Low utilization of prenatal and pre-implantation genetic diagnosis in Huntington disease - risk discounting in preventive genetics.

Huntington disease (HD) is a late-onset, fatal neurodegenerative disorder caused by a (CAG) triplet repeat expansion in the Huntingtin gene that enlarges during male meiosis. In 1996 in this journal, one of us (J. D. S.) presented a methodology to perform pre-implantation genetic diagnosis in families at-risk for HD without revealing the genetic status of the at-risk parent. Despite the introduction of accurate prenatal and pre-implantation genetic testing which can prevent transmission of the abnormal HD gene in the family permanently, utilization of these options is extremely low. In this article, we examine the decision-making process regarding genetic testing in families with HD and discuss the possible reasons for the low uptake among this group.

Pregnancy rates in sequential in vitro fertilization cycles by oocyte donors(1).

OBJECTIVE: To evaluate the clinical outcome of in vitro fertilization (IVF) treatment cycles from individual oocyte donors who underwent multiple sequential donations. METHODS: We reviewed clinical outcome data from sequential anonymous oocyte donation cycles using donors who underwent multiple IVF stimulations. Donors were grouped by the interval between cycles and the cycle number (rank). The primary outcome measure was delivery rate by individual donor per retrieval from the combined derivative fresh and frozen embryo transfers. RESULTS: Duration and amount of gonadotropin therapy and the fertilization rates did not correlate significantly with the interval between cycles or cycle rank. Cumulative delivered pregnancy rates for cycles 1-6 were 51.5%, 54.6%, 50.5%, 51.5%, 51.1%, and 57.6%, respectively. Delivered pregnancy rates did not vary by interval between cycles. CONCLUSION: Young healthy presumed or proven fertile women can reliably donate oocytes for at least six cycles with the expectation of consistently high pregnancy rates.

Preliminary study of the incidence of disomy in sperm fractions after MicroSort flow cytometry.

Using triple colour fluorescent in-situ hybridization (FISH) we have evaluated, on a blind basis, the disomy level for chromosome 21 and the sex chromosomes in flow cytometric sorted sperm samples. There were no statistically significant differences in the disomy rates when comparing the sorted samples (either for X- or Y-bearing spermatozoa) with non-sorted samples. There were no diploid spermatozoa observed in any sample group after MicroSort processing.

Human live birth and sperm-sex ratios compared.

The human secondary sex ratio is compared with the percentage of Y-chromosome bearing spermatozoa in human semen. Live birth sex ratio is about 51.3%, whereas the overall percentage of Y-chromosome bearing spermatozoa in our study samples was 50.3%, i.e. 1% closer to the proportion expected by Mendelian segregation. The observed difference between live birth and sperm-sex ratios was significant (P < 0.0001). A possible effect of male age on the percentage Y-bearing spermatozoa was found to be non-significant.

Percutaneous testicular sperm aspiration: a convenient and effective office procedure to retrieve sperm for in vitro fertilization with intracytoplasmic sperm injection.

PURPOSE: After intracytoplasmic sperm injection was established to facilitate in vitro fertilization in men with the most severe semen abnormalities, the use of testicular sperm to achieve conception became feasible. We investigated the use of a method of percutaneous needle aspiration previously used for diagnostic purposes to obtain testicular sperm for intracytoplasmic sperm injection. MATERIALS AND METHODS: A method of percutaneous aspiration of sperm was developed to facilitate intracytoplasmic sperm injection. A total of 69 testicular aspirations were performed for diagnostic purposes and 179 to obtain sperm on the day of egg retrieval for couples undergoing in vitro fertilization with intracytoplasmic sperm injection. The procedures were performed in an outpatient facility. Most patients received intravenous sedation and a few received only local anesthesia. RESULTS: Sperm adequate for intracytoplasmic sperm injection were obtained in all men with obstructive azoospermia, including those with significant testicular atrophy and those with anejaculation or necrospermia. Adequate numbers of sperm for intracytoplasmic sperm injection were retrieved less reliably in men with nonobstructive azoospermia. The number of sperm correlated positively with testicular size. Morbidity and discomfort were nonexistent. Sperm were obtained from 43 of 69 men undergoing diagnostic and 170 of 179 men undergoing therapeutic aspiration. Sperm motility ranged from 0 to 20% and viability from 55 to 85%. CONCLUSIONS: Percutaneous testicular sperm aspiration is a cost-effective method to retrieve sperm for intracytoplasmic sperm injection in select men with obstructive azoospermia, anejaculation and necrospermia, and some with nonobstructive azoospermia.

Births of normal daughters after MicroSort sperm separation and intrauterine insemination, in-vitro fertilization, or intracytoplasmic sperm injection.

The world's first deliveries of normal babies after use of flow cytometric separated human sperm cells (MicroSort) for preconception gender selection are reported. Offspring were of the desired female gender in 92.9% of the pregnancies. Most of these pregnancies and births were achieved after simple intrauterine insemination.

Screening semen donors for hereditary diseases. The Fairfax cryobank experience.

OBJECTIVE: To study the carrier frequency of hereditary diseases in potential semen donors with no family history of a genetic disease. STUDY DESIGN: Carrier screening was performed on potential semen donors for chromosomal abnormalities, cystic fibrosis, alpha-1-antitrypsin deficiency, hemoglobinopathies, Tay-Sachs disease, Gaucher disease, Canavan disease, and hereditary breast and ovarian cancer (the BRCA1 185delAG mutation). The screening regimen used for each donor was dictated by his ethnic background. RESULTS: Among 361 individuals screened for chromosomal abnormalities, 1 carried an inversion, and 4 were possible mosaics. Fifteen of 407 potential donors carried cystic fibrosis, 18 of 209 carried alpha-1-antitrypsin deficiency, and 2 of 74 carried a hemoglobinopathy. No carriers of Tay-Sachs disease (56 screened), Gaucher disease (32 screened), Canavan disease (22 screened) or the BRCA1 185delAG mutation (22 screened) were found. CONCLUSION: Screening semen donors for a number of genetic diseases that are passed silently from generation to generation is warranted since family history alone cannot identify them.

Efficiency of MicroSort flow cytometry for producing sperm populations enriched in X- or Y-chromosome haplotypes: a blind trial assessed by double and triple colour fluorescent in-situ hybridization.

Using fluorescent in-situ hybridization (FISH) we have evaluated, on a blind basis, the efficiency of flow cytometry to separate human X- and Y-chromosome bearing spermatozoa. Our data demonstrate that human spermatozoa can be sorted to a purity of 80-90% for X spermatozoa and of 60-70% for Y spermatozoa. Our results using triple FISH fully agree with the sorting treatment used in each case and corroborate the efficiency of the flow sorting technique for sperm sex selection. In these limited samples (200-500 sperm/donor), the frequencies of disomic or diploid spermatozoa were not increased when comparing the sorted samples with unselected samples or with our control series.

Preservation of reproductive function before therapy for cancer: new options involving sperm and ovary cryopreservation.

In summary, sperm cryopreservation for future ICSI and ovarian tissue cryopreservation for future autotransplantation are new opportunities to preserve reproductive options of great importance to patients with newly diagnosed cancer. Since patients must utilize these strategies before cancer therapy is initiated, and these patients will not have a future chance to benefit once therapy has damaged gonadal function, awareness of these technologies among oncologists, radiation therapists, and other colleagues who interface with the victims of cancer is a high priority.

Preimplantation genetic diagnosis in Marfan syndrome.

The in vitro fertilization technology coupled with the ability to amplify DNA from a single cell has been used for the preimplantation genetic diagnosis of Marfan syndrome. An intragenic FBN1 gene marker has been used to track the inheritance of this disorder in a family. Marker genotyping was established following two rounds of amplification. Whenever possible, two blastomeres were separately assayed per embryo. The transfer of five embryos resulted in a singleton pregnancy and the birth of a full-term male infant.

The fluctuation of plasma carotenoid concentrations by phase of the menstrual cycle: a controlled diet study.

This is the first controlled diet study to examine the fluctuation of plasma carotenoids, lipoproteins, and serum hormone concentrations by phase of the menstrual cycle. Nonsmoking, premenopausal women (n = 12) with confirmed ovulatory cycles were given a standard diet with 10 mg total carotenoids/d for two cycles under isoenergetic conditions. Blood was drawn for simultaneous measurement of carotenoids, lipoproteins, and hormones on menses days 1-2, 4-6, 11 through 1 d after the luteinizing hormone surge, and 7-8 d after the surge, representing the menses, early and late follicular, and midluteal phases, respectively. Regression modeling with adjustment for plasma cholesterol concentrations was used to compare mean individual and total plasma carotenoid concentrations by phase of the cycle. Plasma carotenoid concentrations were at their lowest at menses and significantly higher thereafter, except for alpha-carotene. Compared with plasma concentrations at menses, beta-carotene peaked (increased by 9%, P = 0.01) in the late follicular phase. Plasma lutein/zeaxanthin and anhydrolutein concentrations were higher by 8-11% (P < or = 0.006) and by 15-31% (P < or = 0.02), respectively, during the last three phases. Plasma lycopene and phytofluene concentrations peaked (increased by 12%, P = 0.004; and by 21%, P = 0.006, respectively) at the midluteal phase. This cyclic fluctuation may affect the estimation of the plasma carotenoid-disease relation in studies of premenopausal women.

Preimplantation genetic testing for Marfan syndrome.

Marfan syndrome (MFS) is an autosomal dominant disease that affects the skeletal, ocular and cardiovascular systems. Defects in the gene that codes for fibrillin (FBN-1) are responsible for MFS. Here we report the world's first use of preimplantation genetic testing (PGT) to achieve a clinical pregnancy and live birth of a baby free of a Marfan mutation. One or two blastomeres from each embryo were tested for a CA repeat within the FBN-1 gene. The prospective mother is homozygous for the CA repeat (2/2) and has two normal copies of the FBN-1 gene, while the prospective father is heterozygous for the CA repeat (1/2), and is affected with the Marfan syndrome. In the father's family, allele 2 segregates with the mutated FBN-1 gene. For PGT, any embryo diagnosed as heterozygous for the CA repeat (1/2) would be presumed to have inherited normal FBN-1 genes from the father and the mother and be unaffected. One in-vitro fertilization (IVF) cycle yielded 12 embryos for preimplantation testing; six of the embryos were heterozygous for the CA repeat (1/2) and presumed to be free of the Marfan mutation. Five of the six embryos were subsequently transferred into the uterus. The fetus was tested by chorionic villus sampling and found to be free of the Marfan mutation by the same linkage analysis, had a normal fetal echocardiogram, and was normal at birth.

Molecular fragile X screening in normal populations.

In December, 1993, we initiated a pilot project in which DNA fragile X (fraX) testing was offered during routine prenatal or genetic counseling to all pregnant women seen at the Genetics & IVF Institute, most of whom were referred for the indication of advanced maternal age. A brochure on fragile X syndrome was sent to each patient prior to her appointment and was reviewed by a counselor or physician during the counseling session. As of June 1995, 3,345 patients were offered testing; 474 women with no identified family history of mental retardation or learning disability and 214 women with a positive family history accepted the test on a self-pay basis. The second population screened was 271 potential donors in our anonymous egg donor program. DNA from blood was tested by Southern blot using EcoRI/EagI and StB12.3. If an expansion was detected, CGG repeat number was determined by PCR-based analysis. Among the 474 patients with unremarkable family histories, three fraX carriers were identified (repeat sizes = 60+), whereas none were found in the 214 patients with a positive family history. Among the potential egg donors, two high borderline patients were identified (repeat sizes = between 50 and 59). Our ongoing study indicates that screening of pregnant or preconceptual populations for fraX carrier status using DNA testing is accepted by many patients and is an important addition to current medical practice.

Comparisons of pregnancy loss patterns after intracytoplasmic sperm injection and other assisted reproductive technologies.

OBJECTIVE: To compare outcome of pregnancies after intracytoplasmic sperm injection (ICSI) with those of other assisted reproductive technologies. DESIGN: Pregnancy outcomes after ICSI were followed prospectively and compared with pregnancy outcomes after IVF with fresh and frozen ETs and donor oocyte cycles. SETTING: A private tertiary referral center for genetics and infertility in Fairfax, Virginia. PATIENTS: One hundred thirty-six couples achieving pregnancy after undergoing ICSI, 71 after IVF, 35 donor oocyte recipients, and 19 after transfer of frozen-thawed embryos. INTERVENTIONS: In vitro fertilization and/or ET for all couples. Dilatation and curettage to obtain products of conception for chromosome analysis in 28 women experiencing spontaneous abortion. MAIN OUTCOME MEASURES: Pregnancy outcomes were classified as preclinical loss, clinical loss, and ongoing pregnancy. RESULTS: The mean frequency of preclinical pregnancy loss was 26% after ICSI, 28% after IVF, 3% after ET using donor oocytes, and 11% after frozen ET. The rate of clinical loss after ICSI (21%) was compared with IVF (18%), donor oocyte cycles (11%), and frozen ETs (21%). CONCLUSIONS: Intracytoplasmic sperm injection is not associated with an increase in pregnancy losses, clinical or preclinical, compared with conventional IVF.