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Biomedical subjects

A N Shelling

Publications and source records attributed to A N Shelling.

At least 19 recordsLinked to original sources
PubMed2006

Ovarian reserve tests for predicting fertility outcomes for assisted reproductive technology: the International Systematic Collaboration of Ovarian Reserve Evaluation protocol for a systematic review of ovarian reserve test accuracy.

BACKGROUND: The presence of a wide range of tests of ovarian reserve suggests that no single test provides a sufficiently accurate result. Many tests are used without reference to an evidence base. So far, individual studies conducted on these tests are too small to give precise estimates of prognostic accuracy. OBJECTIVES: To systematically assess the accuracy of the available tests of ovarian reserve in terms of prediction of fertility outcomes. SEARCH STRATEGY: The search will be conducted using the name of the respective index test being studied (as listed on the MESH database), if more than 2000 citations are listed, 'ovary' and or 'ovarian', 'fertility' and or 'reserve' will be combined with the original search term as required. Studies of the accuracy of tests of ovarian reserve will be obtained without language restrictions from 1980 to 2005 using the following electronic databases and Ovid software: MEDLINE, EMBASE, PUBmed, Biological extracts, Pascal, Cochrane Library (CDSR, DARE, CCTR, HTA), Best Evidence databases, SCISEARCH, Conference Proceedings (ISI Proceedings, Healthstar, Current Contents, Science Citation Index, Cancerlit and Econlit and NHS Economic Evaluation database. The National Research Register, the Medical Research Council's Clinical Trials Register, MEDION, DARE, and the US Clinical Trials register. SELECTION CRITERIA: Studies will be selected if accuracy of tests are compared with a reference standard and include data that can be abstracted into a two-by-two table to calculate sensitivity and specificity. The studies to be included in this review will examine one of the following index 'tests' within a study population of women undergoing assisted reproductive technology: * Clinical variables--age, history of cancelled cycles. * Basal blood tests--follicle-stimulating hormone (FSH), lutenising hormone (LH), FSH:LH ratios, estradiol (E(2)), inhibin A and B, progesterone (P(4)), P(4):E(2) ratios, antimullerian hormone, testosterone, vascular endothelial growth factor, insulin-like growth factor-1:insulin-like growth factor binding protein-1 ratios. * Dynamic tests--clomiphene citrate challenge test, gonadotropin analogue stimulating test, exogenous FSH ovarian reserve test. * Ultrasound tests-antral follicle count, ovarian volume, ovarian stromal peak systolic velocity, including waveform and pulsatility index, ovarian follicular vascularity. * Histology--ovarian biopsy. Data collection and analysis Two independent reviewers will perform quality assessment and data extraction. Prognostic accuracy will be determined by calculating positive and negative likelihood ratios for the following outcomes or reference standards: live birth, ongoing pregnancy, clinical pregnancy, biochemical pregnancy, embryos available for transfer, eggs obtained at oocyte retrieval, cycles cancelled prior to oocyte retrieval. Main results and conclusions N/A.

Data Collection↗
PubMed2005

Expression of TGF-beta1, TGF-beta2, TGF-beta3 and the receptors TGF-betaRI and TGF-betaRII in placentomes of artificially inseminated and nuclear transfer derived bovine pregnancies.

Bovine nuclear transfer pregnancies are characterized by a high incidence of placental abnormalities, notably, increased placentome size and deficiencies in trophoblast cell function and establishment of placental vasculature. Alterations in gene expression during placental growth and development may contribute to the appearance of large placentomes in pregnancies derived from nuclear transfer. The placenta synthesizes a number of cytokines and growth factors, including the transforming growth factor-betas (TGF-betas) that are involved in the establishment, maintenance and/or regulation of pregnancy. All forms of TGF-beta and their receptors are present at the fetal-maternal interface of the bovine placentome, where they are thought to play an important role in regulating growth, differentiation, and function of the placenta. Using real-time RT-PCR, we have examined the expression of TGF-beta1, TGF-beta2, TGF-beta3 and the receptors TGF-betaRI and TGF-betaRII in placentomes of artificially inseminated (AI) and nuclear transfer (NT)-derived bovine pregnancies at days 50, 100 and 150 of gestation. TGF-beta1, TGF-beta2 and TGF-beta3 mRNA expression increased by 2.0-2.8-fold, while TGF-betaRI and TGF-betaRII mRNA expression decreased by 1.7-2.0-fold in NT placentomes compared to AI controls at all gestational ages examined. These findings indicate that NT placentomes may be resistant to the growth suppressive effects of TGF-betas and could contribute to the placental proliferative abnormalities observed in NT-derived placentas. Alternatively, deficiencies in placentation may provide a mechanism whereby TGF-betas are dysregulated in NT pregnancies.

Activin Receptors, Type I↗
PubMed2005

Blepharophimosis and bilateral Duane syndrome associated with a FOXL2 mutation.

This case describes the novel coexistence of sporadic blepharophimosis-ptosis-epicanthus inversus syndrome (BPES) and bilateral type I Duane syndrome in a female infant, with a FOXL2 mutation. Mutational analysis of FOXL2 demonstrated a 30-nucleotide duplication (c.672(-)701dup30) within the polyalanine tract of FOXL2. The association of BPES and Duane syndrome represents a novel phenotype which may suggest a greater pleiotropic effect of FOXL2 in development. During the period of the 4-8th week of embryonic development, the cranial nerves, their nuclei and the corresponding innervation to the extraocular muscles develop, the extraocular muscles undergo development and differentiation. This coincides with the period of time that FOXL2 is expressed strongly in the developing eyelids and the surrounding tissues. Forkhead genes are transcription factors and likely to be involved in signal transduction pathways. This case expands the spectrum of FOXL2 mutations associated with BPES.

Base Sequence↗
PubMed2004

SPRASA, a novel sperm protein involved in immune-mediated infertility.

BACKGROUND: Antisperm antibodies (ASA) may be an important cause of infertility, but current tests for the detection of ASA have poor prognostic value. Identification of the sperm proteins that ASA bind to may aid the development of more useful diagnostic tests. METHODS: One- and two-dimensional PAGE and western blotting analyses, as well as amino acid sequencing, were used to identify a novel sperm protein reactive with ASA (SPRASA) from infertile men. An antiserum reactive with SPRASA was produced by immunizing a rabbit with SPRASA excised from two-dimensional gels. This antiserum was used to demonstrate the localization of SPRASA on the sperm. RESULTS: Amino acid sequences derived from SPRASA matched those of a theoretical protein, XP-085564. This protein is derived from the C-type lysozyme/alpha-lactalbumin gene family. Immunohisto chemistry indicates that SPRASA is localized to the acrosome. Western blot analysis revealed that 50 unselected individuals did not have antibodies that reacted with SPRASA. CONCLUSION: Only ASA from infertile men react with SPRASA, suggesting that this novel protein may be important in the processes of fertility. The identification of SPRASA as the antigen for infertility-associated ASA raises the possibility of developing first, antigen-specific tests for ASA, and secondly, more targeted treatment for immune-mediated infertility.

Acrosome↗
PubMed2004

Epigenetic regulation of inhibin alpha-subunit gene in prostate cancer cell lines.

Inhibin was first identified as a gonad-derived regulator of pituitary FSH; however, it has subsequently been shown to be a tumour suppressor in the gonad and adrenal glands. Whereas non-malignant regions of human primary prostate carcinomas express inhibin alpha-subunit (INHA), malignant tissues lack INHA transcript and protein, which is consistent with epigenetic regulation of the inhibin alpha-subunit gene (INHA) promoter. This study investigated whether methylation of the INHA promoter was responsible for inactivation of INHA transcription and translation in the prostate cancer cell lines, LNCaP, DU145 and PC3. Methylation of the promoter was revealed by bisulphite genomic sequencing and use of inhibitors of methylation and histone deacetylation resulted in reactivation of the INHA transcription and translation. Significant (P<0.05) downregulation of a luciferase reporter gene downstream from a methylated INHA promoter compared with unmethylated INHA promoter occurred in vitro. The data demonstrate that promoter methylation is associated with downregulation of the INHA gene in prostate cancer cell lines, which is consistent with its tumour suppressive role. Therefore INHA has a significant role in prostate tumorigenesis.

Base Sequence↗
PubMed2003

Activity, synthesis, storage, and messenger RNA of cyclooxygenase in intrauterine tissues of guinea pigs near term and during labor.

Whether the reported gestation-dependent increase in cyclooxygenase activity in gestational tissues is due to an accumulation of cyclooxygenase in vivo or an increasing capacity to synthesize cyclooxygenase in vitro is unknown. In this study in guinea pigs, COX activity was estimated from the net production rates of prostaglandins E(2) and F(2alpha) in the presence of optimal substrate concentrations. Cyclooxygenase activity in amnion increased between 45 days of gestation and labor in microsomes (150-fold in relation to PGF(2alpha) production and 116-fold in relation to PGE(2) production) and in tissue explants (42-fold in relation to PGF(2alpha) production). The capacity for de novo synthesis of cyclooxygenase after aspirin treatment increased nine-fold between 45 days of gestation and labor in amnion explants. Comparison of COX activity in amnion explants with or without prior aspirin treatment showed that COX activity is at least three-fold higher in controls than would be expected if the activity was due to de novo synthesis alone. Cyclooxygenase-2 mRNA predominated in amnion but neither cyclooxygenase-2 nor cyclooxygenase-1 mRNA levels (semi-quantitative RT-PCR) changed significantly. This suggests that the gestation-dependent increase in cyclooxygenase activity in guinea pig amnion is due in part to accumulation of cyclooxygenase in vivo, that COX-2 predominates, and that COX activity is not correlated with levels of COX mRNA.

Animals↗
PubMed2002

Identification of novel mutations in FOXL2 associated with premature ovarian failure.

Premature ovarian failure (POF) affects approximately 1% of women and is known to be caused by sex chromosome abnormalities, iatrogenic agents and autoimmune diseases, but in the majority of cases the cause is unknown. However, several families have been identified as having an inherited predisposition to POF, suggesting a genetic component to the condition in these cases. The FOXL2 gene of 70 POF patients from New Zealand and Slovenia was screened for mutations. In a Slovenian POF patient, a novel 30 bp deletion was identified that was predicted to remove 10 out of 14 alanines (A221_A230del), from the polyalanine tract downstream of the winged helix/forkhead domain of the FOXL2 protein. A novel single nucleotide substitution, 772(1009)T>A, which is predicted to change amino acid 258 from tyrosine to asparagine (Y258N), was identified in a New Zealand POF patient. Neither mutation was identified in 200 normal control chromosomes from 100 control samples. Three previously unreported single nucleotide substitutions, considered to be non-functional polymorphisms, were also identified.

Amino Acid Sequence↗
PubMed2001

Analysis of the TGF beta functional pathway in epithelial ovarian carcinoma.

Epithelial ovarian carcinoma is often diagnosed at an advanced stage of disease and is the leading cause of death from gynaecological neoplasia. The genetic changes that occur during the development of this carcinoma are poorly understood. It has been proposed that IGFIIR, TGFbeta1 and TGFbetaRII act as a functional unit in the TGFbeta growth inhibitory pathway, and that somatic loss-of-function mutations in any one of these genes could lead to disruption of the pathway and subsequent loss of cell cycle control. We have examined these 3 genes in 25 epithelial ovarian carcinomas using single-stranded conformational polymorphism analysis and DNA sequence analysis. A total of 3 somatic missense mutations were found in the TGFbetaRII gene, but none in IGFRII or TGFbeta1. An association was found between TGFbetaRII mutations and histology, with 2 out of 3 clear cell carcinomas having TGFbetaRII mutations. This data supports other evidence from mutational analysis of the PTEN and beta-catenin genes that there are distinct developmental pathways responsible for the progression of different epithelial ovarian cancer histologic subtypes.

Carcinoma↗
PubMed2001

Molecular genetics of ovarian cancer.

This is a review of the approaches that can be used to analyze genetic changes in ovarian cancer. Traditional gene localization methods are discussed, followed by a section on gene identification techniques. Once a putative disease-associated gene has been cloned, mutations have to be identified and analyzed. There are numerous mutation detection methods, and the most common ones are outlined. In the penultimate section, the role of immunohistochemistry as a surrogate method for mutation analysis is considered. Finally, the possible use of functional assays is discussed. At present, it would appear that DNA chip technology for the detection of mutations, and microarray analysis of gene expression, are two important techniques likely to have a significant impact on the genetic analysis of ovarian cancer.

Animals↗
PubMed2000

Hierarchical mutation screening protocol for the BRCA1 gene.

The identification of mutations in the BRCA1 gene poses difficulties in achieving a screening outcome that satisfies the twin needs of speed and accuracy. These needs must also take into account the patient's family history and the statistical evaluation of the probability of detecting a mutation. Given the above, we present here a hierarchical mutation screening strategy that comprises two tiers: first, multiplex heteroduplex and exon 13 duplication analysis; second, exon amplification and direct sequencing using a 96-well tray format. The advantages of this strategy are two-fold: first, the division of analytical tools in order to achieve low and high-resolution mutation screening, respectively; second, a streamlined sequencing approach that leads to a sensitive and rapid assay that reduces labor costs and handling errors. The success of this approach is shown by the identification of a novel deletion mutation in exon 14 of the BRCA1 gene, which was not detected by the more conventional protein truncation assay due to the small size of the predicted truncated protein.

Adult↗
PubMed2000

Evaluation of the possible protective role of adeno-associated virus type 2 infection in HPV-associated premalignant disease of the cervix.

OBJECTIVE: Our objective was to examine the prevalence of adeno-associated virus (AAV) infection in women with normal cervical smears and those with HPV-associated cervical intraepithelial neoplasia (CIN). METHODS: HPV typing was performed on DNA from cervical smears of 211 women with CIN (CIN 1 = 83, CIN 3 = 128) and 433 healthy women who had a normal cervical smear. HPV typing was performed on all cases and controls using type-specific oligonucleotide primers (HPV 16, 18, 31, 33). AAV DNA was amplified by nested PCR from the same samples. The amplified DNA were separated on 2% agarose gels, blotted, and hybridized to AAV-2 DNA labeled by random priming with [alpha-(32)P]dCTP to confirm specificity of amplification. RESULTS: A total of 131 cases of CIN were positive for one of the HPV types either alone or in combination. HPV 16 was present in 120 (57%) cases, HPV 18 in 15 (7%), HPV 31 in 27 (13%), and HPV 33 in 15 (7%) and there were multiple HPV types detected in 34 (16%) cases. All of the controls were selected to be negative for HPV. A total of 6/433 (1.4%) control cervical smears and 4/211 (1.9%) of CIN (CIN1 = 2; CIN3 = 2) contained AAV DNA. No correlation between AAV and any clinical feature was observed. CONCLUSIONS: These results are different from some that have been previously published and suggest that AAV DNA is not frequently present in either normal control cervical samples or cervical intraepithelial neoplasia. This does not support the hypothesis that AAV may be protective against cervical cancer. Further research is necessary to understand the natural history of AAV infection and its role in human disease.

Adenovirus Infections, Human↗
PubMed2000

Inhibin: a candidate gene for premature ovarian failure.

Premature ovarian failure (POF) occurs in 1% of all women, and in 0.1% of women under the age of 30 years. The mechanisms that give rise to POF are largely unknown. Inhibin has a role in regulating the pituitary secretion of FSH, and is therefore a potential candidate gene for ovarian failure. Using single-stranded conformation polymorphism (SSCP) and DNA sequencing, DNA samples were screened from 43 women with POF for mutations in the three inhibin genes. Two variants were found: a 1032C-->T transition in the INHssA gene in one patient, and a 769G-->A transition in the INHalpha gene in three patients. The INHssA variant appears to be a polymorphism, as there was no change in the amino acid sequence of the gene product. The INHalpha variant resulted in a non-conservative amino acid change, with a substitution from alanine to threonine. This alanine is highly conserved across species, and has the potential to affect receptor binding. The INHalpha variant is significantly associated with POF (3/43 patients; 7%) compared with control samples (1/150 normal controls; 0.7%) (Fisher's exact test, P < 0.035). Further analysis of the inhibin gene in POF patients and matched controls will determine its role in the aetiology of POF.

Adult↗
PubMed2000

Radiation-induced cell cycle delays and p53 status of early passage melanoma cell lines.

Cell cultures exposed to DNA-damaging agents such as gamma radiation respond by arresting at cell cycle checkpoints, and the p53 tumor suppressor protein is strongly implicated in this behavior. We have investigated the TP53 status and cell cycle response to ionizing radiation of a series of early passage cell lines (designated NZM1 to NZM15) previously developed from patients with metastatic melanoma. The TP53 status of each of the cell lines was determined by single-strand conformation polymorphism and DNA sequence analysis. The majority of the lines appeared to have a wild-type TP53 gene sequence, consistent with published studies. Two lines (NZM4 and NZM7.2) were found to have an identical T-->C transition mutation in nucleotide 721 (exon 7) of the coding region. NZM7.2 (mutant) and NZM7.4 (wild-type) were clonally derived from the same line (NZM7). The existence of radiation-induced cell cycle arrest in G and/or G2M phase was determined 16 h after irradiation (6.3 Gy) by DNA staining and flow cytometric analysis. The mitotic inhibitor paclitaxel was used as a reference compound, with or without irradiation, to assess the efficiency of radiation-induced cell cycle arrest. G1 phase arrest was associated only with the presence of the wild-type TP53 gene, but the efficiency of induced arrest varied among the cell lines and the period of G phase arrest appeared to be short. A significant difference (P < 0.002) was also found between the efficiency of induction of G2 phase arrest and the presence of wild-type TP53 gene. The results provide evidence that although the melanoma cell lines generally had an intact TP53 gene, the efficiency of p53-mediated cycle arrest might be deficient and contribute to the resistance of this tumor to treatment.

Adult↗
PubMed1997

Analysis of loss of heterozygosity and KRAS2 mutations in ovarian neoplasms: clinicopathological correlations.

The molecular events that give rise to ovarian epithelial neoplasms are not well understood. In particular, it is not known whether adenocarcinomas arise from benign or low malignant potential (LMP) precursors. We have examined a large series of benign (25) and LMP (31) ovarian tumors for loss of heterozygosity (LOH) at multiple loci on 17 chromosomes. LOH was observed in benign tumors on chromosomes 6 (14%) and 9 (5%) and on the X chromosome (33%) only. LOH on these chromosomes was also detected in a small number of LMP neoplasms, suggesting that these may derive sometimes from benign precursors. In addition, we examined LOH in 93 adenocarcinomas. Analysis of associations between LOH events showed that LOH on chromosomes 5 and 17 (P = 0.0002) and on chromosomes 17 and 18 (P = 0.00007) were associated significantly with each other, which suggests that these may represent cooperative, progressive events. No novel significant associations were identified between LOH events and stage, grade, or histology, which would indicate the existence of genetic heterogeneity in ovarian neoplasms. KRAS2 mutations were detected more often in LMP neoplasms than in malignant tumors (P = 0.004) and were detected more often in Stage I/II malignant tumors than in Stage III/IV malignant tumors (P = 0.033), suggesting that LMP tumors with KRAS2 mutations are unlikely to progress to frank malignancy. Univariate (but not multivariate) survival analysis showed that LOH of chromosomes 11 (P = 0.039) and 17 (P = 0.04) was associated with a significantly worse prognosis. Replication of these novel findings is necessary, and the identification, isolation, and characterization of the critical genes affected by LOH will determine their importance in the pathogenesis of ovarian malignancies.

Chromosome Deletion↗