Response of β-Glucosidase to Fungal Infections in Seed, Ovary and Fruit

Localization and changes in the activity of -glucosidase were investigated in wheat caryopsis and glumes infected with Stagonospora nodorum as well as in lily ovaries and harvested tomato fruits both inoculated with Botrytis cinerea. It was established that the pathogen invasion caused splitting of wheat seed coat, xylem blocking in lily carpel and decay in tomato fruits. B. cinerea invasion evoked disorders of the embryogenesis accompanied by a decreased activity of -glucosidase in all ovules. The activity of the enzyme was not changed considerably in wheat seeds as the infection occurred in the late embryonal stages and the embryonal processes were not affected. In the seeds of harvested tomatoes distant from the invaded area the enzyme activity was not changed as well.     

Effect of Epidermal Growth Factor on Follicular Development and Steroidogenesis in Perfused Rat Ovary

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Transforming Growth Factor-β Signaling Participates in the Maintenance of the Primordial Follicle Pool in the Mouse Ovary

Physiologically, only a few primordial follicles are activated to enter the growing follicle pool each wave. Recent studies in knock-out mice show that early follicular activation depends on signaling from the tuberous sclerosis complex, the mammalian target of rapamycin complex 1 (mTORC1), phosphatase and tensin homolog deleted on chromosome 10, and phosphatidylinositol 3-kinase (PI3K) pathways. However, the manner in which these pathways are normally regulated, and whether or not TGF-β acts on them are poorly understood. So, this study aims to identify whether or not TGF-β acts on the process. Ovary organ culture experiments showed that the culture of 18.5 days post-coitus (dpc) ovaries with TGF-β1 reduced the total population of oocytes and activated follicles, accelerated oocyte growth was observed in ovaries treated with TGF-βR1 inhibitor 2-(5-chloro-2-fluorophenyl)pteridin-4-yl]pyridin-4-yl-amine (SD208) compared with control ovaries, the down-regulation of TGF-βR1 gene expression also activated early primordial follicle oocyte growth. We further showed that there was dramatically more proliferation of granulosa cells in SD208-treated ovaries and less proliferation in TGF-β1-treated ovaries. Western blot and morphological analyses indicated that TGF-β signaling manipulated primordial follicle growth through tuberous sclerosis complex/mTORC1 signaling in oocytes, and the mTORC1-specific inhibitor rapamycin could partially reverse the stimulated effect of SD208 on the oocyte growth and decreased the numbers of growing follicles. In conclusion, our results suggest that TGF-β signaling plays an important physiological role in the maintenance of the dormant pool of primordial follicles, which functions through activation of p70 S6 kinase 1 (S6K1)/ribosomal protein S6 (rpS6) signaling in mouse ovaries.     

Birth Weight and Polycystic Ovary Syndrome in Adult Life: Is There a Causal Link?

Objectives

Several studies have demonstrated associations of birth weight with metabolic and reproductive abnormalities in adults. The aim of this study was to investigate the birth weight in women with PCOS and its correlation with clinical and biochemical characteristics of the syndrome.

Materials and Methods

We studied 288 women with PCOS according to the NIH criteria and 166 women with normal cycle and without clinical hyperandrogenism. Birth weight and anthropometric characteristics were recorded, and levels of serum androgens, SHBG, insulin and fasting glucose were measured.

Results

Birth weight data were available for 243/288 women with PCOS and age- and BMI-matched 101/166 controls. No differences were found (p> 0.05) in birth weight among women with PCOS and normal controls. Birth weight of PCOS women was negatively correlated with DHEAS levels (p = 0.031, r = -0.143) and positively correlated with waist circumference (p <0.001, r = 0.297) and body mass index (BMI) (p = 0.040, r = 0.132). Birth weight of controls was negatively correlated with SHBG levels (p = 0.021, r = -0.234). Women from both groups were further divided in 6 categories according to birth weight (A. <2.500 gr, B. 2.501-3.000 gr, C. 3.001-3.500 gr, D. 3.501-4.000 gr, E. 4.001-4.500 gr, F. > 4.500 gr). No statistically significant differences were observed in the distribution percentages between PCOS women and controls. (A. 7% vs 7.9%, B. 26.8% vs 20.8%, C. 39.1% vs 48.5%, D. 21.4% vs 20.8%, E. 4.9% vs 2%, F. 0.8% vs 0%), (in all comparisons, p> 0.05).

Conclusions

Women with PCOS do not differ from controls in birth weight distribution. However, birth weight may contribute to subtypes of the syndrome that are characterized by adrenal hyperandrogenism and central obesity.     

Are Dieting and Dietary Inadequacy a Second Hit in the Association with Polycystic Ovary Syndrome Severity?

Background

The composition of the diet is of increasing importance for the development and maturation of the ovarian follicles. In Polycystic Ovary Syndrome (PCOS) healthy dietary interventions improve the clinical spectrum. We hypothesized that dieting and diet inadequacy in the reproductive life course is associated with impaired programming of ovarian follicles and contributes to the severity of the PCOS phenotype.

Methods and Findings

To determine associations between the use of a self-initiated diet and diet inadequacy and the severity of the PCOS phenotype, we performed an explorative nested case control study embedded in a periconception cohort of 1,251 patients visiting the preconception outpatient clinic. 218 patients with PCOS and 799 subfertile controls were selected from the cohort and self-administered questionnaires, anthropometric measurements and blood samples were obtained. The Preconception Dietary Risk Score (PDR score), based on the Dutch dietary guidelines, was used to determine diet inadequacy in all women. The PDR score was negatively associated to cobalamin, serum and red blood cell folate and positively to tHcy. PCOS patients (19.9%), in particular the hyperandrogenic (HA) phenotype (22.5%) reported more often the use of a self-initiated diet than controls (13.1%; p = 0.023). The use of an inadequate diet was also significantly higher in PCOS than in controls (PDR score 3.7 vs 3.5; p = 0.017) and every point increase was associated with a more than 1.3 fold higher risk of the HA phenotype (adjusted OR 1.351, 95% CI 1.09–1.68). Diet inadequacy was independently associated with the anti-Müllerian Hormone (AMH) concentration (β 0.084; p = 0.044; 95% CI 0.002 to 0.165) and free androgen index (β 0.128; p = 0.013; 95% CI 0.028 to 0.229) in PCOS patients.

Conclusions

The use of a self-initiated diet and diet inadequacy is associated with PCOS, in particular with the severe HA phenotype. This novel finding substantiated by the association between diet inadequacy and AMH needs further investigation.     

Integrin ��IIb��3 Activation in Chinese Hamster Ovary Cells and Platelets Increases Clustering Rather than Affinity

Integrin αIIbβ3 affinity regulation by talin binding to the cytoplasmic tail of β3 is a generally accepted model for explaining activation of this integrin in Chinese hamster ovary cells and human platelets. Most of the evidence for this model comes from the use of multivalent ligands. This raises the possibility that the activation being measured is that of increased clustering of the integrin rather than affinity. Using a newly developed assay that probes integrins on the surface of cells with only monovalent ligands prior to fixation, I do not find increases in affinity of αIIbβ3 integrins by talin head fragments in Chinese hamster ovary cells, nor do I observe affinity increases in human platelets stimulated with thrombin. Binding to a multivalent ligand does increase in both of these cases. This assay does report affinity increases induced by either Mn²⁺, a cytoplasmic domain mutant (D723R) in the cytoplasmic domain of β3, or preincubation with a peptide ligand. These results reconcile the previously observed differences between talin effects on integrin activation in Drosophila and vertebrate systems and suggest new models for talin regulation of integrin activity in human platelets.     

Early-to-Mid Gestation Fetal Testosterone Increases Right Hand 2D∶4D Finger Length Ratio in Polycystic Ovary Syndrome-Like Monkeys

A smaller length ratio for the second relative to the fourth finger (2D∶4D) is repeatedly associated with fetal male-typical testosterone (T) and is implicated as a biomarker for a variety of traits and susceptibility to a number of diseases, but no experimental human studies have been performed. The present study utilizes the rhesus monkey, a close relative of humans, and employs discrete gestational exposure of female monkeys to fetal male-typical T levels for 15–35 days during early-to-mid (40–76 days; n = 7) or late (94–139 days; n = 7) gestation (term: 165 days) by daily subcutaneous injection of their dams with 10 mg T propionate. Such gestational exposures are known to enhance male-typical behavior. In this study, compared to control females (n = 19), only early-to-mid gestation T exposure virilizes female external genitalia while increasing 2D∶4D ratio in the right hand (RH) by male-like elongation of RH2D. RH2D length and 2D∶4D positively correlate with androgen-dependent anogenital distance (AG), and RH2D and AG positively correlate with duration of early-to-mid gestation T exposure. Male monkeys (n = 9) exhibit a sexually dimorphic 2D∶4D in the right foot, but this trait is not emulated by early-to-mid or late gestation T exposed females. X-ray determined phalanx measurements indicate elongated finger and toe phalanx length in males, but no other phalanx-related differences. Discrete T exposure during early-to-mid gestation in female rhesus monkeys thus appears to increase RH2D∶4D through right-side biased, non-skeletal tissue growth. As variation in timing and duration of gestational T exposure alter male-like dimensions of RH2D independently of RH4D, postnatal RH2D∶4D provides a complex biomarker for fetal T exposure.     

Activation of NF-��B Protein Prevents the Transition from Juvenile Ovary to Testis and Promotes Ovarian Development in Zebrafish

Testis differentiation in zebrafish involves juvenile ovary to testis transformation initiated by an apoptotic wave. The molecular regulation of this transformation process is not fully understood. NF-κB is activated at an early stage of development and has been shown to interact with steroidogenic factor-1 in mammals, leading to the suppression of anti-Müllerian hormone (Amh) gene expression. Because steroidogenic factor-1 and Amh are important for proper testis development, NF-κB-mediated induction of anti-apoptotic genes could, therefore, also play a role in zebrafish gonad differentiation. The aim of this study was to examine the potential role of NF-κB in zebrafish gonad differentiation. Exposure of juvenile zebrafish to heat-killed Escherichia coli activated the NF-κB pathways and resulted in an increased ratio of females from 30 to 85%. Microarray and quantitative real-time-PCR analysis of gonads showed elevated expression of NF-κB-regulated genes. To confirm the involvement of NF-κB-induced anti-apoptotic effects, zebrafish were treated with sodium deoxycholate, a known inducer of NF-κB or NF-κB activation inhibitor (NAI). Sodium deoxycholate treatment mimicked the effect of heat-killed bacteria and resulted in an increased proportion of females from 25 to 45%, whereas the inhibition of NF-κB using NAI resulted in a decrease in females from 45 to 20%. This study provides proof for an essential role of NF-κB in gonadal differentiation of zebrafish and represents an important step toward the complete understanding of the complicated process of sex differentiation in this species and possibly other cyprinid teleosts as well.     

WNT4/β-Catenin Pathway Maintains Female Germ Cell Survival by Inhibiting Activin βB in the Mouse Fetal Ovary

Female germ cells are essential for organogenesis of the ovary; without them, ovarian follicles do not form and functional and structural characteristics of the ovary are lost. We and others showed previously that when either Wnt4 or β-catenin was inactivated in the fetal ovary, female germ cells underwent degeneration. In this study, we set out to understand whether these two factors belong to the same pathway and how they maintain female germ cell survival. We found that activation of β-catenin in somatic cells in the Wnt4 knockout ovary restored germ cell numbers, placing β-catenin downstream of WNT4. In the absence of Wnt4 or β-catenin, female germ cells entered meiosis properly; however, they underwent apoptosis afterwards. Activin βB (Inhbb), a subunit of activins, was upregulated in the Wnt4 and β-catenin knockout ovaries, suggesting that Inhbb could be the cause for the loss of female germ cells, which are positive for activin receptors. Indeed, removal of Inhbb in the Wnt4 knockout ovaries prevented female germ cells from undergoing degeneration. We conclude that WNT4 maintains female germ cell survival by inhibiting Inhbb expression via β-catenin in the somatic cells. Maintenance of female germ cells hinge upon a delicate balance between positive (WNT4 and β-catenin) and negative (activin βB) regulators derived from the somatic cells in the fetal ovary.     

Sequence and Expression Characteristics of Long Noncoding RNAs in Honey Bee Caste Development – Potential Novel Regulators for Transgressive Ovary Size

Division of labor in social insect colonies relies on a strong reproductive bias that favors queens. Although the ecological and evolutionary success attained through caste systems is well sketched out in terms of ultimate causes, the molecular and cellular underpinnings driving the development of caste phenotypes are still far from understood. Recent genomics approaches on honey bee developmental biology revealed a set of genes that are differentially expressed genes in larval ovaries and associated with transgressive ovary size in queens and massive cell death in workers. Amongst these, two contigs called special attention, both being over 200 bp in size and lacking apparent coding potential. Herein, we obtained their full cDNA sequences. These and their secondary structure characteristics placed in evidence that they are bona fide long noncoding RNAs (lncRNA) differentially expressed in larval ovaries, thus named lncov1 and lncov2. Genomically, both map within a previously identified QTL on chromosome 11, associated with transgressive ovary size in honey bee workers. As lncov1 was over-expressed in worker ovaries we focused on this gene. Real-time qPCR analysis on larval worker ovaries evidenced an expression peak coinciding with the onset of autophagic cell death. Cellular localization analysis through fluorescence in situ hybridization revealed perinuclear spots resembling omega speckles known to regulate trafficking of RNA-binding proteins. With only four lncRNAs known so far in honey bees, two expressed in the ovaries, these findings open a novel perspective on regulatory factors acting in the fine tuning of developmental processes underlying phenotypic plasticity related to social life histories.     

Expression of Estrogen Receptor α 36 (ESR36) in the Hamster Ovary throughout the Estrous Cycle: Effects of Gonadotropins

Estradiol-17β (E) plays an important role in ovarian follicular development. Evidence indicates that some of the effect of E is mediated by the transmembrane estrogen receptor. In this study, we examined the spatio-temporal expression of recently discovered ERα36 (ESR36), a splice variant of Esr1 and a receptor for non-genomic E signaling, in the hamster ovary during the estrous cycle and the role of gonadotropins and ovarian steroid hormones in ESR36 expression. ESR36 expression was high on estrus (D1∶0900 h) and declined precipitously by proestrus (D4∶0900 h) and remained low up to D4∶1600 h. Immunofluorescence findings corroborated immunoblot findings and revealed that ESR36 was expressed only in the cell membrane of both follicular and non-follicular cells, except the oocytes. Ovarian ESR36 was capable of binding to the E-affinity matrix, and have different molecular weight than that of the ESR1 or GPER. Hypophysectomy (Hx) resulted in a marked decline in ESR36 protein levels. FSH and LH, alone or combined, markedly upregulated ESR36 protein in Hx hamsters to the levels observed in D1 hamsters, but neither E nor P had any effect. Inhibition of the gonadotropin surge by phenobarbital treatment on D4∶1100 h attenuated ESR36 expression in D1∶0900 h ovaries, but the decline was restored by either FSH or LH replacement on D4 afternoon. This is the first report to show that ESR36, which is distinct from ESR1 or GPER is expressed in the plasma membrane of ovarian follicular and non-follicular cells, binds to E and its expression is regulated directly by the gonadotropins. In light of our previous findings, the results suggest that ovarian cells contain at least two distinct membrane estrogen receptors, such as GPER and ESR36, and strongly suggest for a non-genomic action of E regulating ovarian follicular functions.     

Injectable and Oral Contraceptive Use and Cancers of the Breast,Cervix, Ovary,and Endometrium in Black South African Women: Case–Control Study

Background

Oral contraceptives are known to influence the risk of cancers of the female reproductive system. Evidence regarding the relationship between injectable contraceptives and these cancers is limited, especially in black South Africans, among whom injectable contraceptives are used more commonly than oral contraceptives.

Methods and Findings

We analysed data from a South African hospital-based case–control study of black females aged 18–79 y, comparing self-reported contraceptive use in patients with breast (n = 1,664), cervical (n = 2,182), ovarian (n = 182), and endometrial (n = 182) cancer, with self-reported contraceptive use in 1,492 control patients diagnosed with cancers with no known relationship to hormonal contraceptive use. We adjusted for potential confounding factors, including age, calendar year of diagnosis, education, smoking, alcohol, parity/age at first birth, and number of sexual partners. Among controls, 26% had used injectable and 20% had used oral contraceptives. For current and more recent users versus never users of oral or injectable contraceptives, the odds ratios (ORs) for breast cancer were significantly increased in users of oral and/or injectable contraceptives (OR 1.66, 95% CI 1.28–2.16, p<0.001) and separately among those exclusively using oral (1.57, 1.03–2.40, p = 0.04) and exclusively using injectable (OR 1.83, 1.31–2.55, p<0.001) contraceptives; corresponding ORs for cervical cancer were 1.38 (1.08–1.77, p = 0.01), 1.01 (0.66–1.56, p = 0.96), and 1.58 (1.16–2.15, p = 0.004). There was no significant increase in breast or cervical cancer risk among women ceasing hormonal contraceptive use ≥10 y previously (p = 0.3 and p = 0.9, respectively). For durations of use ≥5 y versus never use, the ORs of ovarian cancer were 0.60 (0.36–0.99, p = 0.04) for oral and/or injectable contraceptive use and 0.07 (0.01–0.49, p = 0.008) for injectable use exclusively; corresponding ORs for endometrial cancer were 0.44 (0.22–0.86, p = 0.02) and 0.36 (0.11–1.26, p = 0.1).

Conclusions

In this study, use of oral and of injectable hormonal contraceptives was associated with a transiently increased risk of breast and cervical cancer and, for long durations of use, with a reduced risk of ovarian and endometrial cancer. The observed effects of injectable and of oral contraceptives on cancer risk in this study did not appear to differ substantially. Please see later in the article for the Editors'' Summary     

Recent Arguments Against Germ Cell Renewal in the Adult Human Ovary: Is an Absence of Marker Gene Expression Really Acceptable Evidence of an Absence of Oogenesis?

In 2004, a study from our lab published in the journal Nature re-ignited a worldwide debate over the validity of the dogma that mammalian females are incapable of oocyte and follicle production during postnatal life. Amidst widespread skepticism, we forged ahead and published a second study in 2005 in the journal Cell, which not only reaffirmed with different experimental approaches that this dogma is invalid but also identified cells in bone marrow (BM) and blood of adult female mice that could generate oocytes contained within immature follicles in the ovaries of recipient females following transplantation. Although this work has been the subject of extensive critical commentary as well, two recent reports from others have confirmed the germline potential of adult BM-derived cells in mice. Further, independent corroboration of the results and conclusions presented in our earlier Nature paper is also now available. However, two papers have been published that purportedly question our work and conclusions. The first is a paper by Eggan et al. published in the journal Nature, which attempts to draw conclusions about the germline potential of BM-derived cells after focusing solely on ovulated eggs while ignoring what may be occurring at the level of oogenesis in the ovaries. The second is a report from Liu et al. just released in the journal Developmental Biology that claims to provide evidence refuting the possibility that adult female mammals produce new oocytes. However, all of the data presented in this latter report are derived from gene expression studies that the authors say fail to show the occurrence of meiosis or germ cell mitosis in adult human ovaries. Given that more than three years have past since our initial study challenging the dogma was published, it is our belief that continuing arguments against the possibility of postnatal oogenesis in mammals should be based on more rigorous experimental approaches than simply an absence of evidence, especially from gene expression analyses. Further, the interpretations offered by Liu et al. of their results are not as straightforward as they contend since some of their data can also be viewed as supportive of postnatal oogenesis in reproductive age women.     

Association Between Single Nucleotide Polymorphisms of Sterol Regulatory Element Binding Protein-2 and Liver X Receptor α Gene and Risk of Polycystic Ovary Syndrome in a Chinese Han Population

To investigate associations of single nucleotide polymorphisms (SNPs) rs2228314 of sterol regulatory element-binding protein-2 (SREBP-2) or rs11039155 of liver X receptor α (LXRα) with susceptibility to polycystic ovary syndrome (PCOS) in a Chinese Han population. SREBP-2 rs2228314 and LXRα rs11039155 polymorphisms were genotyped in patients with PCOS and age- and sex-matched PCOS-free controls from a Chinese Han population. A total of 605 patients with PCOS and 615 controls were recruited in this study. We found that GC and CC genotypes of rs2228314, and variant C, were associated with a significantly increased risk of PCOS. In addition, GA and AA genotypes of rs11039155, as well as variant A, were also associated with a significantly increased risk of PCOS. Our results showed that SREBP-2 rs2228314 G to C change and variant C genotype as well as LXRα rs11039155 G to A change and variant A may contribute to PCOS in Chinese Han population.