marina baldi - Academia.edu (original) (raw)

Papers by marina baldi

European Journal of Obstetrics & Gynecology and Reproductive Biology, 2006

Objective: The aim of the study was to propose a set of tests to clarify the diagnosis of repeate... more Objective: The aim of the study was to propose a set of tests to clarify the diagnosis of repeated implantation failure in patients undergoing in vitro fertilization (IVF). Study design: Fifty-nine patients with at least two unsuccessful IVF attempts were included in the study. Blood samples were evaluated for the presence of underlying thyroid abnormalities, antiphospholipid antibodies (aPL), increased levels of natural killer cells (NK), inherited thrombophilia and mouse embryo assay factor (MEA-f). The same tests were performed on 20 normal fertile control patients. Results: Seventy-six percent of IVF patients showed at least one abnormal result. This incidence was higher with respect to that found among control patients (45%). The prevalence of thyroid abnormalities, aPL and increased NK level was higher in IVF patients whereas no differences were observed in terms of prevalence of inherited thrombophilias and MEA-f. Conclusions: A better understanding of reproductive failure mechanisms should allow an effective diagnostic flow chart and a focused therapeutic option for patients experiencing repeated IVF failure. With this objective in mind, our data provide two important results: thyroid abnormalities, aPL and increased NK levels are more prevalent in women experiencing IVF failure. No evidence was found for an association between inherited thrombophilia and MEA-f and failure to achieve pregnancy after IVF. #

Human Reproduction, 2005

BACKGROUND: We report on our experience with preimplantation genetic diagnosis (PGD) for single g... more BACKGROUND: We report on our experience with preimplantation genetic diagnosis (PGD) for single gene disorders (SGDs), from 1999 to 2004, describing strategies and overall clinical outcome of 250 cycles in 174 couples for 23 different genetic conditions. METHODS: PGD cycles included 15 for autosomal dominant, 148 for autosomal recessive and 19 for X-linked SGDs. In addition, 68 cycles of PGD for SGDs were performed in combination with HLA matching. The strategy in each case used an initial multiplex PCR, followed by minisequencing to identify the mutation(s) combined with multiplex PCR for closely linked informative markers to increase accuracy. Linkage analysis, using intragenic and/or extragenic polymorphic microsatellite markers, was performed in cases where the disease-causing mutation(s) was unknown or undetectable. RESULTS: In 250 PGD cycles, a total of 1961 cleavage stage embryos were biopsied. PCR was successful in 3409 out of 3149 (92.4%) biopsied blastomeres and a diagnosis was possible in 1849 (94.3%) embryos. Four hundred and twenty-seven embryos were transferred in 211 cycles, resulting in 71 pregnancies (33.6% per embryo transfer), including 15 biochemical pregnancies, six spontaneous miscarriages, two ectopic pregnancies, which were terminated, and nine pregnancies which are still ongoing. The remaining pregnancies were confirmed to be unaffected and went to term without complications, resulting in the birth of 35 healthy babies. CONCLUSIONS: Minisequencing for mutation detection combined with multiplex fluorescence PCR for linkage analysis is an efficient, accurate and widely applicable strategy for PGD of SGDs. Our experience provides a further demonstration that PGD is an effective clinical tool and a useful option for many couples with a high risk of transmitting a genetic disease.

Prenatal Diagnosis, 2005

Objectives Cytogenetic analysis of spontaneous abortion samples can be limited by culture failure... more Objectives Cytogenetic analysis of spontaneous abortion samples can be limited by culture failure. Failure to grow in vitro has traditionally been suspected to be due to in vivo death of tissue associated with spontaneous abortion (SAB) or simply technical factors of growth in culture.

European Journal of Human Genetics, 2005

Recently, preimplantation genetic diagnosis (PGD) has been considered for several indications bey... more Recently, preimplantation genetic diagnosis (PGD) has been considered for several indications beyond its original purpose, not only to test embryos for genetic disease but also to select embryos for a nondisease trait, such as specific human leukocyte antigen (HLA) genotypes, related to immune compatibility with an existing affected child in need of a haematopoetic stem cell (HSC) transplant. We have optimized an indirect single-cell HLA typing protocol based on a multiplex fluorescent polymerase chain reaction (PCR) of short tandem repeat (STR) markers scattered throughout the HLA complex. The assay was clinically applied in 60 cycles from 45 couples. A conclusive HLA-matching diagnosis was achieved in 483/530 (91.1%) of the embryos tested. In total, 74 (15.3%) embryos revealed an HLA match with the affected siblings, 55 (11.4%) of which resulted unaffected and 46 (9.5%) have been transferred to the patients. Nine pregnancies were achieved, five healthy HLA-matched children have already been delivered and cord blood HSCs, were transplanted to three affected siblings, resulting in a successful haematopoietic reconstruction.

Molecular Human Reproduction, 2004

Preimplantation HLA matching has recently emerged as a tool for couples desiring to conceive a po... more 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 minisequencingbased 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 b-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 con®rmed 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.

Molecular Cytogenetics, 2008

Background: Routine cytogenetic investigations for ovarian cancers are limited by culture failure... more Background: Routine cytogenetic investigations for ovarian cancers are limited by culture failure and poor growth of cancer cells compared to normal cells. Fluorescence in situ Hybridization (FISH) application or classical comparative genome hybridization techniques are also have their own limitations in detecting genome imbalance especially for small changes that are not known ahead of time and for which FISH probes could not be thus designed.

Reproductive Biomedicine Online, 2003

The X-linked dominant form of Charcot-Marie-Tooth syndrome (CMTX) is a clinically and genetically... more The X-linked dominant form of Charcot-Marie-Tooth syndrome (CMTX) is a clinically and genetically heterogeneous hereditary disorder of the peripheral nerves caused by mutations in the GJB1 gene that encodes a gap junction protein named connexin 32 (Cx32). Clinically, CMTX is characterized by peripheral motor and sensory deficit with muscle atrophy. A couple with a previous history of pregnancy termination after being diagnosed positive for CMTX by chorionic villus sampling, was referred for preimplantation genetic diagnosis (PGD). The female partner carried the causative H94Q, characterized by a C→G substitution in codon 94 of exon 2 of the GJB1 gene. Embryos obtained after intracytoplasmic sperm injection (ICSI) were evaluated for the presence of the mother's mutation using polymerase chain reaction (PCR), followed by mutation analysis performed using the minisequencing method. Amelogenin sequences on the X and Y chromosomes were also co-amplified to provide a correlation between embryo gender and mutation presence. A single PGD cycle was performed, involving nine fertilized oocytes, five of which developed into good quality embryos useful for biopsy. Two unaffected embryos were transferred, resulting in a singleton pregnancy followed by the birth of a healthy female.

Molecular Human Reproduction, 2003

We have applied a new method of genetic analysis, called`minisequencing', to preimplantation gene... more 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 speci®c 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 bene®t 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 ®brosis, b-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 ef®cient, providing interpretable results in 96.5% (856/887) of the blastomeres tested. Fifteen clinical pregnancies resulted from these PGD cases; conventional prenatal diagnosis con®rmed 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 dif®cult to assess by restriction analysis or other commonly used methods.