Faculty Bibliography

Preimplantation genetic diagnosis was performed in 61 day 3 embryos obtained by in-vitro fertilization from seven patient carriers of haemophilia, Marfan's syndrome, Bloch-Sulzemberg syndrome (incontinentia pigmentosa) or X chromosome-linked immune deficiency, retinitis pigmentosa, and FG syndrome, which is characterized by mental retardation and hypotonia. After multiplex polymerase chain reaction, 16 embryos were diagnosed as being unaffected, and these were transferred to the uterus on the following day (day 4). Of these embryos, six (37.5%) implanted, resulting in the delivery of a singleton and a twin pregnancy, a late second trimester miscarriage (twins at week 20) and a first trimester miscarriage at week 8. All the diagnoses were confirmed by amniocentesis. We report for the first time a late day 4 transfer of biopsied human embryos undergoing preimplantation genetic diagnosis. This transfer schedule allows an extra day to perform genetic analyses on single blastomeres and to monitor any adverse effect of the biopsy procedure

OBJECTIVE: To report our experience with genetic screening of oocyte donor candidates and to determine the frequency with which significant genetic issues are identified. DESIGN: Prospective genetic screening of oocyte donor candidates. SETTING: University hospital oocyte donation program. PATIENT(S): Women presenting consecutively as volunteer oocyte donors. INTERVENTION(S): Genetic screening was performed by pedigree analysis and laboratory studies. MAIN OUTCOME MEASURE(S): Inclusion in the oocyte donor pool based on the results of clinical evaluation and laboratory tests consisting of polymerase chain reaction based mutational analysis for cystic fibrosis carrier status, cytogenetic analysis for karyotype, enzymatic assay for Tay-Sachs disease carrier status, and complete blood count and hemoglobin electrophoresis. RESULT(S): Eight (11%) of 73 oocyte donor candidates were excluded from the donor pool because of a potentially serious genetic finding. Cystic fibrosis mutations were identified in 5 candidates (7%), abnormal karyotypes were found in 2 (3.5%), and an autosomal dominant skeletal dysplasia was identified in 1 (1.4%). CONCLUSION(S): A significant proportion of women who present as candidates for oocyte donation are inappropriate for donation because of their genetic history or genetic testing results. A thorough genetic evaluation, including a history and laboratory screening, is essential to any oocyte donation program to maximize positive outcomes in pregnancies achieved through assisted means

OBJECTIVE: To review the genetics of aging specifically as it pertains to human fertility, as well as the recent advancements in the diagnosis of genetic diseases prior to embryo implantation. METHODS: A review of our own experience as well as the scientific literature with regards to the decline in female fertility with age, the success of IVF in women of older reproductive age, and the role of preimplantation genetic diagnosis (PGD) in the evaluation of the patient at risk for fetal genetic anomalies. RESULTS: The decline in female fertility occurs primarily as a result of a decline in oocyte quality as well as quantity. The frequency of chromosomal anomalies in recognized abortuses increases in parallel with the age-specific rise in the incidence of spontaneous abortions. PGD is an accurate diagnostic tool for exclusion of genetically deficient embryos prior to initiation of pregnancy. CONCLUSION: Reproductive failure in women of older age appears to be directly related to ovarian age. Recent techniques such as cytoplasmic or germinal vesicle transfer are designed to replace the senescent cellular machinery believed to be responsible for genetic errors that occur during early cell division. PGD can accurately identify embryos with genetic deficiencies prior to implantation

Introduction: Round spermatids have been studied extensively in stained and electron microscopy preparations. However, little information is available about their appearance in fresh specimens. We describe a method to identify and collect unstained round spermalids from ejaculates and testicular biopsies. Material and methods: Ejaculates (n=115) and testicular biopsies (n=7) were assessed for the presence of round cells by routine semen analysis and Testsimplets staining. With the aid of a micromanipulator, cells considered to be round spermatids according to cell size and shape, nuclear characteristics, presence of the acrosome, and cytoplasmic characteristics were collected in groups of identical diameter. They were then placed on separate microslides and analyzed by multiprobe fluorescence in situ hybridization (FISH) to test the validity of the selection process and in order to determine the range of spermatid diameter. When no spermatozoa were present in a testicular biopsy, the dissected tissue was smeared and processed by FISH to examine for hapioid cells (spermatids). If hapioid cells were observed selection of individual round cells and FISH analysis were performed. Results: Routine semen analysis and Testsimplets identified >1 × 106 round cells in 33 ejaculates: three of five azoospermic ejaculates had spermatids. Five biopsies contained spermatozoa; one had spermatids and the other showed spermatocytic arrest. The results of the smears examined by FISH confirmed the observations by the Testsimplets staining. Four ejaculates and six biopsies were processed with micromanipulation. Spermatoids chosen with the micromanipulator were 5-7.5 ?, in diameter with a spherical and homogeneous nucleus 4-6.5 ?. in diameter. An acrosome granule was occasionally observed as a light-gray spot; cytoplasm surrounded the nuclear as a thin halo. The surface of the spermalid was smooth or slightly irregular. The 1000 cells (10 slides containing 10 cells each) individually selected were analyzed by FISH!

INTRODUCTION: With the increasing success of advanced assisted reproductive technologies (AART), men with heretofore unbeatable male factor infertility are seeking treatment. However, we do not know the implications of using testicular or ejaculated sperm in AART, from patients with exposure to childhood chemotherapy or from untreated patients with germinal cell cancers. We report a morphological and cytogenetic study of the germinal cells in these patients. MATERIALS AND METHODS: This study involved four patients. Patient 1 had a seminoma with diffuse intraepithelial neoplasia. A biopsy of both testes and a post orchiectomy semen analysis were performed. Patient 2 had a pure embryonal cell carcinoma with intraepithelial neoplasia. An evaluation of the malignant testis and a post orchiectomy semen analysis were performed. Patients 3 and 4 had azoospermia as a result of chemotherapy for childhood leukemia; bilateral testis biopsies were performed to retrieve sperm for an AART cycle. On the biopsies and orchiectomy specimens, cytological analysis, eosin-negrosin viability assessment and morphological analysis by Testsimplets staining were performed. To ascertain presence of spermatids and chromosomal status of spermatozoa and spermatids, multiprobe fluorescence in situ hybridi-zation (FISH) was carried out on smears of the specimens. Electron microscopy and thin section microscopy were additionally performed. RESULTS: Semen analysis of patient 1 and 2 revealed oligo-asthenoteratozoospermia. The nonmalignant testicular biopsy in patient 1 revealed a sperm count of 200,000/ml with 0% motility and 32 % viability. In patient 2 the malignant testis revealed a sperm count of 10,000/ml with 7% motility and 60% viability; electron microscopy revealed a normal germinal cell ultrastructure. Both patients had chromosomally normal spermatozoa when biopsy tissue and semen was evaluated by FISH for chromosomes X, Y, and 18. Patients 3 and 4 had complete azoospermia on semen analysis and histologie exams of the!

Two women had infertility due to a symptomatic unicornuate uterus associated with rudimentary contralateral horn. Both carried successful pregnancies after laparoscopic resection of the horns