Biomedical subjects

M C Magli

Publications and source records attributed to M C Magli.

At least 19 recordsLinked to original sources

PubMed2006

Oocyte euploidy, pronuclear zygote morphology and embryo chromosomal complement.

BACKGROUND: Pronuclear morphology has been proposed as an indicator of embryo development and chromosomal complement. In this study, the morphology of pronuclear zygotes generated from euploid oocytes [diagnosed by first polar body (PB1) analysis] was evaluated and compared with the configurations observed in chromosomally normal embryos (diagnosed by blastomere analysis). MATERIALS AND METHODS: Group 1--238 patients underwent 273 assisted conception cycles in combination with the screening of aneuploidy on PB1 for the chromosomes 13, 15, 16, 18, 21 and 22. Only normal oocytes were inseminated. Group 2--218 patients underwent 318 assisted conception cycles with aneuploidy screening on day 3 embryos. In both groups, oocytes were checked for fertilization and pronuclear morphology at 16 h after insemination. RESULTS: Seventy-three percent of zygotes from Group 1 had the configurations with centralized and juxtaposed pronuclei, large-size aligned or scattered nucleoli and PB located in the longitudinal or perpendicular axis of pronuclei. In Group 2, these configurations corresponded to those with the highest proportion of chromosomally normal embryos. Accordingly, in both groups, these configurations had a higher implantation rate than all the others. CONCLUSIONS: These observations confirm that some patterns of pronuclear morphology are associated with a higher proportion of euploidy and implantation reaffirming the relevance of this scoring system for the prediction of zygote viability.

Adult↗

PubMed2006

Cryopreservation of biopsied embryos at the blastocyst stage.

BACKGROUND: The availability of an efficient cryopreservation program is especially important in the case of embryos that have undergone blastomere biopsy for PGD. Unfortunately, the freezing/thawing of biopsied embryos has given disappointing results when performed at the cleavage stage. In this study, embryos diagnosed as normal after PGD were grown to the blastocyst stage, frozen and thawed for successive frozen embryo transfer. METHODS: A total of 34 patients performed a thawing cycle in which 47 blastocysts were thawed. The cryopreservation solutions were based on HEPES-buffered medium supplemented with human serum albumin (HSA), sucrose and 1,2-propanediol. The same protocol was applied to embryos from 88 IVF/ICSI patients, which underwent 92 thawing cycles with 150 thawed blastocysts. RESULTS: The survival rate was similar in the two groups (53% after PGD and 58% in IVF/ICSI cycles), as well as the cumulative pregnancy rate per patient (59% after PGD versus 47% in IVF/ICSI cycles), despite a higher maternal age and a lower proportion of embryos available for transfer or cryopreservation in the PGD group. CONCLUSIONS: Neither the survival rate nor the subsequent development and chances of implantation, differed between embryos frozen at the blastocyst stage following biopsy and those frozen intact.

Biopsy↗

PubMed2004

ESHRE PGD Consortium data collection IV: May-December 2001.

The ESHRE PGD Consortium was formed in 1997 to survey the practice of preimplantation genetic diagnosis (PGD). Since then, three reports have been published giving an overview on PGD from an ever-increasing number of centres and reporting on an increasing number of PGD cycles and pregnancies and babies born after PGD. After these initial influential publications, important shortcomings were identified primarily on the method of data collection, i.e. with Excel spreadsheets, and in the timing of the collection (cycles were collected in a different time frame from pregnancies and babies, making the follow-up of cycles very difficult). This is why the Steering Committee has made a major investment in developing and implementing a new database in FileMaker Pro 6. It was also decided that cycles would be collected from one calendar year, as well as the pregnancies and babies ensuing from that particular calendar year. This gave us the opportunity to take a closer look at the data collected earlier, and to attempt to improve their quality. This is a report on the corrected data from the first three data collections (I-III) as well as the result of the last data collection (IV) that was completely carried out using the new database.

Data Collection↗

PubMed2004

Development and clinical application of a strategy for preimplantation genetic diagnosis of single gene disorders combined with HLA matching.

Preimplantation HLA matching has recently emerged as a tool for couples desiring to conceive a potential donor progeny for transplantation in a sibling with a life-threatening disorder. In this paper we describe a strategy optimized for preimplantation genetic diagnosis (PGD) of haemoglobinopathies combined with HLA matching. This procedure involves a minisequencing-based genotyping of HLA regions A, B, C and DRB combined with mutation analysis of the gene regions involved by mutation. Analysis of at least eight polymorphic short tandem repeat (STR) markers scattered through the HLA complex has also been included to detect potential contamination and crossing-over occurrences between HLA genes. The above assay can also be used for preimplantation HLA matching as a primary indication. The strategy was clinically applied for HLA matching in 17 cycles (14 for beta-thalassaemia, one for Wiscott-Aldrich syndrome and two for leukaemia). A reliable HLA genotype was achieved in 255/266 (95.9%) of the blastomeres. In total, 22 (14.8%) embryos were obtained that were HLA-matched with the affected siblings, 14 (9.4%) of which were unaffected and transferred back to the patients. Four clinical pregnancies were obtained, three of which (one twin, two singletons) are ongoing and were confirmed as healthy and HLA-identical with the affected children. Minisequencing-based HLA typing combined with HLA STR haplotyping has been shown to be a reliable strategy for preimplantation HLA matching. The major advantage of this approach is that the validation of a single assay can be done once and then used for the majority of the patients, reducing notably time needed for preclinical set-up of each case.

Blastocyst↗

PubMed2004

Prognostic role of preimplantation genetic diagnosis for aneuploidy in assisted reproductive technology outcome.

BACKGROUND: Preimplantation genetic diagnosis (PGD) for aneuploidy is recommended to couples at risk of generating chromosomally abnormal embryos. The aim of this study was to demonstrate that PGD for aneuploidy has an important role in the prognosis of subsequent treatments. METHODS: A total of 389 couples underwent their first PGD for aneuploidy due to either female age >or=38 years (n = 266) or >or=3 previous unsuccessful cycles (n = 123). After the first PGD followed by an unsuccessful treatment cycle, 141 couples underwent 175 subsequent PGD cycles. These patients were divided into three groups depending on the number of euploid embryos available for transfer in their first PGD cycle: group A included patients where no euploid embryos were diagnosed; group B included patients who had only one euploid embryo; and group C included patients with at least two normal embryos resulting from chromosomal analysis. RESULTS: In subsequent cycles, group A patients underwent significantly fewer transfers (45%) compared with group B (69%, P < 0.05) and group C patients (85%, P < 0.001). The pregnancy rate per transfer was significantly decreased in group A (15%) compared with group B (36%; P < 0.02) and group C (30%; P < 0.03). Accordingly, the live birth rate per patient was significantly lower in group A compared with group C (8.5% versus 30%; P < 0.005). CONCLUSIONS: The outcome of the first PGD for aneuploidy may have a predictive role for subsequent attempts.

Adult↗

PubMed2004

Sperm aneuploidy and recurrent pregnancy loss.

Experiments of double target in-situ hybridization were performed separately for chromosomes 1-17, 8-18 and sex chromosomes on sperm samples from 20 couples suffering from three or more recurrent first trimester abortions. For a subset of this study population, additional experiments of multicolour fluorescence in-situ hybridization for chromosomes 4, 7, 12, 13, 15, 18, 21, and 22, were performed on the bases of the available data from abortive tissue karyotyping. A markedly high rate of sperm disomy (14.5-15.5%) was scored in only two cases. For three other patients, the cumulative disomy rates for chromosomes 1, 17, 8, 18, X and Y also increased but at a lower level (7.8-9.5%). For the remaining 15 patients, the frequency of sperm aneuploidy was moderately increased or normal. Men with recurrent pregnancy loss (RPL) and poor semen quality had baseline sperm aneuploidy and diploidy rates higher than men with normal semen parameters (with or without RPL). Using probes for chromosomes 1, 17, 8, 18, X and Y, significantly elevated frequencies of sperm aneuploidy (not diploidy) were found in 10% of men with a history of RPL. Their rate of sperm aneuploidy was 30-34%. For the other men, changes in sperm aneuploidy were not thought to affect RPL. Poor semen quality per se impacted negatively on sperm aneuploidy and diploidy, thus making the interpretation of clinical data more difficult.

Abortion, Habitual↗

PubMed2003

Clinical value of preimplantation genetic diagnosis.

The clinical application of preimplantation genetic diagnosis (PGD) has provided an alternative approach for the prevention of affected pregnancies in couples at high reproductive risk. The frequent contribution of genetic factors to infertility problems makes PGD of particular value for assisted reproductive practices. In addition, the selection of euploid embryos for transfer has a strong impact on IVF efficiency as aneuploidies are the main cause of spontaneous abortions and implantation failures. In this study, the clinical outcome in PGD cycles is presented. The list of monogenic disorders for which PGD is performed is rapidly extending and the safety of the procedure has lead to an increasing interest among couples at high reproductive risk. Following PGD for aneuploidy, a higher implantation rate and a lower incidence of spontaneous abortions are obtained in patient categories where aneuploidy is a prominent cause of reproductive failure. In view of these findings, PGD has become an integral part of assisted reproductive techniques for the prevention of affected pregnancies and improvement of IVF efficiency.

Adult↗

PubMed2003

The minisequencing method: an alternative strategy for preimplantation genetic diagnosis of single gene disorders.

We have applied a new method of genetic analysis, called 'minisequencing', to preimplantation genetic diagnosis (PGD) of monogenic disorders from single cells. This method involves computer-assisted mutation analysis, which allows exact base identity determination and computer-assisted visualization of the specific mutation(s), and thus facilitates data interpretation and management. Sequencing of the entire PCR product is unnecessary, yet the same qualitative characteristics of sequence analysis are maintained. The main benefit of the minisequencing strategy is the use of a mutation analysis protocol based on a common procedure, irrespective of the mutations involved. To evaluate the reliability of this method for subsequent application to PGD, we analysed PCR products from 887 blastomeres including 55 PGD cases of different genetic diseases, such as cystic fibrosis, beta-thalassaemia, sickle cell anaemia, haemophilia A, retinoblastoma, and spinal muscular atrophy. Minisequencing was found to be a useful technique in PGD analysis, due to its elevated sensitivity, automation, and easy data interpretation. The method was also efficient, providing interpretable results in 96.5% (856/887) of the blastomeres tested. Fifteen clinical pregnancies resulted from these PGD cases; conventional prenatal diagnosis confirmed all the PGD results, and 10 healthy babies have already been born. Its applicability to PGD could be helpful, particularly in cases in which the mutation(s) involved are difficult to assess by restriction analysis or other commonly used methods.

Blastomeres↗

PubMed2002

Possible interchromosomal effect in embryos generated by gametes from translocation carriers.

BACKGROUND: The incidence of abnormal pregnancies in carriers of balanced translocations depends strictly on the chromosomes involved in the translocations. The aim of this study was to verify whether conventional aneuploidy screening could be advantageously combined with preimplantation genetic diagnosis (PGD) for translocations. METHODS: Twenty-eight carriers of Robertsonian and reciprocal translocations underwent 43 PGD cycles; specific probes were used to screen the translocation in 172 embryos generated by 35 cycles; most of these embryos were also screened for chromosomes 13, 16, 18, 21, 22 (n = 166), XY (n = 107), 1 (n = 17) and 15 (n = 88). For the remaining eight cycles (carriers of reciprocal translocations) only the chromosomes involved in common aneuploidy screening were investigated on the 40 embryos generated in vitro. RESULTS: In Robertsonian translocations, the proportion of embryos with abnormalities due to the translocation was 21%, common aneuploidies contributed 31% of total abnormalities, whereas the remaining 36% of embryos had abnormalities due to both types of chromosome. For reciprocal translocations, the chromosomes involved in the translocation were responsible for 65% of total abnormalities; only 6% of the embryos were abnormal for common aneuploidies and 16% carried abnormalities due to both the chromosomes involved in the translocation and those not related to the translocation. CONCLUSIONS: An interchromosomal effect seems to play a role in the case of Robertsonian translocations, where the relevant contribution of aneuploidy exposes the couple to an additional risk of abnormal pregnancy.

Aneuploidy↗

PubMed2001

The in vivo and in vitro efficiency and efficacy of PGD for aneuploidy.

Preimplantation genetic diagnosis for aneuploidy was implemented on 1782 morphologically normal embryos generated in vitro by patients with a poor prognosis of pregnancy. Only 592 of them (34%) were diagnosed as chromosomally normal. Embryo transfer was accomplished in 240 cycles resulting in 79 clinical pregnancies (33%) and an implantation rate of 22.6%. The in vitro efficiency of the procedure was established by analysing all the blastomeres obtained from 311 non transferrable embryos and resulted to be 97.1%. The in vivo efficiency of the technique was calculated with the data derived from the prenatal diagnoses by examination of the infants at birth and was 97.8%. In consideration of the reported inaccuracy rate, patients are still recommended to undergo prenatal diagnosis. The transfer of PGD selected embryos in women of advanced reproductive age reduces by half the risk of having a trisomic pregnancy.

Adult↗

PubMed2001

Chromosomal abnormalities in embryos.

Chromosomal analysis was performed on 1620 embryos generated in vitro by patients with a poor prognosis of pregnancy. A diagnosis was yielded in 1596 embryos: 536 (34%) were euploid and 1060 (66%) carried chromosomal abnormalities. The results revealed a strong association between chromosomal abnormalities, cellular stage and percentage of fragmentation. In addition, 92% of embryos with multinucleated cells were diagnosed mosaics, whereas the presence of cytoplasmic concentration was associated to 86% chromosomal abnormalities. The rate of development to expanded blastocysts was dependent on both the cleavage stage at the time of blastomere biopsy and the chromosomal status of the embryos. The highest percentage of blastocyst growth derived from embryos with 7-8 cells on the morning of day 3: 68% were generated from euploid embryos and 43% from chromosomally abnormal embryos. These findings suggest that morphological criteria alone are not sufficient in poor prognosis patients to guarantee the best embryo selection.

Aneuploidy↗

PubMed2001

Double locus analysis of chromosome 21 for preimplantation genetic diagnosis of aneuploidy.

Preimplantation genetic diagnosis (PGD) of numerical chromosome abnormalities significantly reduces spontaneous abortions and may increase pregnancy rates in women of advanced maternal age undergoing in vitro fertilization. However, the technique has an error rate of around 10% and trisomy 21 conceptions have occurred after PGD. To further reduce the risk of transferring trisomy 21 embryos to the patient, we designed a protocol that analyzes chromosome 21 twice by targeting two different loci. This protocol was applied to 388 embryos from 60 cycles of PGD of aneuploidy. The scoring criterion used was based on giving equal importance to both probe results. Of the 242 embryos diagnosed as abnormal, 125 were re-biopsied to assess the rate of false positives and false negatives of the protocol and their clinical relevance. The results of the present study showed no reduction in the overall fluorescent in situ hybridization (FISH) error rate for single cells. However, by using a different scoring criterion, the incidence of false negative can be reduced to 1.6% without missing any trisomy 21. In addition, the present study suggests that if two or more loci from the same chromosome could be simultaneously analyzed in single cells, errors caused by false monosomies could be reduced.

Aneuploidy↗

PubMed2001

Transvaginal ovarian drilling: a new surgical treatment for improving the clinical outcome of assisted reproductive technologies in patients with polycystic ovary syndrome.

OBJECTIVE: To evaluate the efficacy of transvaginal ovarian drilling (TVOD) in patients with polycystic ovary syndrome (PCOS) who were undergoing IVF treatment. DESIGN: Pilot study. SETTING: Reproductive medicine unit. PATIENT(S): Eleven patients with PCOS undergoing treatment with assisted reproductive technology (ART). INTERVENTION(S): Selection criterion for TVOD was repeated poor performance in > or =2 previous IVF cycles. MAIN OUTCOME MEASURE(S): Controlled ovarian hyperstimulation parameters, number of eggs collected, fertilization rate, embryo cleavage rate, implantation rate, pregnancy rate compared with the cycles before TVOD. RESULT(S): In the cycle after TVOD, a significantly higher dosage of FSH was used (33.5 +/- 12 IU vs. 52.2 +/- 15 IU) to collect a higher number of oocytes in the presence of similar E2 values at the day of hCG administration. This resulted in significantly higher fertilization and cleavage rates (27% vs. 66% and 54% vs. 72%, respectively). The pregnancy and the implantation rates after TVOD were similar to those for normovulatory patients undergoing IVF for tubal factor infertility during the study period. CONCLUSION(S): Our data suggest that the TVOD is effective in improving IVF results in difficult to treat patients with PCOS, and it is less invasive and less expensive when compared with laparoscopic ovarian diathermy.

Adult↗

PubMed2000

Female poor responders.

Female poor responders are represented by normovulatory women showing a 'gonadal failure' in term of inadequate number of recruited follicles under conventional controlled ovarian hyperstimulation (COH) for assisted reproductive technologies (ART). ARTs offers today a high chance of pregnancy to infertile couples when a normal ovarian response provides a large choice of embryos for transfer. On the contrary, failure of the ovary to produce enough oocyte for treatment, reduces significantly the likelihood of conceiving in ART, not only in the treatment cycle, but also predicting a poor prognosis in subsequent cycles. Up to date, poor response remains one of the most frequent problems in the field of assisted reproduction. First described in 1981, poor response has been investigated by several authors, but many aspects are still controversial. In this paper definition, pathophysiology and management of poor response are revised and discussed.

Estradiol↗

PubMed2000

Gonadal activity and chromosomal constitution of in vitro generated embryos.

Chromosomal analysis of pre-implantation embryos was carried out in patients with a poor prognosis of full term pregnancy, which underwent induction of multiple follicular growth. In all, 1034 embryos generated from 191 stimulated cycles were screened for nine chromosome aneuploidy by using the multicolour fluorescence in situ hybridisation technique. Thirty-five percent of the diagnosed embryos were chromosomally normal, whereas the remaining presented with numerical abnormalities, which made them not suitable for transfer. The results obtained confirmed that the incidence of abnormalities is mostly dependent on age; however, monosomy and trisomy are more frequent in poor responders. Accordingly, the pregnancy rate per started cycle was significantly higher in women with a normal response to gonadotropic stimulation (33% vs. 8%, P<0. 001). These findings indicate that poor responder patients are physiologically exposed not only to reduced chances of implantation, but also to an increased risk of spontaneous abortion and trisomic pregnancies.

Adult↗

PubMed2000

Preimplantation diagnosis after assisted reproduction techniques for genetically-determined male infertility.

One hundred and thirty-six cycles with a poor prognosis for full-term pregnancy underwent preimplantation genetic diagnosis (PGD) of aneuploidy. The mean maternal age was 31.8 +/- 2.5 years. Only patients younger than 36 years were included in the study with the aim of evaluating whether sperm indices have an effect on the chromosomal constitution of preimplantation embryos. No differences were detected in the percentage of aneuploid embryos; however a higher incidence of monosomies and trisomies was found in MESA-TESE embryos compared to the group of normospermic patients. In addition, an increase in the proportion of gonosomal aneuploidy seemed to be associated with the severity of the male factor parameters. The rate of de-novo chromosomal abnormalities in embryos from patients with a normal karyotype suggested an increased frequency proportional to the severity of the male factor condition, the proportion of monosomic and trisomic embryos, and the percentage of gonosomal aneuploidy increased accordingly. In the case of couples with a male altered karyotype, comparable frequency of chromosomally abnormal embryos, and monosomy and trisomy were observed irrespective of semen indices, gonosomal aneuploidy was only observed in one case where the patient had a karyotype with gonosomal mosaicism. These data confirm that the severe male infertility condition determines an increase in the rate of de-novo abnormalities, as anticipate by the follow-up of the children born after ICSI.

Adult↗

PubMed2000

Developmental potential of somatic stem cells in mammalian adults.

Traditionally, somatic tissue-derived stem cells of mammalian adults have been viewed as pluripotent precursors capable of lifelong maintenance of cellular compartments typical of the tissue in which they reside. However in recent years, in vitro cultures and in vivo transplantation assays have indicated that adult somatic stem cell of various species are capable of adopting multiple fates. Bone marrow cells can give rise to a wide array of phenotypes, including blood, endothelial, bone, cartilage, fat, tendon, lung, liver, muscle, marrow stroma, and even brain cells. Conversely, neural stem cells as well as progenitors present in the muscle may contribute to blood cell production, indicating that adult stem cells present in numerous tissues may generate multiple cell types even of different dermal origin. Therefore, the developmental potential of adult somatic stem cells might be reassessed, although the mechanisms that ultimately lead to determination of cell fate are not completely defined. The successful long-term culturing and expansion of somatic adult stem cells together with their intrinsic versatility leads to future hope of stem cell therapeutic use in a wide spectrum of diseases and disorders of several, even not easily accessible, tissues.

Adult↗

PubMed2000

ESHRE guidelines for good practice in IVF laboratories. Committee of the Special Interest Group on Embryology of the European Society of Human Reproduction and Embryology.

Education has always been a priority for the European Society of Human Reproduction and Embryology (ESHRE). Many efforts have been dedicated to promoting knowledge of techniques, procedures and strategies in order to ensure use of the highest quality practices in reproductive medicine. The need to develop a set of guidelines was a logical consequence that found its first expression in 1990, when Focus on Reproduction (vol. 1, pp. 10-38) published the first guidelines which were distributed among the membership. Five years later a new, more complete edition with several novel techniques and developments appeared in Human Reproduction (vol. 10, pp. 1246-1271). Both have proved to be invaluable references. Five more years have now passed. The necessity to produce current guidelines for good IVF laboratory practice has provided the strongest motivation. This originated from the increasing awareness that embryologists have a duty to prevent unintentional incidents that might result from poor practice in the laboratory. Therefore, the Embryology Special Interest Group (SIG) undertook to draw up guidelines aimed at giving support and guidance to the laboratory staff. All the aspects required to provide a safe working system were taken into consideration by members of the SIG and their effort produced this document. We hope that it will assist staff in achieving the best clinical outcome for their patients.

Cryopreservation↗