adult mice led to decreased epithelial cell proliferation, including reduced proliferation of LGR5 stem cells, and increased apoptosis, similar to the response to global Notch inhibition with DBZ. Less pronounced effects were observed after inhibition of individual receptors. Notch pathway inhibition with DBZ or combined inhibition of Notch1 and Notch2 led to increased differentiation of all gastric antral lineages, with remodelling of cells to express secretory products normally associated with other regions of the GI tract, including intestine. Analysis of mouse and human organoids showed that Notch signalling through Notch1 and Notch2 is intrinsic to the epithelium and required for organoid growth. Conclusions Notch signalling is required to maintain gastric antral stem cells. Notch1 and Notch2 are the primary Notch receptors regulating epithelial cell homoeostasis in mouse and human stomach. PMID:26933171

  5. Reciprocal regulation of antral gastrin and somatostatin gene expression by omeprazole-induced achlorhydria.

    PubMed Central

    Brand, S J; Stone, D

    1988-01-01

    Gastric acid exerts a feedback inhibition on the secretion of gastrin from antral G cells. This study examines whether gastrin gene expression is also regulated by changes in gastric pH. Achlorhydria was induced in rats by the gastric H+/K+ ATPase inhibitor, omeprazole (100 mumol/kg). This resulted in fourfold increases in both serum gastrin (within 2 h) and gastrin mRNA levels (after 24 h). Antral somatostatin D cells probably act as chemoreceptors for gastric acid to mediate a paracrine inhibition on gastrin secretion from adjacent G cells. Omeprazole-induced achlorhydria reduced D-cell activity as shown by a threefold decrease in antral somatostatin mRNA levels that began after 24 h. Exogenous administration of the somatostatin analogue SMS 201-995 (10 micrograms/kg) prevented both the hypergastrinemia and the increase in gastrin mRNA levels caused by omeprazole-induced achlorhydria. Exogenous somatostatin, however, did not influence the decrease in antral somatostatin mRNA levels seen with achlorhydria. These data, therefore, support the hypothesis that antral D cells act as chemoreceptors for changes in gastric pH, and modulates somatostatin secretion and synthesis to mediate a paracrine inhibition on gastrin gene expression in adjacent G cells. Images PMID:2901431

  6. Spatial Characterization of Bioenergetics and Metabolism of Primordial to Preovulatory Follicles in Whole Ex Vivo Murine Ovary.

    PubMed

    Cinco, Rachel; Digman, Michelle A; Gratton, Enrico; Luderer, Ulrike

    2016-12-01

    Previous work characterizing ovarian bioenergetics has defined follicular metabolism by measuring metabolic by-products in culture media. However, culture conditions perturb the native state of the follicle, and these methods do not distinguish between metabolism occurring within oocytes or granulosa cells. We applied the phasor approach to fluorescence lifetime imaging microscopy (phasor FLIM) at 740-nm two-photon excitation to examine the spatial distribution of free and protein-bound nicotinamide adenine dinucleotide hydride (NADH) during primordial through preovulatory stages of follicular development in fresh ex vivo murine neonatal and gonadotropin stimulated prepubertal ovaries. We obtained subcellular resolution phasor FLIM images of primordial through primary follicles and quantified the free/bound NADH ratio (relative NADH/NAD+) separately for oocyte nucleus and oocyte cytoplasm. We found that dynamic changes in oocyte nucleus free/bound NADH paralleled the developmental maturation of primordial to primary follicles. Immunohistochemistry of NAD+-dependent deacetylase SIRTUIN 1 (SIRT1) in neonatal ovary revealed that increasing SIRT1 expression in oocyte nuclei was inversely related to decreasing free/bound NADH during the primordial to primary follicle transition. We characterized oocyte metabolism at these early stages to be NADH producing (glycolysis/Krebs). We extended the results of prior studies to show that cumulus and mural granulosa cell metabolism in secondary through preovulatory follicles is mainly NADH producing (glycolysis/Krebs cycle), while oocyte metabolism is mainly NADH consuming (oxidative phosphorylation). Taken together, our data characterize dynamic changes in free/bound NADH and SIRT1 expression during early follicular development and confirm results from previous studies defining antral and preovulatory follicle metabolism in culture. © 2016 by the Society for the Study of Reproduction, Inc.

  7. Follicle Development of Xenotransplanted Sheep Ovarian Tissue into Male and Female Immunodeficient Rats

    PubMed Central

    Tahaei, Leila Sadat; Eimani, Hussein; Hajmusa, Ghazaleh; Fathi, Rouhollah; Rezazadeh Valojerdi, Mojtaba; Shahverdi, Abdolhossein; Eftekhari-Yazdi, Poopak

    2015-01-01

    Background This study aimed to assess follicle survival after xenotransplantation of sheep ovarian tissue into male and female immunodeficient rats. We evaluated the effects of gonadotropin treatment on follicular development in the transplanted tissue. Materials and Methods In this experimental study, sheep ovarian cortical strips were transplanted into the neck back muscles of 8 male and 8 female immunodeficient, castrated rats. Fourteen days after surgery, each rat was treated with human menopausal gonadotropin (hMG) for 9 weeks. One day after the last injection, ovarian tissues were removed and fixed for histology assessment. Histology analyses were performed before and after grafting. Estradiol (E2) levels were measured before and after gonadectomy, and at the end of the experiment. The control group consisted of 7 male and 7 female noncastrated/non-grafted rats and the sham group comprised 7 male and 7 female castrated/ non-grafted rats for comparison of serum E2 concentrations. Results The percentage of primordial follicles decreased after transplantation in male (25.97%) and female (24.14%) rats compared to the control group (ovarian tissue nongrafted; 37.51%). Preantral follicles increased in the male (19.5%) and female (19.49%) transplanted rats compared to the control group (11.4%). Differences in antral follicles between male (0.06 ± 0.0%) and female (0.06 ± 0.0%) rats were not noticeable compared to control (1.25 ± 0.0%) rats. We observed a significantly higher percent of mean E2 secretion in grafted males compared to grafted females (P˂0.05). Conclusion Despite significant differences in E2 secretion between xenografted male and female rats, we observed no statistical differences in terms of follicular development. PMID:26644859

  8. Follicle Development of Xenotransplanted Sheep Ovarian Tissue into Male and Female Immunodeficient Rats.

    PubMed

    Tahaei, Leila Sadat; Eimani, Hussein; Hajmusa, Ghazaleh; Fathi, Rouhollah; Rezazadeh Valojerdi, Mojtaba; Shahverdi, Abdolhossein; Eftekhari-Yazdi, Poopak

    2015-01-01

    This study aimed to assess follicle survival after xenotransplantation of sheep ovarian tissue into male and female immunodeficient rats. We evaluated the effects of gonadotropin treatment on follicular development in the transplanted tissue. In this experimental study, sheep ovarian cortical strips were transplanted into the neck back muscles of 8 male and 8 female immunodeficient, castrated rats. Fourteen days after surgery, each rat was treated with human menopausal gonadotropin (hMG) for 9 weeks. One day after the last injection, ovarian tissues were removed and fixed for histology assessment. Histology analyses were performed before and after grafting. Estradiol (E2) levels were measured before and after gonadectomy, and at the end of the experiment. The control group consisted of 7 male and 7 female noncastrated/non-grafted rats and the sham group comprised 7 male and 7 female castrated/ non-grafted rats for comparison of serum E2 concentrations. The percentage of primordial follicles decreased after transplantation in male (25.97%) and female (24.14%) rats compared to the control group (ovarian tissue nongrafted; 37.51%). Preantral follicles increased in the male (19.5%) and female (19.49%) transplanted rats compared to the control group (11.4%). Differences in antral follicles between male (0.06 ± 0.0%) and female (0.06 ± 0.0%) rats were not noticeable compared to control (1.25 ± 0.0%) rats. We observed a significantly higher percent of mean E2 secretion in grafted males compared to grafted females (P˂0.05). Despite significant differences in E2 secretion between xenografted male and female rats, we observed no statistical differences in terms of follicular development.

  9. Zygomatic bone graft for oral-antral communication closure and implant placement.

    PubMed

    Peñarrocha-Diago, Miguel; García, Berta; Gomez, Dolores; Balaguer, José

    2007-01-01

    The roots of molar and premolar maxillary teeth are often very close to the floor of the maxillary sinus. As a result, extraction of these teeth can leave an oral-antral communication or lead to a fistula that requires treatment. A woman with an oral-antral communication secondary to extraction of a maxillary molar is presented. The communication was closed by means of a bone graft harvested from the wall of the sinus (zygomatic bone). After 3 months, 2 dental implants were placed, one in the pterygoid area and the other with parasinusal angulation. Rehabilitation followed in the form of a screw-retained, fixed prosthesis 3 months after implant placement. There have been no complications after 1 year of follow-up. This surgical technique allowed closure of an oral-antral communication produced by molar extraction through placement of a zygomatic bone graft and subsequent placement of 2 dental implants.

  10. A permissive role for the vagus nerves in the genesis of antro-antral reflexes in the anaesthetized ferret.

    PubMed Central

    Grundy, D; Hutson, D; Scratcherd, T

    1986-01-01

    1. The role of the vagus nerves in the genesis of antro-antral reflexes was investigated in the urethane-anaesthetized, splanchnectomized ferret. 2. Antral distension stimulated antral contractions with a threshold volume of 3.5 +/- 0.9 ml (corresponding to an intra-antral pressure of 0.27 +/- 0.11 kPa) by a vagal-dependent mechanism as indicated by the attenuated response seen during vagal blockade by cooling. Atropine (1 mg/kg) abolished the antral response to distension. 3. In vagotomized animals, close arterial infusions of acetylcholine at a dose sufficient to return antral motility to basal levels led to the reappearance of the reflex. Low-frequency electrical stimulation of the preganglionic vagal neurones had a similar effect. These effects were also abolished by atropine (1 mg/kg). 4. Hexamethonium (10-25 mg/kg) suppressed the potentiating effect of acetylcholine, indicating a ganglionic site of action. The attenuated response to antral distension seen in vagotomized animals in the absence of exogenous acetylcholine or electrical vagal stimulation was not sensitive to hexamethonium but abolished by atropine (1 mg/kg). 5. The results are consistent with the vagus performing a permissive role in the genesis of antro-antral reflexes mediated through local enteric pathways. PMID:2887649

  11. Short-term hypertonic exposure enhances in vitro follicle growth and meiotic competence of enclosed oocytes while modestly affecting mRNA expression of aquaporin and steroidogenic genes in the domestic cat model.

    PubMed

    Songsasen, N; Thongkittidilok, C; Yamamizu, K; Wildt, D E; Comizzoli, P

    2017-03-01

    Using the domestic cat as a non-rodent, larger animal model, the objective was to determine the impact of a brief incubation in a hypertonic microenvironment on (1) ovarian follicle and oocyte growth in vitro, (2) developmental capacity of the resident oocyte, and (3) expression of aquaporin (AQP) genes in parallel with genes involved in regulation of folliculogenesis. In Study 1: Secondary or early antral follicles encapsulated in 0.5% alginate were allocated to one of three treatment groups: 1) culture in standard medium at 290 mOsm for 15 d (Control); 2) incubation in 350 mOsm medium for 1 h followed by culture in standard medium for 15 d (Hypertonic-1h); or 3) incubation in 350 mOsm medium for 24 h followed by incubation in standard medium for additional 14 d (Hypertonic-24h). After measuring follicle and oocyte diameters on Day 15, in vitro-grown oocytes were incubated for 24 h before assessing nuclear status. In Study 2: secondary or early antral follicles were subjected to one of the three treatments: 1) culture in standard medium at 290 mOsm for 48 h; 2) incubation in 350 mOsm medium for 1 h followed by culture in standard medium for additional 47 h; or 3) incubation in 350 mOsm medium for 24 h followed by culture in standard medium for additional 24 h. At the end of the culture period, all follicles were assessed for mRNA level of Cyp17a1, Cyp19a1, Star, Aqp1, 3, 5, 7 and 8 as well as Fshr using qPCR. Freshly collected follicles also were subjected to gene expression analysis and served as the 'Non-cultured control'. Hypertonic-24h follicles grew larger (P < 0.05) than the control, whereas those in Hypertonic-1h group exhibited intermediate growth, especially when the culture started at the early antral stage. Oocytes in the Hypertonic-24h group were larger and resumed meiosis at a higher rate than in the other treatments. In vitro culture affected (P < 0.05) mRNA expression of Cyp19a1, Star, Aqp1, and Aqp7 in both the secondary and early

  12. Effects of sulglycotide on endogenous prostaglandin synthesis in human antral mucosa.

    PubMed

    Calabrò, A; Orsini, B; Milani, S; Psilogenis, M; Surrenti, C

    1992-12-01

    The effect of sulglycotide, a polysulphated glycopeptide isolated from pig duodenum, on PGE2, 6-ketoPGF1 alpha and TxB2 production from isolated biopsy specimens of human antral mucosa was investigated in vitro, in comparison with carbenoxolone. In addition to a tendency toward increased PGE2 production just short of statistical significance, both sulglycotide and carbenoxolone significantly increased 6-ketoPGF1 alpha accumulation in the incubation medium. TxB2 levels were not significantly modified by the two drugs. Therefore, changes in the prostanoid production by human antral mucosa could, at least partially account for the cytoprotective effects of sulglycotide in man.

  13. Multiplicity Counting

    SciTech Connect

    Geist, William H.

    2015-12-01

    This set of slides begins by giving background and a review of neutron counting; three attributes of a verification item are discussed: 240Pueff mass; α, the ratio of (α,n) neutrons to spontaneous fission neutrons; and leakage multiplication. It then takes up neutron detector systems – theory & concepts (coincidence counting, moderation, die-away time); detector systems – some important details (deadtime, corrections); introduction to multiplicity counting; multiplicity electronics and example distributions; singles, doubles, and triples from measured multiplicity distributions; and the point model: multiplicity mathematics.

  14. Reticulocyte count

    MedlinePlus

    Anemia - reticulocyte ... A higher than normal reticulocytes count may indicate: Anemia due to red blood cells being destroyed earlier than normal ( hemolytic anemia ) Bleeding Blood disorder in a fetus or newborn ( ...

  15. Tower counts

    USGS Publications Warehouse

    Woody, Carol Ann; Johnson, D.H.; Shrier, Brianna M.; O'Neal, Jennifer S.; Knutzen, John A.; Augerot, Xanthippe; O'Neal, Thomas A.; Pearsons, Todd N.

    2007-01-01

    Counting towers provide an accurate, low-cost, low-maintenance, low-technology, and easily mobilized escapement estimation program compared to other methods (e.g., weirs, hydroacoustics, mark-recapture, and aerial surveys) (Thompson 1962; Siebel 1967; Cousens et al. 1982; Symons and Waldichuk 1984; Anderson 2000; Alaska Department of Fish and Game 2003). Counting tower data has been found to be consistent with that of digital video counts (Edwards 2005). Counting towers do not interfere with natural fish migration patterns, nor are fish handled or stressed; however, their use is generally limited to clear rivers that meet specific site selection criteria. The data provided by counting tower sampling allow fishery managers to determine reproductive population size, estimate total return (escapement + catch) and its uncertainty, evaluate population productivity and trends, set harvest rates, determine spawning escapement goals, and forecast future returns (Alaska Department of Fish and Game 1974-2000 and 1975-2004). The number of spawning fish is determined by subtracting subsistence, sport-caught fish, and prespawn mortality from the total estimated escapement. The methods outlined in this protocol for tower counts can be used to provide reasonable estimates ( plus or minus 6%-10%) of reproductive salmon population size and run timing in clear rivers. 

  16. Alphafetoprotein and atretic follicles in the ovary of the pregnant rat.

    PubMed

    Seralini, G E; Lafaurie, M; Krebs, B; Stora, C

    1986-01-01

    Previous experiments have been conducted concerning the role of alphafetoprotein in genital system blockade in several cases: during adult rat N2-fluorenylacetamide hepatocarcinogenesis, after alphafetoprotein injections into normal adult female rats, during fetal life, and during postnatal and prepuberal development. In these conditions, alphafetoprotein is present at high plasma levels, and the normal cyclic ovarian function is stopped or nonexistent. The degenerating oocytes observed in the ovaries are often AFP-positive by histo-immunolocalization. Pregnancy corresponds to a physiological state in which alphafetoprotein levels are high while the gonadal activity is not characterized by ovulatory cycles. In order to assess our hypothesis, alpha-fetoprotein was studied in the ovary of pregnant rats from day 18 to 21 of gestation by an immunofluorescent technique, and alpha-fetoprotein was assayed in plasma samples. The results of this work show that, during pregnancy, follicular maturation is blocked at the antral stage, and the follicles contain degenerating oocytes that are AFP-positive in immunofluorescence. In conclusion, we suggest that the alpha-fetoprotein produced by the fetal liver and the yolk sac is disseminated in the amniotic fluid and passes through the placenta, and then reaches the ovarian follicles and the oocytes. The possible role of alphafetoprotein in follicular atresia is discussed.

  17. A Taenia crassiceps metacestode factor enhances ovarian follicle atresia and oocyte degeneration in female mice.

    PubMed

    Solano, S; Zepeda, N; Copitin, N; Fernandez, A M; Tato, P; Molinari, J L

    2015-01-01

    The histopathological effects of Taenia crassiceps infection or T. crassiceps metacestode factor inoculation on the mouse ovary were determined using six female mice in three groups: infected mice, mice inoculated with the metacestode factor and control mice. The control group was subcutaneously inoculated with healthy peritoneal fluid. The infected group was intraperitoneally inoculated with 40 T. crassiceps metacestodes, and the metacestode factor group was subcutaneously inoculated with T. crassiceps metacestode factor (MF). Light and electron microscopy and TUNEL (terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labelling) assays revealed a significant increase in ovarian follicular atresia (predominantly in antral/preovulatory stages of development), oocyte degeneration (P< 0.05), and a decrease in the amount of corpus luteum in follicles of mice infected and inoculated with MF compared with the control group. Significant abnormalities of the granulosa cells and oocytes of the primordial, primary and secondary ovarian follicles occurred in both treated mouse groups (P< 0.05) compared with no degeneration in the control group. These pathological changes in female mice either infected with T. crassiceps metacestodes or inoculated with T. crassiceps MF may have consequences for ovulation and fertility.

  18. The significance of small intestinal epithelium in gastric antral biopsies in children.

    PubMed

    Weinberg, Arthur G

    2012-01-01

    Intestinal metaplasia of the gastric antrum is common in adults with chronic gastritis and occurs in Helicobacter -associated gastritis in children. This study examined the frequency and clinical correlates of intestinal epithelium in 1690 consecutive antral biopsies obtained from children over a 2-year period in a tertiary pediatric care facility. Intestinal epithelium in gastric glands not associated with overlying villi was present in 22 (1.3%) biopsies. These came from 20 patients, 2-17 years of age, none of whom had clinical or histologic evidence of Helicobacter infection or significant chronic gastritis. Eight (40%) had an antral pancreatic rest, 8 had some other localized antral abnormality, and 4 were endoscopically normal. Four additional patients with a pancreatic rest had no intestinal epithelium. Six surgically resected rests and 2 rests found at autopsy were also reviewed. Heterotopic intestinal epithelium was present in 1 of the 2 postmortem specimens but was absent from all 6 surgically resected lesions. No intestinal epithelium was present in 67 antral biopsies with Helicobacter gastritis observed during this same period. Although the intestinal epithelium in these patients could be metaplastic, it more likely represents inadvertent sampling of the gastroduodenal junction induced by a lesion in the distal antrum or a focus of heterotopic epithelium and might best be addressed in the surgical pathology report by a comment to this effect. The distinction from metaplasia is more than semantic, because a diagnosis of intestinal metaplasia can have adverse clinical implications and should be made with caution in a child.

  19. Remote ischemic preconditioning on neovascularization and follicle viability on ovary autotransplantation in rats.

    PubMed

    Damous, L L; Silva, S M; Simões, R S; Morello, R J; Carbonel, A P F; Simões, M J; Montero, E F S

    2008-04-01

    Verify the optimum remote vascular occlusion time to reduce ovarian injury in autologous transplants in rats. Twenty-four adult female rats were assigned to four groups: GC (control group): bilateral oophorectomy followed by ovary transplant; GIPC (ischemic preconditioning group): remote ischemic preconditioning at the iliac artery for 5, 10, and 15 minutes (GIPC-5, GIPC-20, and GIPC-15) previous to bilateral oophorectomy and ovarian transplantation. The right ovary was fixed in the retroperitoneum. Euthanasia was performed 4 days after the surgical procedure. The follicles were counted and classified as developing versus atretic. The immunohistochemical assay identified vascular factor of endothelial growth (VEGF) in the ovarian stroma and assessed the proliferation capacity by means of the Ki-67 in the ovarian follicles. Every group showed an inflammatory infiltrate, luteous body, and ovarian follicles in several phases of development. The ischemic preconditioning groups displayed greater amounts of viable ovarian follicles and increased vascularization and vasodilatation than the control group. GIPC-15 showed the highest amount of viable follicles compared to the others (P < .05 GIPC-15 vs GC; GIPC-15 vs GIPC-5). More VEGF-labeled cells were observed in GIPC-10 than the control group (P < .05, GIPC-10 vs GC). The proliferation index assessed by Ki-67 marking showed GC: 80%; GIPC-5: 76%; GIPC-10: 67%; and GIPC-15: 64% (P > .05). The PCI-15 cohort seem to be the most adequate timing to achieve functional support and preservation of a greater number of viable ovarian follicles.

  20. Immunohistochemical study of hair follicle stem cells in regenerated hair follicles induced by Wnt10b

    PubMed Central

    Zhang, Yiming; Xing, Yizhan; Guo, Haiying; Ma, Xiaogen; Li, Yuhong

    2016-01-01

    The regulation of the periodic regeneration of hair follicles is complicated. Although Wnt10b has been reported to induce hair follicle regeneration, the characteristics of induced hair follicles, especially the target cells of Wnt10b, have not yet been clearly elucidated. Thus, we systematically evaluated the expression and proliferation patterns of Wnt10b-induced hair follicles. We found that Wnt10b promoted the proliferation of hair follicle stem cells from 24 hours after AdWnt10b injection. Seventy-two hours after AdWnt10b injection, cells outside of bulge area began to proliferate. When the induced hair follicle entered full anagen, although the hair follicle stem cells were normal, canonical Wnt signaling was maintained in the hair precortex cells. Our results reveal that the target cells that overexpressed Wnt10b included hair follicle stem cells, hair precortex cells, and matrix cells. PMID:27766026

  1. In vitro production of a caprine embryo from a preantral follicle cultured in media supplemented with growth hormone.

    PubMed

    Magalhães, D M; Duarte, A B G; Araújo, V R; Brito, I R; Soares, T G; Lima, I M T; Lopes, C A P; Campello, C C; Rodrigues, A P R; Figueiredo, J R

    2011-01-01

    The objective was to evaluate the effects of growth hormone (GH) on the survival, growth, maturation, and fertilization of oocytes derived from caprine preantral ovarian follicles cultured in vitro. Preantral follicles were isolated from the cortex of caprine ovaries and individually cultured for 18 d in the absence (control) or presence of bovine GH at concentrations of 10 or 50 ng/mL (GH10 and GH50, respectively). Follicle development was evaluated on the basis of survival, antral cavity formation, diameter increase, and the presence of healthy cumulus-oocyte complexes and mature oocytes. After culture, oocytes were subjected to in vitro maturation (IVM) and in vitro fertilization (IVF). The rate of antrum formation after Day 6 of culture was higher in both GH10 and GH50 than in the control (81.0, 92.7, and 47.6%, respectively, P < 0.05). Percentages of grown oocytes that were acceptable for IVM were also higher (P < 0.05) in GH-treated groups than in the control (54.8, 48.8, and 11.9% for GH10, GH50, and Control). A higher percentage of oocytes in the GH50 treatment underwent meiotic resumption (50.0%), produced mature oocytes, and enabled production of an embryo after IVF than in the control group (0.0%; P < 0.05). In conclusion, GH promoted in vitro growth and maturation of goat preantral follicle oocytes and enabled production of an embryo. Furthermore, this study was apparently the first to produce a caprine embryo by in vitro fertilization of oocytes derived from preantral follicles grown in vitro.

  2. Altered expression of Bcl-2 and Bax in follicles within dehydroepiandrosterone-induced polycystic ovaries in rats.

    PubMed

    Bas, Diana; Abramovich, Dalhia; Hernandez, Fatima; Tesone, Marta

    2011-05-01

    PCOS (polycystic ovary syndrome) is a heterogeneous disease characterized by hyperandrogenaemia, hirsutism, oligo- or amenorrhea, insulin resistance and anovulation. The aim of the present study was to evaluate if the balance between the ovarian expression of Bax (proapoptotic protein) and Bcl-2 (antiapoptotic protein) is altered in a PCOS model developed in rats by DHEA (dehydroepiandrosterone) administration. In addition, the ovarian morphology and the circulating progesterone levels were evaluated. Histological studies confirmed the presence of follicular cysts, atretic follicles and the absence of corpora lutea in the ovaries from the PCOS group and a significant decrease in circulating progesterone levels. Immunohistochemical studies showed that the expression of Bcl-2 and Bax were mainly localized in granulosa cells of AFs (antral follicles) in both groups. Bax expression was greater in preantral and AFs from PCOS ovarian sections than in the controls. In contrast, intense Bcl-2 immunostaining was observed in the control AFs, while Bcl-2 protein was either absent in PFs (preantral follicles) or weakly expressed in AFs from PCOS rats. These results were partially confirmed by Western studies. Data revealed that the ovarian level of Bcl-2 protein was lower in PCOS than in the control and that there were no differences in Bax ovarian levels between groups. However, Bax/Bcl-2 ratio was significantly higher in PCOS group than in the control group. In conclusion, an increase in ovarian apoptosis through an imbalance among the Bcl-2 family members may be involved in the transformation of growing follicles in cystic follicles in the ovaries from DHEA-induced PCOS rats.

  3. Relative mRNA expression and immunolocalization for transforming growth factor-beta (TGF-β) and their effect on in vitro development of caprine preantral follicles.

    PubMed

    Rodrigues, G Q; Bertoldo, M J; Brito, I R; Silva, C M G; Sales, A D; Castro, S V; Duffard, N; Locatelli, Y; Mermillod, P; Lobo, C H; Campello, C C; Rodrigues, A P R; Freitas, V J F; Figueiredo, J R

    2014-09-01

    This study aimed to evaluate the immunolocalization and messenger RNA (mRNA) expression for transforming growth factor-beta (TGF-β) and its receptors (TGF-βRI and RII), as well as mRNA expression for P450 aromatase and FSH receptor in caprine preantral follicles. The effects of TGF-β, FSH alone, or in association on the in vitro follicular development were also assessed. Immunohistochemical analyses showed the expression of TGF-β and its receptors in oocytes of all follicle stages and granulosa cells of primary and secondary follicles. mRNA for TGF-β receptors and for FSH receptor (FSHR) was present in preantral follicles as well as in oocytes and granulosa cells of antral follicles. Isolated secondary follicles were cultured in α-minimum essential medium (MEM) alone or supplemented with either FSH (100 ng/ml), TGF-β (10 ng/ml), or TGF-β + FSH for 18 d. TGF-β increased significantly oocyte diameter when compared to FSH alone and control. After 18 d of culture, all groups showed a significant reduction in P450 aromatase and FSHR mRNA levels in comparison to fresh control. In contrast, treatment with FSH significantly increased the mRNA expression for TGF-β in comparison to fresh control and other treatments. In conclusion, the findings showed that TGF-β and its receptors are present in caprine ovarian follicles. Furthermore, they showed a positive effect on oocyte growth in vitro.

  4. Microinjection of Follicle-Enclosed Mouse Oocytes

    NASA Astrophysics Data System (ADS)

    Jaffe, Laurinda A.; Norris, Rachael P.; Freudzon, Marina; Ratzan, William J.; Mehlmann, Lisa M.

    The mammalian oocyte develops within a complex of somatic cells known as a follicle, within which signals from the somatic cells regulate the oocyte, and signals from the oocyte regulate the somatic cells. Because isolation of the oocyte from the follicle disrupts these communication pathways, oocyte physiology is best studied within an intact follicle. Here we describe methods for quantitative microinjection of follicle-enclosed mouse oocytes, thus allowing the introduction of signaling molecules as well as optical probes into the oocyte within its physiological environment.

  5. Morphological evidence of apoptosis and the prevalence of apoptotic versus mitotic cells in the membrana granulosa of ovarian follicles during spontaneous and induced atresia in ewes.

    PubMed

    Jolly, P D; Smith, P R; Heath, D A; Hudson, N L; Lun, S; Still, L A; Watts, C H; McNatty, K P

    1997-04-01

    Apoptosis is a process by which granulosa cells are thought to be deleted during ovarian follicular atresia. The aims of the present studies, using sheep as the experimental model, were to determine 1) whether morphological changes in cells composing the membrana granulosa during the process of atresia conformed with the general criteria of apoptotic cell death as assessed using tissue sections stained with hematoxylin and eosin; 2) whether cells classified as apoptotic on the basis of their morphology contained fragmented DNA using an in situ 3' end-labeling technique; and 3) the degree of apoptosis and mitosis within the granulosa cell populations of large antral follicles (> or = 3 mm in diameter) during both spontaneous and experimentally induced atresia using stereological methods. The results showed that most degenerate granulosa cells in follicles undergoing atresia display the morphological characteristics of apoptosis, suggesting that this is the most common pathway of cell deletion. Typical features were cells containing nuclei with marginated chromatin; cells with a single small densely staining nucleus (pyknotic appearance); cells with multiple smaller, densely staining nuclear fragments; and densely staining membrane-bound bodies (apoptotic bodies) either singly or in clusters. Cells with morphological features more typical of oncosis or necrosis were sometimes observed, but mainly during the later stages of atresia. All cells classified as apoptotic on the basis of morphological criteria contained fragmented DNA as measured by 3' end-labeling. Apoptotic bodies and/or cells were found in all follicles examined, including those classified as healthy. The overall prevalence of apoptotic cells plus apoptotic bodies expressed as a percentage of the total granulosa cell number per follicle varied from 0.02% to 0.20% in healthy follicles, varied from 0.21% to 2.00% in follicles in early (primary) atresia, and was > 2.0% in follicles in later (secondary

  6. Notch1 and Notch2 receptors regulate mouse and human gastric antral epithelial cell homoeostasis.

    PubMed

    Gifford, Gail B; Demitrack, Elise S; Keeley, Theresa M; Tam, Andrew; La Cunza, Nilsa; Dedhia, Priya H; Spence, Jason R; Simeone, Diane M; Saotome, Ichiko; Louvi, Angeliki; Siebel, Christian W; Samuelson, Linda C

    2017-06-01

    We tested the ability of Notch pathway receptors Notch1 and Notch2 to regulate stem and epithelial cell homoeostasis in mouse and human gastric antral tissue. Mice were treated with the pan-Notch inhibitor dibenzazepine (DBZ) or inhibitory antibodies targeting Notch1 and/or Notch2. Epithelial proliferation, apoptosis and cellular differentiation were measured by histological and molecular approaches. Organoids were established from mouse and human antral glands; growth and differentiation were measured after treatment with Notch inhibitors. Notch1 and Notch2 are the predominant Notch receptors expressed in mouse and human antral tissue and organoid cultures. Combined inhibition of Notch1 and Notch2 in adult mice led to decreased epithelial cell proliferation, including reduced proliferation of LGR5 stem cells, and increased apoptosis, similar to the response to global Notch inhibition with DBZ. Less pronounced effects were observed after inhibition of individual receptors. Notch pathway inhibition with DBZ or combined inhibition of Notch1 and Notch2 led to increased differentiation of all gastric antral lineages, with remodelling of cells to express secretory products normally associated with other regions of the GI tract, including intestine. Analysis of mouse and human organoids showed that Notch signalling through Notch1 and Notch2 is intrinsic to the epithelium and required for organoid growth. Notch signalling is required to maintain gastric antral stem cells. Notch1 and Notch2 are the primary Notch receptors regulating epithelial cell homoeostasis in mouse and human stomach. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

  7. Ultrasonographic assessment of intragastric volume in neonates: factors affecting the relationship between intragastric volume and antral cross-sectional area.

    PubMed

    Tomomasa, T; Tabata, M; Nako, Y; Kaneko, H; Morikawa, A

    1996-11-01

    Measuring antral cross-sectional area by ultrasonography can be an ideal way to evaluate intragastric milk volume in infants. Technical details, however, remain to be established before its clinical application. We investigated the effects of posture and ultrasonographic plane on the correlation between milk volume and antral cross-sectional area. After gastric aspiration through a nasogastric tube, healthy newborns were fed 0, 10, 20, and 40 ml of milk cumulatively, and antral cross-sectional area was measured in either upright, sitting, or right lateral position. To determine the best sonographic plane, subjects were put in the right lateral position and antral cross-sectional area was measured in the plane of the aorta and either the superior mesenteric artery, the superior mesenteric vein, the midline of the abdominal surface, 1 cm right of midline, or 2 cm right of midline. The results showed that antral cross-sectional area reflects intragastric milk volume most accurately, with minimal gas interference when measured in the right lateral position. The area correlates well with milk volume in the plane of the aorta and either the superior mesenteric artery, the superior mesenteric vein, or the midline. Next, we studied the effect of intragastric gas on the antral cross-sectional area in subjects who were given 40 ml of milk followed by an injection of air. More than 20 ml of intragastric gas increases antral cross-sectional area significantly. Ultrasonographic evaluation of intragastric volume requires attention to the above factors.

  8. Age-Specific Serum Anti-Mullerian Hormone and Follicle Stimulating Hormone Concentrations in Infertile Iranian Women

    PubMed Central

    Raeissi, Alireza; Torki, Alireza; Moradi, Ali; Mousavipoor, Seyed Mehdi; Pirani, Masoud Doosti

    2015-01-01

    Background Anti-Müllerian hormone (AMH) is secreted by the granulosa cells of growing follicles during the primary to large antral follicle stages. Abnormal levels of AMH and follicle stimulating hormone (FSH) may indicate a woman’s diminished ability or inability to conceive. Our aim is to investigate the changes in serum AMH and FSH concentrations at different age groups and its correlation with ovarian reserves in infertile women. Materials and Methods This cross-sectional study analyzed serum AMH and FSH levels from 197 infertile women and 176 healthy controls, whose mean ages were 19-47 years. Sample collection was performed by random sampling and analyzed with SPSS version 16 software. Results There were significantly lower mean serum AMH levels among infertile women compared to the control group. The mean AMH serum levels from different ages of infertile and control group (fertile women) decreased with increasing age. However, this reduction was greater in the infertile group. The mean FSH serum levels of infertile women were significantly higher than the control group. Mean serum FSH levels consistently increased with increasing age in infertile women; however mean luteinizing hormone (LH) levels were not consistent. Conclusion We have observed increased FSH levels and decreased AMH levels with increasing age in women from 19 to 47 years of age. Assessments of AMH and FSH levels in combination with female age can help in predicting ovarian reserve in infertile women. PMID:25918589

  9. Age-specific serum anti-mullerian hormone and follicle stimulating hormone concentrations in infertile Iranian women.

    PubMed

    Raeissi, Alireza; Torki, Alireza; Moradi, Ali; Mousavipoor, Seyed Mehdi; Pirani, Masoud Doosti

    2015-01-01

    Anti-Müllerian hormone (AMH) is secreted by the granulosa cells of growing follicles during the primary to large antral follicle stages. Abnormal levels of AMH and follicle stimulating hormone (FSH) may indicate a woman's diminished ability or inability to conceive. Our aim is to investigate the changes in serum AMH and FSH concentrations at different age groups and its correlation with ovarian reserves in infertile women. This cross-sectional study analyzed serum AMH and FSH levels from 197 infertile women and 176 healthy controls, whose mean ages were 19-47 years. Sample collection was performed by random sampling and analyzed with SPSS version 16 software. There were significantly lower mean serum AMH levels among infertile women compared to the control group. The mean AMH serum levels from different ages of infertile and control group (fertile women) decreased with increasing age. However, this reduction was greater in the infertile group. The mean FSH serum levels of infertile women were significantly higher than the control group. Mean serum FSH levels consistently increased with increasing age in infertile women; however mean luteinizing hormone (LH) levels were not consistent. We have observed increased FSH levels and decreased AMH levels with increasing age in women from 19 to 47 years of age. Assessments of AMH and FSH levels in combination with female age can help in predicting ovarian reserve in infertile women.

  10. Comparison of automated and manual follicle monitoring in an unrestricted population of 100 women undergoing controlled ovarian stimulation for IVF.

    PubMed

    Ata, Baris; Seyhan, Ayse; Reinblatt, Shauna Leigh; Shalom-Paz, Einat; Krishnamurthy, Srinivasan; Tan, Seang Lin

    2011-01-01

    Ovarian response to gonadotrophin stimulation is monitored with serial ultrasound (US) examinations. Sonography-based Automated Volume Count (SonoAVC) is a relatively new three-dimensional (3D) US technology, which automatically generates a set of measurements including the mean follicular diameter (MFD) and a volume-based diameter (d(V)) for each follicle in the ovaries. The present study aimed to assess the applicability and reproducibility of this automated follicle measurement method in an IVF programme. For this prospective method comparison study, 100 women undergoing US monitoring of a controlled ovarian stimulation cycle were recruited. Each follicle was manually measured by taking the mean of maximal diameters on three orthogonal planes with two-dimensional (2D) US. A 3D volume of each ovary was then captured. The ovarian volumes were later analysed using SonoAVC. The agreement between the two methods for the numbers of follicles and the size of the leading follicle was assessed with the Bland-Altman method. The reproducibility of SonoAVC measurements was assessed with the intraclass correlation coefficient (ICC). Both SonoAVC-generated MFD and d(V)-based follicle counts, as well as the leading follicle diameter, had good agreement with conventional 2D US measurements. SonoAVC measurements had very good reproducibility, with ICC ≥0.8 for most evaluations. Automated follicle monitoring with SonoAVC can replace or be used interchangeably with conventional 2D measurements. Automated follicle monitoring can save time, provide a method of quality control and create opportunities for developing HCG criteria based on follicular volume or for monitoring patients from a distance.

  11. Xenobiotic Effects on Ovarian Preantral Follicles1

    PubMed Central

    Mark-Kappeler, Connie J.; Hoyer, Patricia B.; Devine, Patrick J.

    2011-01-01

    Women are born with a finite population of ovarian follicles, which are slowly depleted during their reproductive years until reproductive failure (menopause) occurs. The rate of loss of primordial follicles is determined by genetic and environmental influences, but certain toxic exposures can accelerate this process. Ionizing radiation reduces preantral follicle numbers in rodents and humans in a dose-dependent manner. Cigarette smoking is linked to menopause occurring 1–4 yr earlier than with nonsmokers, and components of smoke, polycyclic aromatic hydrocarbons, can cause follicle depletion in rodents or in ovaries in vitro. Chemotherapeutic agents, such as alkylating drugs and cisplatin, also cause loss of preantral ovarian follicles. Effects depend on dose, type, and reactivity of the drug, and the age of the individual. Evidence suggests DNA damage may underlie follicle loss induced by one common alkylating drug, cyclophosphamide. Occupational exposures have also been linked to ovarian damage. In an industrial setting, 2-bromopropane caused infertility in men and women, and it can induce ovarian follicle depletion in rats. Solvents, such as butadiene, 4-vinylcyclohexene, and their diepoxides, can also cause specific preantral follicle depletion. The mechanism(s) underlying effects of the latter compound may involve alterations in apoptosis, survival factors such as KIT/Kit Ligand, and/or the cellular signaling that maintains primordial follicle dormancy. Estrogenic endocrine disruptors may alter follicle formation/development and impair fertility or normal development of offspring. Thus, specific exposures are known or suspected of detrimentally impacting preantral ovarian follicles, leading to early ovarian failure. PMID:21697514

  12. White Blood Cell Count

    MedlinePlus

    ... limited. Home Visit Global Sites Search Help? White Blood Cell Count Share this page: Was this page helpful? ... Count; Leukocyte Count; White Count Formal name: White Blood Cell Count Related tests: Complete Blood Count , Blood Smear , ...

  13. Counting Penguins.

    ERIC Educational Resources Information Center

    Perry, Mike; Kader, Gary

    1998-01-01

    Presents an activity on the simplification of penguin counting by employing the basic ideas and principles of sampling to teach students to understand and recognize its role in statistical claims. Emphasizes estimation, data analysis and interpretation, and central limit theorem. Includes a list of items for classroom discussion. (ASK)

  14. Counting Populations

    ERIC Educational Resources Information Center

    Damonte, Kathleen

    2004-01-01

    Scientists use sampling to get an estimate of things they cannot easily count. A population is made up of all the organisms of one species living together in one place at the same time. All of the people living together in one town are considered a population. All of the grasshoppers living in a field are a population. Scientists keep track of the…

  15. Counting Penguins.

    ERIC Educational Resources Information Center

    Perry, Mike; Kader, Gary

    1998-01-01

    Presents an activity on the simplification of penguin counting by employing the basic ideas and principles of sampling to teach students to understand and recognize its role in statistical claims. Emphasizes estimation, data analysis and interpretation, and central limit theorem. Includes a list of items for classroom discussion. (ASK)

  16. Counting Populations

    ERIC Educational Resources Information Center

    Damonte, Kathleen

    2004-01-01

    Scientists use sampling to get an estimate of things they cannot easily count. A population is made up of all the organisms of one species living together in one place at the same time. All of the people living together in one town are considered a population. All of the grasshoppers living in a field are a population. Scientists keep track of the…

  17. The lack of significant changes in scalp hair follicle density with advancing age.

    PubMed

    Sinclair, R; Chapman, A; Magee, J

    2005-04-01

    Age-related reduction in hair is seen in the axillary and pubic regions as well as the scalp; however, it has not been investigated qualitatively on the scalp. Horizontally sectioned scalp biopsy is an ideal tool to investigate the impact of advancing age on scalp hair follicle density and morphology. To examine the effect of age and follicle miniaturization on total hair count in 1666 horizontally sectioned mid-scalp biopsies from 928 women aged between 13 and 84 years with hair loss. Specialist hair loss referral clinic in a teaching hospital. Analysis of data set. Methods: All scalp biopsies were 4 mm in diameter and taken from the crown. Miniaturization was assessed by calculating the ratio of terminal to vellus-like hairs (T/V) at the mid-isthmus level and considered significant if the ratio was < or = 4 : 1. Fibrosis was documented when present. Linear regression was used to examine the association between total hair count, age and miniaturization. The average number of hair follicles per biopsy was 39.6 (SD +/- 10.8). A highly significant negative association (P < 0.0001) was found between age and total follicle number, although the predictive value of age in total hair count was found to be small [root error mean square (R2) < 2%]. Controlling for T/V < or = 4 : 1, the association was weakened, but remained significant. The relationship unconfounded by T/V < or = 4 : 1 shows that for every additional year of ageing, 0.077 total hair follicles (0.22%) are lost per biopsy. Age and follicular miniaturization were found to be extremely weak predictors of total hair count in women with hair loss.

  18. Isolation and culture of chicken primordial follicles.

    PubMed

    Leghari, Imdad Hussain; Zhao, Dan; Mi, Yuling; Zhang, Caiqiao

    2015-10-01

    The establishment of a primordial follicle culture system is important for the study of follicular development. Hence, the objective of this study was to isolate chicken primordial follicles and establish culture methods. Ovaries from 2-wk-old chickens were treated with trypsin-EDTA, collagenase II, or collagenase type IA, along with a mechanical isolation technique. Isolated follicles were cultured under different conditions. Results showed a significant difference in the follicular recovery and survival rates among different enzymes and methods used. The maximal follicular yield was obtained by trypsin+EDTA and collagenase II digestion, followed by collagenase type IA digestion. However, the highest follicular viability rate was observed in groups of collagenase type IA digestion and the mechanical isolation method. Enzymatic treatment resulted in higher misshapen oocytes or follicles, though the diameters of the follicles were not significantly changed. In addition, our follicle culture results for different conditions showed maximal survival rates of primordial follicles in alginate hydrogel beads after 12 d of culture. Thus, we successfully established methods for isolating and culturing chicken primordial follicles. The present method will greatly facilitate investigation of the regulation of follicular development. © 2015 Poultry Science Association Inc.

  19. Ion beam microanalysis of human hair follicles

    NASA Astrophysics Data System (ADS)

    Kertész, Zs.; Szikszai, Z.; Pelicon, P.; Simčič, J.; Telek, A.; Bíró, T.

    2007-07-01

    Hair follicle is an appendage organ of the skin which is of importance to the survival of mammals and still maintains significance for the human race - not just biologically, but also through cosmetic and commercial considerations. However data on composition of hair follicles are scarce and mostly limited to the hair shaft. In this study we provide detailed information on the elemental distribution in human hair follicles in different growth phases (anagen and catagen) using a scanning proton microprobe. The analysis of skin samples obtained from human adults undergoing plastic surgery and of organ-cultured human hair follicles may yield a new insight into the function, development and cyclic activity of the hair follicle.

  20. Immunohistochemical and Ultrastructural Study of the Lamellae of Oocytes in Atretic Follicles in Relation to Different Processes of Cell Death

    PubMed Central

    Escobar, M.L.; Echeverría, O.M.; García, G.; Ortiz, R.; Vázquez-Nin, G.H.

    2015-01-01

    Atresia is the process through which non-selectable oocytes are eliminated; it involves apoptosis and/or autophagy. This study used immunohistochemical and ultrastructural techniques to characterize the lamellae present in the cytoplasm of oocytes in follicles in the process of atresia in prepubertal and adult Wistar rats. The results indicate that the lamellae are positive to tubulin and myosin immunodetection under light and electron microscopy. Labeling is greater with anti-tubulin and lesser with anti-myosin. Our observations indicate that lamellae are present in oocytes at the initial antral stage in prepubertal rats; that is, from day 14 post-birth to adult age. We were able to determine that the increase in altered lamellae principally occurs in the apoptotic cells rather than in the autophagic cells. PMID:26428888

  1. Effects of ionizing radiation and pretreatment with (D-Leu6,des-Gly10) luteinizing hormone-releasing hormone ethylamide on developing rat ovarian follicles

    SciTech Connect

    Jarrell, J.; YoungLai, E.V.; McMahon, A.; Barr, R.; O'Connell, G.; Belbeck, L.

    1987-10-01

    To assess the effects of a gonadotropin-releasing hormone agonist, (D-Leu6,des-Gly10) luteinizing hormone-releasing hormone ethylamide, in ameliorating the damage caused by ionizing radiation, gonadotropin-releasing hormone agonist was administered to rats from day 22 to 37 of age in doses of 0.1, 0.4, and 1.0 microgram/day or vehicle and the rats were sacrificed on day 44 of age. There were no effects on estradiol, progesterone, luteinizing, or follicle-stimulating hormone, nor an effect on ovarian follicle numbers or development. In separate experiments, rats treated with gonadotropin-releasing hormone agonist in doses of 0.04, 0.1, 0.4, or 1.0 microgram/day were either irradiated or sham irradiated on day 30 and all groups sacrificed on day 44 of age. Irradiation produced a reduction in ovarian weight and an increase in ovarian follicular atresia. Pretreatment with the agonist prevented the reduction in ovarian weight and numbers of primordial and preantral follicles but not healthy or atretic antral follicles. Such putative radioprotection should be tested on actual reproductive performance.

  2. A new surgical management for oro-antral communication: the resorbable guided tissue regeneration membrane--bone substitute sandwich technique.

    PubMed

    Ogunsalu, C

    2005-09-01

    This paper describes a new technique for the closure of oro-antral fistula/communication, in which both hard tissue (bone) and soft tissue closure is achieved. The sandwich technique utilizes a suitable bone grafting material sandwiched between two sheaths of Biogide (a resorbable membrane) for the hard tissue closure of oro-antral communication post traumatic exodontia. The bone grafting material utilized for this case was Bio-oss. The result obtained was excellent with regeneration of sufficient bony tissue to allow placement of an endosseous implant. This sandwich technique is a simple and excellent technique for the closure of oro-antral communication, especially when subsequent placement of endosseous implant is considered without the need of donor site surgery for bone grafting. The otorhinolaryngologists and oral and maxillofacial surgeons should find this technique very useful in the closure of oro-antral fistulae.

  3. Increased abundance of aromatase and follicle stimulating hormone receptor mRNA and decreased insulin-like growth factor-2 receptor mRNA in small ovarian follicles of cattle selected for twin births.

    PubMed

    Echternkamp, S E; Aad, P Y; Eborn, D R; Spicer, L J

    2012-07-01

    Cattle genetically selected for twin ovulations and births (Twinner) exhibit increased ovarian follicular development, increased ovulation rate, and greater blood and follicular fluid IGF-1 concentrations compared with contemporary cattle not selected for twins (Control). Experimental objectives were to 1) assess relationships among aromatase (CYP19A1), IGF-1 (IGF1), IGF-2 receptor (IGF2R), and FSH receptor (FSHR) mRNA expression in small (≤5 mm) antral follicles and 2) determine their association with increased numbers of developing follicles in ovaries of Twinner females. Ovaries were collected from mature, cyclic (d 3 to 6) Twinner (n = 11), and Control (n = 12) cows at slaughter and pieces of cortical tissue were fixed and embedded in paraffin. Expression of mRNA was evaluated by in situ hybridization using (35)S-UTP-labeled antisense and sense probes for CYP19A1, FSHR, IGF1, and IGF2R mRNA. Silver grain density was quantified within the granulosa and theca cells of individual follicles (2 to 7 follicles/cow) by Bioquant image analysis. Follicles of Twinners tended to be smaller in diameter than Controls (1.9 ± 0.1 vs. 2.3 ± 0.1 mm; P = 0.08), but thickness of granulosa layer did not differ (P > 0.1) by genotype. Relative abundance of CYP19A1 (P < 0.01) and FSHR (P < 0.05) mRNA was greater in granulosa cells of Twinners vs. Controls, respectively, whereas IGF2R mRNA expression was less in both granulosa (P < 0.01) and theca (P < 0.05) cells in follicles of Twinners vs. Controls, respectively. Abundance of CYP19A1 mRNA in granulosa cells was correlated negatively with IGF2R mRNA expression in both granulosa (r = -0.33; P < 0.01) and theca (r = -0.21; P = 0.05) cells. Expression of IGF1 mRNA was primarily in granulosa cells, including cumulus cells, and its expression did not differ between Twinners vs. Controls (P > 0.10). Detected increases in CYP19A1 and FSHR, but not IGF1, mRNA expression along with decreases in IGF2R mRNA expression in individual

  4. Tooth follicle extirpation and uvulectomy.

    PubMed

    Johnston, N L; Riordan, P J

    2005-12-01

    Migration is not only the movement of people, but also of their culture, customs and beliefs. As more people from developing countries in Africa migrate to industrialized countries, the more likely health professionals will find themselves providing care for people of whose customs and practices they have little knowledge. This review of the literature suggests that removal of deciduous canine follicles and uvulectomy are frequently practised in some African and neighbouring countries. Reasons given for deciduous canine extirpation include the prevention of vomiting, fever and diarrhoea. The indications for uvulectomy appear widespread, including treatment for persistent fever, coughing and growth retardation. The practices are usually performed by traditional healers. Risks for children who undergo these procedures are extensive, including septicaemia, potential for HIV transmission, numerous dental complications and death. With improved understanding between Western health teams and local, traditional people, an improved system may develop whereby the two systems can work together in providing improved health outcomes for the people.

  5. Erythropoietin resistance in end-stage renal disease patient with gastric antral vascular ectasia.

    PubMed

    Lee, Desiree Ji Re; Fragata, Juliana; Pestana, José Osmar Medina; Draibe, Sergio; Canziani, Maria Eugênia; Cendoroglo, Miguel; Góes, Miguel Ângelo de

    2015-01-01

    We observed a case of recombinant human erythropoietin resistance caused by Gastric Antral Vascular Ectasia in a 40-year-old female with ESRD on hemodialysis. Some associated factors such as autoimmune disease, hemolysis, heart and liver disease were discarded on physical examination and complementary tests. The diagnosis is based on the clinical history and endoscopic appearance of watermelon stomach. The histologic findings are fibromuscular proliferation and capillary ectasia with microvascular thrombosis of the lamina propria. However, these histologic findings are not necessary to confirm the diagnosis. Gastric Antral Vascular Ectasia is a serious condition and should be considered in ESRD patients on hemodialysis with anemia and resistance to recombinant human erythropoietin because GAVE is potentially curable with specific endoscopic treatment method or through surgical procedure.

  6. Alveolar antral artery isolation during sinus lift procedure with the double window technique.

    PubMed

    Maridati, Paolo; Stoffella, Enrico; Speroni, Stefano; Cicciu, Marco; Maiorana, Carlo

    2014-01-01

    The sinus lift technique, introduced in 1976 by Tatum and subsequently described by Boyne in 1980, is nowadays considered a safe and reliable procedure for the rehabilitation of the atrophic upper posterior maxilla. The alveolar antral artery (AAA) is anastomoses between the posterior superior alveolar artery (PSAA) and the infraorbital artery (IOA) and may be present in the sinusal antrostomy. The haemorrhage of this vascular bundle represents the second intra-operatory complication in term of frequency during sinus lift procedure. Purpose of this study was to illustrate and describe a new technique allowing the AAA isolation during sinus lift procedure in cases in which the artery is clearly present inside the surgical area, detectable through CT scan exam. Presence, course and possible identification of the alveolar antral artery are also discussed, according to the studies present in the literature.

  7. Altered Theca and Cumulus Oocyte Complex Gene Expression, Follicular Arrest and Reduced Fertility in Cows with Dominant Follicle Follicular Fluid Androgen Excess

    PubMed Central

    Summers, Adam F.; Pohlmeier, William E.; Sargent, Kevin M.; Cole, Brizett D.; Vinton, Rebecca J.; Kurz, Scott G.; McFee, Renee M.; Cushman, Robert A.; Cupp, Andrea S.; Wood, Jennifer R.

    2014-01-01

    Aspiration of bovine follicles 12–36 hours after induced corpus luteum lysis serendipitously identified two populations of cows, one with High androstenedione (A4; >40 ng/ml; mean = 102) and another with Low A4 (<20 ng/ml; mean = 9) in follicular fluid. We hypothesized that the steroid excess in follicular fluid of dominant follicles in High A4 cows would result in reduced fertility through altered follicle development and oocyte maternal RNA abundance. To test this hypothesis, estrous cycles of cows were synchronized and ovariectomy was performed 36 hours later. HPLC MS/MS analysis of follicular fluid showed increased dehydroepiandrosterone (6-fold), A4 (158-fold) and testosterone (31-fold) in the dominant follicle of High A4 cows. However, estrone (3-fold) and estradiol (2-fold) concentrations were only slightly elevated, suggesting a possible inefficiency in androgen to estrogen conversion in High A4 cows. Theca cell mRNA expression of LHCGR, GATA6, CYP11A1, and CYP17A1 was greater in High A4 cows. Furthermore, abundance of ZAR1 was decreased 10-fold in cumulus oocyte complexes from High A4 cows, whereas NLRP5 abundance tended to be 19.8-fold greater (P = 0.07). There was a tendency for reduction in stage 4 follicles in ovarian cortex samples from High A4 cows suggesting that progression to antral stages were impaired. High A4 cows tended (P<0.07) to have a 17% reduction in calving rate compared with Low A4 cows suggesting reduced fertility in the High A4 population. These data suggest that the dominant follicle environment of High A4 cows including reduced estrogen conversion and androgen excess contributes to infertility in part through altered follicular and oocyte development. PMID:25330369

  8. Conditional deletion of menin results in antral G cell hyperplasia and hypergastrinemia

    PubMed Central

    Veniaminova, Natalia A.; Hayes, Michael M.; Varney, Jessica M.

    2012-01-01

    Antral gastrin is the hormone known to stimulate acid secretion and proliferation of the gastric corpus epithelium. Patients with mutations in the multiple endocrine neoplasia type 1 (MEN1) locus, which encodes the protein menin, develop pituitary hyperplasia, insulinomas, and gastrinomas in the duodenum. We previously hypothesized that loss of menin leads to derepression of the gastrin gene and hypergastrinemia. Indeed, we show that menin represses JunD induction of gastrin in vitro. Therefore, we examined whether conditional deletion of Men1 (Villin-Cre and Lgr5-EGFP-IRES-CreERT2), with subsequent loss of menin from the gastrointestinal epithelium, increases gastrin expression. We found that epithelium-specific deletion of Men1 using Villin-Cre increased plasma gastrin, antral G cell numbers, and gastrin expression in the antrum, but not the duodenum. Moreover, the mice were hypochlorhydric by 12 mo of age, and gastric somatostatin mRNA levels were reduced. However, duodenal mRNA levels of the cyclin-dependent kinase inhibitor p27Kip1 were decreased, and cell proliferation determined by Ki67 staining was increased. About 11% of the menin-deficient mice developed antral tumors that were negative for gastrin; however, gastrinomas were not observed, even at 12 mo of age. No gastrinomas were observed with conditional deletion of Men1 in the Lgr5 stem cells 5 mo after Cre induction. In summary, epithelium-specific deletion of the Men1 locus resulted in hypergastrinemia due to antral G cell hyperplasia and a hyperproliferative epithelium, but no gastrinomas. This result suggests that additional mutations in gene targets other than the Men1 locus are required to produce gastrin-secreting tumors. PMID:22766853

  9. Management of Oro-antral Communication and Fistula: Various Surgical Options

    PubMed Central

    Khandelwal, Pulkit; Hajira, Neha

    2017-01-01

    Oro-antral communication and fistula can occur as a result of inadequate and improper treatment. Inadvertent communication with the maxillary sinus can occur during certain surgical procedures in the maxillary posterior region. Though, spontaneous healing may occur in defects which are smaller than 2 mm but larger communications require immediate attention and should be treated without delay, in order to avoid sinusitis and further complications leading to patient discomfort. PMID:28289607

  10. Control of ovarian primordial follicle activation

    PubMed Central

    2012-01-01

    The ovarian follicles develop initially from primordial follicles. The majority of ovarian primordial follicles are maintained quiescently as a reserve for the reproductive life span. Only a few of them are activated and develop to an advanced follicular stage. The maintenance of dormancy and activation of primordial follicles are controlled by coordinated actions of a suppressor/activator with close communications with somatic cells and intra-oocyte signaling pathways. Many growth factors and signaling pathways have been identified and the transforming growth factor-beta superfamily plays important roles in early folliculogenesis. However, the mechanism of maintaining the dormancy and survival of primordial follicles has remained unknown for decades. Recently, since the first finding that all primordial follicles are activated prematurely in mice deficient forkhead box O3a, phosphatidylinositol 3 kinase/phosphatase and tensin homolog (PTEN) signaling pathway was reported to be important in the regulation of dormancy and initial follicular activation. With these informations on early folliculogenesis, clinical application can be expected such as in vitro maturation of immature oocytes or in vitro activation of follicles by PTEN inhibitor in cryopreserved ovarian cortical tissues for fertility preservation. PMID:22563545

  11. Effects of frequency on the wave form of propagated slow waves in canine gastric antral muscle.

    PubMed Central

    Publicover, N G; Sanders, K M

    1986-01-01

    Experiments were performed to test the effects of frequency on the wave form of electrical slow waves in canine antral circular muscle. At frequencies between 3.0 and 5.6 cycles per minute antral slow waves revealed an alternating wave form pattern. At physiological frequencies antral muscle was incapable of consistently propagating mechanically productive slow waves. Two components of the slow wave were identified on the basis of propagation refractory period. At inter-slow-wave intervals of 3-14 s, the amplitude and duration of the plateau phase wave decreased, but the upstroke phase of the slow wave was minimally affected. Intervals of 2.5-4 s resulted in a normal upstroke event but abolished the plateau. At shorter intervals the upstroke phase of the slow wave was greatly reduced or abolished. The absolute propagation refractory period averaged 2.8 +/- 0.9 s (n = 7) following repolarization of a 'conditioning' slow wave. Slow waves failed to propagate within the absolute propagation refractory period. Acetylcholine decreased the interval required for the plateau phase of the slow wave to recover and permitted conduction of mechanically productive slow waves at or above physiological frequencies. The data presented suggest that gastric motility is modulated by extrinsic and intrinsic factors which regulate slow-wave frequency. PMID:3701649

  12. Isolation and Characterization of Mouse Antral Oocytes Based on Nucleolar Chromatin Organization.

    PubMed

    Monti, Manuela; Redi, Carlo Alberto

    2016-01-07

    This protocol describes a simple and quick method to isolate and characterize mouse antral GV (Germinal Vesicle) oocytes as able (SN, Surrounded Nucleolus) or unable (NSN, Not Surrounded Nucleolus) to develop to the blastocyst stage after in vitro maturation (IVM) and in vitro fertilization (IVF). It makes use of Hoeschst33342 (or any other DNA intercalating dye) able to bind to the heterochromatin of the nucleolus showing a ring in the SN oocytes or not, like in the NSN oocytes. This represents the easiest and quickest way to sort both antral oocytes that can be eventually used for IVM or IVF procedures. Briefly, the protocol consists of the following steps: hormone injection to stimulate follicular growth; isolation of the oocytes at the GV stage from the antral compartment by puncturing the ovary with a sterile needle; preparation of thin glass pipettes for mouth pipetting of the oocytes; sorting of the oocytes with Hoechst33342 prepared at a supravital concentration; IVM, IVF or any other molecular/cellular analysis. Unfortunately there are still few evidences to sort SN and NSN oocytes using less invasive techniques. If and once they will be identified, they could be potentially applied to human assisted reproductive technologies, although with several aspects that should be modified. To date, this technique has potential implications to dramatically increase IVM and IVF successful procedures in both endangered and species with economic interest.

  13. Postprandial water intake inhibits gastric antral motility with increase of cholecystokinin in humans.

    PubMed

    Kusano, Motoyasu; Minashi, Keiko; Maeda, Masaki; Shimoyama, Yasuyuki; Kuribayashi, Shikou; Higuchi, Tatsuya; Sugimoto, Sayaka; Kawamura, Osamu; Mori, Masatomo

    2005-10-01

    The effects of postprandial water intake on the gastrointestinal tract have not been systematically investigated in humans. In 8 healthy volunteers, the gastric antral pressure was measured with a strain gauge transducer, while the esophageal and lower esophageal sphincter pressures were measured with an infused catheter with a Dent sleeve. The esophageal pH at 5 cm above the lower sphincter was measured with a microglass electrode. A standard test meal (560 kcal) was eaten and 500 ml water was ingested 1 h later. The plasma cholecystokinin level was assessed at 4-min intervals. As a control, the same study was done on another day with sham water intake. At 4 min after water intake, there was a significant decrease in gastric antral motility and a significant increase in the plasma cholecystokinin level. Water intake also significantly increased the occurrence of gastroesophageal reflux. The rapid increase in cholecystokinin after water intake may be initiated by a feedback mechanism related to inflow of fatty chyme into the duodenum that inhibits gastric antral activity.

  14. Stem cells from human hair follicles: first mechanical isolation for immediate autologous clinical use in androgenetic alopecia and hair loss.

    PubMed

    Gentile, Pietro; Scioli, Maria G; Bielli, Alessandra; Orlandi, Augusto; Cervelli, Valerio

    2017-01-01

    Hair follicles are known to contain a well-characterized niche for adult stem cells: the bulge, which contains epithelial and melanocytic stem cells. Stem cells in the hair bulge, a clearly demarcated structure within the lower permanent portion of hair follicles, can generate the interfollicular epidermis, hair follicle structures, and sebaceous glands. The bulge epithelial stem cells can also reconstitute in an artificial in vivo system to a new hair follicle. In this study, we have developed a new method to isolate human adult stem cells by mechanical centrifugation of punch biopsy from human hair follicles without culture condition. Here, we used human follicle stem cells (HFSCs), to improve the hair density in 11 patients (38 to 61 years old) affected by AGA in stage 3-5 as determined by the Norwood-Hamilton classification scale. The primary outcomes were microscopic identification and counting of HFSCs. The secondary outcomes were clinical preliminary results and safety and feasibility in HFSCs-treated scalp. Each scalp tissue suspension contained about 3,728.5±664.5 cells. The percentage of hair follicle-derived mesenchymal stem cells CD44+ [from dermal papilla (DP)] was about 5%+0.7% whereas the percentage of hair follicle epithelial stem cells CD200+ (from the bulge) was about 2.6%+0.3%. In total, 23 weeks after the last treatment with HFSCs mean hair count and hair density increases over baseline values. In particular, a 29%±5% increase in hair density for the treated area and less than a 1% increase in hair density for the placebo area. We have shown that the isolated cells are capable to improve the hair density in patients affected by androgenetic alopecia (AGA). These cells appear to be located in the bulge area of human.

  15. Primordial follicle assembly was regulated by Notch signaling pathway in the mice.

    PubMed

    Chen, Chun-Lei; Fu, Xia-Fei; Wang, Lin-Qing; Wang, Jun-Jie; Ma, Hua-Gang; Cheng, Shun-Feng; Hou, Zhu-Mei; Ma, Jin-Mei; Quan, Guo-Bo; Shen, Wei; Li, Lan

    2014-03-01

    Notch signaling pathway, a highly conserved cell signaling system, exists in most multicellular organisms. The objective of this study was to examine Notch signaling pathway in germ cell cyst breakdown and primordial follicle formation. The receptor and ligand genes of Notch pathway (Notch1, Notch2, Jagged1, Jagged2 and Hes1) were extremely down-regulated after newborn mouse ovaries were cultured then exposed to DAPT or L-685,458 in vitro (P < 0.01). Since DAPT or L-685,548 inhibits Notch signaling pathway, the expression of protein LHX8 and NOBOX was significantly reduced during the formation of the primordial follicles. Down-regulated mRNA expression of specific genes including Lhx8, Figla, Sohlh2 and Nobox, were also observed. The percentages of female germ cells in germ cell cysts and primordial follicles were counted after culture of newborn ovaries for 3 days in vitro. The result showed female germ cells in cysts was remarkably up-regulated while as the oocytes in primordial follicles was significantly down-regulated (P < 0.05). In conclusion, Notch signaling pathway may regulate the formation of primordial follicle in mice.

  16. FSH (Follicle-Stimulating Hormone) Test

    MedlinePlus

    ... follicle-stimulating hormone (FSH), a hormone associated with reproduction and the development of eggs in women and ... FSH and LH with the development of secondary sexual characteristics at an unusually young age are an ...

  17. Ovarian follicle selection and granulosa cell differentiation.

    PubMed

    Johnson, A L

    2015-04-01

    The reproductive strategy for avian species that produce a sequence (or clutch) of eggs is dependent upon the maintenance of a small cohort of viable, undifferentiated (prehierarchal) follicles. It is from this cohort that a single follicle is selected on an approximate daily basis to initiate rapid growth and final differentiation before ovulation. This review describes a working model in which follicles within this prehierarchal cohort are maintained in an undifferentiated state by inhibitory cell signaling until the time of selection. Ultimately, follicle selection represents a process in which a single undifferentiated follicle per day is predicted to escape such inhibitory mechanisms to begin rapid growth and final maturation before ovulation. Several processes initiated within the granulosa cell layer at selection are dependent upon G protein-coupled receptors signaling via cyclic adenosine monophosphate (cAMP), and several critical processes are described herein. Finally, reference is made to several practical outcomes that can result from understanding the process of selection, including applications within the poultry industry. Proximal factors and processes that mediate follicle selection can either extend or decrease the length of the laying sequence, and thus directly influence overall egg production. In particular, any aberration that results in the selection of more than one follicle per day will result in decreased egg production. More generally, in wild birds these processes are modified by prevailing environmental conditions and by social interactions to influence clutch size. The elucidation of cellular processes that regulate follicle selection can assist in the development of assisted reproductive technologies for application in threatened and endangered avian species. © 2014 Poultry Science Association Inc.

  18. Automated follicle analysis in ovarian ultrasound

    NASA Astrophysics Data System (ADS)

    Krivanek, Anthony; Liang, Weidong; Sarty, Gordon E.; Pierson, Roger A.; Sonka, Milan

    1998-06-01

    For women undergoing assisted reproductive therapy, ovarian ultrasound has become an invaluable tool for monitoring the growth and assessing the physiological status of individual follicles. Measurements of the size and shape of follicles are the primary means of evaluation by physicians. Currently, follicle wall segmentation is achieved by manual tracing which is time consuming and susceptible to inter- operator variation. We are introducing a completely automated method of follicle wall isolation which provides faster, more consistent analysis. Our automated method is a 4-step process which employs watershed segmentation and a knowledge-based graph search algorithm which utilizes a priori information about follicle structure for inner and outer wall detection. The automated technique was tested on 36 ultrasonographic images of woman's ovaries. Five images from this set were omitted due to poor image quality. Validation of the remaining 31 ultrasound images against manually traced borders has shown an average rms error of 0.61 +/- 0.40 mm for inner border and 0.61 +/- 0.31 mm for outer border detection. Quantitative comparison of the computer-defined borders and the user-defined borders advocates the accuracy of our automated method of follicle analysis.

  19. The dynamics of the primordial follicle reserve.

    PubMed

    Kerr, Jeffrey B; Myers, Michelle; Anderson, Richard A

    2013-12-01

    The female germline comprises a reserve population of primordial (non-growing) follicles containing diplotene oocytes arrested in the first meiotic prophase. By convention, the reserve is established when all individual oocytes are enclosed by granulosa cells. This commonly occurs prior to or around birth, according to species. Histologically, the 'reserve' is the number of primordial follicles in the ovary at any given age and is ultimately depleted by degeneration and progression through folliculogenesis until exhausted. How and when the reserve reaches its peak number of follicles is determined by ovarian morphogenesis and germ cell dynamics involving i) oogonial proliferation and entry into meiosis producing an oversupply of oocytes and ii) large-scale germ cell death resulting in markedly reduced numbers surviving as the primordial follicle reserve. Our understanding of the processes maintaining the reserve comes primarily from genetically engineered mouse models, experimental activation or destruction of oocytes, and quantitative histological analysis. As the source of ovulated oocytes in postnatal life, the primordial follicle reserve requires regulation of i) its survival or maintenance, ii) suppression of development (dormancy), and iii) activation for growth and entry into folliculogenesis. The mechanisms influencing these alternate and complex inter-related phenomena remain to be fully elucidated. Drawing upon direct and indirect evidence, we discuss the controversial concept of postnatal oogenesis. This posits a rare population of oogonial stem cells that contribute new oocytes to partially compensate for the age-related decline in the primordial follicle reserve.

  20. Proteomic Analysis of Hair Follicles

    NASA Astrophysics Data System (ADS)

    Ishioka, Noriaki; Terada, Masahiro; Yamada, Shin; Seki, Masaya; Takahashi, Rika; Majima, Hideyuki J.; Higashibata, Akira; Mukai, Chiaki

    2013-02-01

    Hair root cells actively divide in a hair follicle, and they sensitively reflect physical conditions. By analyzing the human hair, we can know stress levels on the human body and metabolic conditions caused by microgravity environment and cosmic radiation. The Japan Aerospace Exploration Agency (JAXA) has initiated a human research study to investigate the effects of long-term space flight on gene expression and mineral metabolism by analyzing hair samples of astronauts who stayed in the International Space Station (ISS) for 6 months. During long-term flights, the physiological effects on astronauts include muscle atrophy and bone calcium loss. Furthermore, radiation and psychological effects are important issue to consider. Therefore, an understanding of the effects of the space environment is important for developing countermeasures against the effects experienced by astronauts. In this experiment, we identify functionally important target proteins that integrate transcriptome, mineral metabolism and proteome profiles from human hair. To compare the protein expression data with the gene expression data from hair roots, we developed the protein processing method. We extracted the protein from five strands of hair using ISOGEN reagents. Then, these extracted proteins were analyzed by LC-MS/MS. These collected profiles will give us useful physiological information to examine the effect of space flight.

  1. Follicle characteristics and follicle developmental related Wnt6 polymorphism in Chinese indigenous Wanxi-white goose.

    PubMed

    Chen, Xingyong; Bai, Haichen; Li, Li; Zhang, Wei; Jiang, Runshen; Geng, Zhaoyu

    2012-11-01

    In birds, downy feather quantity mainly affected by the follicles. Wnt6, a secreted cysteine-rich protein, plays a key role in follicular development as an intercellular signaling molecule. The present study was to investigate the follicle development and Wnt6 polymorphism in Wanxi-white geese, a Chinese indigenous breed. In total, 300 fertilized eggs were hatched. At embryonic stage and on early birth goslings, the diameter and density of follicles from different sites were examined after sectioning and staining. The results showed that the diameter of primary feather follicles in thorax, venter, dorsum and flank had no difference at embryonic stage. In contrast, after birth, thorax and ventral feather follicles had greater diameter than those on dorsum and flank. Similarly, the primary feather follicle density was higher in thorax and venter than in dorsum and flank at embryonic stage. The secondary feather follicle diameter in flank was greater than that in other sites examined. The secondary follicle showed lush growth in E27 with thickest in ventral and thorax. Overall, follicle formed consistently in dorsal and flank, and follicle in thorax and ventral formed in another consistent way. The polymorphism study showed 2 single nucleotide polymorphisms of Wnt6 and 3 genotypes identified. Sequencing revealed two nucleotide transitions, T451C and A466G, which were synonymous mutations causing codons for aspartate and lysine to change from GAU to GAC and from AAA to AAG, respectively. This information about follicle development and Wnt6 polymorphisms would provide potential utilization in marker-assisted selection program for down feather selection.

  2. Selection of the dominant follicle in cattle and horses.

    PubMed

    Ginther, O J

    2000-07-02

    The nature of selection of the dominant follicle is reviewed by comparing research results between cattle and horses. In both species, emergence of a follicular wave is stimulated by an FSH surge. The surge reaches a peak by the time the follicles attain 4 mm in diameter in cattle and 13 mm in mares. In cattle, all of the growing follicles >/=5 mm contribute to the decline in FSH concentrations. However, the declining FSH concentrations are still needed by the growing follicles. Several days after the peak of the FSH surge and emergence of the wave, the two largest follicles reach means of 8.5 and 7.7 mm in cattle and 22 and 19 mm in horses. At this approximate time, the follicles begin to undergo deviation in follicle diameters, which is characterized by continued growth of the largest follicle to become the dominant follicle and reduced or terminated growth of the remaining follicles to become subordinate follicles. In both species, on average, the future dominant follicle emerges before the future largest subordinate follicle, and the two follicles grow in parallel until deviation. The difference in diameter between the two largest follicles at the beginning of deviation is equivalent in growth to approximately 8 h in cattle and 24 h in mares. Apparently, this is adequate time for the largest follicle to establish the deviation process before the second-largest follicle reaches a similar diameter. During this time, the largest follicle plays the primary role in further suppressing circulating FSH concentrations to below the requirements of the smaller follicles, which causes their regression. The follicle-produced FSH suppressants appear to be estradiol and inhibin. In addition to enhancing its FSH-suppressing ability, the largest follicle also develops the ability to utilize the reduced concentrations of FSH for its continued growth. It is therefore postulated that the essence of selection of a dominant follicle in these two species is a close two

  3. Curcumin Blocks Naproxen-Induced Gastric Antral Ulcerations through Inhibition of Lipid Peroxidation and Activation of Enzymatic Scavengers in Rats.

    PubMed

    Kim, Jeong-Hwan; Jin, Soojung; Kwon, Hyun Ju; Kim, Byung Woo

    2016-08-28

    Curcumin is a polyphenol derived from the plant Curcuma longa, which is used for the treatment of diseases associated with oxidative stress and inflammation. The present study was undertaken to determine the protective effect of curcumin against naproxen-induced gastric antral ulcerations in rats. Different doses (10, 50, and 100 mg/kg) of curcumin or vehicle (curcumin, 0 mg/kg) were pretreated for 3 days by oral gavage, and then gastric mucosal lesions were caused by 80 mg/kg naproxen applied for 3 days. Curcumin significantly inhibited the naproxen-induced gastric antral ulcer area and lipid peroxidation in a dose-dependent manner. In addition, curcumin markedly increased activities of radical scavenging enzymes, such as superoxide dismutase (SOD), catalase, and glutathione peroxidase in a dose-dependent manner. Specifically, 100 mg/kg curcumin completely protected the gastric mucosa against the loss in the enzyme, resulting in a drastic increase of activities of radical scavenging enzymes up to more than the level of untreated normal rats. Histological examination obviously showed that curcumin prevents naproxen-induced gastric antral ulceration as a result of direct protection of the gastric mucosa. These results suggest that curcumin blocks naproxen-induced gastric antral ulcerations through prevention of lipid peroxidation and activation of radical scavenging enzymes, and it may offer a potential remedy of gastric antral ulcerations.

  4. Depletion of CD200+ Hair Follicle Stem Cells in Human Prematurely Gray Hair Follicles

    PubMed Central

    Mohanty, Sujata; Kumar, Anil; Dhawan, Jyoti; Sharma, Vinod K; Gupta, Somesh

    2013-01-01

    Introduction: Melanocyte stem cells (MelSCs) are known to be depleted in gray hair follicles. Hair follicle stem cells (HFSCs) are important for maintenance of stemness of MelSCs. Methods: We compared the proportion of CD200+ (Cluster of Differentiation 200 positive) stem cells in the outer root sheath cell suspension of gray and pigmented hair follicles of three patients with the premature graying of hair. In addition, explants culture for HFSCs was also carried out from gray and pigmented hair follicles. Cultured HFSCs were also differentiated into melanocytes. Results: The mean ± SD CD200+ HFSCs population were 9.4 ± 1.4% and 3.5 ± 0.5% for pigmented and gray hair follicles, respectively (P = 0.002). In explants culture, the growth of HFSCs from the gray hair follicle stopped at around day 20–22, whereas the growth of the cells from the pigmented follicle continued. Conclusion: CD200+ HFSCs are depleted in prematurely gray hair in the humans. CD200+ hair follicle stem cell yield is poorer in gray hair explant culture than pigmented hair explant culture. PMID:24023430

  5. Follicle size and volume is less indicative of development of a persistent follicle in beef heifers

    USDA-ARS?s Scientific Manuscript database

    We have established that feeding Melengestrol Acetate (MGA) for 14 d 0.5mg/h/d (< 1 ng/ml of P4) develops persistent follicles with increased follicle size, follicular fluid volume, theca cell weight, and follicular fluid androstenedione concentration in mature cows. Therefore, we hypothesized that...

  6. Effects of gremlin-2 on the transition of primordial follicles during early folliculogenesis in the human ovary.

    PubMed

    Ikeda, Yuki; Hasegawa, Akiko; Tsubamoto, Hiroshi; Wakimoto, Yu; Kumamoto, Kanako; Shibahara, Hiroaki

    2016-08-01

    To investigate the localization and function of gremlin-2 during human ovarian folliculogenesis. Ovarian tissue from a gynecologic cancer patient was cultured in the presence or absence of gremlin-2 and then analyzed histologically. Growing follicles were counted by the microscopic observations of ovarian histological sections. Immunocytochemical staining was carried out to detect the expression of bone morphogenetic protein (BMP) 4 and phosphorylated Smad 1/5/8 (p-Smad 1/5/8). Gremlin-2 was detected in human primordial, primary, and early growing follicles before culture. By day 4 of culture, the follicle growth rate in the presence of gremlin-2 (13.7%; 24/175) was significantly lower than that of the control (54.8%; 92/175; p<0.01). BMP4 expression was similar in the presence and absence of gremlin-2, whereas the p-Smad 1/5/8 signal was noticeably stronger in the absence of gremlin-2 in primordial and early-stage growing follicles. Gremlin-2 maintains the follicle store as primordial follicles by suppressing Smad 1/5/8 signaling in the human ovary. The data presented here provide potential insight into reproductive medicine for cases of intractable infertility, such as premature ovarian insufficiency and cancer survivors. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  7. Growth differentiation factor-9 mediates follicle-stimulating hormone-thyroid hormone interaction in the regulation of rat preantral follicular development.

    PubMed

    Kobayashi, Noriko; Orisaka, Makoto; Cao, Mingju; Kotsuji, Fumikazu; Leader, Arthur; Sakuragi, Noriaki; Tsang, Benjamin K

    2009-12-01

    FSH regulates follicular growth in a stage-development fashion. Although preantral follicle stage is gonadotropin responsive, FSH is not required for preantral follicular growth. With the antrum, the follicles continue growing under the influence of FSH and become gonadotropin dependent. Although thyroid hormone is important for normal female reproductive function, its role and interaction with FSH in the regulation of preantral ovarian follicular growth is yet to be defined. In the present study, we have examined the action and interaction of FSH and T(3) in the regulation of the growth of preantral follicles, especially in their transition from preantral to early antral stage, using an established follicle culture system and evaluated the involvement of growth differentiation factor-9 (GDF-9) in this process in vitro. We have demonstrated that although T(3) alone had no effect on follicular development, it markedly enhanced FSH-induced preantral follicular growth. Although FSH alone significantly down-regulated FSH receptor (FSHR) mRNA abundance in the preantral follicles and T(3) alone was ineffective, expression of the message was significantly increased in the presence of both hormones. In addition, intra-oocyte injection of GDF-9 antisense oligonucleotides (GDF-9 morpholino) induced follicular cell apoptosis and suppressed follicular growth induced by FSH and T(3). These responses were attenuated by exogenous GDF-9. Our findings support the concept that thyroid hormone regulates ovarian follicular development through its direct action on the ovary and that promotes FSH-induced preantral follicular growth through up-regulation of FSHR, a mechanism dependent on the expression and action of oocyte-derived GDF-9.

  8. Scintigraphic measurement of gastric emptying and ultrasonographic assessment of antral area: relation to appetite.

    PubMed Central

    Hveem, K; Jones, K L; Chatterton, B E; Horowitz, M

    1996-01-01

    BACKGROUND: Ultrasound measurement of gastric emptying has potential advantages over scintigraphy, but there is little information about its accuracy. AIMS: The relation between ultrasonographic measurements of antral area and (a) scintigraphic measurements of gastric emptying and intragastric distribution of liquids (b) postprandial satiation, were evaluated. SUBJECTS: Seven normal volunteers were studied. METHOD: Each subject drank 75 g dextrose dissolved in 350 ml of water (300 kcal) or beef soup (20 kcal), both labelled with technetium-99m sulphur colloid on separate days and had measurement of gastric emptying by scintigraphy and ultrasound. RESULTS: Scintigraphic and ultrasound 50% emptying times (T50s) were comparable and longer (p < 0.001) for dextrose than soup mean (SEM) (dextrose 107 (16) min v 108 (18) min, soup 24 (4) min v 23 (5) min). There were close correlations between scintigraphic and ultrasound T50s (dextrose r = 0.94, p < 0.005, soup r = 0.97, p < 0.001) and between the time at which the distal stomach content decreased from its maximum value by 50% (measured scintigraphically) and the ultrasound T50 (dextrose r = 0.95, p < 0.005, soup r = 0.99, p < 0.0001). In contrast, there was no significant relation between the distal stomach content when expressed as a percentage of the maximum content in the total stomach and the ultrasound T50. After dextrose, fullness was related (r = 0.92, p < 0.01) to the postprandial increase in antral area measured by ultrasound. CONCLUSIONS: Ultrasound measurements of gastric emptying are: (a) of comparable sensitivity to scintigraphy in quantifying emptying of both low and high nutrient liquids (b) correlate with postprandial satiation, suggesting that the latter may be mediated by antral distension. PMID:8984016

  9. A quantitative cytochemical study of glucose-6-phosphate dehydrogenase and delta 5-3 beta-hydroxysteroid dehydrogenase activity in the membrana granulosa of the ovulable type of follicle of the rat.

    PubMed

    Zoller, L C; Weisz, J

    1979-08-01

    During the last four days of follicular development prior to ovulation, the activities of delta 5-3 beta-hydroxysteroid dehydrogenase (3 beta OHD) and glucose-6-phosphate dehydrogenase (G-6-PD) were quantified in cryostat sections of the rat ovary. The product of the enzyme reactions were measured using a scanning and integrating microdensitometer. The enzyme activity was measured in the peripheral region, the antral region and the cumulus of the membrana granulosa (MG) of these follicles on the morning of each of the four days of the estrous cycle. G-6-PD activity was measured in the presence and absence of an intermediate hydrogen acceptor, phenazine methosulphate, to provide a measure of the quantity of Type I and Type II Hydrogen (H) generated: Type I H is considered to be related to hydroxylating reactions such as those of steroids and Type II H to other general biosynthetic activities of cells. In all three regions of the MG of follicles of the ovulable type, 3 beta OHD activity was lowest in estrus and diestrus-1, increased on diestrus-2 and peaked in proestrus. In estrus and diestrus-1, the level of 3 beta OHD activity in the three regions was comparable. However, by diestrus-2, and even more conspicuously in proestrus, enzyme activity was significantly greater in the peripheral region than in the antral region or in the cumulus. During the same period, the level of enzyme activity remained comparable in the last two regions. Throughout the estrous cycle, both Type I and Type II H generation from G-6-PD was greatest in the peripheral region, less in the antral region and least in the cumulus. In the eripheral region, Type I H generation increased progressively after diestrus-1, to reach a maximum in prestrus. In the antral region, Type I H generation increased between diestrus-1 and diestrus-2 and then remained unchanged through proestrus. In the cumulus, Type I H generation remained at levels seen in estrus throughout the remainder of the cycle. Generation

  10. Sodium pertechnetate Tc 99m antral scan in the diagnosis of retained gastric antrum

    SciTech Connect

    Lee, C.H.; P'eng, F.K.; Yeh, P.H.

    1984-03-01

    Retained gastric antrum (RGA) is a major factor in recurrent peptic ulcer. We studied 121 patients with proven anastomotic ulcers following subtotal gastrectomy and Billroth II reconstruction with sodium pertechnetate Tc 99m to determine the presence of RGA. Of the patients, 59 required surgery, 22 had RGA, and 16 had a positive scan for RGA. This noninvasive examination has 100% specificity. If RGA is identified before operation, minimal time is wasted in exploration of the abdomen, which is especially important in dealing with emergency cases. When an antral scan is negative for RGA, the surgeon is still advised to search for this condition.

  11. Maxillary Reconstruction for Sinus Lift Complications With Oro-Antral Fistula: The Le Fort I Approach.

    PubMed

    Pigache, Pénélope; Anavekar, Namrata; Raoul, Gwénaël; Ferri, Joël

    2016-03-01

    Although sinus lift procedures are reliable, some complications can lead to serious maxillary sequelae, including the development of oro-antral fistula (OAF). Maxillary reconstruction in such patients presents a challenge owing to sinus floor alterations, graft remnants, chronic infection, and morbidity from the original sinus lift approach. The current study describes our technique of maxillary reconstruction using a Le Fort 1 approach following major sinus lift complications with associated residual OAF. This technique provides excellent access for sinus curettage, OAF closure, and osseous reconstruction. It allowed a successful rehabilitation in our patients, with no implant loss and good functional and esthetic results.

  12. Short-term exposure of human ovarian follicles to cyclophosphamide metabolites seems to promote follicular activation in vitro.

    PubMed

    Lande, Yechezkel; Fisch, Benjamin; Tsur, Abraham; Farhi, Jacob; Prag-Rosenberg, Roni; Ben-Haroush, Avi; Kessler-Icekson, Gania; Zahalka, Muayad A; Ludeman, Susan M; Abir, Ronit

    2017-01-01

    How chemotherapy affects dormant ovarian primordial follicles is unclear. The 'burnout' theory, studied only in mice, suggests cyclophosphamide enhances primordial follicle activation. Using 4-hydroperoxycyclophosphamide (4hc) and phosphoramide mustard (PM), this study assessed how the active cyclophosphamide metabolites 4-hydroxycyclophosphamide (4-OHC) and PM, affect human primordial follicles. Frozen-thawed human ovarian samples were sliced and cultured with basic culture medium (cultured controls) or with 4hc/PM (3 µmol/l/10 µmol/l) (treated samples) for 24-48 h. Follicular counts and classification, Ki67 and anti-Müllerian hormone (AMH) immunohistochemistry and an apoptosis assay were used for evaluation, and 17β-oestradiol and AMH were measured in spent media samples. Generally, there was primordial follicle decrease and elevated developing follicle rates in treated samples compared with cultured (P = 0.04 to P < 0.0005) and uncultured controls (P < 0.05 to P < 0.0001). No traces of apoptosis were found. There were almost twicethe levels of AMH and 17β-oestradiol in treated compared with untreated samples (AMH with 4hc 3 µmol/l; P = 0.04). All follicles stained positively for AMHincluded treated samples. Ki67 positive staining was noted in all samples. Cyclophosphamide metabolites seem to enhance human primordial follicle activation to developing follicles, in vitro. Study findings support the 'burnout' theory as the mechanism of chemotherapy-induced ovarian toxicity. Copyright © 2016 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

  13. Biology of the hair follicle: the basics.

    PubMed

    Krause, Karoline; Foitzik, Kerstin

    2006-03-01

    The mammalian hair follicle represents a unique, highly regenerative neuroectodermal-mesodermal interaction system that contains numerous stem cells. It is the only organ in the mammalian organism that undergoes life-long cycles of rapid growth (anagen), regression (catagen), and resting periods (telogen). These transformations are controlled by changes in the local signaling milieu, based on changes in expression/activity of a constantly growing number of cytokines, hormones, neurotransmitters, and their cognate receptors as well as of transcription factors and enzymes that have become recognized as key mediators of hair follicle cycling. Transplantation experiments have shown that the driving force of cycling, the "hair cycle clock," is located in the hair follicle itself. However, the exact underlying molecular mechanisms that drive this oscillator system remain unclear. These controls of hair follicle cycling are of great clinical interest because hair loss or unwanted hair growth largely reflect undesired changes in hair follicle cycling. To develop therapeutic agents for the management of these hair cycle abnormalities, it is critical to decipher and pharmacologically target the key molecular controls that underlie the enigmatic "hair cycle clock."

  14. Follicle Microstructure and Innervation Vary between Pinniped Micro- and Macrovibrissae.

    PubMed

    Mattson, Erin E; Marshall, Christopher D

    2016-01-01

    Histological data from terrestrial, semiaquatic, and fully aquatic mammal vibrissa (whisker) studies indicate that follicle microstructure and innervation vary across the mystacial vibrissal array (i.e. medial microvibrissae to lateral macrovibrissae). However, comparative data are lacking, and current histological studies on pinniped vibrissae only focus on the largest ventrolateral vibrissae. Consequently, we investigated the microstructure, medial-to-lateral innervation, and morphometric trends in harp seal (Pagophilus groenlandicus) vibrissal follicle-sinus complexes (F-SCs). The F-SCs were sectioned either longitudinally or in cross-section and stained with a modified Masson's trichrome stain (microstructure) or Bodian's silver stain (innervation). All F-SCs exhibited a tripartite blood organization system. The dermal capsule thickness, the distribution of major branches of the deep vibrissal nerve, and the hair shaft design were more symmetrical in medial F-SCs, but these features became more asymmetrical as the F-SCs became more lateral. Overall, the mean axon count was 1,221 ± 422.3 axons/F-SC and mean axon counts by column ranged from 550 ± 97.4 axons/F-SC (medially, column 11) to 1,632 ± 173.2 axons/F-SC (laterally, column 2). These values indicate a total of 117,216 axons innervating the entire mystacial vibrissal array. The mean axon count of lateral F-SCs was 1,533 ± 192.9 axons/ F-SC, which is similar to values reported in the literature for other pinniped F-SCs. Our data suggest that conventional studies that only examine the largest ventrolateral vibrissae may overestimate the total innervation by ∼20%. However, our study also accounts for variation in quantification methods and shows that conventional analyses likely only overestimate innervation by ∼10%. The relationship between axon count and cross-sectional F-SC surface area was nonlinear, and axon densities were consistent across the snout. Our data indicate that harp seals exhibit

  15. Transglutaminase from Hair Follicle of Guinea Pig

    PubMed Central

    Chung, S. I.; Folk, J. E.

    1972-01-01

    Two transglutaminases are found in homogenates of the inner root sheaths of guinea pig hair-follicles. One is indistinguishable from the well-characterized liver transglutaminase [J. Biol. Chem., 246, 1093 (1971)]. The other, which is present in far greater quantity, has not been detected in other organs or tissues. Gel filtration and polyacrylamide gel electrophoresis studies indicate that the native hair-follicle enzyme, of molecular weight 54,000, is composed of two subunits of identical molecular weight. Specificity studies suggest that the intermolecular cross-linking of fibrin and fibrinogen that is catalyzed by this enzyme is a result of the formation of ε(γ-glutamyl)lysine bonds. The probable participation of hair-follicle transglutaminase in the formation of these cross-links in the proteins of hair is discussed. Images PMID:4501114

  16. Administration of follicle-stimulating hormone induces autophagy via upregulation of HIF-1α in mouse granulosa cells

    PubMed Central

    Zhou, Jilong; Yao, Wang; Li, Chengyu; Wu, Wangjun; Li, Qifa; Liu, Honglin

    2017-01-01

    Recent studies reported the important role of autophagy in follicular development. However, the underlying molecular mechanisms remain elusive. In this study, we investigated the effect of follicle-stimulating hormone (FSH) on mouse granulosa cells (MGCs). Results indicated that autophagy was induced by FSH, which is known to be the dominant hormone regulating follicular development and granulosa cell (GC) proliferation. The activation of mammalian target of rapamycin (mTOR), a master regulator of autophagy, was inhibited during the process of MGC autophagy. Moreover, MHY1485 (an agonist of mTOR) significantly suppressed autophagy signaling by activating mTOR. The expression of hypoxia-inducible factor 1-alpha (HIF-1α) was increased after FSH treatment. Blocking hypoxia-inducible factor 1-alpha attenuated autophagy signaling. In vitro, CoCl2-induced hypoxia enhanced cell autophagy and affected the expression of beclin1 and BCL2/adenovirus E1B interacting protein 3 (Bnip3) in the presence of FSH. Knockdown of beclin1 and Bnip3 suppressed autophagy signaling in MGCs. Furthermore, our in vivo study demonstrated that the FSH-induced increase in weight was significantly reduced after effectively inhibiting autophagy with chloroquine, which was correlated with incomplete mitophagy process through the PINK1-Parkin pathway, delayed cell cycle, and reduced cell proliferation rate. In addition, chloroquine treatment decreased inhibin alpha subunit, but enhanced the expression of 3 beta-hydroxysteroid dehydrogenase. Blocking autophagy resulted in a significantly lower percentage of antral and preovulatory follicles after FSH stimulation. In conclusion, our results indicate that FSH induces autophagy signaling in MGCs via HIF-1α. In addition, our results provide evidence that autophagy induced by FSH is related to follicle development and atresia. PMID:28817115

  17. The effects of microwave emitted by cellular phones on ovarian follicles in rats.

    PubMed

    Gul, Abdulaziz; Celebi, Hacer; Uğraş, Serdar

    2009-11-01

    The aim of this study was to investigate whether there were any toxic effects of microwaves of cellular phones on ovaries in rats. In this study, 82 female pups of rats, aged 21 days (43 in the study group and 39 in the control group) were used. Pregnant rats in the study group were exposed to mobile phones that were placed beneath the polypropylene cages during the whole period of pregnancy. The cage was free from all kinds of materials, which could affect electromagnetic fields. A mobile phone in a standby position for 11 h and 45 min was turned on to speech position for 15 min every 12 h and the battery was charged continuously. On the 21st day after the delivery, the female rat pups were killed and the right ovaries were removed. The volumes of the ovaries were measured and the number of follicles in every tenth section was counted. The analysis revealed that in the study group, the number of follicles was lower than that in the control group. The decreased number of follicles in pups exposed to mobile phone microwaves suggest that intrauterine exposure has toxic effects on ovaries. We suggest that the microwaves of mobile phones might decrease the number of follicles in rats by several known and, no doubt, countless unknown mechanisms.

  18. Rescue from dominant follicle atresia by follicle-stimulating hormone in mice.

    PubMed

    Zhou, X L; Teng, Y; Cao, R; Fu, H; Xiong, K; Sun, W X; Zhu, C C; Huang, X J; Xiao, P; Liu, H L

    2013-08-12

    We investigated the effects of follicle-stimulating hormone (FSH) on atresia of the dominant follicle and changes in relevant apoptosis genes in granulosa cells of dominant follicles regulated by FSH in vivo. Four-week-old mice were administered FSH by intraperitoneal injection to induce follicular maturation. Granulosa cells of dominant follicles were collected at 48, 72, and 96 h after the first FSH injection. Phosphate-buffered saline was injected as a control. The mRNA levels of relevant granulosa cell apoptosis genes were examined by real-time quantitative polymerase chain reaction, and apoptosis of granulosa cells in dominant ovarian follicles was determined by the terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay. Apoptosis in granulosa cells of dominant follicles was almost TUNEL-negative at 48, 72-66, 72, and 96-90 h after the first FSH injection, but granulosa cell apoptosis in dominant follicles was clearly detected at 96, 102, and 102-96 h by TUNEL. The BIM, caspase-3, and caspase-9 mRNA expression levels were significantly lower after FSH treatment at 72-66 and 96-90 h, compared with that at 72 and 96 h (P < 0.05). Caspase-8 and FasL mRNA expressions did not respond to FSH. FSH rescued granulosa cells from apoptosis when the relevant apoptosis genes were upregulated in early atretic follicles. FSH did not rescue granulosa cells from apoptosis if the DNA was cut into fragments by endonucleases. Thus, the rescue by FSH of granulosa cells from apoptosis and dominant follicle atresia may be accomplished by inhibition of apoptosis in mitochondria.

  19. Occurrence and characteristics of residual follicles formed after transvaginal ultrasound-guided follicle aspiration in cattle.

    PubMed

    Viana, J H M; Dorea, M D; Siqueira, L G B; Arashiro, E K N; Camargo, L S A; Fernandes, C A C; Palhão, M P

    2013-01-15

    Ultrasound-guided transvaginal follicle aspiration is used to recover cumulus-oocyte complexes (for IVF) and to synchronize follicular wave emergence (ablation of dominant follicle). Although aspirated follicles are generally supposed to undergo immediate atresia, there are indications that they may remain active. The objective was to evaluate the occurrence and characteristics of residual follicles (RF) after transvaginal follicle aspiration in cattle. Ovarian follicular wave emergence was synchronized in Holstein cows (N = 13) in the presence (groups 1 and 3) or absence (groups 2 and 4) of norgestomet implants. The largest follicle was aspirated at a diameter of 8 mm (groups 1 and 2) or 12 mm (groups 3 and 4). Ovarian follicles were visualized (transrectal ultrasonography) every 12 h after wave emergence. Follicular fluid samples were collected from the largest follicle and from the ensuing RF and concentrations of estradiol and progesterone were determined. After aspiration, 73.2% (52/71) of the follicles refilled with fluid, and a new antrum was detected 12 to 24 h later. Norgestomet did not affect (P > 0.05) RF occurrence or diameter, but in RF from group 4, concentrations of estradiol decreased (-530.7 ± 133.9 ng/mL; P < 0.01) whereas progesterone increased (+429.6 ± 171.7 ng/mL; P < 0.05) relative to preaspiration. In RF, there were three steroidogenesis patterns: (1) high estradiol concentration and high estradiol:progesterone ratio (estradiol-active RF); (2) low estradiol, but high progesterone concentrations (luteinized RF); and (3) low estradiol and low progesterone concentrations (inactive RF). Estradiol-active RF were more likely (P < 0.05) from follicles with high estradiol concentrations (regardless of diameter). In conclusion, fluid-filled structures (RF) with variable steroid production patterns are frequently formed after ultrasound-guided follicle aspiration. The occurrence and features of these RF depended on the diameter and status of these

  20. mRNA expression profile of the TNF-α system in LH-induced bovine preovulatory follicles and effects of TNF-α on gene expression, ultrastructure and expansion of cumulus-oocyte complexes cultured in vitro.

    PubMed

    Silva, A W B; Bezerra, F T G; Glanzner, W G; Dos Santos, J T; Dau, A M P; Rovani, M T; Ilha, G F; Costa, J J N; Cunha, E V; Donato, M A M; Peixoto, C A; Gonçalves, P B D; Bordignon, V; Silva, J R V

    2017-03-01

    This study evaluated (1) the effects of in vivo GnRH treatment on mRNA expression of TNF-α system (TNF-α, TNFR1 and TNFR2) in granulosa cells of bovine preovulatory follicles, (2) the in vitro influence of gonadotropins on mRNA expression of TNF-α system in cultured cumulus cells, (3) the protein expression of the TNF-α system in late antral follicles and, (4) the influence of TNF-α on cumulus cells expansion, ultrastructure and on expression of HAS2, CASP3 and CASP6 in follicular cells cultured for 24 h. An increased expression of TNF-α and TNFR1 was observed after 3, 6 and 12 h of GnRH treatment when compared to 0 and 24h. Higher TNFR2 mRNA levels were observed 3, 6 and 12 h after GnRH, when compared to 0 and 24 h. Proteins of TNF-α system were also expressed in late antral follicles. In vitro, TNF-α did not affect cumulus cells expansion, but reduced the HAS2, CASP3 and CASP6 mRNA levels in cumulus cells after 12 h. After 24 h of culture, TNF-α increased the mRNA levels for CASP6 in mural granulosa cells, while the TNF-α, TNFR1 and TNFR2 mRNA levels were increased in cumulus-oocyte complexes (COCs) cultured for 12 h with gonadotropins, but not after 24 h. Ultrastructural analysis confirmed the integrity of COCs cultured in presence of TNF-α. In conclusion, TNF-α system members are present in bovine antral follicles and expression of TNF-α is influenced by gonadotropins in vivo and in vitro. In vitro, TNF-α maintained cumulus cells ultrastructure during COC culture.

  1. Aging of the Hair Follicle Pigmentation System

    PubMed Central

    Tobin, Desmond J

    2009-01-01

    Skin and hair phenotypes are powerful cues in human communication. They impart much information, not least about our racial, ethnic, health, gender and age status. In the case of the latter parameter, we experience significant change in pigmentation in our journey from birth to puberty and through to young adulthood, middle age and beyond. The hair follicle pigmentary unit is perhaps one of our most visible, accessible and potent aging sensors, with marked dilution of pigment intensity occurring long before even subtle changes are seen in the epidermis. This dichotomy is of interest as both skin compartments contain melanocyte subpopulations of similar embryologic (i.e., neural crest) origin. Research groups are actively pursuing the study of the differential aging of melanocytes in the hair bulb versus the epidermis and in particular are examining whether this is in part linked to the stringent coupling of follicular melanocytes to the hair growth cycle. Whether some follicular melanocyte subpopulations are affected, like epidermal melanocytes, by UV irradiation is not yet clear. A particular target of research into hair graying or canities is the nature of the melanocyte stem compartment and whether this is depleted due to reactive oxygen species-associated damage, coupled with an impaired antioxidant status, and a failure of melanocyte stem cell renewal. Over the last few years, we and others have developed advanced in vitro models and assay systems for isolated hair follicle melanocytes and for intact anagen hair follicle organ culture which may provide research tools to elucidate the regulatory mechanisms of hair follicle pigmentation. Long term, it may be feasible to develop strategies to modulate some of these aging-associated changes in the hair follicle that impinge particularly on the melanocyte populations. PMID:20927229

  2. The developmental biology of feather follicles

    PubMed Central

    Yu, Mingke; Yue, Zhicao; Wu, Ping; Wu, Da-Yu; Mayer, Julie-Ann; Medina, Marcus; Widelitz, Randall B.; Jiang, Ting-Xin; Chuong, Cheng-Ming

    2015-01-01

    The feather is a complex epidermal organ with hierarchical branches and represents a multi-layered topological transformation of keratinocyte sheets. Feathers are made in feather follicles. The basics of feather morphogenesis were previously described (Lucas and Stettenheim, 1972). Here we review new molecular and cellular data. After feather buds form (Jiang et al., 2004), they invaginate into the dermis to form feather follicles. Above the dermal papilla is the proliferating epidermal collar. Distal to it is the ramogenic zone where the epidermal cylinder starts to differentiate into barb ridges or rachidial ridge. These neoptile feathers tend to be downy and radially symmetrical. They are replaced by teleoptile feathers which tend to be bilateral symmetrical and more diverse in shapes. We have recently developed a “transgenic feather” protocol that allows molecular analyses: BMPs enhance the size of the rachis, Noggin increases branching, while anti-SHH causes webbed branches. Different feather types formed during evolution (Wu et al., 2004). Pigment patterns along the body axis or intra-feather add more colorful distinctions. These patterns help facilitate the analysis of melanocyte behavior. Feather follicles have to be connected with muscles and nerve fibers, so they can be integrated into the physiology of the whole organism. Feathers, similarly to hairs, have the extraordinary ability to go through molting cycles and regenerate. Some work has been done and feather follicles might serve as a model for stem cell research. Feather phenotypes can be modulated by sex hormones and can help elucidate mechanisms of sex hormone-dependent growth control. Thus, the developmental biology of feather follicles provides a multi-dimension research paradigm that links molecular activities and cellular behaviors to functional morphology at the organismal level. PMID:15272383

  3. Interaction between melatonin and follicle-stimulating hormone promotes in vitro development of caprine preantral follicles.

    PubMed

    Rocha, R M P; Lima, L F; Alves, A M C V; Celestino, J J H; Matos, M H T; Lima-Verde, I B; Bernuci, M P; Lopes, C A P; Báo, S N; Campello, C C; Rodrigues, A P R; Figueiredo, J R

    2013-01-01

    The aim of this study was to investigate the effects of melatonin and follicle-stimulating hormone (FSH) on the in vitro culture of goat preantral follicles. Ovarian fragments were cultured for 7 d in α-minimum essential medium (α-MEM(+)) containing melatonin (100, 250, 500, or 1,000 pM), FSH (50 ng/mL), or a combination of the 2 hormones and further analyzed by histology and transmission electron and fluorescent microscopy. The results showed that after 7 d of culture, tissues cultured in α-MEM(+) alone or supplemented with FSH alone, melatonin (500 and 1,000 pM), or the combination of FSH and melatonin (1,000 pM) maintained percentages of normal preantral follicles similar to the fresh control. In contrast to the noncultured tissues, the percentage of developing follicles was increased under all culture conditions after 7 d (P < 0.05). The addition of 1,000 pM melatonin associated with FSH to the culture medium increased follicular and oocyte diameters compared with α-MEM(+) alone after 7 d of culture (P < 0.05). Ultrastructural and fluorescent analyses confirmed the integrity of follicles cultured with 1,000 pM of melatonin plus FSH for 7 d. In conclusion, this study demonstrated that the interaction between melatonin and FSH maintains ultrastructural integrity and stimulates further growth of cultured caprine preantral follicles.

  4. Transforming growth factor-beta: its role in ovarian follicle development.

    PubMed

    Rosairo, Davina; Kuyznierewicz, Ileana; Findlay, Jock; Drummond, Ann

    2008-12-01

    Ovarian follicular growth and differentiation in response to transforming growth factor-beta (TGFB) was investigated using postnatal and immature ovarian models. TGFB ligand and receptor mRNAs were present in the rat ovary 4-12 days after birth and at day 25. In order to assess the impact of TGFB1 on follicle growth and transition from the primordial through to the primary and preantral stages of development, we established organ cultures with 4-day-old rat ovaries. After 10 days in culture with FSH, TGFB1, or a combination of the two, ovarian follicle numbers were counted and an assessment of atresia was undertaken using TUNEL. Preantral follicle numbers declined significantly when treated with the combination of FSH and TGFB1, consistent with our morphological appraisal suggesting an increase in atretic primary and preantral follicles. To investigate the mechanisms behind the actions of TGFB1, we isolated granulosa cells and treated them with FSH and TGFB1. Markers of proliferative, steroidogenic, and apoptotic capacity were measured by real-time PCR. Cyclin D2 mRNA expression by granulosa cells was significantly increased in response to the combination of FSH and TGFB. The expression of forkhead homolog in rhabdomyosarcoma (Foxo1) mRNA by granulosa cells was significantly reduced in the presence of both FSH and TGFB1, individually and in combination regimes. By contrast, the expression of steroidogenic enzymes/proteins was largely unaffected by TGFB1. These data suggest an inhibitory role for TGFB1 (in the presence of FSH) in follicle development and progression.

  5. Gene and stem cell therapy of the hair follicle.

    PubMed

    Hoffman, Robert M

    2005-01-01

    The hair follicle is a highly complex appendage of the skin containing a multiplicity of cell types. The follicle undergoes constant cycling through the life of the organism including growth and resorption with growth dependent on specific stem cells. The targeting of the follicle by genes and stem cells to change its properties, in particular, the nature of the hair shaft is discussed. Hair follicle delivery systems are described such as liposomes and viral vectors for gene therapy. The nature of the hair follicle stem cells is discussed, in particular, its pluripotency.

  6. Granulosa cell responsiveness to follicle stimulating hormone during early growth of hen ovarian follicles.

    PubMed

    Johnson, A L; Lee, Jeeyoung

    2016-01-01

    In the laying hen ovary, the cyclic recruitment of a follicle represents a process in which a single follicle is selected to enter the rapid growth phase and undergo final maturation prior to ovulation. Published data support the proposal that final differentiation of the granulosa cell (GC) layer commences at the time of follicle selection. This process is characterized by the enhanced capacity for FSH-induced cell signaling via the protein kinase A/cyclic adenosine monophosphate (cAMP) pathway. One consequence of such signaling within the GC layer is the initial capacity for steroidogenesis (predominantly progesterone production) mediated by increased expression of mRNA encoding steroidogenic acute regulatory protein (STAR) and the cholesterol side-chain cleavage enzyme (CYP11A). Prior to selection, the GC layer remains minimally responsive to a 3 h challenge with FSH (10 ng/mL), in vitro, compared to that from the most recently selected 9- to 12-mm follicle. By comparison, when the duration of the cell culture prior to FSH challenge is increased to 18 h, GCs collected from 1- to 2-mm, 3- to 5-mm, and 6- to 8-mm follicles respond to a 3 h FSH challenge by increasing STAR expression and progesterone production, with the greatest response from GCs collected from 6- to 8-mm follicles. Culture with Bone Morphogenetic Protein 6 (BMP6) enhances both CYP11A expression and FSH responsiveness at each stage of development, with the greatest response again occurring in GCs from 6- to 8-mm follicles. Significantly, factors that activate mitogen activated protein kinase (MAPK) or protein kinase C (PKC) signaling prevent the ability of prolonged culture or culture with BMP6 to induce FSH-responsiveness and the initiation of GC differentiation at each stage of development. Collectively, these results provide further support for the hypothesis that prior to follicle selection, inhibitory cell signaling (e.g., MAPK, PKC) maintains the GC layer in an undifferentiated state in

  7. [Endorphin-containing cells in the gastric antral mucosa in duodenal ulcer].

    PubMed

    Zverkov, I V; Vinogradov, V A; Smagin, V G

    1983-10-01

    Immunohistochemical staining with the use of peroxidase-antiperoxidase was applied to study cells producing gamma- and alpha-endorphines in the gastric antral mucosa in duodenal ulcer. The cells producing gamma-endorphines were discovered to be mainly located in the epithelium of the cervical and upper third of the pyloric glands and to be alike G-cells producing gastrin. The cells producing alpha-endorphine were found both in the epithelium of the upper third of the gastric pyloric glands and in the gastric mucosa lamina proper. In peptic ulcer, there was an almost two-fold increase in the amount of gamma-endorphine-producing cells and diminution of epithelial endocrine cells producing alpha-endorphine.

  8. External-beam radiotherapy: A realistic therapeutic option for the gastric antral vascular ectasia

    PubMed Central

    Montero, Angel; Fernández-Lizarbe, Eva; Rodríguez, Miguel-Ángel; Hernanz, Raúl; Polo, Alfredo; Ramos, Alfredo

    2012-01-01

    The gastric antral vascular ectasia (GAVE) is a well recognizable endoscopic entity characterized by the presence of multiple linear angioectatic vessels predominantly located in the antrum, with a typical appearance of “watermelon stomach”. This condition typically affects elderly females presenting as iron-deficiency anaemia due to chronic gastric bleeding. Standard treatment is endoscopic ablation of the gastric mucosa. For non-responders, radical surgery is considered a curative treatment but with considerable morbidity and mortality. Radiation therapy is a well-known alternative for many benign diseases, including anomalous vascular hyperproliferative diseases, although its role has not been defined for GAVE. The present case illustrates the efficacy and tolerance of radiotherapy in the treatment of symptomatic gastric watermelon. PMID:24377029

  9. Cyclic nucleotides of canine antral smooth muscle. Effects of acetylcholine, catecholamines and gastrin.

    PubMed

    Baur, S; Grant, B; Wooton, J

    1981-01-07

    1. The effects of acetylcholine, catecholamines and gastrin on the intracellular content of cyclic AMP and cyclic GMP in antral circular muscle have been determined. 2. Acetylcholine results in a significant but transient increase in intracellular cyclic GMP. 3. Isoproterenol and norepinephrine increase intracellular cyclic AMP. Based on half-maximal effective doses, isoproterenol is 2.7-times more effective than norepinephrine. The increase in intracellular cyclic AMP by both agents is inhibited by propranolol but not phentolamine, indicating that both agents act on the muscle cell by a beta-receptor-coupled mechanism. 4. Gastrin has no demonstrable effect on either cyclic AMP or cyclic GMP. This suggests that while gastrin and acetylcholine can produce a like myoelectric response in the muscle cell, the action of gastrin is mediated by a separate receptor, presumably on the muscle cell, and not by a release of acetylcholine.

  10. Loss of local capture of the pulmonary vein myocardium after antral isolation: prevalence and clinical significance.

    PubMed

    Squara, Fabien; Liuba, Ioan; Chik, William; Santangeli, Pasquale; Zado, Erica S; Callans, David J; Marchlinski, Francis E

    2015-03-01

    Capture of the myocardial sleeves of the pulmonary veins (PV) during PV pacing is mandatory for assessing exit block after PV isolation (PVI). However, previous studies reported that a significant proportion of PVs failed to demonstrate local capture after PVI. We designed this study to evaluate the prevalence and the clinical significance of loss of PV capture after PVI. Thirty patients (14 redo) undergoing antral PVI were included. Before and after PVI, local PV capture was assessed during circumferential pacing (10 mA/2 milliseconds) with a circular multipolar catheter (CMC), using EGM analysis from each dipole of the CMC and from the ablation catheter placed in ipsilateral PV. Pacing output was varied to optimize identification of sleeve capture. All PVs demonstrated sleeve capture before PVI, but only 81% and 40% after first time and redo PVI, respectively (P < 0.001 vs. before PVI). In multivariate analysis, absence of spontaneous PV depolarizations after PVI and previous PVI procedures were associated with less PV sleeve capture after PVI (40% sleeve capture, P < 0.001 for both). Loss of PV local capture by design was coincident with the development of PV entrance block and importantly predicted absence of acute reconnection during adenosine challenge with 96% positive predictive value (23% negative predictive value). Loss of PV local capture is common after antral PVI resulting in entrance block, and may be used as a specific alternate endpoint for PV electrical isolation. Additionally, loss of PV local capture may identify PVs at very low risk of acute reconnection during adenosine challenge. © 2014 Wiley Periodicals, Inc.

  11. Size-specific follicle selection improves mouse oocyte reproductive outcomes

    PubMed Central

    Xiao, Shuo; Duncan, Francesca E.; Bai, Lu; Nguyen, Catherine T.; Shea, Lonnie D.; Woodruff, Teresa K.

    2015-01-01

    Encapsulated in vitro follicle growth (eIVFG) has great potential to provide an additional fertility preservation option for young women and girls with cancer or other reproductive health threatening diseases. Currently, follicles are cultured for a defined period of time and analyzed as a cohort. However, follicle growth is not synchronous, and culturing follicles for insufficient or excessive times can result in compromised gamete quality. Our objective is to determine whether the selection of follicles based on size, rather than absolute culture time, better predict follicle maturity and oocyte quality. Multilayer secondary mouse follicles were isolated and encapsulated in 0.25% alginate. Follicles were cultured individually either for defined time periods or up to specific follicle diameter ranges, at which point several reproductive endpoints were analyzed. The metaphase II (MII) percentage after oocyte maturation on day 6 was the highest (85%) when follicles were cultured for specific days. However, if follicles were cultured to a terminal diameter of 300–350 μm irrespective of absolute time in culture, 93% of the oocytes reached MII. More than 90% of MII oocytes matured from follicles with diameters of 300–350 μm showed normal spindle morphology and chromosome alignment, 85% of oocytes showed 2 pronuclei after in vitro fertilization (IVF), 81% developed into the 2-cell embryo stage, and 38% developed to the blastocyst stage, all significantly higher than the percentages in the other follicle size groups. Our study demonstrates that size-specific follicle selection can be used as a non-invasive marker to identify high quality oocytes and improve reproductive outcomes during eIVFG. PMID:26116002

  12. All about Carbohydrate Counting

    MedlinePlus

    Toolkit No. 14 All About Carbohydrate Counting What is carbohydrate counting? Carbohydrate counting is a way to plan your meals. It can help ... Diabetes Association, Inc. 2/14 Toolkit No. 14: All About Carbohydrate Counting continued The chart at the ...

  13. Cellular and molecular regulation of the activation of mammalian primordial follicles: somatic cells initiate follicle activation in adulthood.

    PubMed

    Zhang, Hua; Liu, Kui

    2015-01-01

    The first small follicles to appear in the mammalian ovaries are primordial follicles. The initial pool of primordial follicles serves as the source of developing follicles and oocytes for the entire reproductive lifespan of the animal. Although the selective activation of primordial follicles is critical for female fertility, its underlying mechanisms have remained poorly understood. A search of PubMed was conducted to identify peer-reviewed literature pertinent to the study of mammalian primordial follicle activation, especially recent reports of the role of primordial follicle granulosa cells (pfGCs) in regulating this process. In recent years, molecular mechanisms that regulate the activation of primordial follicles have been elucidated, mostly through the use of genetically modified mouse models. Several molecules and pathways operating in both the somatic pfGCs and oocytes, such as the phosphatidylinositol 3 kinase (PI3K) and the mechanistic target of rapamycin complex 1 (mTORC1) pathways, have been shown to be important for primordial follicle activation. More importantly, recent studies have provided an updated view of how exactly signaling pathways in pfGCs and in oocytes, such as the KIT ligand (KL) and KIT, coordinate in adult ovaries so that the activation of primordial follicles is achieved. In this review, we have provided an updated picture of how mammalian primordial follicles are activated. The functional roles of pfGCs in governing the activation of primordial follicles in adulthood are highlighted. The in-depth understanding of the cellular and molecular mechanisms of primordial follicle activation will hopefully lead to more treatments of female infertility, and the current progress indicates that the use of existing primordial follicles as a source for obtaining fertilizable oocytes as a new treatment for female infertility is just around the corner. © The Author 2015. Published by Oxford University Press on behalf of the European Society of

  14. Hierarchical patterning modes orchestrate hair follicle morphogenesis

    PubMed Central

    Glover, James D.; Wells, Kirsty L.; Matthäus, Franziska; Painter, Kevin J.; Ho, William; Riddell, Jon; Johansson, Jeanette A.; Ford, Matthew J.; Jahoda, Colin A. B.; Klika, Vaclav; Mort, Richard L.

    2017-01-01

    Two theories address the origin of repeating patterns, such as hair follicles, limb digits, and intestinal villi, during development. The Turing reaction–diffusion system posits that interacting diffusible signals produced by static cells first define a prepattern that then induces cell rearrangements to produce an anatomical structure. The second theory, that of mesenchymal self-organisation, proposes that mobile cells can form periodic patterns of cell aggregates directly, without reference to any prepattern. Early hair follicle development is characterised by the rapid appearance of periodic arrangements of altered gene expression in the epidermis and prominent clustering of the adjacent dermal mesenchymal cells. We assess the contributions and interplay between reaction–diffusion and mesenchymal self-organisation processes in hair follicle patterning, identifying a network of fibroblast growth factor (FGF), wingless-related integration site (WNT), and bone morphogenetic protein (BMP) signalling interactions capable of spontaneously producing a periodic pattern. Using time-lapse imaging, we find that mesenchymal cell condensation at hair follicles is locally directed by an epidermal prepattern. However, imposing this prepattern’s condition of high FGF and low BMP activity across the entire skin reveals a latent dermal capacity to undergo spatially patterned self-organisation in the absence of epithelial direction. This mesenchymal self-organisation relies on restricted transforming growth factor (TGF) β signalling, which serves to drive chemotactic mesenchymal patterning when reaction–diffusion patterning is suppressed, but, in normal conditions, facilitates cell movement to locally prepatterned sources of FGF. This work illustrates a hierarchy of periodic patterning modes operating in organogenesis. PMID:28700594

  15. Mechanisms of organogenesis of primary lymphoid follicles.

    PubMed

    Beyer, Tilo; Meyer-Hermann, Michael

    2008-04-01

    Primary lymphoid follicles (PLFs) in secondary lymphoid tissue (SLT) of mammals are the backbone for the formation of follicular dendritic cell (FDC) networks. These are important for germinal center reactions during which affinity maturation creates optimized antibodies in adaptive immune responses. In the context of organogenesis, molecular requirements for the formation of follicles have been identified. The present study complements these findings with a simulation of the dynamics of the PLF formation, and a critical analysis of the relevant molecular interactions. In contrast to other problems of pattern formation, the homeostasis of cell populations in SLTs is not governed by a growth-death balance but by a flow equilibrium of migrating cells. The influx of cells into these tissues has been extensively studied. However, less information is available about the efflux of lymphocytes from SLTs. This study formulates the minimal requirements for cell efflux that guarantee a flow equilibrium and, thus, a stable PLF. The model predicts that in addition to already identified regulatory mechanisms, a negative regulation of the generation of FDCs is required. Furthermore, a comparison with data concerning the microanatomy of SLTs yields the conclusion that dynamical changes of the lymphatic endothelium during the formation of FDC networks are necessary to understand the genesis and maintenance of follicles.

  16. Hair follicle transplantation on scar tissue.

    PubMed

    Jung, Soyeon; Oh, Suk Joon; Hoon Koh, Sung

    2013-07-01

    Hair transplantation is a continuously evolving field. The procedure was originally developed by Dr. Orentreich in 1959, but he applied it only to the androgenic alopecia. Potential applications for hair grafting extend beyond treatment of hair loss. Our study group consisted of 25 cases of 23 patients. The causes of scar resulting to hair loss were burns, operation, and trauma. The scalp strips or follicular unit extracts were harvested from occipital, posterior auricular, dog-eared scalp, adjacent scalp area, and nuchal area. The recipient sites were scalp, eyebrow, lip, and eyelid. The follow-up cases over 6 months after operation were 18 among total 25 cases. The result after hair follicle transplantation was excellent (44.4%), good (38.9%), fair (11.1%), and poor (5.6%). The hair follicle transplantation on the scar tissue is more difficult than grafting on normal tissue because the scar is accompanied by poor blood circulation and stiffness of tissue. The patients with burned scar achieved more favorable result than did others. Incision scars are deeper than burned scars, and their success rates are poor. We should recommend the patients that hair follicle transplantation on the scar may need secondary or more operations for the aesthetically better result.

  17. Reduced quality and accelerated follicle loss with female reproductive aging - does decline in theca dehydroepiandrosterone (DHEA) underlie the problem?

    PubMed

    Ford, Judith H

    2013-12-13

    Infertility, spontaneous abortion and conception of trisomic offspring increase exponentially with age in mammals but in women there is an apparent acceleration in the rate from about age 37. The problems mostly commonly occur when the ovarian pool of follicles is depleted to a critical level with age but are also found in low follicular reserve of other etiologies. Since recent clinical studies have indicated that dehydroepiandrosterone (DHEA) supplementation may reverse the problem of oocyte quality, this review of the literature was undertaken in an attempt to find an explanation of why this is effective? In affected ovaries, oxygenation of follicular fluid is low, ultrastructural disturbances especially of mitochondria, occur in granulosa cells and oocytes, and considerable disturbances of meiosis occur. There is, however, no evidence to date that primordial follicles are compromised. In females with normal fertility, pre-antral ovarian theca cells respond to stimulation by inhibin B to provide androgen-based support for the developing follicle. With depletion of follicle numbers, inhibin B is reduced with consequent reduction in theca DHEA. Theca cells are the sole ovarian site of synthesis of DHEA, which is both a precursor of androstenedione and an essential ligand for peroxisome proliferator-activated receptor alpha (PPARα), the key promoter of genes affecting fatty acid metabolism and fat transport and genes critical to mitochondrial function. As well as inducing a plethora of deleterious changes in follicular cytoplasmic structure and function, the omega 9 palmitate/oleate ratio is increased by lowered activity of PPARα. This provides conditions for increased ceramide synthesis and follicular loss through ceramide-induced apoptosis is accelerated. In humans critical theca DHEA synthesis occurs at about 70 days prior to ovulation thus effective supplementation needs to be undertaken about four months prior to intended conception; timing which is also

  18. Reduced quality and accelerated follicle loss with female reproductive aging - does decline in theca dehydroepiandrosterone (DHEA) underlie the problem?

    PubMed Central

    2013-01-01

    Infertility, spontaneous abortion and conception of trisomic offspring increase exponentially with age in mammals but in women there is an apparent acceleration in the rate from about age 37. The problems mostly commonly occur when the ovarian pool of follicles is depleted to a critical level with age but are also found in low follicular reserve of other etiologies. Since recent clinical studies have indicated that dehydroepiandrosterone (DHEA) supplementation may reverse the problem of oocyte quality, this review of the literature was undertaken in an attempt to find an explanation of why this is effective? In affected ovaries, oxygenation of follicular fluid is low, ultrastructural disturbances especially of mitochondria, occur in granulosa cells and oocytes, and considerable disturbances of meiosis occur. There is, however, no evidence to date that primordial follicles are compromised. In females with normal fertility, pre-antral ovarian theca cells respond to stimulation by inhibin B to provide androgen-based support for the developing follicle. With depletion of follicle numbers, inhibin B is reduced with consequent reduction in theca DHEA. Theca cells are the sole ovarian site of synthesis of DHEA, which is both a precursor of androstenedione and an essential ligand for peroxisome proliferator-activated receptor alpha (PPARα), the key promoter of genes affecting fatty acid metabolism and fat transport and genes critical to mitochondrial function. As well as inducing a plethora of deleterious changes in follicular cytoplasmic structure and function, the omega 9 palmitate/oleate ratio is increased by lowered activity of PPARα. This provides conditions for increased ceramide synthesis and follicular loss through ceramide-induced apoptosis is accelerated. In humans critical theca DHEA synthesis occurs at about 70 days prior to ovulation thus effective supplementation needs to be undertaken about four months prior to intended conception; timing which is also

  19. Dental implants after the use of bichat's buccal fat pad for the sealing of oro-antral communications. A case report and literature review.

    PubMed

    Galletti, Cosimo; Cammaroto, Giovanni; Galletti, Francesco; Camps-Font, Octavi; Gay-Escoda, Cosme; Bara-Casaus, José-Javier

    2016-12-01

    Oro-antral communications are frequent complications in oral surgery, and generally occur after molar extractions, maxillary sinus elevations or dental implant procedures. The presence of these defects may increase the morbidity and often need a surgical approach. The present report describes an oro-antral communication in a 52-year-old female who presented a 2 week-course of painless nasal obstruction and rhinorrea after a right maxillary sinus floor elevationwith simultaneous dental implant placement. Based on the anamnesis, clinical examination and a computed tomographyof the paranasal sinuses, a diagnosis of odontogenic rhinosinusitis associated with a 1.5 cm diameter oro-antral communicationwas establishedand its surgical closure using Bichat's buccal fat pad was planned.After 15 months, the patient was successfully rehabilitated with an implant-supported 3 unit fixed partial denture. Key words:Dental implants, buccal fat pad, oro-antral communications.

  20. Dental implants after the use of bichat’s buccal fat pad for the sealing of oro-antral communications. A case report and literature review

    PubMed Central

    Cammaroto, Giovanni; Galletti, Francesco; Camps-Font, Octavi; Gay-Escoda, Cosme; Bara-Casaus, José-Javier

    2016-01-01

    Oro-antral communications are frequent complications in oral surgery, and generally occur after molar extractions, maxillary sinus elevations or dental implant procedures. The presence of these defects may increase the morbidity and often need a surgical approach. The present report describes an oro-antral communication in a 52-year-old female who presented a 2 week-course of painless nasal obstruction and rhinorrea after a right maxillary sinus floor elevationwith simultaneous dental implant placement. Based on the anamnesis, clinical examination and a computed tomographyof the paranasal sinuses, a diagnosis of odontogenic rhinosinusitis associated with a 1.5 cm diameter oro-antral communicationwas establishedand its surgical closure using Bichat’s buccal fat pad was planned.After 15 months, the patient was successfully rehabilitated with an implant-supported 3 unit fixed partial denture. Key words:Dental implants, buccal fat pad, oro-antral communications. PMID:27957285

  1. Mouse ovarian follicles secrete factors affecting the growth and development of like-sized ovarian follicles in vitro.

    PubMed

    Spears, Norah; Baker, Stuart; Srsen, Vlastimil; Lapping, Rebecca; Mullan, Julie; Nelson, Robert; Allison, Vivian

    2002-12-01

    A series of experiments have been carried out to determine whether follicles secrete factors able to affect the growth and development of other, like-sized follicles. Late preantral mouse ovarian follicles were either cocultured or cultured in media conditioned by previously cultured follicles. In particular, the experiments examined whether follicles do secrete such factors, whether the level of FSH in the culture media can affect that process, and what the nature of such secretory factor(s) might be. First, pairs of follicles were cocultured across a polycarbonate membrane containing pores. This showed that communication between the follicles resulted in the stimulation of growth and that the stimulation was due, at least in part, to the production of secretory factor(s). In subsequent experiments, follicles were cultured in media that had been preconditioned by previously cultured follicles. The concentration of FSH in the cultures determined the effect of the conditioned media: conditioned media was stimulatory to follicle growth when levels of FSH remained high throughout the culture, but inhibitory when FSH levels were dropped midway through the cultures. Heat inactivation removed this inhibitory effect, showing that the factor was likely to be a protein; addition of follistatin to the conditioned media did not alter its effect, indicating that the factor was unlikely to be activin. We have shown through a series of culture experiments that mouse follicles secrete factor(s) that can affect the development of other like-sized follicles when cultured from the late preantral to Graafian stages. Furthermore, we have shown that the effect (or production) of such factors is dependent on the FSH environment of the follicles.

  2. Reflectance spectroscopy for noninvasive evaluation of hair follicle stage

    NASA Astrophysics Data System (ADS)

    Liu, Caihua; Guan, Yue; Wang, Jianru; Zhong, Xiewei; Liu, Xiuli; Zhu, Dan

    2015-05-01

    Hair follicle offers an excellent model for systems biology and regenerative medicine. So far, the stages of hair follicle growth have been evaluated by histological examination. In this work, a noninvasive spectroscopy was proposed by measuring the diffuse reflectance of mouse skin and analyzing the melanin value. Results show that the skin diffuse reflectance was relatively high when hair follicles were at the telogen stage and at the beginning of the anagen stage, and decreased with the progression of the anagen stage. When the hair follicle entered into the catagen stage, the diffuse reflectance gradually increased. The changes in the melanin content of skin had contrary dynamics. Substages of the hair follicle cycle could be distinguished by comparing the changes in melanin value with the histological examination. This study provided a new method for noninvasive evaluation of the hair follicle stage, and should be valuable for basic and therapeutic investigations on hair regeneration.

  3. Impairing follicle-stimulating hormone (FSH) signaling in vivo: targeted disruption of the FSH receptor leads to aberrant gametogenesis and hormonal imbalance.

    PubMed

    Dierich, A; Sairam, M R; Monaco, L; Fimia, G M; Gansmuller, A; LeMeur, M; Sassone-Corsi, P

    1998-11-10

    Pituitary gonadotropins follicle-stimulating hormone (FSH) and luteinizing hormone stimulate the gonads by regulating germ cell proliferation and differentiation. FSH receptors (FSH-Rs) are localized to testicular Sertoli cells and ovarian granulosa cells and are coupled to activation of the adenylyl cyclase and other signaling pathways. Activation of FSH-Rs is considered essential for folliculogenesis in the female and spermatogenesis in the male. We have generated mice lacking FSH-R by homologous recombination. FSH-R-deficient males are fertile but display small testes and partial spermatogenic failure. Thus, although FSH signaling is not essential for initiating spermatogenesis, it appears to be required for adequate viability and motility of the sperms. FSH-R-deficient females display thin uteri and small ovaries and are sterile because of a block in folliculogenesis before antral follicle formation. Although the expression of marker genes is only moderately altered in FSH-R -/- mice, drastic sex-specific changes are observed in the levels of various hormones. The anterior lobe of the pituitary gland in females is enlarged and reveals a larger number of FSH- and thyroid-stimulating hormone (TSH)-positive cells. The phenotype of FSH-R -/- mice is reminiscent of human hypergonadotropic ovarian dysgenesis and infertility.

  4. Intrauterine Exposure to Paracetamol and Aniline Impairs Female Reproductive Development by Reducing Follicle Reserves and Fertility.

    PubMed

    Holm, Jacob Bak; Mazaud-Guittot, Severine; Danneskiold-Samsøe, Niels Banhos; Chalmey, Clementine; Jensen, Benjamin; Nørregård, Mette Marie; Hansen, Cecilie Hurup; Styrishave, Bjarne; Svingen, Terje; Vinggaard, Anne Marie; Koch, Holger Martin; Bowles, Josephine; Koopman, Peter; Jégou, Bernard; Kristiansen, Karsten; Kristensen, David Møbjerg

    2016-03-01

    Studies report that fetal exposure to paracetamol/acetaminophen by maternal consumption can interfere with male reproductive development. Moreover, recent biomonitoring data report widespread presence of paracetamol in German and Danish populations, suggesting exposure via secondary (nonpharmaceutical) sources, such as metabolic conversion from the ubiquitous industrial compound aniline. In this study, we investigated the extent to which paracetamol and aniline can interfere with female reproductive development. Intrauterine exposure to paracetamol by gavage of pregnant dams resulted in shortening of the anogenital distance in adult offspring, suggesting that fetal hormone signaling had been disturbed. Female offspring of paracetamol-exposed mothers had ovaries with diminished follicle reserve and reduced fertility. Fetal gonads of exposed animals had also reduced gonocyte numbers, suggesting that the reduced follicle count in adults could be due to early disruption of germ cell development. However, ex vivo cultures of ovaries from 12.5 days post coitum fetuses showed no decrease in proliferation or expression following exposure to paracetamol. This suggests that the effect of paracetamol occurs prior to this developmental stage. Accordingly, using embryonic stem cells as a proxy for primordial germ cells we show that paracetamol is an inhibitor of cellular proliferation, but without cytotoxic effects. Collectively, our data show that intrauterine exposure to paracetamol at levels commonly observed in pregnant women, as well as its precursor aniline, may block primordial germ cell proliferation, ultimately leading to reduced follicle reserves and compromised reproductive capacity later in life.

  5. Protective Effect of 4-(3,4-Dihydroxyphenyl)-3-Buten-2-One from Phellinus linteus on Naproxen-Induced Gastric Antral Ulcers in Rats.

    PubMed

    Kim, Jeong-Hwan; Kwon, Hyun Ju; Kim, Byung Woo

    2016-05-28

    The present study investigated the protective effect of naturally purified 4-(3,4- dihydroxyphenyl)-3-buten-2-one (DHP) from Phellinus linteus against naproxen-induced gastric antral ulcers in rats. To verify the protective effect of DHP on naproxen-induced gastric antral ulcers, various doses (1, 5, and 10 μg/kg) of DHP were pretreated for 3 days, and then gastric damage was caused by 80 mg/kg naproxen applied for 3 days. DHP prevented naproxen-induced gastric antral ulcers in a dose-dependent manner. In particular, 10 μg/kg DHP showed the best protective effect against naproxen-induced gastric antral ulcers. Moreover, DHP significantly attenuated the naproxen-induced lipid peroxide level in gastric mucosa and increased the activities of radical scavenging enzymes, such as superoxide dismutase, catalase, and glutathione peroxidase, in a dose-dependent manner. A histological examination clearly demonstrated that the gastric antral ulcer induced by naproxen nearly disappeared after the pretreatment of DHP. These results suggest that DHP can inhibit naproxen-induced gastric antral ulcers through prevention of lipid peroxidation and activation of radical scavenging enzymes.

  6. Follicle-stimulating hormone treatment of male infertility.

    PubMed

    Foresta, Carlo; Selice, Riccardo; Garolla, Andrea; Ferlin, Alberto

    2008-11-01

    Treatment with gonadotrophins is very effective in patients affected by hypogonadotrophic hypogonadism. The success of follicle-stimulating hormone (FSH) treatment in these men has brought the utilization of the same therapy in infertile oligozoospermic patients, aimed at obtaining a quantitative increase in sperm count. FSH plays a crucial role in human reproduction. This physiological role in spermatogenesis has induced various attempts to treat idiopathic oligozoospermic men with FSH, often inducing the restoration of normal spermatogenesis and spontaneous pregnancy. However, the results obtained so far are still controversial. In this research, attention is focused on the possible criteria able to predict a seminal response to the specific hormonal treatment. Moreover, we have correlated different polymorphisms of FSH receptor gene with the outcome of FSH treatment. In this article, the literature is reviewed, and the authors' experience on using FSH treatment in oligozoospermic patients is discussed. FSH treatment may represent a valid tool for infertile men. However, it should be performed on selected patients utilizing some predictive parameters able to identify a priori responder patients with high probability.

  7. Ovarian follicle dynamics of female Greater Scaup during egg production

    USGS Publications Warehouse

    Gorman, K.B.; Flint, P.L.; Esler, Daniel; Williams, T.D.

    2007-01-01

    Studies of female waterfowl nutrient reserve use during egg production require a precise understanding of ovarian follicle dynamics to correctly interpret breeding status, and, therefore, derive proper inference. Concerns over numerical declines of North American scaup have increased the need to better understand the role of female condition in reproductive performance. We quantified ovarian follicle dynamics of female Greater Scaup (Aythya marila) breeding on the Yukon–Kuskokwim Delta, Alaska, using a method that accounts for within day variation in follicle size. We considered several models for describing changes in follicle growth with the best supported model estimating the duration of rapid follicle growth (RFG) to be 5.20 ± 0.52 days (±95% confidence intervals) for each developing follicle. Average diameter and dry mass of preovulatory follicles were estimated to be 9.36 mm and 0.26 g, respectively, at the onset of RFG, and these follicle characteristics were 41.47 mm and 15.57 g, respectively, at ovulation. The average diameter of postovulatory follicles immediately following ovulation was estimated to be 17.35 mm, regressing quickly over several days. In addition, we derived predictive equations using diameter and dry mass to estimate the number of days before, and after, ovulation for pre- and postovulatory follicles, as well as an equation to estimate dry mass of damaged follicles. Our results allow precise definition of RFG and nest initiation dates, clutch size, and the daily energetic and nutritional demands of egg production at the individual level. This study provides the necessary foundation for additional work on Greater Scaup reproductive energetics and physiology, and offers an approach for quantifying ovarian follicle dynamics in other species.

  8. Introduction to Hair-Follicle-Associated Pluripotent Stem Cells.

    PubMed

    Hoffman, Robert M

    2016-01-01

    Nestin-expressing stem cells of the hair follicle, discovered by our laboratory, have been shown to be able to form outer-root sheaths of the follicle as well as neurons and many other non-follicle cell types. We have termed the nestin-expressing stem cells of the hair follicle as hair-follicle-associated pluripotent (HAP) stem cells. We have shown that the HAP stem cells from the hair follicle can effect the repair of peripheral nerve and spinal cord injury. The hair follicle stem cells differentiate into neuronal and glial cells after transplantation to the injured peripheral nerve and spinal cord, and enhance injury repair and locomotor recovery. When the excised hair follicle with its nerve stump was placed in Gelfoam(®) 3D histoculture, HAP stem cells grew and extended the hair follicle nerve which consisted of βIII-tubulin-positive fibers with F-actin expression at the tip. These findings indicate that βIII-tubulin-positive fibers elongating from the whisker follicle sensory nerve stump were growing axons. The growing whisker sensory nerve was highly enriched in HAP stem cells, which appeared to play a major role in its elongation and interaction with other nerves in 3D Gelfoam(®) histoculture, including the sciatic nerve, the trigeminal nerve, and the trigeminal nerve ganglion. These results suggest that a major function of the HAP stem cells in the hair follicle is for growth of the follicle sensory nerve. Recently, we have shown that HAP stem cells can differentiate into beating cardiac muscle cells. HAP stem cells have critical advantages for regenerative medicine over embryonic stem (ES) cells and induced pluripotent stem (iPS) cells in that they are highly accessible from each patient, thereby eliminating immunological issues since they are autologous, require no genetic manipulation, are non-tumorigenic, and do not present ethical issues.

  9. [Maxillary sinus augmentation and implant placement following removal of a maxillary antral cyst: a preliminary clinical study].

    PubMed

    Wang, Xinyu; Di, Ping; Li, Jianhui; Hu, Xiulian; Lin, Ye

    2015-09-01

    To evaluate the clinical outcome of maxillary sinus augmentation and implant placement following removal of a maxillary antral cyst. In this study 32 patients with 33 maxillary antral cysts were enrolled. The partial wall of cyst was removed through a small lateral sinus approach and cyst spontaneous shrink was expected when wall of cyst was destroyed for open drainage. Three to six months later the secondary sinus approach 1.5 mm circling the first approach for augmentation was undertaken. Dental implants were placed simultaneously or later. All patients finished prosthetic rehabilitation in the study and were followed up for (30.9 ± 11.5) months. The intraoperative and postoperative complications were recorded. The survival rate of the implants was calculated. In this study, 33 maxillary antral cysts of 32 patients were removed. Maxillary sinus augmentation was performed after a mean of (4.5 ± 1.5) months (range, 2-8 months). Sixty-two implants were inserted and all patients finished prosthetic rehabilitation after a mean of (10.8 ± 2.7) months (range, 5-17 months). The survival rate of implants was 95% (59/62). Three implants failed before their prosthesis delivered due to failure of osseointegration and were reinserted later. No recurrence of cyst was observed until the last recall. In this study, 24 specimens were mucosal cysts. Five specimens were mucoceles and 4 specimens were not certain in pathology. Maxillary sinus cysts have a negative effect on maxillary sinus augumentation if not removed. The present preliminary data of the study suggests that the clinical result of maxillary sinus augmentation and implant placement following removal of a maxillary antral cyst is predictable.

  10. Estimate of the population of preantral follicles in the ovaries of Bos taurus indicus and Bos taurus taurus cattle.

    PubMed

    Silva-Santos, K C; Santos, G M G; Siloto, L S; Hertel, M F; Andrade, E R; Rubin, M I B; Sturion, L; Melo-Sterza, F A; Seneda, M M

    2011-10-01

    The number of oocytes recovered from Bos taurus indicus females subjected to ovum pick-up averaged two to four times greater compared to Bos taurus taurus females. The objective of the present study was to test the hypothesis that this difference in oocyte yield was due to more preantral follicles in the ovaries of Bos indicus females. Ovaries (n = 64) from Nelore (Bos indicus) fetuses (n = 10), heifers (n = 12), and cows (n = 10), and Aberdeen Angus (Bos taurus) fetuses (n = 10), heifers (n = 12), and cows (n = 10) were cut longitudinally into halves, fixed, and processed for histological evaluation. The number of preantral follicles was estimated by counting them in each histological section, using the oocyte nucleus as a marker and employing a correction factor. The average number of preantral follicles in the ovaries of Bos indicus vs Bos taurus was (mean ± SD) 143,929 ± 64,028 vs 285,155 ± 325,195 for fetuses, 76,851 ± 78,605 vs 109,673 ± 86,078 for heifers, and 39,438 ± 31,017 vs 89,577 ± 86,315 for cows (P > 0.05). The number of preantral follicles varied greatly among individual animals within the same category, as well as between breeds. In conclusion, we inferred that the higher oocyte yield from Bos indicus females was not due to a greater ovarian reserve of preantral follicles. Therefore, mechanisms controlling follicle development after the preantral stage likely accounted for differences between Bos indicus and Bos taurus females in number of oocytes retrieved at ovum pick-up. Copyright © 2011 Elsevier Inc. All rights reserved.

  11. The hair follicle as a target for gene therapy.

    PubMed

    Gupta, S; Domashenko, A; Cotsarelis, G

    2001-01-01

    The hair follicle possesses progenitor cells for continued hair follicle cycling and for epidermal keratinocytes, melanocytes and Langerhans cells. These different cell types can be targeted by topical gene delivery to mouse skin. Using a combination of liposomes and DNA, we demonstrated the feasibility of targeting hair follicle cells in human scalp xenografts as well. We defined liposome composition and stage of the hair cycle as important parameters influencing transfection of human hair follicles. Transfection occurred only during anagen onset. Considerations and obstacles for using gene therapy to treat alopecias and skin disease are discussed. A theoretical framework for future gene therapy treatments for cutaneous and systemic disorders is presented.

  12. Alginate: A Versatile Biomaterial to Encapsulate Isolated Ovarian Follicles.

    PubMed

    Vanacker, Julie; Amorim, Christiani A

    2017-02-28

    In vitro culture of ovarian follicles isolated or enclosed in ovarian tissue fragments and grafting of isolated ovarian follicles represent a potential alternative to restore fertility in cancer patients who cannot undergo cryopreservation of embryos or oocytes or transplantation of frozen-thawed ovarian tissue. In this regard, respecting the three-dimensional (3D) architecture of isolated follicles is crucial to maintaining their proper follicular physiology. To this end, alginate hydrogel has been widely investigated using follicles from numerous animal species, yielding promising results. The goal of this review is therefore to provide an overview of alginate applications utilizing the biomaterial as a scaffold for 3D encapsulation of isolated ovarian follicles. Different methods of isolated follicle encapsulation in alginate are discussed in this review, as its use of 3D alginate culture systems as a tool for in vitro follicle analysis. Possible improvements of this matrix, namely modification with arginine-glycine-aspartic acid peptide or combination with fibrin, are also summarized. Encouraging results have been obtained in different animal models, and particularly with isolated follicles encapsulated in alginate matrices and grafted to mice. This summary is designed to guide the reader towards development of next-generation alginate scaffolds, with enhanced properties for follicle encapsulation.

  13. Regulation and 3 dimensional culture of tertiary follicle growth.

    PubMed

    Cheon, Yong-Pil

    2012-09-01

    It has been revealed that multiple cohorts of tertiary follicles develop during some animal estrous cycle and the human menstrual cycle. To reach developmental competence, oocytes need the support of somatic cells. During embryogenesis, the primordial germ cells appear, travel to the gonadal rudiments, and form follicles. The female germ cells develop within the somatic cells of the ovary, granulosa cells, and theca cells. How the oocyte and follicle cells support each other has been seriously studied. The latest technologies in genes and proteins and genetic engineering have allowed us to collect a great deal of information about folliculogenesis. For example, a few web pages (http://www.ncbi.nlm.nih.gov; http://mrg.genetics.washington.edu) provide access to databases of genomes, sequences of transcriptomes, and various tools for analyzing and discovering genes important in ovarian development. Formation of the antrum (tertiary follicle) is the final phase of folliculogenesis and the transition from intraovarian to extraovian regulation. This final step coordinates with the hypothalamic-pituitary-ovarian axis. On the other hand, currently, follicle physiology is under intense investigation, as little is known about how to overcome women's ovarian problems or how to develop competent oocytes from in vitro follicle culture or transplantation. In this review, some of the known roles of hormones and some of the genes involved in tertiary follicle growth and the general characteristics of tertiary follicles are summarized. In addition, in vitro culture of tertiary follicles is also discussed as a study model and an assisted reproductive technology model.

  14. Intra-antral application of an anti-fungal agent for recurrent maxillary fungal rhinosinusitis: a case report

    PubMed Central

    2012-01-01

    Introduction Fungal infection of the paranasal sinuses is an increasingly recognized entity both in immunocompetent and immunocompromised individuals. Treatment has been via use of either surgical or medical modalities, or a combination of the two. Here, we present a case of utilization of intra-antral application of an anti-fungal agent in the management of recurrent fungal sinusitis in an indigent Nigerian patient. Case presentation We present the case of a 30-year-old West African Yoruba man, an indigent Nigerian clergyman, who presented to our facility with a history of recurrent nasal discharge (about one year), recurrent nasal blockage (about five months), and right facial swelling (about one week). After intra-nasal antrostomy for debulking with a systemic anti-fungal agent, our patient had a recurrence after four months. Our patient subsequently had an intra-antral application of flumetasone and clioquinol (Locacorten®-Vioform®) weekly for six weeks with improvement of symptoms and no recurrence after six months of follow-up. Conclusions We conclude that topical intra-antral application of anti-fungal agents is effective in patients with recurrent fungal maxillary sinusitis after surgical debulking. PMID:22905703

  15. Molecular forms of gastrin in antral mucosa, plasma and gastric juice during vagal stimulation of anesthetized cats.

    PubMed

    Uvnäs-Wallensten, K; Rehfeld, J F

    1976-10-01

    Gastrin was released by electrical vagal stimulation in anesthetized cats. Antral mucosa, blood and gastric juice samples collected during vagal stimulation were subjected to gel filtration in order to characterize the different molecular forms of gastrin. In antral mucosa component III (gastrin-17) predominated. Besides, the antrum contained 5 per cent component II (gastrin-34, "big" gastrin), 1 per cent component I and trace amounts of component IV (gastrin-14 or "mini" gastrin). Immediately after vagal stimulation, component III (gastrin-17) appeared in the gastric venous effluent followed within a few minutes by component IV (gastrin-14). Component I and II (gastrin-34) were not detectable in any of the plasma samples. We suggest that component III (gastrin-17) is released from the antral mucosa and is then rapidly metabolized to component IV (gastrin-14) possibly to a significant extent in the fundic region of the stomach. Large amounts of component III (gastrin-17) were found in the vagally-induced gastric juice. Only very small amounts of degradation products were present, indicating that cat gastrin is relatively resistant to peptic degradation and acid hydrolysis.

  16. Complex morphology of gastrin-releasing G-cells in the antral region of the mouse stomach.

    PubMed

    Frick, Claudia; Rettenberger, Amelie Therese; Lunz, Malena Luisa; Breer, Heinz

    2016-11-01

    Gastrin-releasing enteroendocrine cells (G-cells) are usually described as flask-shaped cells with a large base and a small apical pole, integrated in the epithelium lining the basal region of the antral invaginations in the stomach. By means of a transgenic mouse line in which the enhanced version of GFP is endogenously expressed under the control of a gastrin promoter, we have analyzed the spatial distribution and morphological features of G-cells. We found that G-cells were not only located at the basal region of the invagination but to a lesser extent also at the upper region. Visualization of the entire cellular morphology revealed that G-cells show complex morphologies. Basally located G-cells are roundish-shaped cells which project a prominent apical process towards the lumen and extend basal protrusions containing the hormone gastrin that were frequently found in close proximity to blood vessels and occasionally in the vicinity of nerve fibers. Inspection of G-cells in the upper region of antral invaginations disclosed a novel population of G-cells. These cells have a spindle-like contour and long apical and basal processes which extend vertically along the antral invagination, parallel to the lumen. This G-cell population seems to be in contact with a network of nerve fibers. While the functional role of these untypical G-cells is still elusive, the results of this study provide some useful indications to possible roles of these G-cells.

  17. Epithelial cell proliferation in human fundic and antral mucosae. Influence of superselective vagotomy and relationship with gastritis

    SciTech Connect

    Gutierrez, O.; Lehy, T.; Rene, E.; Gres, L.; Bonfils, S.

    1985-11-01

    Epithelial cell proliferation in the fundic and antral mucosae was studied in 19 duodenal ulcer patients, 11 patients having undergone fundic superselective vagotomy for duodenal ulcer, and 10 controls. This was achieved through in vitro incorporation of tritiated thymidine in mucosal biopsies and radioautography. Except for increased fundic mucosal height, duodenal ulcer patients did not differ from controls for all parameters studied. In vagotomized patients, as compared to the other two groups, the labeling index was significantly enhanced in the innervated antral mucosa where atrophic gastritis developed, but there was no change in the labeling index and no worsening of mucosal inflammation in the denervated fundic mucosa. The only abnormality in the latter was a striking expansion, towards the surface, of the proliferative area within the fundic pit. The labeling indices and the degree of gastritis in gastric mucosae are significantly correlated in control and duodenal ulcer patients. If findings in antral mucosa, after superselective vagotomy, seemed related to gastritis lesions, those in fundic mucosa were not and may indicate an alteration due to the vagotomy per se.

  18. The Big Pumpkin Count.

    ERIC Educational Resources Information Center

    Coplestone-Loomis, Lenny

    1981-01-01

    Pumpkin seeds are counted after students convert pumpkins to jack-o-lanterns. Among the activities involved, pupils learn to count by 10s, make estimates, and to construct a visual representation of 1,000. (MP)

  19. The Big Pumpkin Count.

    ERIC Educational Resources Information Center

    Coplestone-Loomis, Lenny

    1981-01-01

    Pumpkin seeds are counted after students convert pumpkins to jack-o-lanterns. Among the activities involved, pupils learn to count by 10s, make estimates, and to construct a visual representation of 1,000. (MP)

  20. Effects of BMP4/SMAD signaling pathway on mouse primordial follicle growth and survival via up-regulation of Sohlh2 and c-kit.

    PubMed

    Ding, Xiangyun; Zhang, Xiaoli; Mu, Yulan; Li, Yi; Hao, Jing

    2013-01-01

    Bone morphogenetic protein 4 (BMP4) is essential for the development of primordial follicles, although its underlying mechanism remains largely unknown. By using cultured ovaries, the effects of BMP4 and the potential signal transduction pathways were investigated. Ovaries from 3-day-old female mouse pups were maintained in organ culture in the absence (control) or presence of BMP4 (100 ng/ml). At different culture time, the effects of BMP4 on primordial follicle growth and survival were assayed by follicle count and TUNEL labeling. The expression of phospho-SMAD1/5/8, Sohlh2, and c-kit were measured by immunohistochemistry, RT-PCR, and Western blotting. Immunohistochemistry was also performed to determine the expression pattern of BMP4, pSMAD1/5/8, Sohlh2, and c-kit in vivo during ovarian development. The results showed treatments of ovaries with BMP4 resulted in a significant (P < 0.05) increase on the primordial-to-primary follicle transition. The oocytes of primordial follicles treated with BMP4 were also less likely to undergo apoptosis. BMP4 enhanced the phosphorylation of SMAD1/5/8 and up-regulated the expression of Sohlh2 and c-kit in primordial follicles. During ovarian development in vivo, Sohlh2, and c-kit exhibited similar expression patterns to BMP4 and pSMAD1/5/8 in primordial follicles. The present studies suggest that BMP4/SMAD signaling pathway initiate primordial follicle growth and prevented oocyte apoptosis via up-regulation of Sohlh2 and c-kit. Copyright © 2012 Wiley Periodicals, Inc.

  1. Stability of cultured dental follicle cells.

    PubMed

    Yao, Shaomian; Norton, Jolanna; Wise, Gary E

    2004-06-01

    Because the dental follicle is required for tooth eruption, establishment of dental follicle cell (DFC) lines is needed for experimentation to determine how the cells regulate eruption. Thus, it is critical that the follicle cells in culture remain stable and neither become transformed nor differentiate. To determine the stability of rat DFC cultures in terms of exhibiting contact inhibition of growth when confluent (no transformation), DFC at different passages were analysed using flow cytometry. Gene expression of cyclin E was determined by reverse transcription polymerase chain reaction as a further method to determine if growth was occurring when the cells were confluent. Alkaline phosphatase and von Kossa staining were also performed as a means of determining stability in terms of differentiation; that is, are the DFC maintaining their phenotype or are they differentiating into osteoblasts and osteocytes? After plating cells of a given passage, they initially underwent a rapid phase of growth with 30-40% of the cells in S, G(2) and M (dividing track) as determined by flow cytometry. The number of such cells declined to only 7-15% at preconfluency. At late confluency, only 2 and 5% of the cells were in the dividing track in passages 6 and 9, respectively, but in passage 12 this had risen to 15%. For a given passage of cells, cyclin E gene expression significantly declined in late confluency as compared to the early growth phase. However, in passage 12, the gene expression of cyclin E at late confluency was higher than the expression at late confluency in passage 6. Thus, the DFC were remarkably stable through passage 9, but by passage 12 it appeared that a small percentage of the cells had become transformed and had lost their contact inhibition growth properties. Alkaline phosphatase and von Kossa staining were negative for all passages, suggesting that the cells remained stable in terms of differentiation and did not differentiate into either osteoblasts or

  2. Assessment of antral grinding of a model solid meal with echo-planar imaging.

    PubMed

    Marciani, L; Gowland, P A; Fillery-Travis, A; Manoj, P; Wright, J; Smith, A; Young, P; Moore, R; Spiller, R C

    2001-05-01

    Mathematical modeling of how physical factors alter gastric emptying is limited by lack of precise measures of the forces exerted on gastric contents. We have produced agar gel beads (diameter 1.27 cm) with a range of fracture strengths (0.15-0.90 N) and assessed their breakdown by measuring their half-residence time (RT(1/2)) using magnetic resonance imaging. Beads were ingested either with a high (HV)- or low (LV)-viscosity liquid nutrient meal. With the LV meal, RT(1/2) was similar for bead strengths ranging from 0.15 to 0.65 N but increased from 22 +/- 2 min (bead strength <0.65 N) to 65 +/- 12 min for bead strengths >0.65 N. With the HV meal, emptying of the harder beads was accelerated. The sense of fullness after ingesting the LV meal correlated linearly (correlation coefficient = 0.99) with gastric volume and was independently increased by the harder beads, which were associated with an increased antral diameter. We conclude that the maximum force exerted by the gastric antrum is close to 0.65 N and that gastric sieving is impaired by HV meals.

  3. Induced pluripotent stem cells from human hair follicle keratinocytes as a potential source for in vitro hair follicle cloning

    PubMed Central

    Lim, Sheng Jye; Ho, Shu Cheow; Mok, Pooi Ling; Tan, Kian Lee; Ong, Alan H.K.

    2016-01-01

    Background Human hair follicles are important for the renewal of new hairs and their development. The generation of induced pluripotent stem cells (iPSCs) from hair follicles is easy due to its accessibility and availability. The pluripotent cells derived from hair follicles not only have a higher tendency to re-differentiate into hair follicles, but are also more suited for growth in hair scalp tissue microenvironment. Methods In this study, human hair follicular keratinocytes were used to generate iPSCs, which were then further differentiated in vitro into keratinocytes. The derived iPSCs were characterised by using immunofluorescence staining, flow cytometry, and reverse-transcription PCR to check for its pluripotency markers expression. Results The iPSC clones expressed pluripotency markers such as TRA-1-60, TRA-1-81, SSEA4, OCT4, SOX2, NANOG, LEFTY, and GABRB. The well-formed three germ layers were observed during differentiation using iPSCs derived from hair follicles. The successful formation of keratioctyes from iPSCs was confirmed by the expression of cytokeratin 14 marker. Discussion Hair follicles represent a valuable keratinocytes source for in vitro hair cloning for use in treating hair balding or grafting in burn patients. Our significant findings in this report proved that hair follicles could be used to produce pluripotent stem cells and suggested that the genetic and micro-environmental elements of hair follicles might trigger higher and more efficient hair follicles re-differentiation. PMID:27867768

  4. Surgical Management of Oro-Antral Communications Using Resorbable GTR Membrane and FDMB Sandwich Technique: A Clinical Study.

    PubMed

    Sandhya, G; Reddy, P Bal; Kumar, K A Jeevan; Sridhar Reddy, B; Prasad, N; Kiran, G

    2013-09-01

    The paper describes a new technique for closure of the oro-antral communication, in which both hard (bone) and soft tissue closure was achieved. The technique uses a Guided Tissue Regeeration (GTR) membrane and Freez Dried Mineralized Bone (FDMB) allograft for closure of the defect. Aim of the study was to assess the advantages of the surgical management of oro-antral communications using resorbable GTR membrane and FDMB sandwich technique. A total 10 patients were selected in whom dental extractions were complicated by formation of oro-antral communication (OAC). The resorbable guided tissue regeneration membrane (PERIOCOL-GTR) and freeze dried mineralized bone allograft material was used. Some cancellous granules of freeze dried bone allograft was sandwiched between sheaths of appropriately trimmed collagen membrane which was previously sutured together on three sides using 3/0 resorbable polyglycolic acid suture (vicryl). The fourth side was then adequately closed using the same suture after the bone graft had been inserted, thus creating a closed sandwich. The prepared sandwich was then tucked into the OAC in such a way that it formed a convexity towards the sinus and a concavity towards the alveolar bone. The rough surface of the sandwich is faced to the alveolar bone and additional bone graft is filled into this concavity. Suturing done without tension. Post-operative orthopantomogram was taken to radiologically quantify the amount of bone grafting/augmentation and closure of oro-antral fistula. There was an average of 11.84 mm bone formation after 6 months, the average width preserved and obtained was 6.9 mm. By the end of 4 months there was evidence of bone formation in 7 subjects and in three subjects bony trabeculae formed was almost similar to the adjacent bone. By the end of 6 months follow-up of 7 subjects showed trabeculae indistinguishable from the adjacent bone. The study was done in 10 patients with a follow-up period of 6 months and found to

  5. Incidence of gastroesophageal reflux disease and positive maxillary antral cultures in children with symptoms of chronic rhinosinusitis.

    PubMed

    Nation, Javan; Kaufman, Michael; Allen, Meredith; Sheyn, Anthony; Coticchia, James

    2014-02-01

    Studies have shown that gastroesophageal reflux disease occurs more frequently than expected in children with chronic rhinosinusitis. The objective of this study is to further understand the relationship of pediatric chronic rhinosinusitis and gastroesophageal reflux disease in children with symptoms of rhinorrhea, nasal congestion, and chronic cough. A retrospective chart review of 63 children, ages 6 months to 10 years old with rhinorrhea, nasal congestion, and chronic cough. The patients underwent maxillary cultures, adenoidectomy, and distal third esophageal biopsies. Children with esophageal biopsies showing esophagitis were classified as positive for gastroesophageal reflux disease, and maxillary antral swabs growing a high density of bacteria were classified as positive for chronic rhinosinusitis. Six months to 5 years old children (n=43), 6 (14%) had simultaneous positive maxillary antral cultures and positive esophageal biopsies, 11 (26%) had positive esophageal biopsies alone, 23 (53%) had positive maxillary antral cultures alone, and 3 (7%) had neither. Six to 10 years old children (n=20), 9 (45%) had simultaneous positive maxillary antral cultures and positive esophageal biopsies, 1 (5%) patient had positive esophageal biopsies alone, 3 (15%) patients had positive maxillary antral cultures alone, and 7 (35%) patients had neither. Twenty-seven (42%) of the patients from the whole study had gastroesophageal reflux positive biopsies. The younger children were statistically likely to have chronic rhinosinusitis and gastroesophageal reflux disease independently of each other (p=0.0002). A direct group comparison found the younger group to have independent chronic rhinosinusitis and gastroesophageal reflux disease and the older group to have simultaneous chronic rhinosinusitis and gastroesophageal reflux disease (p=0.0006). In children with the presenting symptoms of rhinorrhea, nasal congestion, and chronic cough, younger children tend to have either chronic

  6. Involvement of miRNAs in equine follicle development.

    PubMed

    Schauer, S N; Sontakke, S D; Watson, E D; Esteves, C L; Donadeu, F X

    2013-09-01

    Previous evidence from in vitro studies suggests specific roles for a subset of miRNAs, including miR-21, miR-23a, miR-145, miR-503, miR-224, miR-383, miR-378, miR-132, and miR-212, in regulating ovarian follicle development. The objective of this study was to determine changes in the levels of these miRNAs in relation to follicle selection, maturation, and ovulation in the monovular equine ovary. In Experiment 1, follicular fluid was aspirated during ovulatory cycles from the dominant (DO) and largest subordinate (S) follicles of an ovulatory wave and the dominant (DA) follicle of a mid-cycle anovulatory wave (n=6 mares). Follicular fluid levels of progesterone and estradiol were lower (P<0.01) in S follicles than in DO follicles, whereas mean levels of IGF1 were lower (P<0.01) in S and DA follicles than in DO follicles. Relative to DO and DA follicles, S follicles had higher (P≤0.01) follicular fluid levels of miR-145 and miR-378. In Experiment 2, follicular fluid and granulosa cells were aspirated from dominant follicles before (DO) and 24 h after (L) administration of an ovulatory dose of hCG (n=5 mares/group). Relative to DO follicles, L follicles had higher follicular fluid levels of progesterone (P=0.05) and lower granulosa cell levels of CYP19A1 and LHCGR (P<0.005). Levels of miR-21, miR-132, miR-212, and miR-224 were increased (P<0.05) in L follicles; this was associated with reduced expression of the putative miRNA targets, PTEN, RASA1, and SMAD4. These novel results may indicate a physiological involvement of miR-21, miR-145, miR-224, miR-378, miR-132, and miR-212 in the regulation of cell survival, steroidogenesis, and differentiation during follicle selection and ovulation in the monovular ovary.

  7. Gene bionetworks that regulate ovarian primordial follicle assembly.

    PubMed

    Nilsson, Eric; Zhang, Bin; Skinner, Michael K

    2013-07-23

    Primordial follicle assembly is the process by which ovarian primordial follicles are formed. During follicle assembly oocyte nests break down and a layer of pre-granulosa cells surrounds individual oocytes to form primordial follicles. The pool of primordial follicles formed is the source of oocytes for ovulation during a female's reproductive life. The current study utilized a systems approach to detect all genes that are differentially expressed in response to seven different growth factor and hormone treatments known to influence (increase or decrease) primordial follicle assembly in a neonatal rat ovary culture system. One novel factor, basic fibroblast growth factor (FGF2), was experimentally determined to inhibit follicle assembly. The different growth factor and hormone treatments were all found to affect similar physiological pathways, but each treatment affected a unique set of differentially expressed genes (signature gene set). A gene bionetwork analysis identified gene modules of coordinately expressed interconnected genes and it was found that different gene modules appear to accomplish distinct tasks during primordial follicle assembly. Predictions of physiological pathways important to follicle assembly were validated using ovary culture experiments in which ERK1/2 (MAPK1) activity was increased. A number of the highly interconnected genes in these gene networks have previously been linked to primary ovarian insufficiency (POI) and polycystic ovarian disease syndrome (PCOS). Observations have identified novel factors and gene networks that regulate primordial follicle assembly. This systems biology approach has helped elucidate the molecular control of primordial follicle assembly and provided potential therapeutic targets for the treatment of ovarian disease.

  8. Gene bionetworks that regulate ovarian primordial follicle assembly

    PubMed Central

    2013-01-01

    Background Primordial follicle assembly is the process by which ovarian primordial follicles are formed. During follicle assembly oocyte nests break down and a layer of pre-granulosa cells surrounds individual oocytes to form primordial follicles. The pool of primordial follicles formed is the source of oocytes for ovulation during a female’s reproductive life. Results The current study utilized a systems approach to detect all genes that are differentially expressed in response to seven different growth factor and hormone treatments known to influence (increase or decrease) primordial follicle assembly in a neonatal rat ovary culture system. One novel factor, basic fibroblast growth factor (FGF2), was experimentally determined to inhibit follicle assembly. The different growth factor and hormone treatments were all found to affect similar physiological pathways, but each treatment affected a unique set of differentially expressed genes (signature gene set). A gene bionetwork analysis identified gene modules of coordinately expressed interconnected genes and it was found that different gene modules appear to accomplish distinct tasks during primordial follicle assembly. Predictions of physiological pathways important to follicle assembly were validated using ovary culture experiments in which ERK1/2 (MAPK1) activity was increased. Conclusions A number of the highly interconnected genes in these gene networks have previously been linked to primary ovarian insufficiency (POI) and polycystic ovarian disease syndrome (PCOS). Observations have identified novel factors and gene networks that regulate primordial follicle assembly. This systems biology approach has helped elucidate the molecular control of primordial follicle assembly and provided potential therapeutic targets for the treatment of ovarian disease. PMID:23875758

  9. Disruption of the ovarian follicle reservoir of prepubertal rats following prenatal exposure to a continuous 900-MHz electromagnetic field.

    PubMed

    Türedi, Sibel; Hancı, Hatice; Çolakoğlu, Serdar; Kaya, Haydar; Odacı, Ersan

    2016-06-01

    The effects on human health of electromagnetic field (EMF) have begun to be seriously questioned with the entry into daily life of devices establishing EMF, such as cell phones, wireless fidelity, and masts. Recent studies have reported that exposure to EMF, particularly during pregnancy, affects the developing embryo/fetus. The aim of this study was therefore to examine the effects of exposure to continuous 900-Megahertz (MHz) EMF applied in the prenatal period on ovarian follicle development and oocyte differentiation. Six pregnant Sprague Dawley rats were divided equally into a non-exposed control group (CNGr) and a group (EMFGr) exposed to continuous 900-MHz EMF for 1 h daily, at the same time every day, on days 13-21 of pregnancy. New groups were established from pups obtained from both groups after birth. One group consisting of female pups from CNGr rats was adopted as newborn CNGr (New-CNGr, n = 6), and another group consisting of female pups from EMFGr rats was adopted as newborn EMFGr (New-EMFGr, n = 6). No procedure was performed on New-CNGr or New-EMFGr rats. All rat pups were sacrificed on the postnatal 34th day, and their ovarian tissues were removed. Follicle count, histological injury scoring and morphological assessment with apoptotic index criteria were performed with sections obtained following routine histological tissue preparation. Follicle count results revealed a statistically significant decrease in primordial and tertiary follicle numbers in New-EMFGr compared to New-CNGr (p < 0.05), while atretic follicle numbers and apoptotic index levels increased significantly (p < 0.05). Histopathological examination revealed severe follicle degeneration, vasocongestion, a low level of increased stromal fibrotic tissue and cytoplasmic vacuolization in granulosa cell in New-EMFGr. Prenatal exposure to continuous 900-MHz EMF for 1 h each day from days 13-21 led to a decrease in ovarian follicle reservoirs in female rat pups at the

  10. Transcriptome response to hormonal manipulation of follicle-enclosed oocytes in rainbow trout

    USDA-ARS?s Scientific Manuscript database

    Captive fish often display reproductive dysfunction associated with follicle maturation. Gonadotropins and the progestogen maturation-inducing hormones (MIH) are important regulators of follicle maturation; however, their actions including regulating follicle maturation are not fully understood. The...

  11. Movento influences development of granulosa cells and ovarian follicles and FoxO1 and Vnn1 gene expression in BALB/c mice

    PubMed Central

    Kafshgiri, Sakineh Kaboli; Parivar, Kazem; Baharara, Javad; Kerachian, Mohammad Amin; Roodbari, Nasim Hayati

    2016-01-01

    Objective(s): Pesticides has wide range of infertility in female reproductive. This study was done to evaluate the effect of movento pesticide on development of granulosa cells and ovarian follicles and FoxO1 and Vnn1 gene expression in BALB/c mice. Materials and Methods: In this study 40 healthy BALB/c mice 5-6 weeks age were used. Animals were randomly allocated into four groups. Control (without any intervention), three experimental groups received 25, 50 and 100 mg/kg movento dissolved in PBS by gavage for 21 days. Animals scarified after three weeks. For determining the effects of movento on granulosa cells in culture, treatments were conducted to movento (125, 250, 500 μg/ml) for 24 hr. We surveyed the expression of the FoxO1 and Vnn1 in granulosa cells in vitro, and its relation to cell death by flowcytometer and DAPI. Levels of FoxO1 and Vnn1 were analyzed by real-time PCR. Results: Exposure to movento significantly decreased ovarian weight and the number of primary, secondary and antral follicles. Further, treatment with different concentration of movento induced apoptosis on granulosa cells. Gene expression analysis showed the transcriptional expression of FoxO1 and vnn1 in granulosa cells. Level of Vnn1 mRNA in granulosa cells was decreased in granulosa cells and expression of FoxO1 significantly increased in treated groups in compare to controls (P-value <0.05). Conclusion: Exposure to movento significantly reduced the number of follicles and increased apoptosis of granulosa cells leading disruption of the reproductive system. Also movento reduced expression of Vnn1 and increased FoxO1 genes in a dose dependent manner. PMID:27917277

  12. Conjunctival lymphoid follicles in new world rodents.

    PubMed

    Astley, Roger A; Chodosh, James; Caire, William; Wilson, Gregory M

    2007-09-01

    We report for the first time, the detection of conjunctival lymphoid follicles (CLF) in the eyes of New World rodents. CLF were found in 7 of the 15 species examined, 6 of the 10 genera, and in at least one individual in four families of rodents. These follicles are dense collections of leukocytes in the conjunctival substantia propria with a thinned overlying epithelium lacking in goblet cells. Although the precise location of CLF within the conjunctiva varied from species to species, all CLF were found in the fornix of the conjunctival sac. In general, size and complexity of CLF varied with the size of the eye; the larger the eye, the larger and more complex the CLF. Our findings also reveal that some species of New World rodents, like the majority of Old World rodents examined in this and previous studies might lack CLF. However, until larger samples are examined, this is difficult to state with certainty. Consequently, the presence/absence of CLF at this point might not be informative for phylogenetic comparisons. Our findings also suggest the deer mouse, Peromyscus maniculatus, might serve as a useful model species for studying ocular infections and immunology of the eye.

  13. 1 Secondary follicle growth and oocyte maturation by culture in alginate hydrogel following cryopreservation of the ovary or individual follicles

    PubMed Central

    Xu, Min; Banc, Anna; Woodruff, Teresa K.; Shea, Lonnie D.

    2009-01-01

    An option for fertility preservation for women facing a cancer diagnosis involves the cryopreservation of ovarian tissue for later re-transplantation or in vitro culture, with in vitro culture preferred to avoid reintroduction of the cancer. Small, immature follicles survive the freeze-thaw process, and can be matured through in follicle maturation (IFM) that involves an initial growth of the follicle and subsequent maturation of the oocyte. The ovarian tissue can be cryopreserved in two forms: (i) cortical strips consisting of follicles and surrounding stroma (Cryo-Ov) or (ii) individually isolated follicles (Cryo-In). The aim of this study was to assess the follicle growth and oocyte maturation for follicles that were cryopreserved either as strips or individually using a slow-freezing cryopreservation method. The two follicle groups, together with non-cryopreserved control follicles, were grown in an alginate-based three-dimensional culture system for twelve days. The overall survival, size increase and antrum formation rates were comparable among the three groups. At day 12 of culture, Androstenedione levels were decreased in the Cryo-Ov group relative to the other two, and the ratio of progesterone to estradiol was increased in the two cryopreserved groups relative to the control. Both Gja1 (known as connexin 43) and Gja4 (known as connexin 37) mRNA expression were decreased at day 6 in the cryopreserved groups relative to controls, and by day 12, Gja1 was similar for all 3 groups. Moreover, Cryo-In resulted in lower GVBD rate indicating some impaired oocyte development. Overall, the present study demonstrated that mouse preantral follicles, either within ovarian tissues or individually isolated, could be successfully cryopreserved by the slow-freezing method, as evidenced by post-thaw follicle development and steroidgenesis, oocyte maturation and molecular markers for oocyte and/or granulosa cells connection. PMID:19191350

  14. Involvement of Notch signaling in early chick ovarian follicle development.

    PubMed

    Li, Jun; Zhao, Dan; Guo, Changquan; Li, Jian; Mi, Yuling; Zhang, Caiqiao

    2016-01-01

    The formation of primordial follicles is a crucial process in the establishment of follicle pools required for the female's reproductive life span. For laying hens, ample follicles are a prerequisite for high laying performance. Notch signaling plays critical roles in germ cell cysts breakdown and in the formation of primordial follicles. Here, we investigated the role of Notch signaling in the ovarian development of post-hatch chicks. Results showed that around post-hatch day 4 (H4), the germ cell cysts broke apart, oocytes became surrounded by squamous pregranulosa cells, and the primordial follicles were then formed. Subsequently, we detected the expression of Notch signaling-related genes including Notch receptors (Notch1, 2), ligands (Jag1, 2 and Dll1, 4), and target genes (Hes1, Hey1). These genes all showed expression at H4 and some of these genes were up-regulated during primordial follicle formation. To evaluate the Notch signaling requirement for early follicular development, we adopted an in vitro ovary culture system. Suppression of Notch signaling by γ-secretase inhibitor induced a decrease of primordial follicles and an increase of germ cells in cysts. Attenuating Notch signaling also inhibited the phosphatidylinositol 3-kinase/protein kinase B pathways and suppressed cadherin expression. These results suggest that Notch signaling is endowed with an indispensable role in primordial follicle formation in post-hatch chicks.

  15. Superficially located enlarged lymphoid follicles characterise nodular gastritis.

    PubMed

    Okamura, Takuma; Sakai, Yasuhiro; Hoshino, Hitomi; Iwaya, Yugo; Tanaka, Eiji; Kobayashi, Motohiro

    2015-01-01

    Nodular gastritis is a form of chronic Helicobacter pylori gastritis affecting the gastric antrum and characterised endoscopically by the presence of small nodular lesions resembling gooseflesh. It is generally accepted that hyperplasia of lymphoid follicles histologically characterises nodular gastritis; however, quantitative analysis in support of this hypothesis has not been reported. Our goal was to determine whether nodular gastritis is characterised by lymphoid follicle hyperplasia.The number, size, and location of lymphoid follicles in nodular gastritis were determined and those properties compared to samples of atrophic gastritis. The percentages of high endothelial venule (HEV)-like vessels were also evaluated.The number of lymphoid follicles was comparable between nodular and atrophic gastritis; however, follicle size in nodular gastritis was significantly greater than that seen in atrophic gastritis. Moreover, lymphoid follicles in nodular gastritis were positioned more superficially than were those in atrophic gastritis. The percentage of MECA-79 HEV-like vessels was greater in areas with gooseflesh-like lesions in nodular versus atrophic gastritis.Superficially located hyperplastic lymphoid follicles characterise nodular gastritis, and these follicles correspond to gooseflesh-like nodular lesions observed endoscopically. These observations suggest that MECA-79 HEV-like vessels could play at least a partial role in the pathogenesis of nodular gastritis.

  16. Steroid signaling in mature follicles is important for Drosophila ovulation

    PubMed Central

    Knapp, Elizabeth

    2017-01-01

    Although ecdysteroid signaling regulates multiple steps in oogenesis, it is not known whether it regulates Drosophila ovulation, a process involving a matrix metalloproteinase-dependent follicle rupture. In this study, we demonstrated that ecdysteroid signaling is operating in mature follicle cells to control ovulation. Moreover, knocking down shade (shd), encoding the monooxygenase that converts ecdysone (E) to the more active 20-hydroxyecdysone (20E), specifically in mature follicle cells, blocked follicle rupture, which was rescued by ectopic expression of shd or exogenous 20E. In addition, disruption of the Ecdysone receptor (EcR) in mature follicle cells mimicked shd-knockdown defects, which were reversed by ectopic expression of EcR.B2 but not by EcR.A or EcR.B1 isoforms. Furthermore, we showed that ecdysteroid signaling is essential for the proper activation of matrix metalloproteinase 2 (Mmp2) for follicle rupture. Our data strongly suggest that 20E produced in follicle cells before ovulation activates EcR.B2 to prime mature follicles to be responsive to neuronal ovulatory stimuli, thus providing mechanistic insights into steroid signaling in Drosophila ovulation. PMID:28069934

  17. 21 CFR 522.1002 - Follicle stimulating hormone.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Follicle stimulating hormone. 522.1002 Section 522....1002 Follicle stimulating hormone. (a)(1) Specifications. Each package contains 2 vials. One vial... hormone. The other vial contains 10 milliliters of aqueous diluent. (2) Sponsor. See 059521 in §...

  18. 21 CFR 522.1002 - Follicle stimulating hormone.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Follicle stimulating hormone. 522.1002 Section 522....1002 Follicle stimulating hormone. (a)(1) Specifications. Each package contains 2 vials. One vial... hormone. The other vial contains 10 milliliters of aqueous diluent. (2) Sponsor. See No. 052923 in §...

  19. 21 CFR 522.1002 - Follicle stimulating hormone.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Follicle stimulating hormone. 522.1002 Section 522....1002 Follicle stimulating hormone. (a)(1) Specifications. Each package contains 2 vials. One vial... hormone. The other vial contains 10 milliliters of aqueous diluent. (2) Sponsor. See No. 052923 in §...

  20. 21 CFR 522.1002 - Follicle stimulating hormone.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Follicle stimulating hormone. 522.1002 Section 522....1002 Follicle stimulating hormone. (a)(1) Specifications. Each package contains 2 vials. One vial... hormone. The other vial contains 10 milliliters of aqueous diluent. (2) Sponsor. See No. 052923 in §...

  1. 21 CFR 522.1002 - Follicle stimulating hormone.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Follicle stimulating hormone. 522.1002 Section 522....1002 Follicle stimulating hormone. (a)(1) Specifications. Each package contains 2 vials. One vial... hormone. The other vial contains 10 milliliters of aqueous diluent. (2) Sponsor. See No. 052923 in §...

  2. Progesterone Regulation of Primordial Follicle Assembly In Bovine Fetal Ovaries

    PubMed Central

    Nilsson, Eric E.; Skinner, Michael K.

    2009-01-01

    Fertility in mammals is dependant on females having an adequate primordial follicle pool to supply oocytes for fertilization. The formation of primordial follicles is called ovarian follicular assembly. In rats and mice progesterone and estradiol have been shown to inhibit follicle assembly with assembly occurring after birth when the pups are removed from the high-steroid maternal environment. In contrast, primordial follicle assembly in other species, such as cattle and humans, occurs during fetal development before birth. The objective of the current study is to determine if progesterone levels regulate primordial follicle assembly in fetal bovine ovaries. Ovaries and blood were collected from bovine fetuses. Interestingly, ovarian progesterone and estradiol concentrations were found to decrease with increasing fetal age and correlated to increased primordial follicle assembly. Microarray analysis of fetal ovary RNA suggests that progesterone membrane receptor and estrogen nuclear receptor are expressed. Treatment of fetal bovine ovary cultures with a higher progesterone concentration significantly decreased primordial follicle assembly. Observations indicate that progesterone affects ovarian primordial follicle assembly in cattle, as it does in rats and mice. PMID:19747959

  3. Interrelationships between ovarian follicles grown in culture and possible mediators.

    PubMed

    Sirotkin, Alexander V; Florkovičová Koničková, Iveta; Schaeffer, Hans-Jorg; Laurincik, Jozef; Harrath, Abdel Halim

    2017-03-01

    Dominance or cooperation between ovarian follicles can determine the number of ovulations and fecundity, but interrelationships between follicles in mono- and poly-ovulatory species and their mechanisms are poorly understood. The goals of this work were to determine the existence and compare the character of mutual influence of cultured ovarian follicles from a mono-ovulatory species (cow) with established follicular dominance with those from a poly-ovulatory species (pig), in which interrelationship between follicles remain unknown, and to examine the role of ovarian cell proliferation, the insulin-like growth factor I (IGF-I)- oxytocin (OT) system, and steroid hormones in mediating interrelationships among ovarian follicles. Bovine and porcine ovarian follicles were isolated and cultured alone and in pairs, and the percentage of growing follicles was calculated. Porcine follicles were cultured alone and in pairs after addition of exogenous OT and IGF-I (100ngmL(-1)) or inactivation of endogenous OT and IGF-I by antisera against these hormones (1%). Proliferation of porcine follicular cells was assessed by SDS PAGE-Western immunoblotting, the release of IGF-I, progesterone, androstenedione and estradiol by cultured porcine ovarian follicles was analyzed by RIA/EIA. Overall, our observations suggest (1) competition/dominance (mutual suppression of growth) in bovine ovarian follicles, (2) cooperation (mutual support of growth) in porcine ovarian follicles, (3) that this mutual growth of porcine ovarian follicles was caused by the promotion of cell proliferation, (4) that this mechanism was probably not involved in bovine follicular dominance, (5) that communication between both porcine and bovine follicles affects their secretory activity, and (6) that both follicular dominance in cows and cooperation of follicles in pigs can be mediated by either down- or up-regulation of the IGF-I-OT system, which in turn affects follicular steroidogenesis and promotes follicular

  4. Mesenchymal-epithelial interactions during hair follicle morphogenesis and cycling

    PubMed Central

    Sennett, Rachel; Rendl, Michael

    2012-01-01

    Embryonic hair follicle induction and formation are regulated by mesenchymal-epithelial interactions between specialized dermal cells and epidermal stem cells that switch to a hair fate. Similarly, during postnatal hair growth, communication between mesenchymal dermal papilla cells and surrounding epithelial matrix cells coordinates hair shaft production. Adult hair follicle regeneration in the hair cycle again is thought to be controlled by activating signals originating from the mesenchymal compartment and acting on hair follicle stem cells. Although many signaling pathways are implicated in hair follicle formation and growth, the precise nature, timing, and intersection of these inductive and regulatory signals remains elusive. The goal of this review is to summarize our current understanding and to discuss recent new insights into mesenchymal-epithelial interactions during hair follicle morphogenesis and cycling. PMID:22960356

  5. Piscine follicle-stimulating hormone triggers progestin production in gilthead seabream primary ovarian follicles.

    PubMed

    Zapater, Cinta; Chauvigné, François; Scott, Alexander P; Gómez, Ana; Katsiadaki, Ioanna; Cerdà, Joan

    2012-11-01

    Ovarian growth (vitellogenesis) in most lower vertebrates is mediated by estradiol-17beta (E2) secreted by the follicles in response to follicle-stimulating hormone (Fsh), whereas oocyte maturation and ovulation are mediated by progestins, such as 17alpha,20beta-dihydroxypregn-4-en-3-one (17,20beta-P), produced in response to luteinizing hormone (Lh). In teleosts, follicular synthesis of 17,20beta-P at the time of maturation is due primarily to up-regulation of the enzymes P450c17-II (Cyp17a2) and 20beta-hydroxysteroid dehydrogenase (Cbr1). Here, we show that follicular cells associated with primary growth (previtellogenic) oocytes of the gilthead seabream also express cyp17a2 and cbr1, in addition to P450c17-I (cyp17a1) and aromatase (cyp19a1), enzymes required for E2 synthesis. Ovaries containing only oogonia and early primary ovarian follicles had a 60-fold higher concentration of 17,20beta-P than ovaries in the succeeding stages and had a higher expression of cbr1 and Fsh receptor (fshra). Stimulation of explants of primary follicles in vitro with recombinant piscine Fsh (rFsh), which specifically activates the seabream Fshra, promoted a rapid accumulation of 17,20beta-P, and synthesis was sustained by an external supply of 17alpha-hydroxyprogesterone. In the presence of Cbr1 inhibitors, rFsh-mediated 17,20beta-P production was reduced, with a concomitant increase in testosterone and E2 synthesis. In primary explants, rFsh up-regulated cyp17a2 and cbr1 transcription and simultaneously down-regulated cyp17a1 and cyp19a1 steady-state mRNA levels within 24 h. In contrast, in explants containing vitellogenic follicles, rFsh had no effect on cyp17a2 and cbr1 expression, but increased that of cyp17a1 and cyp19a1. These data suggest a functional Fshra-activated Cyp17a2/Cbr1 steroidogenic pathway in gilthead seabream primary ovarian follicles triggering the production of 17,20beta-P.

  6. Basic fibroblast growth factor promotes macaque follicle development in vitro.

    PubMed

    Lu, C L; Yan, J; Zhi, X; Xia, X; Wang, T R; Yan, L Y; Yu, Y; Ding, T; Gao, J M; Li, R; Qiao, J

    2015-05-01

    Fertility preservation is an important type of frontier scientific research in the field of reproductive health. The culture of ovarian cortices to i) initiate primordial follicle growth and ii) procure developing follicles for later oocyte maturation is a promising fertility preservation strategy, especially for older women or cancer patients. At present, this goal remains largely unsubstantiated in primates because of the difficulty in attaining relatively large follicles via ovarian cortex culture. To overcome this hurdle, we cultured macaque monkey ovarian cortices with FSH, kit ligand (KL), basic fibroblast growth factor (bFGF), and/or epidermal growth factor (EGF). The various factors and factor combinations promoted primordial follicle development to different extents. Notably, both bFF (bFGF, 100 ng/ml and FSH, 50 ng/ml) and KF (KL, 100 ng/ml and FSH, 50 ng/ml) contributed to the activation of primordial follicles at day 12 (D12) of culture, whereas at D18, the proportions of developing follicles were significantly higher in the bFF and KF groups relative to the other treatment groups, particularly in the bFF group. Estradiol and progesterone production were also highest in the bFF group, and primary follicle diameters were the largest. Up until D24, the bFF group still exhibited the highest proportion of developing follicles. In conclusion, the bFGF-FSH combination promotes nonhuman primate primordial follicle development in vitro, with the optimal experimental window within 18 days. These results provide evidence for the future success of human ovarian cortex culture and the eventual acquisition of mature human follicles or oocytes for fertility restoration. © 2015 Society for Reproduction and Fertility.

  7. Ultrasonographic characterization of follicle deviation in follicular waves with single dominant and codominant follicles in dromedary camels (Camelus dromedarius).

    PubMed

    Manjunatha, B M; Al-Bulushi, S; Pratap, N

    2014-04-01

    Follicular wave emergence was synchronized by treating camels with GnRH when a dominant follicle (DF) was present in the ovaries. Animals were scanned twice a day from day 0 (day of GnRH treatment) to day 10, to characterize emergence and deviation of follicles during the development of the follicular wave. Follicle deviation in individual animals was determined by graphical method. Single DFs were found in 16, double DFs in 9 and triple DFs in two camels. The incidence of codominant (double and triple DFs) follicles was 41%. The interval from GnRH treatment to wave emergence, wave emergence to deviation, diameter and growth rate of F1 follicle before or after deviation did not differ between the animals with single and double DFs. The size difference between future DF(s) and the largest subordinate follicle (SF) was apparent from the day of wave emergence in single and double DFs. Overall, interval from GnRH treatment to wave emergence and wave emergence to the beginning of follicle deviation was 70.6 ± 1.4 and 58.6 ± 2.7 h, respectively. Mean size of the DF and largest SF at the beginning of deviation was 7.4 ± 0.2 and 6.3 ± 0.1 mm, respectively. In conclusion, the characteristics of follicle deviation are similar between the animals that developed single or double DFs.

  8. Impact of growth hormone (GH) and follicle stimulating hormone (FSH) on in vitro canine preantral follicle development and estradiol production.

    PubMed

    Serafim, M K B; Duarte, A B G; Silva, G M; Souza, C E A; Magalhães-Padilha, D M; Moura, A A A; Silva, L D M; Campello, C C; Figueiredo, J R

    2015-04-01

    Evaluate the effect of different concentrations of growth hormone (GH) on the in vitro development of domestic dog (Canis lupus familiaris) preantral follicles in the presence or absence of follicle stimulating hormone (FSH). Secondary preantral follicles, isolated by microdissection, were cultured in a medium composed of αMEM with bovine serum albumin (BSA), glutamine, hypoxanthine, insulin, transferrin, selenium and ascorbic acid (αMEM(+)-control) added at different concentrations of GH (GH10 ng/ml or GH50 ng/ml) and FSH (GH10+FSH, GH50+FSH). Follicle development was evaluated based on the percentage of intact follicles, antrum formation, follicular diameter, follicular viability using fluorescent markers and estradiol production. GH50 was the only treatment that maintained the same percentage of normal morphologically follicles from day 0 to day 18 of culture (P<0.05). For all treatments, except the control, follicles were viable throughout the 18 days of culture (P<0.05). GH50 supplemented with FSH (GH50+FSH) resulted in the highest average follicular diameter (P<0.05) from day 12 to 18. Follicles from both the control and the GH50+FSH treatment groups actively and increasingly secreted estradiol from day 6 to 18 of culture (P<0.05). Our study demonstrates that GH benefits the maintenance of follicular morphology in a dose-dependent manner and, in association with FSH, stimulates in vitro follicular growth and estradiol production. Copyright © 2015 Elsevier Ltd. All rights reserved.

  9. Developmental effects of imatinib mesylate on follicle assembly and early activation of primordial follicle pool in postnatal rat ovary.

    PubMed

    Asadi-Azarbaijani, Babak; Santos, Regiane R; Jahnukainen, Kirsi; Braber, Saskia; van Duursen, Majorie B M; Toppari, Jorma; Saugstad, Ola D; Nurmio, Mirja; Oskam, Irma C

    2017-03-01

    Imatinib mesylate is an anti-cancer agent that competitively inhibits several receptor tyrosine kinases (RTKs). RTKs play important roles in the regulation of primordial follicle formation, the recruitment of primordial follicles into the pool of growing follicles and maturation of the follicles. In the present study, we investigated the effects of the tyrosine kinase inhibitor imatinib on primordial follicle assembly and early folliculogenesis in postnatal rats. Female Sprague-Dawley rats were treated with either imatinib (150mg/kg) or placebo (water) on postnatal days 2-4. Bilateral ovariectomy was performed on postnatal day 2 and 5. Histology, immunohistochemistry, and mRNA analysis were performed. Imatinib treatment was associated with increased density of the multi-oocyte follicles (P<0.01), oogonia (p<0.01) and germline clusters (P<0.05), decreased activation of primordial follicles, increased expression of c-Kit and AMH, and decreased protein expression of Kit-ligand and GDF9 when compared to age-matched controls. In conclusion, imatinib affects folliculogenesis in postnatal rat ovaries by delaying the cluster breakdown, follicular assembly and early activation of the primordial follicle pool. Copyright © 2016 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

  10. Morphogenesis of polycystic ovaries as assessed by pituitary-ovarian androgenic function.

    PubMed

    Mori, Takahide; Nonoguchi, Kousuke; Watanabe, Hirohiko; Ishikawa, Hironobu; Tamura, Izuru; Kinoshita, Katsuji

    2009-05-01

    Despite polycystic ovaries (PCO) being a common morphology in women with polycystic ovary syndrome and regular menstruation, the regulatory principles in the morphogenesis of antral follicles have not yet been elucidated. In recognition of the complementary interaction between androgen-induced expression of the FSH receptor and FSH-augmented expression of the androgen receptor in granulose cells of antral follicles, a possible correlation of antral follicle count (AFC) and pituitary-ovarian androgenic function was investigated in 180 infertile women over days 3-5 of the menstrual cycle. Six discrete types of PCO with decreasing pituitary-ovarian androgenic function were identified: Type I (classical Stein-Leventhal syndrome), Type II (hyperandrogenemism), Type III (singular hyper-LH), Type IV (cryptic hyperandrogenism), Type V (relative LH dominancy) and Type VI (relative FSH dominancy), in parallel to a diminishing number of AFC from Type I to Type VI. Because during the early follicular phase of the cycle until the selection of the dominant follicle, antral follicles are composed of newly emerged healthy follicles plus atretic antral follicles that remain non-ovulated from previous cycles, it is proposed that the six types of PCO may represent the folliculogenetic spectra along which PCO morphogenesis proceeds.

  11. Randomized comparison between pulmonary vein antral isolation versus complex fractionated electrogram ablation for paroxysmal atrial fibrillation.

    PubMed

    Chen, Minglong; Yang, Bing; Chen, Hongwu; Ju, Weizhu; Zhang, Fengxiang; Tse, Hung-Fat; Cao, Kejiang

    2011-09-01

    Circumferential pulmonary vein antral isolation (PVAI) and atrial complex fractionated electrograms (CFEs) are both ablative techniques for the treatment of paroxysmal atrial fibrillation (PAF). However, data on the comparative value of these 2 ablation strategies are very limited. We randomized 118 patients with drug-refractory PAF to receive PVAI ablation (n = 60) or CFE ablation (n = 58). For CFE group, spontaneous/induced AF was mapped using validated, automated software to guide ablation until all CFE areas were eliminated. For PVAI group, all 4 pulmonary vein antra were electrically isolated as confirmed by circular mapping catheter. Patients with spontaneous/inducible AF after the initial ablation procedure were crossed over to the other arms. After initial ablation procedure, AF persisted/inducible in 24/59 patients (41%), and 34/58 patients (59%) assigned to PVAI and CFE ablation, respectively (P = 0.05). Then 58 patients underwent PVAI + CFE ablation. After 22.6 ± 6.4 months, PVAI ablation group was more likely than CFE ablation group to achieve control of any AF/atrial tachycardia (AT) off drugs (43/60, 72% vs 33/58, 57%, P = 0.075) and lower recurrence rate of AT (11.9% vs 34.5%, P = 0.004). Patients who received CFE ablation alone (38%) had significantly lower overall success rate to achieve control of AF/AT off drugs compared with patients who received PVAI ablation (77%, P = 0.002) alone or PVAI + CFE ablation (69%, P = 0.008) due to higher recurrence rate of AT (50% vs 6% vs 13%, P < 0.01). CFE ablation in PAF patients was associated with higher occurrence rate of postprocedure AT compared with PVAI ablation, whereby making it less likely to be a sole ablation strategy for PAF patients.  © 2011 Wiley Periodicals, Inc.

  12. Treatment of gastropathy and gastric antral vascular ectasia in patients with portal hypertension.

    PubMed

    Ripoll, Cristina; Garcia-Tsao, Guadalupe

    2007-12-01

    Portal hypertensive gastropathy (PHG) and gastric antral vascular ectasia (GAVE) are two distinct gastric mucosal lesions that may cause acute and/or chronic upper gastrointestinal hemorrhage in patients with cirrhosis. Whereas PHG is associated with portal hypertension, GAVE may present in patients without portal hypertension or liver disease. Diagnosis is made upon visualization of the characteristic lesions with upper gastrointestinal endoscopy, although the differential may be difficult at times. PHG is characterized endoscopically by a mosaic pattern with or without red signs and a proximal distribution. PHG mainly causes chronic blood loss and anemia in patients with cirrhosis but also can cause acute hemorrhage. First-line therapy for chronic hemorrhage from PHG is a nonselective beta-blocker (propranolol or nadolol) and iron supplementation. If bleeding/anemia are not controlled with these measures and the patient is transfusion-dependent, shunt therapy (transjugular intrahepatic portosystemic shunt or shunt surgery) should be considered. Management of acute bleeding from PHG, an infrequent event, should be accomplished with a vasoactive drug, somatostatin (or its analogues) or terlipressin. If bleeding responds, the patient must be switched to a nonselective beta-blocker. Shunt therapy should be considered in patients who rebleed or continue to bleed despite adequate beta-blocker therapy. GAVE is less common than PHG. It is characterized by red spots without a background mosaic pattern, typically in the gastric antrum. When lesions have a linear distribution, the lesion is called "watermelon stomach." GAVE is a cause of chronic gastrointestinal bleeding and anemia in patients with cirrhosis. If lesions are localized, first-line therapy is argon plasma coagulation. In more diffuse lesions, therapy with argon plasma coagulation is more complicated. Preliminary data suggest that cryotherapy may be a reasonable option for diffuse GAVE lesions. Neither beta

  13. Effect of luteinizing hormone overstimulation on equine follicle maturation.

    PubMed

    Schauer, S N; Guillaume, D; Decourt, C; Watson, E D; Briant, C; Donadeu, F X

    2013-02-01

    There is evidence in several species that high circulating LH concentrations can interfere with normal follicle development and ovulation. In the mare, high LH levels after induction of luteolysis with PGF(2α) have been temporally associated with an increased incidence of anovulatory follicles. We hypothesized that a premature increase in LH levels during a follicular wave in mares would disrupt normal follicle maturation leading to ovulatory dysfunction. In experiment 1, all follicles >10 mm were ablated at midestrous cycle in pony mares followed by twice daily administration of equine LH (eLH; 1.6 μg/kg body weight) or saline (vehicle; N = 8 mares per group). When a dominant follicle reached >32 mm, an ovulatory dose of hCG was given. Treatment with eLH had no effects on ovulatory responses or progesterone levels during the posttreatment luteal phase. In experiment 2, after follicle ablation, mares were treated with eLH or vehicle (as above) or were given a single injection of PGF(2α) (N = 7 mares per group), followed by aspiration of a dominant follicle when it reached >32 mm. Administration of eLH induced an increase in circulating LH levels similar to that after PGF(2α) injection. Neither PGF(2α) nor eLH administration had significant effects on follicle growth or total number of follicles in the postablation wave. However, compared with mares treated with vehicle, the preovulatory follicle in the eLH and PGF(2α) groups had lower levels of androstenedione (P = 0.03) and higher levels of insulin-like growth factor I (P = 0.03). Further, levels of prostaglandin E2 in preovulatory follicles tended to be lower in the eLH and PGF(2α) groups (P = 0.06). In conclusion, exposure of developing follicles to high LH in mares did not have apparent effects on ovulation but it induced changes in follicular fluid factor levels which might reflect a disruption in follicle and/or oocyte maturation, indicating the need to further study the implications of using PGF(2

  14. Hair counts from normal scalp biopsy in Taiwan.

    PubMed

    Ko, Jui-Hung; Huang, Yu-Huei; Kuo, Tseng-Tong

    2012-09-01

    Horizontal scalp biopsy is a better method to use when evaluating hair disorders, but an accurate diagnosis requires quantitative data regarding normal hair counts to be used as a control, which are currently available only for Caucasians, African Americans, Koreans, and Iranians. To evaluate scalp hair counts in the Taiwanese population. Two 4-mm punch biopsies were taken from clinically normal-looking scalp skin of 31 patients who underwent scalp surgery for non-hair-loss disorders. Punch biopsy specimens were sectioned horizontally at the sebaceous gland level and the number of follicular units, terminal hairs including anagen and telogen hairs, and vellus hairs were counted and compared with other published data. The numbers of hairs were also compared between the sexes, different age groups, and different anatomical locations of the scalp. The average counts of total hairs, terminal hairs, vellus hairs, and follicular units per 4-mm punch scalp skin were 21.3, 20.5, 0.8, and 9.4, respectively. The mean ratio of anagen hair to telogen hair was 91.6:8.4 and of terminal hair to vellus hair was 25.3:1. The average density of hair follicles was 1.69 mm(2). The mean counts of terminal hairs, total hairs, follicular units, and hair follicles/mm(2) were significantly different between different age groups. Differences in hair count parameters were recognized between different ethnic groups. Parameters of hair density established in the Taiwanese population will provide clinicians with useful reference data for the evaluation of hair disorders in Asians. © 2012 by the American Society for Dermatologic Surgery, Inc. Published by Wiley Periodicals, Inc.

  15. Dental follicle infection following a dog bite.

    PubMed

    Wright, G; Muir, M L; Bryan, R; Smith, A J; Hosey, M T

    2006-03-01

    Animal bite wounds and their subsequent infection are relatively common. Incidence rates for dog bites are significantly higher among children aged 0-9 years, especially among boys. Although bite wounds may initially look innocuous, they frequently lead to serious infection with a potential for life-threatening complications. The microbiology of dog bite wounds is usually polymicrobial, typically including anaerobes, Staphylococcus aureus and Pasteurella species. A case is described of a 22-month-old boy who, subsequent to a dog bite over the left maxilla, suffered infection of the dental follicle of the primary maxillary canine with Pasteurella multocida. The infection proved difficult to treat, requiring several attempts at incision and drainage of the abscess together with systemic antibiotics, and resulted in the eventual loss of the tooth.

  16. Isosmotic modulation of Ca2+-regulated exocytosis in guinea-pig antral mucous cells: role of cell volume.

    PubMed

    Fujiwara, S; Shimamoto, C; Katsu, K i; Imai, Y; Nakahari, T

    1999-04-01

    1. Exocytotic events and changes of cell volume in mucous cells from guinea-pig antrum were examined by video-enhanced optical microscopy. 2. Acetylcholine (ACh) evoked exocytotic events following cell shrinkage, the frequency and extent of which depended on the ACh concentration. ACh actions were mimicked by ionomycin and thapsigargin, and inhibited by Ca2+-free solution and Ca2+ channel blockers (Ni2+, Cd2+ and nifedipine). Application of 100 microM W-7, a calmodulin inhibitor, also inhibited the ACh-induced exocytotic events. These results indicate that ACh actions are mediated by intracellular Ca2+ concentration ([Ca2+]i) in antral mucous cells. 3. The effects of ion channel blockers on exocytotic events and cell shrinkage evoked by ACh were examined. Inhibition of KCl release (quinine, Ba2+, NPPB or KCl solution) suppressed both the exocytotic events and cell shrinkage evoked by ACh. 4. Bumetanide (inhibition of NaCl entry) or Cl--free solution (increasing Cl- release and inhibition of NaCl entry) evoked exocytotic events following cell shrinkage in unstimulated antral mucous cells and caused further cell shrinkage and increases in the frequency of exocytotic events in ACh-stimulated cells. However, Cl--free solution did not evoke exocytotic events in unstimulated cells in the absence of extracellular Ca2+, although cell shrinkage occurred. 5. To examine the effects of cell volume on ACh-evoked exocytosis, the cell volume was altered by increasing the extracellular K+ concentration. The results showed that cell shrinkage increases the frequency of ACh-evoked exocytotic events and cell swelling decreases them. 6. Osmotic shrinkage or swelling caused the frequency of ACh-evoked exocytotic events to increase. This suggests that the effects of cell volume on ACh-evoked exocytosis under anisosmotic conditions may not be the same as those under isosmotic conditions. 7. In antral mucous cells, Ca2+-regulated exocytosis is modulated by cell shrinkage under isosmotic

  17. Transcriptome based identification of mouse cumulus cell markers that predict the developmental competence of their enclosed antral oocytes

    PubMed Central

    2013-01-01

    Background The cumulus cells (CCs) enveloping antral and ovulated oocytes have been regarded as putative source of non-invasive markers of the oocyte developmental competence. A number of studies have indeed observed a correlation between CCs gene expression, embryo quality, and final pregnancy outcome. Here, we isolated CCs from antral mouse oocytes of known developmental incompetence (NSN-CCs) or competence (SN-CCs) and compared their transcriptomes with the aim of identifying distinct marker transcripts. Results Global gene expression analysis highlighted that both types of CCs share similar transcriptomes, with the exception of 422 genes, 97.6% of which were down-regulated in NSN-CCs vs. SN-CCs. This transcriptional down-regulation in NSN-CCs was confirmed by qRT-PCR analysis of CC-related genes (Has2, Ptx3, Tnfaip6 and Ptgs2). Only ten of the 422 genes were up-regulated with Amh being the most up-regulated in NSN-CCs, with an average 4-fold higher expression when analysed by qRT-PCR. Conclusions The developmental incompetence (NSN) or competence (SN) of antral oocytes can be predicted using transcript markers expressed by their surrounding CCs (i.e., Has2, Ptx3, Tnfaip6, Ptgs2 and Amh). Overall, the regulated nature of the group of genes brought out by whole transcriptome analysis constitutes the molecular signature of CCs associated either with developmentally incompetent or competent oocytes and may represent a valuable resource for developing new molecular tools for the assessment of oocyte quality and to further investigate the complex bi-directional interaction occurring between CCs and oocyte. PMID:23758669

  18. Human hair growth ex vivo is correlated with in vivo hair growth: selective categorization of hair follicles for more reliable hair follicle organ culture.

    PubMed

    Kwon, Oh Sang; Oh, Jun Kyu; Kim, Mi Hyang; Park, So Hyun; Pyo, Hyun Keol; Kim, Kyu Han; Cho, Kwang Hyun; Eun, Hee Chul

    2006-02-01

    Of the numerous assays used to assess hair growth, hair follicle organ culture model is one of the most popular and powerful in vitro systems. Changes in hair growth are commonly employed as a measurement of follicular activity. Hair cycle stage of mouse vibrissa follicles in vivo is known to determine subsequent hair growth and follicle behavior in vitro and it is recommended that follicles be taken at precisely the same cyclic stage. This study was performed to evaluate whether categorization of human hair follicles by the growth in vivo could be used to select follicles of the defined anagen stage for more consistent culture. Occipital scalp samples were obtained from three subjects, 2 weeks later after hair bleaching. Hair growth and follicle length of isolated anagen VI follicles were measured under a videomicroscope. Follicles were categorized into four groups according to hair growth and some were cultured ex vivo for 6 days. Follicles showed considerable variations with respect to hair growth and follicle length; however, these two variables were relatively well correlated. Hair growth in culture was closely related with hair growth rate in vivo. Moreover, minoxidil uniquely demonstrated a significant increase of hair growth in categorized hair follicles assumed at a similar early anagen VI stage of hair cycle. Selection of follicles at a defined stage based on hair-growth rate would permit a more reliable outcome in human hair follicle organ culture.

  19. In vitro activation of dormant follicles for fertility preservation.

    PubMed

    Adhikari, Deepak

    2013-01-01

    Recent advances in radiotherapy and chemotherapy have led to higher cure rates for female children and adolescents with cancer. However, these treatments adversely affect germ cell survival, and ovarian failure is thus a probable side effect of these anticancer therapies. Moreover, an increasing number of women are choosing to postpone childbearing until later in life, but their primordial follicle reserves degenerate with advancing age. Thus there is a pressing need for the development of fertility preservation methods for these individuals. Ovarian tissue cryopreservation prior to loss of the primordial follicle population either due to cancer treatments or normal aging is a promising option for safeguarding fertility. A complete in vitro maturation (IVM) system could help generate mature eggs for later use without the patient having to undergo the cumbersome process involved in current assisted reproduction methods to generate mature eggs. Cryopreserved ovarian cortical tissues have attracted the attention of reproductive biologists and clinicians because of the large number of safely frozen primordial follicles in them, and it is theoretically possible to use these follicles for in vitro activation (IVA) and subsequent IVM. Ovarian tissue collection is independent of patient age and social or personal conditions. Despite being widely accepted potential techniques for fertility preservation, IVA and IVM of human primordial follicles to obtain fertilizable eggs remains far from reality. This chapter highlights the current achievements and obstacles in obtaining growing follicles through activation of dormant follicles.

  20. Novel Cl- currents elicited by follicle stimulating hormone and acetylcholine in follicle-enclosed Xenopus oocytes

    PubMed Central

    1993-01-01

    Voltage-clamp techniques were used to study the membrane currents elicited by follicle stimulating hormone (FSH) and acetylcholine (ACh) in follicle-enclosed oocytes of Xenopus laevis (follicles). Both agonists caused complex responses that were more evident when the follicles were in hypotonic Ringer solution (HR; 190.4 mosM). In this medium, currents activated by FSH regularly showed three phases whereas currents activated by ACh displayed three to six phases. At a holding potential of -60 mV, FSH, and ACh responses involved combinations of inward and outward currents. Both FSH and ACh responses included a slow smooth inward component that was associated with an increase in membrane conductance, mainly to Cl- (S(in)). This current was strongly dependent on the osmolarity of the external solution: an increase in osmolarity of the HR solution of 18-20 mosM caused a 50% decrease in S(in). In contrast, a fast and transient Cl- current (F(in)) specifically elicited by ACh was not dependent on osmolarity. Both, F(in) and S(in) currents required the presence of follicular cells, since defolliculation using three different methods abolished all the response to FSH and at least four components of the ACh responses. The membrane channels carrying F(in) and oscillatory Cl- currents elicited by stimulation of ACh or serum receptors, were much more permeable to I- and Br- than Cl-, whereas S(in) channels were equally permeable to these anions. Unlike the oscillatory Cl- currents generated in the oocyte itself, S(in) and F(in) currents in follicle-enclosed oocytes were not abolished by chelation of intracellular Ca2+, either with EGTA or BAPTA, which suggests that intracellular Ca2+ does not play a critical role in the activation of these currents. Our experiments show that S(in) and F(in) currents are quite distinct from the previously characterized oscillatory Cl- responses of oocytes. Moreover, the results strongly suggest that the FSH and ACh receptors, the Cl- channels

  1. Follicle Viability after Vitrification of Bovine Ovarian Tissue.

    PubMed

    Guedes, Janaína de Souza; Rodrigues, Jhenifer Kliemchen; Campos, Ana Luisa Menezes; Moraes, Camila Cruz de; Caetano, João Pedro Junqueira; Marinho, Ricardo Mello

    2017-08-31

    Purpose The present study aimed to evaluate the impact of vitrification on the viability of follicles using a three-dimensional (3D) in vitro culture. Methods Bovine ovarian tissue samples (n = 5) obtained from slaughterhouses were utilized. The cortex was cut into small fragments of 2 × 3 × 0.5 mm using a tissue slicer. From these fragments, secondary follicles were first isolated by mechanical and enzymatic methods, then encapsulated in alginate gel and individually cultured for 20 days. Additional fragments of the same ovarian tissue were vitrified in a solution containing 25% glycerol and 25% ethylene glycol. After warming, the follicles underwent the same follicular isolation process that was performed for the fresh follicles. Results A total of 61 follicles were isolated, 51 from fresh ovarian tissue, and 10 from vitrified tissue. After the culture, the vitrified and fresh follicles showed 20% and 43.1% survival rates respectively (p = 0.290), with no significant differences. At the end of the culture, there were no significant differences in follicular diameter between the vitrified (422.93 ± 85.05 µm) and fresh (412.99 ± 102.55 µm) groups (p = 0.725). Fresh follicles showed higher mean rate of antrum formation when compared with vitrified follicles (47.1% and 20.0% respectively), but without significant difference (p = 0.167). Conclusions The follicles were able to develop, grow and form antrum in the 3D system after vitrification, despite the lower results obtained with the fresh tissue. Thieme Revinter Publicações Ltda Rio de Janeiro, Brazil.

  2. Anarthria impairs subvocal counting.

    PubMed

    Cubelli, R; Nichelli, P; Pentore, R

    1993-12-01

    We studied subvocal counting in two pure anarthric patients. Analysis showed that they performed definitively worse than normal subjects free to articulate subvocally and their scores were in the lower bounds of the performances of subjects suppressing articulation. These results suggest that subvocal counting is impaired after anarthria.

  3. Complexities of Counting.

    ERIC Educational Resources Information Center

    Stake, Bernadine Evans

    This document focuses on one child's skip counting methods. The pupil, a second grade student at Steuben School, in Kankakee, Illinois, was interviewed as she made several attempts at counting twenty-five poker chips on a circular piece of paper. The interview was part of a larger study of "Children's Conceptions of Number and Numeral,"…

  4. Counting Sheep in Basque

    ERIC Educational Resources Information Center

    Araujo, Frank P.

    1975-01-01

    Demonstrates the interplay of a cognitive system, the Basque numerative system, and a behavioral one, counting sheep. The significant features of the Basque numerative system are analyzed; then it is shown how use of these features facilitates the counting of sheep on open ranges by Basque sheep farmers in California. (Author/RM)

  5. Averaging of TNTC counts.

    PubMed Central

    Haas, C N; Heller, B

    1988-01-01

    When plate count methods are used for microbial enumeration, if too-numerous-to-count results occur, they are commonly discarded. In this paper, a method for consideration of such results in computation of an average microbial density is developed, and its use is illustrated by example. PMID:3178211

  6. Counting Sheep in Basque

    ERIC Educational Resources Information Center

    Araujo, Frank P.

    1975-01-01

    Demonstrates the interplay of a cognitive system, the Basque numerative system, and a behavioral one, counting sheep. The significant features of the Basque numerative system are analyzed; then it is shown how use of these features facilitates the counting of sheep on open ranges by Basque sheep farmers in California. (Author/RM)

  7. Ovarian aging and the activation of the primordial follicle reserve in the long-lived Ames dwarf and the short-lived bGH transgenic mice.

    PubMed

    Saccon, Tatiana D; Moreira, Fabiana; Cruz, Luis A; Mondadori, Rafael G; Fang, Yimin; Barros, Carlos C; Spinel, L; Bartke, A; Masternak, Michal M; Schneider, A

    2017-11-05

    The aim of this study was to evaluate the effect of growth hormone (GH) in the maintenance of the ovarian primordial follicle reserve. Ovaries from 16 mo old GH-deficient Ames Dwarf (df/df) and Normal (N/df) mice were used. A subgroup of df/df and N mice received GH or saline injections for six weeks starting at 14 mo of age. In addition, ovaries from 12 mo old mice overexpressing bovine GH (bGH) and controls were used. df/df mice had higher number of primordial and total follicles than N/df mice (p < 0.05), while GH treatment decreased follicle counts in both genotypes (p < 0.05). In addition, bGH mice had lower number of primordial and total follicles than the controls (p < 0.05). pFoxO3a levels were higher in mice treated with GH and in bGH mice (p < 0.05) when comparing with age match controls. These results indicate that increased circulating GH is associated with a reduced ovarian primordial follicle reserve and increased pFoxO3a content in oocytes. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  8. Time-dependent changes in Ca2+ sensitivity during phasic contraction of canine antral smooth muscle.

    PubMed Central

    Ozaki, H; Gerthoffer, W T; Publicover, N G; Fusetani, N; Sanders, K M

    1991-01-01

    phasic contractions which were superimposed upon the Ca(2+)-independent contracture. In the presence of calyculin-A, relaxation occurred in two phases. The initial, rapid phase of relaxation was not significantly affected by calyculin-A, but the slow phase was significantly decreased. 7. These results suggest that the relationship between [Ca2+]cyt, MLC phosphorylation and contraction changes as a function of [Ca2+]cyt in canine antral muscles. This may be due to a Ca(2+)-and time-dependent phosphatase that regulates the level of myosin phosphorylation. PMID:1804961

  9. Effects of Steroid Hormone in Avian Follicles

    PubMed Central

    Caicedo Rivas, R. E.; Nieto, M. Paz-Calderón; Kamiyoshi, M.

    2016-01-01

    The aim of the present study was to examine the effects of testosterone (T) and estradiol-17β (E2) on the production of progesterone (P4) by granulosa cells, and of the E2 on the production of P4 and T by theca internal cells. In the first experiment, granulosa cells isolated from the largest (F1) and third largest (F3) preovulatory follicle were incubated for 4 h in short-term culture system, P4 production by granulosa cells of both F1 and F3 was increased in a dose-dependent manner by ovine luteinizing hormone (oLH), but not T or E2. In the second experiment, F1 and F3 granulosa cells cultured for 48 h in the developed monolayer culture system were recultured for an additional 48 h with increasing doses of various physiological active substances existing in the ovary, including T and E2. Basal P4 production for 48 h during 48 to 96 h of the cultured was about nine fold greater by F1 granulosa cells than by F3 granulosa cells. In substances examined oLH, chicken vasoactive intestinal polypeptide (cVIP) and T, but not E2, stimulated in a dose-dependent manner P4 production in both F1 and F3 granulosa cells. In addition, when the time course of P4 production by F1 granulosa cells in response to oLH, cVIP, T and E2 was examined for 48 h during 48 to 96 h of culture, although E2 had no effect on P4 production by granulosa cells of F1 during the period from 48 to 96 h of culture, P4 production with oLH was found to be increased at 4 h of the culture, with a maximal 9.14 fold level at 6 h. By contrast, P4 production with cVIP and T increased significantly (p<0.05) from 8 and 12 h of the culture, respectively, with maximal 6.50 fold response at 12 h and 6, 48 fold responses at 36 h. Furthermore, when F1 granulosa cells were precultured with E2 for various times before 4 h culture with oLH at 96 h of culture, the increase in P4 production in response to oLH with a dose-related manner was only found at a pretreatment time of more than 12 h. In the third experiment, theca

  10. Extensive Hair Shaft Growth after Mouse Whisker Follicle Isolation, Cryopreservation and Transplantation in Nude Mice

    PubMed Central

    Cao, Wenluo; Li, Lingna; Tran, Benjamin; Kajiura, Satoshi; Amoh, Yasuyuki; Liu, Fang; Hoffman, Robert M.

    2015-01-01

    We previously demonstrated that whole hair follicles could be cryopreserved to maintain their stem-cells differentation potential. In the present study, we demonstrated that cryopreserved mouse whisker hair follicles maintain their hair growth potential. DMSO better cryopreserved mouse whisker follicles compared to glycerol. Cryopreserved hair follicles also maintained the hair follicle-associated-pluripotent (HAP) stem cells, evidenced by P75NTR expression. Subcutaneous transplantation of DMSO-cryopreserved hair follicles in nude mice resulted in extensive hair fiber growth over 8 weeks, indicating the functional recovery of hair shaft growth of cryopreserved hair follicles. PMID:26716690

  11. AUTOMATIC COUNTING APPARATUS

    DOEpatents

    Howell, W.D.

    1957-08-20

    An apparatus for automatically recording the results of counting operations on trains of electrical pulses is described. The disadvantages of prior devices utilizing the two common methods of obtaining the count rate are overcome by this apparatus; in the case of time controlled operation, the disclosed system automatically records amy information stored by the scaler but not transferred to the printer at the end of the predetermined time controlled operations and, in the case of count controlled operation, provision is made to prevent a weak sample from occupying the apparatus for an excessively long period of time.

  12. [The hair follicle as a target for gene therapy].

    PubMed

    Cotsarelis, G

    2002-05-01

    The hair follicle possesses progenitor cells required for continuous hair follicle cycling and for epidermal keratinocytes, melanocytes and Langerhans cells. These different cell types can be the target of topical gene delivery in the skin of the mouse. Using a combination of liposomes and DNA, we demonstrate the feasibility of targeting hair follicle cells in human scalp xenografts. We consider liposome composition and stage of the hair cycle as important parameters influencing transfection of human hair follicles. Transfection is possible only during the early anagen phase. Factors and obstacles for the use of gene therapy in treating alopecia and skin diseases are discussed. A theoretical framework for future treatment of cutaneous and systemic disorders using gene therapy is presented.

  13. Inhibin-non-steroidal regulation of follicle stimulating hormone secretion

    SciTech Connect

    Burger, H.G.; Findlay, J.K. ); de Kretser, D.M. ); Igarashi, M. )

    1987-01-01

    This book contains the proceedings of inhibin non-steroidal regulation of follicle stimulating hormone secretion. Topics covered include: FSH regulation, Molecular biology, Radioimmunoassay, Physiology - Testocular inhibin, Physiology - ovarian inhibin, and local actions.

  14. Putting the Human Hair Follicle Cycle on the Map.

    PubMed

    Panteleyev, Andrey A

    2016-01-01

    A detailed characterization of the normal (in situ) human hair follicle cycle, supplemented with expressional data on specific hair follicle markers, has been awaited by basic hair researchers and dermatologists. Combining this hair cycle guide, together with a thorough analysis of the human-on-mouse hair xenograft model, provides solid ground for examining human hair cycle biology and pathology and for hair cycle-related pharmacological testing.

  15. Role of the vitamin D receptor in hair follicle biology.

    PubMed

    Demay, Marie B; MacDonald, Paul N; Skorija, Kristi; Dowd, Diane R; Cianferotti, Luisella; Cox, Megan

    2007-03-01

    The vitamin D receptor (VDR) is expressed in numerous cells and tissues, including the skin. The critical requirement for cutaneous expression of the VDR has been proven by investigations in mice and humans lacking functional receptors. These studies demonstrate that absence of the VDR leads to the development of alopecia. The hair follicle is formed by reciprocal interactions between an epidermal placode, which gives rise to the hair follicle keratinocytes and the underlying mesoderm which gives rise to the dermal papilla. Hair follicle morphogenesis ends the second week of life in mice. Studies in VDR null mice have failed to demonstrate a cutaneous abnormality during this period of hair follicle morphogenesis. However, VDR null mice are unable to initiate a new hair cycle after the period of morphogenesis is complete, therefore, do not grow new hair. Investigations in transgenic mice have demonstrated that restricted expression of the VDR to keratinocytes is capable of preventing alopecia in the VDR null mice, thus demonstrating that the epidermal component of the hair follicle requires VDR expression to maintain normal hair follicle homeostasis. Studies were then performed to determine which regions of the VDR were required for these actions. Investigations in mice lacking the first zinc finger of the VDR have demonstrated that they express a truncated receptor containing an intact ligand binding and AF2 domain. These mice are a phenocopy of mice lacking the VDR, thus demonstrate the critical requirement of the DNA binding domain for hair follicle homeostasis. Transgenic mice expressing VDRs with mutations in either the ligand-binding domain or the AF2 domain were generated. These investigations demonstrated that mutant VDRs incapable of ligand-dependent transactivation were able to prevent alopecia. Investigations are currently underway to define the mechanism by which the unliganded VDR maintains hair follicle homeostasis.

  16. Toxicity of Gossypol from Cottonseed Cake to Sheep Ovarian Follicles

    PubMed Central

    Câmara, Antônio Carlos Lopes; Gadelha, Ivana Cristina Nunes; Borges, Pedro Augusto Cordeiro; de Paiva, Silvano Alves; Melo, Marília Martins; Soto-Blanco, Benito

    2015-01-01

    Gossypol, a polyphenol compound produced by cotton plant, has proven reproductive toxicity, but the effects of gossypol on sheep ovaries are unknown. This study was aimed to determine the in vitro and in vivo effects of gossypol on the ovarian follicles of sheep. This trial was divided into two experiments. In the first one, we used twelve non-pregnant, nulliparous, Santa Inês crossbred ewes, which were randomly distributed into two equal groups and fed diets with and without cottonseed cake. Feed was offered at 1.5% of the animal’s body weight for 63 days. The concentrations of total and free gossypol in the cottonseed cake were 3.28 mg/g and 0.11 mg/g, respectively. Throughout the trial period, no animal showed clinical signs of toxicity and no effects on body weight were observed. However, there was a significantly lower number of viable ovarian follicles (20.6%) and higher number of atretic follicles (79.4%) in the gossypol-fed sheep compared to the control (85.1 and 34.9%, respectively). These findings were observed at all stages of follicular development. In the second experiment, eight ovaries from slaughterhouse were cultured with different concentrations of gossypol acetic acid (0, 5, 10 and 20 μg/mL) for 24 hours or seven days. The in vitro action of gossypol resulted in a significant decrease in viable ovarian follicles, especially the primary and transition follicles, and a significant increase in the number of atretic follicles after 24 hours of culture. These follicles were greatly affected when cultured with gossypol for seven days. It is concluded that gossypol present in cotton seeds directly acts on ovarian follicles in sheep to increase atresia. PMID:26600470

  17. Method to identify Demodex in the eyelash follicle without epilation.

    PubMed

    Mastrota, Katherine M

    2013-06-01

    The ubiquitous skin mite Demodex has been implicated as an instigator of anterior blepharitis. To date, overpopulation of the organism has been verified by cilia epilation (Coston or modified Coston method). This monograph describes an alternate technique to isolate Demodex organisms by eyelash rotation within the follicle. Via this method, without epilation, mites are made visible in follicles that may or may not display the cylindrical cuffing characteristic of demodicidosis.

  18. Inventory count strategies.

    PubMed

    Springer, W H

    1996-02-01

    An important principle of accounting is that asset inventory needs to be correctly valued to ensure that the financial statements of the institution are accurate. Errors is recording the value of ending inventory in one fiscal year result in errors to published financial statements for that year as well as the subsequent fiscal year. Therefore, it is important that accurate physical counts be periodically taken. It is equally important that any system being used to generate inventory valuation, reordering or management reports be based on consistently accurate on-hand balances. At the foundation of conducting an accurate physical count of an inventory is a comprehensive understanding of the process coupled with a written plan. This article presents a guideline of the physical count processes involved in a traditional double-count approach.

  19. Counting Knights and Knaves

    ERIC Educational Resources Information Center

    Levin,Oscar; Roberts, Gerri M.

    2013-01-01

    To understand better some of the classic knights and knaves puzzles, we count them. Doing so reveals a surprising connection between puzzles and solutions, and highlights some beautiful combinatorial identities.

  20. Blood Count Tests

    MedlinePlus

    ... white blood cells (WBC), and platelets. Blood count tests measure the number and types of cells in ... helps doctors check on your overall health. The tests can also help to diagnose diseases and conditions ...

  1. Understanding Blood Counts

    MedlinePlus

    ... Heart, Kidneys, Liver and Lung Function Infections Iron Overload Low Blood Counts Pain Disease- and Treatment-Related ... cancer research around the world and provides free information and support services. Privacy Policy Security Copyright Link ...

  2. Human hair follicle: reservoir function and selective targeting.

    PubMed

    Blume-Peytavi, U; Vogt, A

    2011-10-01

    Penetration of topically applied compounds may occur via the stratum corneum, skin appendages and hair follicles. The follicular infundibulum increases the surface area, disrupts the epidermal barrier towards the lower parts of the follicle, and serves as a reservoir. Topical delivery of active compounds to specific targets within the skin, especially to distinct hair follicle compartments or cell populations, may help to treat local inflammatory reactions selectively, with reduced systemic side-effects. Various in vitro and in vivo methods exist for studying the hair follicle structure and follicular penetration pathways. These include cyanoacrylate skin surface stripping, confocal microscopy and cyanoacrylate scalp follicle biopsy. The complex anatomical structure as well as the cyclical activity of the hair follicle must be taken into consideration when designing delivery systems. In addition, delivery into and retention inside the infundibular reservoir are controlled by, for example, molecule or particle size, their polarity and the type of preparation. Preferred penetration depth and storage time must also be considered. Particles with release mechanisms should be preferred; however, the release of drugs from nanoparticles still requires further investigations.

  3. A delay differential equation model of follicle waves in women.

    PubMed

    Panza, Nicole M; Wright, Andrew A; Selgrade, James F

    2016-01-01

    This article presents a mathematical model for hormonal regulation of the menstrual cycle which predicts the occurrence of follicle waves in normally cycling women. Several follicles of ovulatory size that develop sequentially during one menstrual cycle are referred to as follicle waves. The model consists of 13 nonlinear, delay differential equations with 51 parameters. Model simulations exhibit a unique stable periodic cycle and this menstrual cycle accurately approximates blood levels of ovarian and pituitary hormones found in the biological literature. Numerical experiments illustrate that the number of follicle waves corresponds to the number of rises in pituitary follicle stimulating hormone. Modifications of the model equations result in simulations which predict the possibility of two ovulations at different times during the same menstrual cycle and, hence, the occurrence of dizygotic twins via a phenomenon referred to as superfecundation. Sensitive parameters are identified and bifurcations in model behaviour with respect to parameter changes are discussed. Studying follicle waves may be helpful for improving female fertility and for understanding some aspects of female reproductive ageing.

  4. Gene Bionetwork Analysis of Ovarian Primordial Follicle Development

    PubMed Central

    Nilsson, Eric E.; Savenkova, Marina I.; Schindler, Ryan; Zhang, Bin; Schadt, Eric E.; Skinner, Michael K.

    2010-01-01

    Ovarian primordial follicles are critical for female reproduction and comprise a finite pool of gametes arrested in development. A systems biology approach was used to identify regulatory gene networks essential for primordial follicle development. Transcriptional responses to eight different growth factors known to influence primordial follicles were used to construct a bionetwork of regulatory genes involved in rat primordial follicle development. Over 1,500 genes were found to be regulated by the various growth factors and a network analysis identified critical gene modules involved in a number of signaling pathways and cellular processes. A set of 55 genes was identified as potential critical regulators of these gene modules, and a sub-network associated with development was determined. Within the network two previously identified regulatory genes were confirmed (i.e., Pdgfa and Fgfr2) and a new factor was identified, connective tissue growth factor (CTGF). CTGF was tested in ovarian organ cultures and found to stimulate primordial follicle development. Therefore, the relevant gene network associated with primordial follicle development was validated and the critical genes and pathways involved in this process were identified. This is one of the first applications of network analysis to a normal developmental process. These observations provide insights into potential therapeutic targets for preventing ovarian disease and promoting female reproduction. PMID:20661288

  5. Spectroscopic characterization of recombinant follicle stimulating hormone

    NASA Astrophysics Data System (ADS)

    Groen, B. H.; Bloemendal, M.; Mulders, J. W. M.; Hadden, J. M.; Chapman, D.; Van Stokkum, I. H. M.; Van Grondelle, R.

    1996-09-01

    Recombinant follicle stimulating hormone (recFSH, Org. 32489) has been characterized by absorption (UV and IR), (polarized) fluorescence, linear-dichroism (LD) and circular-dichroism (CD) spectroscopy. Absorption and fluorescence spectra of the isolated subunits have also been measured. From the spectra the extinction coefficient, fluorescence quantum yield and anisotropy have been calculated. Global analysis is used to characterize the bands in the spectra. The adsorption, CD, LD and fluorescence excitation spectra all contain a band around 300 nm that appears to be a sensitive indicator for the intactness of the protein. Evidence is provided for the involvement of tyrosinate in the fluorescence, and for a close contact between the tryptophan (in the β subunit) with at least one tyrosine of the α subunit. The overall secondary structure of recFSH has been determined from its far-UV CD and its IR absorption spectrum. The secondary structure of recFSH is estimated to contain 15-25% α-helix, 15-25% β-turn and 30-40% β-sheet. The β-sheet in recFSH is almost exclusively antiparallel. The results confirm that recFSH contains significantly more α-helix than the closed related human glycoproteins, chorionic gonadotropin and lutropin; however, the α-helices may be short and distorted.

  6. Integral hair lipid in human hair follicle.

    PubMed

    Lee, Won-Soo

    2011-12-01

    Integral hair lipid (IHL) is bound to the keratinized cell surface to make an environmentally resistant lipid envelope. It is mainly positioned on the hair cuticle and inner root sheath. IHL in the hair follicle may regard as hair barrier to be similar to the epidermal lipid layer functioning as skin barrier. Major constituents of IHL are fatty acid, phytosphingosine, ceramide in decreasing order. Minor constituents of IHL are cholesterol, cholesterol sulfate and cholesterol oleate. Cuticle or cortical cell surface in hair are abundant in fatty acids unlike the keratinized area of epidermis or sebaceous gland, and about 30-40% of such fatty acids are composed of 18-methyl-eicosanoic acid which is known to be bound to proteins by ester or thioester bond. Various factors including moisture, solvent, oxidative damage during bleaching or permanent waving affect IHL. Photochemical changes also can occur in IHL as well as in hair protein and hair pigment. Lipid metabolism is thought to play an essential role in lipid envelope of hair, but also involvement in hair development and function. Copyright © 2011 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.

  7. Alterations in Hair Follicle Dynamics in Women

    PubMed Central

    Piérard-Franchimont, Claudine; Piérard, Gérald E.

    2013-01-01

    Endocrine changes supervening after parturition and menopause participate in the control of sebum production and hair growth modulation. The ensuing conditions include some peculiar aspects of hair loss (effluvium), alopecia, and facial hirsutism. The hair cycling is of major clinical relevance because most hair growth disorders result from disturbances in this chronobiological feature. Of note, any correlation between a biologic abnormality and hair cycling disturbance does not prove a relationship of causality. The proportion of postmenopausal women is rising in the overall population. Therefore, the prevalence of these hair follicle disturbances is globally on the rise. Current therapies aim at correcting the underlying hormonal imbalances, and at improving the overall cosmetic appearance. However, in absence of pathogenic diagnosis and causality criteria, chances are low that a treatment given by the whims of fate will adequately control hair effluvium. The risk and frequency of therapeutic inertia are further increased. When the hair loss is not controlled and/or compensated by growth of new hairs, several clinical aspects of alopecia inexorably develop. Currently, there is little evidence supporting any specific treatment for these endocrine hair disorders in post-partum and postmenopausal women. Current hair treatment strategies are symptomatic and nonspecific so current researchers aim at developing new, targeted methods. PMID:24455742

  8. Alterations in hair follicle dynamics in women.

    PubMed

    Piérard-Franchimont, Claudine; Piérard, Gérald E

    2013-01-01

    Endocrine changes supervening after parturition and menopause participate in the control of sebum production and hair growth modulation. The ensuing conditions include some peculiar aspects of hair loss (effluvium), alopecia, and facial hirsutism. The hair cycling is of major clinical relevance because most hair growth disorders result from disturbances in this chronobiological feature. Of note, any correlation between a biologic abnormality and hair cycling disturbance does not prove a relationship of causality. The proportion of postmenopausal women is rising in the overall population. Therefore, the prevalence of these hair follicle disturbances is globally on the rise. Current therapies aim at correcting the underlying hormonal imbalances, and at improving the overall cosmetic appearance. However, in absence of pathogenic diagnosis and causality criteria, chances are low that a treatment given by the whims of fate will adequately control hair effluvium. The risk and frequency of therapeutic inertia are further increased. When the hair loss is not controlled and/or compensated by growth of new hairs, several clinical aspects of alopecia inexorably develop. Currently, there is little evidence supporting any specific treatment for these endocrine hair disorders in post-partum and postmenopausal women. Current hair treatment strategies are symptomatic and nonspecific so current researchers aim at developing new, targeted methods.

  9. Neutron counting with cameras

    SciTech Connect

    Van Esch, Patrick; Crisanti, Marta; Mutti, Paolo

    2015-07-01

    A research project is presented in which we aim at counting individual neutrons with CCD-like cameras. We explore theoretically a technique that allows us to use imaging detectors as counting detectors at lower counting rates, and transits smoothly to continuous imaging at higher counting rates. As such, the hope is to combine the good background rejection properties of standard neutron counting detectors with the absence of dead time of integrating neutron imaging cameras as well as their very good spatial resolution. Compared to Xray detection, the essence of thermal neutron detection is the nuclear conversion reaction. The released energies involved are of the order of a few MeV, while X-ray detection releases energies of the order of the photon energy, which is in the 10 KeV range. Thanks to advances in camera technology which have resulted in increased quantum efficiency, lower noise, as well as increased frame rate up to 100 fps for CMOS-type cameras, this more than 100-fold higher available detection energy implies that the individual neutron detection light signal can be significantly above the noise level, as such allowing for discrimination and individual counting, which is hard to achieve with X-rays. The time scale of CMOS-type cameras doesn't allow one to consider time-of-flight measurements, but kinetic experiments in the 10 ms range are possible. The theory is next confronted to the first experimental results. (authors)

  10. Extracellular-like matrices and leukaemia inhibitory factor for in vitro culture of human primordial follicles.

    PubMed

    Younis, Assiel J; Lerer-Serfaty, Galit; Stav, Dana; Sabbah, Bethsabee; Shochat, Tzippy; Kessler-Icekson, Gania; Zahalka, Muayad A; Shachar-Goldenberg, Michal; Ben-Haroush, Avi; Fisch, Benjamin; Abir, Ronit

    2017-02-01

    The possibility of maturing human primordial follicles in vitro would assist fertility restoration without the danger of reseeding malignancies. Leukaemia inhibitory factor (LIF) and certain culture matrices may promote human follicular growth. The present study compared human primordial follicular growth on novel culture matrices, namely human recombinant vitronectin (hrVit), small intestine submucosa (SIS), alginate scaffolds and human recombinant virgin collagen bioengineered in tobacco plant lines (CollPlant). The frozen-thawed ovarian samples that were used had been obtained from girls or young women undergoing fertility preservation. In the first part of the study, 20 samples were cultured for 6 days on hrVit or SIS with basic culture medium alone or supplemented with one of two concentrations of LIF (10ngmL-1 and 100ngmL-1), with and without LIF-neutralising antibody. In the second part of the study, 15 samples were cultured for 6 days on alginate scaffolds or CollPlant matrices with basic culture medium. Follicular development was assessed by follicular counts and classification, Ki67 immunohistochemistry and 17β-oestradiol and anti-Müllerian hormone measurements in spent media samples. Primordial follicular growth was not enhanced by LIF. Despite some significant differences among the four matrices, none appeared to have a clear advantage, apart from significantly more Ki67-stained follicles on alginate and CollPlant matrices. Further studies of other culture matrices and medium supplements are needed to obtain an optimal system.

  11. A primer for studying cell cycle dynamics of the human hair follicle.

    PubMed

    Purba, Talveen S; Brunken, Lars; Hawkshaw, Nathan J; Peake, Michael; Hardman, Jonathan; Paus, Ralf

    2016-09-01

    The cell cycle is of major importance to human hair follicle (HF) biology. Not only is continuously active cell cycling required to facilitate healthy hair growth in anagen VI HFs, but perturbations in the cell cycle are likely to be of significance in HF pathology (i.e. in scarring, non-scarring, chemotherapy-induced and androgenic alopecias). However, cell cycle dynamics of the human hair follicle (HF) are poorly understood in contrast to what is known in mouse. The current Methods Review aims at helping to close this gap by presenting a primer that introduces immunohistological/immunofluorescent techniques to study the cell cycle in the human HF. Moreover, this primer encourages the exploitation of the human HF as a powerful and clinically relevant tool to investigate mammalian cell cycle biology in situ. To achieve this, we describe methods to study markers of general 'proliferation' (nuclei count, Ki-67 expression), apoptosis (terminal deoxynucleotidyl transferase dUTP nick-end labelling, cleaved caspase 3), mitosis (phospho-histone H3, 'pS780'), DNA synthesis (5-ethynyl-2'-deoxyuridine) and cell cycle regulation (cyclins) in the human HF. In addition, we provide specific examples of dual immunolabelling for instructive cell cycle analyses and for investigating the cell cycle behaviour of specific HF keratinocyte subpopulations, such as keratin 15+ stem/progenitor cells. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  12. Effect of epidermal growth factor on follicle-stimulating hormone-induced proliferation of granulosa cells from chicken prehierarchical follicles.

    PubMed

    Lin, Jin-xing; Jia, Yu-dong; Zhang, Cai-qiao

    2011-11-01

    The development of ovarian follicular cells is controlled by multiple circulating and local hormones and factors, including follicle-stimulating hormone (FSH) and epidermal growth factor (EGF). In this study, the stage-specific effect of EGF on FSH-induced proliferation of granulosa cells was evaluated in the ovarian follicles of egg-laying chickens. Results showed that EGF and its receptor (EGFR) mRNAs displayed a high expression in granulosa cells from the prehierarchical follicles, including the large white follicle (LWF) and small yellow follicle (SYF), and thereafter the expression decreased markedly to the stage of the largest preovulatory follicle. SYF represents a turning point of EGF/EGFR mRNA expression during follicle selection. Subsequently the granulosa cells from SYF were cultured to reveal the mediation of EGF in FSH action. Cell proliferation was remarkably increased by treatment with either EGF or FSH (0.1-100 ng/ml). This result was confirmed by elevated proliferating cell nuclear antigen (PCNA) expression and decreased cell apoptosis. Furthermore, EGF-induced cell proliferation was accompanied by increased mRNA expressions of EGFR, FSH receptor, and the cell cycle-regulating genes (cyclins D1 and E1, cyclin-dependent kinases 2 and 6) as well as decreased expression of luteinizing hormone receptor mRNA. However, the EGF or FSH-elicited effect was reversed by simultaneous treatment with an EGFR inhibitor AG1478. In conclusion, EGF and EGFR expressions manifested stage-specific changes during follicular development and EGF mediated FSH-induced cell proliferation and retarded cell differentiation in the prehierarchical follicles. These expressions thus stimulated follicular growth before selection in the egg-laying chicken.

  13. On the granulosa cells of ovarian follicles. II. Identification of different morphological patterns of granulosa cells in evolutive follicles.

    PubMed

    Zecchi, S; Repice, F; Balboni, G C

    1981-03-15

    An attempt has been made for identifying different types of granulosa cells in the wall of cavitary ovarian follicles. Human, porcine and rat ovaries have been examined at the light and electron microscopes. Some smears of granulosa cells as well as human foetal ovaries have been also studied. These preliminary results seem to confirm that in the granulosa layer of evolutive follicles the cells may present some different morphological and histochemical features.

  14. Ultrasound characteristics of experimentally induced luteinized unruptured follicles (LUF) and naturally occurring hemorrhagic anovulatory follicles (HAF) in the mare.

    PubMed

    Cuervo-Arango, J; Newcombe, J R

    2012-02-01

    The development of hemorrhagic anovulatory follicles (HAF) involves luteinization and hemorrhage of the follicle. This is observed on ultrasound as an increase in the echogenicity of the granulosa layer and formation of echoic particles in the antrum. The inhibition of prostaglandin synthesis with flunixin meglumine (FM) during the periovulatory period induces ovulatory failure with development of luteinized unruptured follicles (LUF). These two types of anovulatory follicles appear to share similar ultrasound features but they have not been compared critically. The following endpoints: follicle diameter, follicular contents score, interval from hCG administration to beginning of follicular hemorrhage, interval from hemorrhage to organization of follicular contents, and cycle length were studied and compared in mares with HAF (n = 11) and LUF (n = 13). The objective of this study was to elucidate whether these two unruptured follicles have a consistent clinical pattern of development and therefore can be considered as part of the same anovulatory syndrome. None of the endpoints analyzed differed significantly between HAF and LUF. However, there was a greater individual variation in HAF as compared with LUF in regards to interval from hCG to hemorrhage, follicular diameter at the administration of hCG, and beginning of hemorrhage. In conclusion, HAF share a similar cascade of ultrasound characteristics with the experimentally induced LUF. This finding may provide new insights in elucidating the pathogenesis of HAF.

  15. Antral Helicobacter Pylori Infection, Hypergastrinemia and Peptic Ulcers: Effect of Eradicating the Organism

    PubMed Central

    Park, Sill Moo; Yoo, Byung Chul; Lee, Hyo Rang; Yoon, Joon Hyun; Cha, Young Joo

    1993-01-01

    Background: A randomized prospective study on the response of fasting serum gastrin concentrations in peptic ulcer patients was performed in order to test the hypothesis that H. pylori infection in the gastric antrum increases gastrin release, and to examine whether the high fasting serum gastrin concentrations respond to treatment that eradicates H. pylori. Methods: One hundred and twenty-seven patients with gastric or duodenal ulcer were included in this study. Patients were divided into three groups on the basis of antral H. pylori status and therapeutic modalities. The first group, 58 patients infected by H. pylori, was treated with metronidazole and tripotassium dicitrato bismuthate combined with ranitidine and mylanta. The second group, 40 patients also infected by H. Pylori, was treated with ranitidine and mylanta. The third group, 29 patients, free of H. pylori infection, was designed to evaluate the influence of H2-receptor antagonist on the change of gastrin. When ulcers were completely healed, changes of gastrin concentrations and H. pylori status were re-examined. Results: H. pylori was eradicated in all patients who have received antibacterial therapy in 4 weeks, and serum gastrin concentrations were significantly decreased after eradication of the organism both in gastric and in duodenal ulcer diseases. (Gastric ulcer: 129.3±47.0 pg/ml before and 63.7±21.6 pg/ml after treatment. Duodenal ulcer: 108.3±35.0 pg/ml and 66.5±21.9 pg/ml, respectively. Total: 112.7±38.2 pg/ml vs 66.0±21.6 pg/ml) (p<0.01). In contrast, H. pylori-positive patients who have not received antibacterial therapy were still infected at the completion of the study, and serum gastrin concentrations increased even though the difference was not significant. (Gastric ulcer: 118.4±51.2 pg/ml vs 124.0±56.5 pg/ml. Duodenal ulcer: 85.4±35.1 pg/ml vs 104.6±43.5. Total: 99.5±45.3 vs 112.9±48.7 pg/ml.) (p>0.05) None of the patients who were initially H. pylori-negative has been

  16. The effect of Helicobacter pylori eradication therapy on gastric antral myoelectrical activity and gastric emptying in patients with non-ulcer dyspepsia.

    PubMed

    Miyaji, H; Azuma, T; Ito, S; Abe, Y; Ono, H; Suto, H; Ito, Y; Yamazaki, Y; Kohli, Y; Kuriyama, M

    1999-10-01

    Dysmotility of the gastroduodenal region and delayed gastric emptying have been considered to play roles in non-ulcer dyspepsia (NUD). Helicobacter pylori-induced inflammation of the gastric mucosa may affect gastric motility. To evaluate the effects of H. pylori eradication therapy on gastrointestinal motility and symptoms in NUD patients. : Forty-six NUD patients were examined for gastric emptying, antral myoelectrical activity, H. pylori infection, and symptom scores. In H. pylori-positive NUD patients, gastric emptying, antral myoelectrical activity, and symptom scores were also analysed 2 months after cure of H. pylori infection. Sixty-seven per cent of NUD patients were H. pylori-positive. Both abnormal gastric emptying and antral myoelectrical activity were observed in NUD patients. H. pylori-positive NUD patients were divided into three groups according to their gastric emptying: the delayed group, the normal group, and the rapid group. In the delayed and rapid gastric emptying groups, the emptying and symptom scores were improved significantly by eradication. There was no improvement in symptom scores in the normal gastric emptying NUD group by the eradication therapy. Disturbed gastric emptying and antral myoelectrical activity play roles in NUD. H. pylori-induced disturbed gastric emptying may cause some NUD symptoms. Gastric emptying and symptom scores are improved by H. pylori eradication therapy in NUD patients with disturbed gastric emptying; H. pylori eradication therapy is effective in H. pylori-positive NUD patients with disturbed gastric emptying.

  17. Immunoglobulin negative follicle centre cell lymphoma.

    PubMed Central

    Gregg, E. O.; Al-Saffar, N.; Jones, D. B.; Wright, D. H.; Stevenson, F. K.; Smith, J. L.

    1984-01-01

    Immunoglobulin (Ig) could not be detected on the surface or in the cytoplasm of neoplastic cells from five cases of follicle centre cell lymphoma with centroblastic/centrocytic follicular histology when examined by immunohistology of frozen or wax embedded sections. Examination by fluorescein labelled antibodies of cells in suspensions prepared from the biopsies revealed a monotypic surface Ig positive population in one case and a surface or cytoplasmic Ig kappa:lambda light chain imbalance in a further two cases consistent with neoplastic B cell involvement: in all cases the proportion of cells failing to express Ig or T cell markers ranged from 24 to 75%. The monoclonal antibodies B1 (Pan B cell), FMC4 (HLA class II) and J5 (cALL antigen) stained the majority of cells in suspension with residual cells staining with UCHT1 or OKT11 (T cell monoclonal antibodies). In frozen sections, neoplastic follicular cells did not stain with UCHT1. However, in the one case tested these cells stained with the antibodies B1 and FMC4. In paraffin sections J chain could be demonstrated in the cytoplasm of three out of five cases. Cells from four cases were cultured in vitro for Ig production: two failed to produce Ig and monotypic light chains were the sole Ig product of the remaining two cases. The failure to express Ig by the majority of the neoplastic cells from the cases described in this report is at variance with the follicular histology of these neoplasms. Mechanisms responsible for this failure are discussed with reference to current models of B cell differentiation. Images Figure 1 PMID:6437429

  18. Fast counting electronics for neutron coincidence counting

    DOEpatents

    Swansen, James E.

    1987-01-01

    An amplifier-discriminator is tailored to output a very short pulse upon an above-threshold input from a detector which may be a .sup.3 He detector. The short pulse output is stretched and energizes a light emitting diode (LED) to provide a visual output of operation and pulse detection. The short pulse is further fed to a digital section for processing and possible ORing with other like generated pulses. Finally, the output (or ORed output ) is fed to a derandomizing buffer which converts the rapidly and randomly occurring pulses into synchronized and periodically spaced-apart pulses for the accurate counting thereof. Provision is also made for the internal and external disabling of each individual channel of amplifier-discriminators in an ORed plurality of same.