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M Wilding

Publications and source records attributed to M Wilding.

At least 19 recordsLinked to original sources
PubMed2006

Intra-follicular leptin concentration as a predictive factor for in vitro oocyte fertilization in assisted reproductive techniques.

BACKGROUND: Granulosa-cells are able to produce and store leptin, suggesting that this hormone is locally involved in the regulation of follicular growth. In this study, the role of follicular fluid (FF) leptin concentration in predicting oocyte fertilization and embryo quality was evaluated in 35 normogonadotrophic women undergoing controlled ovarian stimulation (COS) for assisted reproductive techniques. MATERIALS AND METHODS: Leptin concentration was measured in 47 consecutively collected FF in which a mature oocyte had been found during the ovum pick-up. Embryos deriving from fertilized oocytes were submitted to quality scoring systems. RESULTS: Mean leptin concentration was significantly higher in FF whose oocytes showed 2 pronuclei (no. 25) when compared with those with no evidence of fertilization (no. 22) at the 16-18 h check (26.0+/-6.1 vs 15.3+/-10.6 ng/ml, respectively, p<0.01). Follicular mean diameters were similar in the two groups (21.4+/-3.4 and 21.0+/-5.1 mm, respectively). Logistic regression analysis identified FF leptin levels as the best predictive parameter for oocyte fertilization (p<0.001). When receiving operating characteristics curve was employed, a FF leptin concentration of 20.25 ng/ml was the most reliable cut-off in predicting fertilization of oocytes. FF with leptin concentrations higher than this value (no. 27) had an oocyte fertilization rate of 85.7%. In contrast, FF levels < or =20.25 ng/ml (no. 20) were associated with a rate of 16.7% (p<0.05). No correlation emerged between FF leptin and the score attributed to 15 valuable embryos at the zygote stage (r=-0.01) and at 48 h after insemination (r=0.1). CONCLUSIONS: FF leptin levels are a better predictor of oocyte fertilization success rates than follicular diameter. These results underline the relevance of FF variables in developing methods for oocyte selection.

Adult↗
PubMed2006

Fetal gonadotrope cell origin of FSH-secreting pituitary adenoma - insight into human pituitary tumour pathogenesis.

OBJECTIVE: The pathogenesis of human pituitary adenomas remains unclear, but we report a case of FSH-secreting pituitary adenoma whose monohormonal phenotype suggests it was of fetal origin. PATIENTS: A 28-year-old woman presented with abdominal discomfort and irregular menses, enlarged multicystic ovaries and elevated serum oestradiol, with sustained high-normal FSH and low LH levels. MEASUREMENTS: Endocrine studies were performed before and after curative surgery, with assessment of tumour hormone secretion in vitro, and immunostaining of tumour tissue for a series of gonadotrope proteins. RESULTS: Immunocytochemistry showed that tumour cells were monohormonal for FSH. Normal components of gonadotrope signalling pathways were expressed, including oestrogen receptor-alpha, activin receptors, secretogranin-II and chromogranin-A. beta-glycan, the putative inhibin coreceptor, was absent. Tumour culture in vitro confirmed secretion of FSH with minimal LH, that was unsuppressed by oestradiol or inhibin-A. Human fetal pituitary tissue contained FSH-only cells at 18 weeks gestation, whereas normal adult pituitary tissue contained only bihormonal gonadotropes. CONCLUSIONS: We propose that this pituitary adenoma represents an indolent tumour of monohormonal fetal gonadotrope cells that originated early in gestation. Pituitary tumours may therefore arise from abnormal persistence of fetal cell types, with extremely slow growth over many years until reaching a size threshold to generate an endocrine syndrome. Understanding fetal pituitary architecture and function may be more informative for new insights into pituitary tumour pathogenesis than classical theories of cancer biology that invoke unrestrained cell proliferation.

Adenoma↗
PubMed2005

Phosphorylation of p27(BBP)/eIF6 and its association with the cytoskeleton are developmentally regulated in Xenopus oogenesis.

p27BBP/eIF6 is an evolutionarily conserved regulator of ribosomal function. It is necessary for 60S biogenesis and impedes improper joining of 40S and 60S subunits, regulated by protein kinase C or Efl1p. No data on p27BBP/eIF6 during early development of Metazoa are available. We studied the distribution, post-translational changes and association with the cytoskeleton of p27BBP/ eIF6 during Xenopus oogenesis and early development. Results indicate that p27BBP/eIF6 is present throughout oogenesis, partly associated with 60S subunits, partly free and with little cytoskeleton bound. During prophase I, p27BBP/eIF6 is detected as a single band of 27-kDa. Upon maturation induced by progesterone or protein kinase C, a serine-phosphorylated 29 kDa isoform appears and is kept throughout development to the neurula stage. Confocal microscopy showed that the distribution of p27BBP/eIF6 and its association with the cytoskeleton varies according to oogenesis stages. Briefly, in stage 6 oocytes, p27BBP/eIF6 has a limited dot-like distribution, and does not co-localize with cytokeratin, whereas upon maturation it spreads throughout the cytoplasm. After fertilization, a large fraction coalesces around cytomembranes and a cytochalasin B-sensitive co-localization with cytokeratin occurs. RNAse removes p27BBP/eIF6 from the cytokeratin fibres. Developmental data suggest a role of p27BBP/eIF6 in controlling ribosomal availability or regulating cross-talk between ribosomes and the cytoskeleton.

Animals↗
PubMed2004

Effects of recombinant LH (rLH) supplementation during controlled ovarian hyperstimulation (COH) in normogonadotrophic women with an initial inadequate response to recombinant FSH (rFSH) after pituitary downregulation.

BACKGROUND: This study was aimed to evaluate the effect of different recombinant LH (rLH) doses on the ovarian outcome of normogonadotrophic women with an initial inadequate response to recombinant FSH (rFSH) after pituitary downregulation. METHODS: Only women undergoing a 'long protocol' with a GnRH-agonist followed by rFSH administration were enrolled. On the eighth day of stimulation, 46 patients with serum E2 levels < 180 pg/ml and with no follicle > 10 mm were randomized in two groups to receive a supplementation with a daily rLH dose of 75 (group A) or 150 IU (group B), respectively. Forty-six normal responders continuing their monotherapy with rFSH formed the control group (C). RESULTS: The mean number of oocytes retrieved and the percentage of mature oocytes in the group B (9.65 +/- 2.16, 79.0%) were comparable with those observed in the group C (10.65 +/- 2.8, 82.5%) and significantly higher when compared with the group A (6.39 +/- 1.53, 65.7%). The mean number of ampoules of rLH was significantly higher in the group B (14.4 +/- 2.0 vs. 9.65 +/- 1.1), whereas these patients received a significantly lower mean number of rFSH ampoules (44.6 +/- 7.4 vs. 36.1 +/- 3.8). Seven (30.4%), 9 (39.1%) and 22 (47.8%) pregnancies were achieved in the groups A, B and C, respectively. CONCLUSIONS: These results suggest that patients with initial inadequate responses to rFSH after pituitary downregulation benefit from the addition of a daily dose of 150 IU of rLH, starting from the eighth day of stimulation.

Adult↗
PubMed2002

The effect of ease of transfer and type of catheter used on pregnancy and implantation rates in an IVF program.

PURPOSE: To test the effects of type of embryo transfer catheter, transfer difficulty, and observations after the transfer procedure on pregnancy and implantation rates in an IVF programme. METHODS: Patients were prepared for IVF using standard protocols. Embryo transfer was performed using either Edwards-Wallace or TDT catheter. The difficulty of transfer was graded by a clinician and biologist. Blood observed inside the catheter after the transfer procedure was scored as endometrial damage. Pregnancy and implantation rates were scored. RESULTS: Type of embryo transfer catheter and the observation of blood did not significantly affect pregnancy and implantation rates when transfer was performed by a single operator. CONCLUSIONS: In the hands of experienced, skilled operators, neither choice of transfer catheter and difficulty of transfer nor observations of blood on the transfer catheter caused any significant reduction in outcome to the patient.

Adult↗
PubMed2001

Pressure-induced amorphization and an amorphous-amorphous transition in densified porous silicon.

Crystalline and amorphous forms of silicon are the principal materials used for solid-state electronics and photovoltaics technologies. Silicon is therefore a well-studied material, although new structures and properties are still being discovered. Compression of bulk silicon, which is tetrahedrally coordinated at atmospheric pressure, results in a transition to octahedrally coordinated metallic phases. In compressed nanocrystalline Si particles, the initial diamond structure persists to higher pressure than for bulk material, before transforming to high-density crystals. Here we report compression experiments on films of porous Si, which contains nanometre-sized domains of diamond-structured material. At pressures larger than 10 GPa we observed pressure-induced amorphization. Furthermore, we find from Raman spectroscopy measurements that the high-density amorphous form obtained by this process transforms to low-density amorphous silicon upon decompression. This amorphous-amorphous transition is remarkably similar to that reported previously for water, which suggests an underlying transition between a high-density and a low-density liquid phase in supercooled Si (refs 10, 14, 15). The Si melting temperature decreases with increasing pressure, and the crystalline semiconductor melts to a metallic liquid with average coordination approximately 5 (ref. 16).

Journal Article↗
PubMed2001

Enzyme activity in anuran spermatozoa upon induction of the acrosome reaction.

At the time of sperm-egg fusion in Discoglossus pictus, a large amount of electron-dense material of an unknown nature is liberated from the sperm. In the present work we studied this material in D. pictus sperm, using an assay utilising strips of autoradiographic film as a gelatin substrate for proteolytic enzymes. Upon treatment with A23187, D. pictus sperm produced a large halo on the gelatin substrate, indicating the presence of enzymes released by the sperm at the time of the acrosome reaction. In contrast, Xenopus laevis sperm did not produce halos upon treatment with A23187. The use of protease inhibitors such as TLCK, leupeptin, chymostatin, SBTI and EACA strongly suggests that the D. pictus whole acrosome contains trypsin and chymotrypsin activity while an SBTI-sensitive activity is absent in a small portion of the acrosome, possibly the anteriormost region. Furthermore, the material released at the acrosome reaction also contains an EACA-inhibited activity, indicating the presence of plasminogen activator. We conclude that D. pictus sperm release proteolytic enzyme(s) that may act at the egg surface at the time of gamete fusion.

Acrosome↗
PubMed2001

Confocal microscopy analysis of the activity of mitochondria contained within the 'mitochondrial cloud' during oogenesis in Xenopus laevis.

We have used ratiometric confocal microscopy and three fluorescence techniques to study the distribution and activity of mitochondria in frog oocytes during the early stages of oogenesis. Mitochondria in frog oocytes during oogenesis were characterised by a high ratio in the 'mitochondrial cloud' and perinuclear region and a low ratio in mitochondria freely dispersed within the cytoplasm. We tested whether the high ratio visualised by the three techniques represented mitochondrial membrane potential by perturbing the mitochondrial membrane potential. Carbonyl cyanide p-(trifluoromethyl)phenylhydrazone (FCCP) caused the immediate destruction of the membrane potential, and consequent loss of fluorescence from the membrane-potential-sensitive confocal channel. In contrast, nigericin caused an increase in membrane potential represented by a steady increase in fluorescence ratio. These data demonstrate that mitochondrial activity can be measured during oogenesis in frog oocytes, and suggest that the mitochondrial cloud and perinuclear regions are characterised by highly active mitochondria.

Animals↗
PubMed2001

Mitochondrial aggregation patterns and activity in human oocytes and preimplantation embryos.

Mitochondria play a vital role in the metabolism of energy-containing compounds in the oocyte cytoplasm to provide adenosine trisphosphate for fertilization and preimplantation embryo development. In this study, ratiometric confocal microscopy with the mitochondrion-specific membrane potential-sensitive fluorescence dye JC-1 (5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolyl-carbocyanine iodide) was used to measure the activity of mitochondria in human oocytes and developing preimplantation embryos. Mitochondria in oocytes and embryos were characterized by distinct localized aggregation patterns. These patterns however did not determine localized regions of heterogeneity in mitochondrial activity. Mitochondrial activity was analysed during oocyte maturation and after fertilization. The activity of mitochondria in fresh metaphase II oocytes was negatively correlated with maternal age. This trend continued when the activity of developing embryos was analysed. Mitochondrial activity was strongly correlated with the rate of embryo development on day 3 after fertilization, but not on day 2. Partial regression analysis showed that the rate of cleavage of preimplantation embryos was more highly correlated with embryo mitochondrial activity than maternal age. These data suggest that the efficiency of mitochondrial respiration in oocytes and preimplantation embryos is closely correlated with the programmed rate of embryo development, and suggest that maternal age further influences this factor. The loss of mitochondrial activity in oocytes obtained from ageing couples may therefore contribute to lower embryo development and pregnancy rates observed during cycles of IVF.

Benzimidazoles↗
PubMed2001

The peritoneal fluid concentration of leptin is increased in women with peritoneal but not ovarian endometriosis.

This study was designed to measure leptin concentrations in the peritoneal fluid (PF) of women with different aspects of pelvic endometriosis. Among 36 consecutive women undergoing laparoscopy, nine were diagnosed as having minimal-mild endometriosis (stage I-II). Among nine other subjects with advanced stage (III-IV) disease, six showed one or more ovarian endometriotic cysts as the only operative finding. The remaining 18 unaffected women constituted the control group. Patients with endometriosis had significantly higher PF leptin concentrations (32.6 +/- 16.2 versus 17.1 +/- 6.6 ng/ml, P = 0.002); this difference remained significant when corrected for body mass index (BMI) (PF leptin/BMI ratio 1.41 +/- 0.67 versus 0.76 +/- 0.28, P = 0.001). Furthermore, the PF leptin/BMI ratio was significantly higher in women with peritoneal implants than in those in whom no implant was found at laparoscopy (1.6 +/- 0.7 versus 0.83 +/- 0.33, P = 0.007). Conversely, patients with one or more ovarian endometriomata as the only finding, had a PF leptin/BMI ratio comparable with that in women where no cyst was found (1.05 +/- 0.4 versus 1.1 +/- 0.65). In women with stage I-II endometriosis, a higher mean PF leptin/BMI ratio was found compared with those affected by stage III-IV (1.78 +/- 0.68 versus 1.05 +/- 0.43, P = 0.01). These results show that during endometriosis the presence of peritoneal disease, and not of ovarian endometriotic cysts, influences leptin concentrations in PF. The data suggest that leptin may play a role in the development of peritoneal endometriosis, and that different biochemical phenomena might be involved in the pathogenesis of the ovarian form of the disease.

Adult↗
PubMed2001

Pregnancy after cytoplasmic transfer in a couple suffering from idiopathic infertility: case report.

Couples suffering from idiopathic infertility are among the most difficult to treat in IVF cycles due to the unknown cause of failure to reproduce. This can lead to years of unsuccessful attempts to conceive and often the abandonment of treatment. In some couples, the only observable problem is the development of poor quality embryos. In this case report, we describe the successful use of cytoplasmic donation in a couple where the high level of embryo fragmentation and poor embryo development was thought to be the unique cause of failure to conceive after intracytoplasmic sperm injection (ICSI). We noted a strong reduction in embryo fragmentation and a consequent increase in embryo quality after the treatment. Transfer of four of the embryos receiving donor cytoplasm led to a pregnancy with two gestational sacs and the birth of healthy twins.

Adult↗
PubMed2001

Rescue of IVF cycles by HMG in pituitary down-regulated normogonadotrophic young women characterized by a poor initial response to recombinant FSH.

BACKGROUND: The aim of this study was to investigate the effects of adding human menopausal gonadotrophin (HMG) during controlled ovarian stimulation in normoovulatory normogonadotrophic patients showing an initial suboptimal response to a standardized long protocol therapy with recombinant FSH (rFSH) (300 IU/day). METHODS: A total of 43 such patients were randomized in two groups. In Group A, 150 IU rFSH was substituted by 150 IU HMG after day 8 of stimulation. The stimulation protocol of Group B involved a simple increase of the daily rFSH dose to 375 IU after day 8. A total of 40 BMI and age matched patients with an optimal ovarian response formed the control group (Group C). RESULTS: The mean Group A serum concentration of oestradiol on the day of HCG administration and average number of oocytes retrieved were significantly higher than Group B (P < 0.001) and equivalent to Group C. A total of 10 pregnancies (50%) in Group A, 8 (34.8%) in Group B and 19 (47.5%) in the control group were achieved. CONCLUSIONS: The data suggest that LH supplementation improves the ovarian outcome in patients characterized by an inadequate initial response to rFSH therapy in a long protocol.

Adult↗
PubMed2000

Interaction of cell cycle kinases, microtubules, and chromatin in ascidian oocytes during meiosis.

We used kinase assays and confocal microscopy to study the interaction of cell cycle proteins with microtubule organising centres (MTOC) and chromatin in ascidian oocytes during meiosis. The activity of maturation promoting factor (MPF) and mitogen activated protein kinase (MAPK) appear not to be correlated in control oocytes. MPF activity peaks during metaphase I and II of the meiotic cell cycle whereas the activity of MAPK peaks at telophase I and is subsequently degraded to remain at low levels for the remainder of meiosis. The protein synthesis inhibitor emetine induces the degradation in MPF activity in unfertilized metaphase-I (M-I) oocytes, while MAPK is unaffected. Emetine does not alter the activities of these cell cycle kinases in fertilized oocytes during meiosis I but MPF activity remains low while MAPK activity is high for an elongated time period and oocytes do not complete meiosis I. Emetine induces maternal MTOC duplication in unfertilized M-I oocytes and prevents sperm aster growth in fertilized oocytes, but it does not alter the M-I meiotic apparatus in unfertilized oocytes. These experiments suggest that neither MPF alone nor emetine-sensitive proteins are responsible for M-I arrest in ascidian oocytes, MAPK may ensure this stability. In addition, we showed that the maternal MTOC is present at M-I but suppressed from duplicating in an emetine-sensitive manner.

Animals↗
PubMed2000

Meiosis-associated calcium waves in ascidian oocytes are correlated with the position of the male centrosome.

We have used confocal microscopy to measure calcium waves and examine the distribution of tubulin in oocytes of the ascidian Ciona intestinalis during meiosis. We show that the fertilisation calcium wave in these oocytes originates in the vegetal pole. The sperm penetration site and female meiotic apparatus are found at opposite poles of the oocyte at fertilisation, confirming that C. intestinalis sperm enter in the vegetal pole of the oocyte. Following fertilisation, ascidian oocytes are characterised by repetitive calcium waves. Meiosis I-associated waves originate at the vegetal pole of the oocyte, and travel towards the animal pole. In contrast, the calcium waves during meiosis II initiate at the oocyte equator, and cross the oocyte cytoplasm perpendicular to the point of emission of the polar body. Immunolocalisation of tubulin during meiosis II reveals that the male centrosome is also located between animal and vegetal poles prior to initiation of the meiosis II-associated calcium waves, suggesting that the male centrosome influences the origin of these calcium transients. Ascidians are also characterised by an increase in sensitivity to intracellular calcium release after fertilisation. We show that this is not simply an effect of oocyte activation. The data strongly suggest a role for the male centrosome in controlling the mechanism and localisation of post-fertilisation intracellular calcium waves.

Animals↗
PubMed1999

The ins and outs of meiosis.

During oogenesis the oocyte is arrested in meiosis twice. First at prophase I, then a second time at metaphase I in many invertebrates and in metaphase II in the vast majority of vertebrates. Meiosis resumption is triggered by the sperm. This article examines mechanisms that cause oocytes' arrest in meiosis and how spermatozoa help the oocyte to get out of this cellular predicament. J. Exp. Zool. (Mol. Dev. Evol.) 285:226-236, 1999.

Animals↗
PubMed1999

Nicotinamide alters the calcium release pattern and the degradation of MPF activity after fertilisation in ascidian oocytes.

Fertilisation in ascidian oocytes triggers a plasma membrane current, the release of intracellular calcium and the degradation of Maturation Promoting Factor (MPF) activity leading to the completion of meiosis and the initiation of embryo development. We have previously shown that the fertilisation current in ascidians is produced through the metabolism of nicotinamide nucleotide (NN) metabolites to ADP ribose. In this study we have used nicotinamide to test whether NN metabolism plays additional roles in fertilisation in ascidians. Nicotinamide treatment blocked calcium-induced calcium release (CICR) and arrested the cell cycle prior to the completion of meiosis I. Nicotinamide further prevented the abolition of MPF activity after fertilisation. Interestingly, nicotinamide treatment caused ascidian oocytes to form interphase-like pronuclei after fertilisation, despite the high MPF activity. The data demonstrate that NN metabolism is involved in calcium signalling through CICR and further suggest that a NN metabolite acts as a messenger connecting MPF activity to the formation of the meiotic apparatus.

Animals↗
PubMed1999

Pregnancies after activated oocyte transfer: a new option for infertility treatment.

In this preliminary report, we describe a new technique involving the same-day transfer of activated oocytes to the uterus after intracytoplasmatic sperm injection (ICSI). The technique, termed activated oocyte transfer (AOT), offered to 19 couples, yielded a pregnancy rate per cycle of about 30%, equivalent to traditional in-vitro fertilization (IVF) and ICSI in a laboratory setting. AOT is performed 4 h after oocyte retrieval, permitting the patient to undergo treatment as an out-patient procedure.

Adult↗