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Biomedical subjects

Karen Sermon

Publications and source records attributed to Karen Sermon.

10 recordsLinked to original sources
PubMed2006

Preimplantation genetic diagnosis for Marfan syndrome.

OBJECTIVE: To develop and apply efficient and reliable protocols for preimplantation genetic diagnosis (PGD) for Marfan syndrome. DESIGN: Two mutation-specific protocols were developed, and the markers D15S1028, D15S992, D15S196, D15S576, D15S123, and D15S143 were used to set up four multiplex polymerase chain reactions (PCRs). SETTING: Research Center Reproduction and Genetics. PATIENT(S): Ten couples carrying mutations in the FBN1 gene. INTERVENTION(S): Six PGD protocols were developed for 10 couples, and 7 of them underwent a total of 16 clinical cycles. MAIN OUTCOME MEASURE(S): Amplification, allele drop-out (ADO), and contamination rates during the preclinical assays. DNA analyses of blastomeres from embryos biopsied during PGD cycles. RESULT(S): Six different protocols were set up, with the main objective being to to use one protocol for several couples. A total of 16 PGD cycles were performed, which resulted in the delivery of an unaffected boy and three ongoing pregnancies. CONCLUSION(S): The development of single-cell multiplex PCRs for linked markers and its use in PGD reduce the workload of the genetic diagnostic laboratory as well as the average waiting time for patients. This approach also allows for the simultaneous and accurate detection of recombination, contamination, and ADO, thereby increasing the reliability of the diagnosis.

Adult↗
PubMed2006

Single-cell chromosomal imbalances detection by array CGH.

Genomic imbalances are a major cause of constitutional and acquired disorders. Therefore, aneuploidy screening has become the cornerstone of preimplantation, prenatal and postnatal genetic diagnosis, as well as a routine aspect of the diagnostic workup of many acquired disorders. Recently, array comparative genomic hybridization (array CGH) has been introduced as a rapid and high-resolution method for the detection of both benign and disease-causing genomic copy-number variations. Until now, array CGH has been performed using a significant quantity of DNA derived from a pool of cells. Here, we present an array CGH method that accurately detects chromosomal imbalances from a single lymphoblast, fibroblast and blastomere within a single day. Trisomy 13, 18, 21 and monosomy X, as well as normal ploidy levels of all other chromosomes, were accurately determined from single fibroblasts. Moreover, we showed that a segmental deletion as small as 34 Mb could be detected. Finally, we demonstrated the possibility to detect aneuploidies in single blastomeres derived from preimplantation embryos. This technique offers new possibilities for genetic analysis of single cells in general and opens the route towards aneuploidy screening and detection of unbalanced translocations in preimplantation embryos in particular.

Aneuploidy↗
PubMed2006

The interface between assisted reproductive technologies and genetics: technical, social, ethical and legal issues.

The interface between assisted reproductive technologies (ART) and genetics comprises several sensitive and important issues that affect infertile couples, families with severe genetic diseases, potential children, professionals in ART and genetics, health care, researchers and the society in general. Genetic causes have a considerable involvement in infertility. Genetic conditions may also be transmitted to the offspring and hence create transgenerational infertility or other serious health problems. Several studies also suggest a slightly elevated risk of birth defects in children born following ART. Preimplantation genetic diagnosis (PGD) has become widely practiced throughout the world for various medical indications, but its limits are being debated. The attitudes towards ART and PGD vary substantially within Europe. The purpose of the present paper was to outline a framework for development of guidelines to be issued jointly by European Society of Human Genetics and European Society of Human Reproduction and Embryology for the interface between genetics and ART. Technical, social, ethical and legal issues of ART and genetics will be reviewed.

Ethics, Medical↗
PubMed2006

Selection of reference genes in mouse embryos and in differentiating human and mouse ES cells.

Embryonic Stem (ES) cells have the potential to form every cell of the body and thus are of great promise for tissue transplantation. One of the rising techniques that allows studying the differentiation state of ES cells is quantitative RT-PCR (qRT-PCR). When relative quantification by qRT-PCR is applied, accurate normalization is necessary, since differentiated embryonic stem cells and developing embryos contain heterogeneous cell populations. Corrections for variations in the qRT-PCR reaction are needed to allow comparisons between different samples. We applied the normalization tools geNorm and Normfinder to ten reference genes identifying the most stable ones for relative quantification of gene expression during differentiation of human ES cells, as well as in differentiated mouse ES cells and in the developing mouse embryo. For relative quantification by qRT-PCR in these systems, we advise to use normalization factors based on multiple stable reference genes. However, when the use of several reference genes would be unpractical, a single reference gene in each experimental setup could be sufficient. When looking for single stable reference genes, beta-actin works best in both mouse embryo and ES cell experiments and glyceraldehyde-3-phosphate-dehydrogenase can be applied in both mouse and human ES cell experiments.

Actins↗
PubMed2004

Intergenerational instability of the expanded CTG repeat in the DMPK gene: studies in human gametes and preimplantation embryos.

The CTG repeat at the 3' untranslated region of the dystrophia myotonica protein kinase (DMPK) gene shows marked intergenerational and somatic instability in patients with myotonic dystrophy (DM1), when the repeat is expanded to more than approximately 55 repeats. Intensive research has yielded some insights into the timing and mechanism of these intergenerational changes: (1) increases in expansion sizes occur during gametogenesis but probably not during meiosis, (2) the marked somatic mosaicism becomes apparent from the 2nd trimester of development onward and increases during adult life, and (3) DNA repair mechanisms are involved. We have performed preimplantation genetic diagnosis for DM1 since 1995, which has given us the unique opportunity to study the expanded CTG repeat in affected embryos and in gametes from affected patients. We were able to demonstrate significant increases in the number of repeats in embryos from female patients with DM1 and in their immature and mature oocytes, whereas, in spermatozoa and embryos from male patients with DM1, smaller increases were detected. These data are in concordance with data on other tissues from adults and fetuses and fill a gap in our knowledge of the behavior of CTG triplet expansions in DM1.

Blastocyst↗
PubMed2004

Preimplantation genetic diagnosis.

Preimplantation genetic diagnosis (PGD) was introduced at the beginning of the 1990s as an alternative to prenatal diagnosis, to prevent termination of pregnancy in couples with a high risk for offspring affected by a sex-linked genetic disease. At that time, embryos obtained in vitro were tested to ascertain their sex, and only female embryos were transferred. Since then, techniques for genetic analysis at the single-cell level, involving assessment of first and second polar bodies from oocytes or blastomeres from cleavage-stage embryos, have evolved. Fluorescence in-situ hybridisation (FISH) has been introduced for the analysis of chromosomes and PCR for the analysis of genes in cases of monogenic diseases. In-vitro culture of embryos has also improved through the use of sequential media. Here, we provide an overview of indications for, and techniques used in, PGD, and discuss results obtained with the technique and outcomes of pregnancies. A brief review of new technologies is also included.

Chromosome Aberrations↗
PubMed2002

Preimplantation genetic diagnosis for Huntington's disease with exclusion testing.

Huntington's disease is an autosomal dominant, late-onset disorder, for which the gene and the causative mutation have been known since 1993. Some at-risk patients choose for presymptomatic testing and can make reproductive choices accordingly. Others however, prefer not to know their carrier status, but may still wish to prevent the birth of a carrier child. For these patients, exclusion testing after prenatal sampling has been an option for many years. A disadvantage of this test is that unaffected pregnancies may be terminated if the parent at risk (50%) has not inherited the grandparental Huntington gene, leading to serious moral and ethical objections. As an alternative, preimplantation genetic diagnosis (PGD) on embryos obtained in vitro may be proposed, after which only embryos free of risk are replaced. Embryos can then be selected, either by the amplification of the CAG repeat in the embryos without communicating results to the patients (ie non-disclosure testing), which brings its own practical and moral problems, or exclusion testing. We describe here the first PGD cycles for exclusion testing for Huntington's disease in five couples. Three couples have had at least one PGD cycle so far. One pregnancy ensued and a healthy female baby was delivered.

Female↗
PubMed2002

Preimplantation genetic diagnosis for fragile Xa syndrome: difficult but not impossible.

BACKGROUND: In this paper, we review our clinical preimplantation genetic diagnosis (PGD) programme for fragile Xa syndrome, analysing if PGD for these couples is still a valuable option, as it is particularly difficult for two reasons. First, the couples have to be informative (the number of triplet repeats on the healthy FMR-1 allele of the mother has to be different from the number of repeats on the healthy FMR-1 allele of the father) and second, women with a premutation are at increased risk of premature ovarian failure. METHODS: A total of 34 couples attended our genetics department between December 1998 and July 2001, requesting information about PGD for fragile Xa syndrome. RESULTS: Eight couples decided not to go further with the procedure and of the 26 remaining couples, 16 were informative (61.5%). Four couples have so far not started ovarian stimulation, one patient was totally refractive to stimulation and 11 couples have had a total of 19 oocyte retrievals. From these, there have been 13 embryo transfers with a clinical pregnancy rate per embryo transfer of 23%; the implantation rate was 13.6% and the live birth rate per couple was 27.3%. CONCLUSIONS: PGD for fragile Xa is feasible for a number of couples. A pre-PGD work-up should include a determination of the premutation or mutation carrier status, the maternal or paternal origin of the premutation and an estimation of the ovarian reserve of the patient. Fragile Xa premutation carriers should be advised not to postpone reproduction for too long.

Adult↗
PubMedDate not supplied

Current concepts in preimplantation genetic diagnosis (PGD): a molecular biologist's view.

The first clinically applied preimplantation genetic diagnosis (PGD) was reported more than a decade ago and since then PGD has known an exponential growth. This first report described the use of PCR to sex embryos from couples at risk for X-linked diseases. Not surprisingly, in the first years, the development of PCR-based tests led to PGD for well-known monogenic diseases such as cystic fibrosis and thalassaemia. When fluorescent in-situ hybridization (FISH) was introduced it quickly replaced PCR-based methods, which had led to misdiagnoses, for sexing of embryos. FISH was also quickly introduced for aneuploidy screening, which has as its main aim the improvement of IVF results in patients with poor reproductive outcome, and later for PGD in translocation carriers. In this review, PGD for patients with a pre-existing genetic risk will be discussed, i.e. the monogenic diseases and the translocations, as well as different biopsy methods and promising new developments.

Biopsy↗