'Why have women not returned to use their frozen oocytes?': a 5-year follow-up of women after planned oocyte cryopreservation.

RESEARCH QUESTION: What are the reproductive choices and retrospective reflections of women at least 4 years after planned oocyte cryopreservation (POC)? DESIGN: This was an internet survey, using the REDCap application, of women who underwent POC, at a single-centre university-affiliated IVF unit, 4-8 years before the survey. The questionnaire addressed reproductive choices and outcomes following POC. RESULTS: Seventy-nine women who underwent POC during 2011-2014 were invited to participate, and 70 (89%) responded. Mean age at cryopreservation was 37.1 ± 2.4 (range 30-41) years, mean age at study participation 42.6 ± 2.6 (range 35-48) years, and mean time from first cryopreservation cycle to study participation 5.5 ± 1.3 (range 4-8) years. The main retrospectively reported reason for POC was not wanting to become pregnant without a partner (59, 84%). During the follow-up period, 44 women (63%) attempted to conceive either naturally or by assisted reproductive technology using fresh or cryopreserved oocytes. Of those, 28 women achieved a live birth (64% of those who tried to conceive). Fourteen respondents (20% of all respondents) reported using their cryopreserved oocytes, and three (21%) achieved a birth using those oocytes. Fifteen women (34%) of those who tried to conceive used donor spermatozoa. CONCLUSIONS: The most common reasons for not using frozen oocytes were achieving pregnancy without frozen oocytes or preferring not to have a child without a partner. A considerable proportion of women who had POC and were not interested in being a single parent by choice eventually try to conceive using donor spermatozoa several years later.

Anthropometric adjustments are helpful in the interpretation of BMD and BMC Z-scores of pediatric patients with Prader-Willi syndrome.

Anthropometric adjustments of bone measurements are necessary in Prader-Willi syndrome patients to correctly assess the bone status of these patients. This enables physicians to get a more accurate diagnosis of normal versus abnormal bone, allow for early and effective intervention, and achieve better therapeutic results. INTRODUCTION: Bone mineral density (BMD) is decreased in patients with Prader-Willi syndrome (PWS). Because of largely abnormal body height and weight, traditional BMD Z-scores may not provide accurate information in this patient group. The goal of the study was to assess a cohort of individuals with PWS and characterize the development of low bone density based on two adjustment models applied to a dataset of BMD and bone mineral content (BMC) from dual-energy X-ray absorptiometry (DXA) measurements. METHODS: Fifty-four individuals, aged 5-20 years with genetically confirmed PWS, underwent DXA scans of spine and hip. Thirty-one of them also underwent total body scans. Standard Z-scores were calculated for BMD and BMC of spine and total hip based on race, sex, and age for all patients, as well as of whole body and whole-body less head for those patients with total-body scans. Additional Z-scores were generated based on anthropometric adjustments using weight, height, and percentage body fat and a second model using only weight and height in addition to race, sex, and age. RESULTS: As many PWS patients have abnormal anthropometrics, addition of explanatory variables weight, height, and fat resulted in different bone classifications for many patients. Thus, 25-70 % of overweight patients, previously diagnosed as normal, were subsequently diagnosed as below normal, and 40-60 % of patients with below-normal body height changed from below normal to normal depending on bone parameter. CONCLUSIONS: This is the first study to include anthropometric adjustments into the interpretation of BMD and BMC in children and adolescents with PWS. This enables physicians to get a more accurate diagnosis of normal versus abnormal BMD and BMC and allows for early and effective intervention.

Sexual dichotomy of gonadal function in Prader-Willi syndrome from early infancy through the fourth decade.

STUDY QUESTION: At what age does the type of hypogonadism, namely hypothalamic or primary gonadal defect, become established in men and women with Prader-Willi syndrome (PWS)? SUMMARY ANSWER: The type of hypogonadism becomes established only in late adolescence and early adulthood. WHAT IS KNOWN ALREADY: The etiology of hypogonadism in PWS is heterogeneous and the clinical expression is variable. Primary testicular failure is common in PWS men, while combinations of ovarian dysfunction and gonadotrophin deficiency are seen in women. STUDY DESIGN, SIZE, DURATION: This is a prospective study of a cohort of 106 PWS patients followed for a mean duration of 4.5 years. Serial blood samples were obtained and assayed for gonadotrophins, inhibin B, anti-Mullerian hormone (AMH), dehydroepiandrosterone sulfate (DHEAS), testosterone (males), and estradiol (females). Results were compared with normal reference values obtained from the literature. For the purpose of this study, we defined the following age groups: infants <1 year; children 1-10 years; adolescents 11-20 years and adults >20 years. PARTICIPANTS/MATERIALS, SETTING, METHODS: Study participants were 49 males (aged 2 months to 36 years) and 57 females (aged 1 month to 37 years) with genetically confirmed diagnoses of PWS (deletions 60, uniparental disomy 54, imprinting center defect 2) followed in the Israel national multidisciplinary PWS clinic. MAIN RESULTS AND THE ROLE OF CHANCE: Serum LH levels were in the normal range (1.0-6.0 mIU/ml) for 7/10 adult men, and high in 3, while FSH (normal range 1.0-6.1 mIU/ml) was elevated (34.4 ± 11.5 mIU/ml) in 6 and normal (3.5 ± 1.6 mIU/ml) in 4 men. Testosterone was low (5.7 ± 3.4 nmol/l) compared with the normal range of 12.0-34.5 nmol/l in the reference population in all men >20 years. AMH showed a normal decrease with age, despite low testosterone levels. Inhibin B was normal (241 ± 105 pg/ml) in infant boys, but low or undetectable in most adult men. Hormonal profiles were more heterogeneous in women than in men. Estradiol was consistently detectable in only 7/13 adult women. Inhibin B was low or undetectable in all PWS females although occasional samples showed levels within the normal range of 15-95 pg/ml. Vaginal bleeding was reported to occur for the first time in eight women at a median age of 20 years (13-34 years), but only one had regular monthly menses. The type of hypogonadism (primary or secondary) in PWS can be determined only after age 20 years. LIMITATIONS, REASONS FOR CAUTION: The study cohort was heterogeneous, showing variability in BMI, cognitive disability and medical treatment. WIDER IMPLICATIONS OF THE FINDINGS: Demonstration of the natural history of reproductive hormone development in PWS suggests that androgen replacement may be indicated for most PWS boys in mid-adolescence. Recommendations for hormone replacement in PWS women need to be individually tailored, serial measurements of inhibin B should be performed, and contraception should be considered in those women who may have the potential for fertility.

Ovarian reserve and PGD treatment outcome in women with myotonic dystrophy.

Myotonic dystrophy (DM) is the most common form of muscular dystrophy in adults. There are conflicting reports about its effect on female fertility. This study investigated ovarian reserve and IVF-preimplantation genetic diagnosis (PGD) outcome in women with DM1. A total of 21 women undergoing PGD for DM1 were compared with 21 age- and body mass index-matched women undergoing PGD for other diseases. Ovarian reserve markers, response to stimulation, embryo quality and clinical pregnancy and live birth rates were compared. Day-3 FSH concentration was higher, while anti-Müllerian hormone concentration and antral follicle count were lower in the DM1 group (median, range: 6.9 (1.8-11.3) versus 5.7 (1.5-10.7)IU/l; 0.9 (0.17-5.96) versus 2.68 (0.5-9.1)ng/ml; and 13 (0-63) versus 23 (8-40) follicles, respectively, all P < 0.05). Total FSH dose was higher (5200 versus 2250 IU, P = 0.004), while the numbers of oocytes retrieved (10 versus 16, P < 0.04) and metaphase-II oocytes (9 versus 12, P < 0.03) were lower in the DM1 group. The number of cycles with top-quality embryos and the clinical pregnancy rate were lower in the DM1 group. In conclusion, there is evidence of diminished ovarian reserve and less favourable IVF-PGD outcome in women with DM1. Myotonic Dystrophy (DM) is the most common form of muscular dystrophy in adults. There is evidence of subfertility in males affected with the disease but conflicting reports about the effect of the disease on female fertility. The aim of our study was to investigate ovarian reserve and IVF-PGD results in women with DM. Twenty-one women undergoing preimplantation genetic diagnosis (PGD) treatment for DM were compared to 21 age- and BMI matched women undergoing PGD treatment for other diseases. The two groups were compared for antral follicle count (AFC) and serum anti-Mullerian hormone (AMH) levels (the best known markers of ovarian reserve and fertility potential), ovarian response, embryo quality and pregnancy and live birth rates. AFC and the AMH levels were statistically significant lower in the DM group. Total medication dose needed for ovarian stimulation was higher, the number of oocytes and mature oocytes retrieved, and the number of cycles with top quality embryos were lower in the DM group compared to the controls. In conclusion, there is evidence of diminished ovarian reserve, and less favorable IVF-PGD outcome in women with DM. Therefore, we recommend advising these women about the possibility of early decreasing ovarian function in order to prevent any delay in reproductive planning.

Familial haplotyping and embryo analysis for Preimplantation genetic diagnosis (PGD) using DNA microarrays: a proof of principle study.

PURPOSE: Development of PGD assays for molecular disorders is based on analysis of a familial mutation together with linked polymorphic STR markers; a process which is lengthy and requires the identification of multiple informative markers prior to PGD analysis. On the other hand, whole genome amplification (WGA), in conjunction with microarray platforms, allows the use of a universal assay for the analysis of a very large number of SNP markers at once. The aim of this study was to test high throughput pre-PGD familial haplotyping for in-case blastomere analysis in order to eliminate time-consuming pre-case preparations for each family. METHODS: A PGD cycle was performed for a couple with paternal Charcot Marie Tooth 1A (CMT1A) using a classic multiplex nested PCR approach. Mutant embryos from the case were blindly reanalyzed, as single or multi-cell biopsies, using a multiple displacement amplification-based WGA protocol and microarray SNP analysis. In parallel, relevant genomic DNA samples from the family were also analyzed by SNP microarray. RESULTS: After applying a 'unique informative allele' selection algorithm to the data, this array-based assay reconfirmed the initial diagnosis in all samples. CONCLUSIONS: We describe a PGD method that is both accurate and feasible during the time-frame required for embryo transfer. This strategy greatly reduces the time for pre-case haplotype preparation.

Prevalence of low serum cobalamin in infertile couples.

A high prevalence of low levels of cobalamin had been found in a survey of multi-ethnic normal individuals in Israel. The purpose of this study was to investigate the incidence of cobalamin deficiency among Israeli couples suffering from infertility. All couples seen at the in vitro fertilization clinic at an urban hospital (Shaare Zedek Medical Center) in Jerusalem for a 6-month period were invited. Mean cobalamin levels were 259.2 pg ml(-1) in males and 275.1 pg ml(-1) in females (normal >200 pg ml(-1)), 35.5% of 172 men and 23.3% of 223 females had cobalamin deficiency (P = 0.01). There were 171 couples with complete demographic questionnaires and cobalamin values for each partner. In 74 couples (43.3%), one partner was cobalamin deficient, with no significant difference between those with unexplained infertility versus those with explained infertility; and in 13 couples, both partners were cobalamin deficient. Thirty-nine per cent of all men with an abnormal semen analysis had cobalamin deficiency, a finding that requires further investigation. This study questions whether higher rates of male infertility in Israel are partially ascribable to cobalamin deficiency. Recommendation for supplementation in both males and females to achieve high-normal levels of cobalamin would be prudent.

Polar body-based preimplantation genetic diagnosis for N-acetylglutamate synthase deficiency.

OBJECTIVE: We describe a sensitive and highly reliable preimplantation genetic diagnosis (PGD) assay for N-acetylglutamate synthetase (NAGS) deficiency using polar body (PB) analysis in conjunction with multiple markers flanking the gene. This rare autosomal recessive mitochondrial disorder is characterized by hyperammonemia, uncontrollable movements, developmental delay, visual impairment, failure to thrive and vomiting and is caused by mutations in the NAGS gene located on chromosome 17q21.31. METHODS: For a family with an affected child we have developed a multiplex fluorescent PCR protocol that included detection of the specific familial mutation (2729insC) in conjunction with the analysis of five informative polymorphic markers flanking the gene: D17S902, D17S965, D17S1861, D17S791 and D17S1868. Following successful amplification in single-cell fibroblasts, this protocol was used in the couple carriers of NAGS mutation. RESULTS: Of 18 retrieved eggs, 16 were at the M2 stage and 9 fertilized. 12 polar body 1s (PB1) were heterozygotes, 1 homozygote wild-type, 1 total amplification failure, and two showed inconclusive results. Three oocytes that had heterozygote PB1s showed mutant polar body 2 (PB2) indicating a wild-type oocyte. Despite the fact that the specific 2729insC mutation did not amplify in the PGD cycle, analysis of linked markers in PBs was sufficient to ensure an accurate diagnosis in 5 out of 9 oocytes. This cycle resulted in the transfer of 3 embryos originating from oocytes diagnosed as wild-type by PB analysis, with the subsequent birth of healthy twin girls. Postnatal genetic testing revealed that both girls harbored the healthy maternal allele and carried the mutant paternal allele. CONCLUSIONS: Our multiplex-nested PCR protocol based on several linked microsatellite markers offers an efficient and accurate method for PGD for NAGS syndrome even when the mutation is not amplified.

Differentiation of choriocarcinoma cell line (JAr).

Choriocarcinoma, a highly malignant tumor arising from the trophoblast, comprises a heterogenous population of cells including cytotrophoblasts, intermediate trophoblasts, and syncytiotrophoblasts. In order to investigate trophoblast differentiation, we used centrifugal elutriation to separate cells from the JAr choriocarcinoma cell line according to their size and to further show that the resultant cell populations differ in their stage of differentiation. Two % of the cell population consists of large, multinuclear cells, which display the highest level of chorionic gonadotropin (CG) mRNAs. The increase in the CG beta mRNA with cell size is a consequence of the transcriptional mechanism, since agents which induce differentiation in JAr cells, i.e., methotrexate, increase the level of CG alpha and CG beta transcripts, cause a shift in cell size, and result in the formation of multinuclear cells. The multinuclear cells in the JAr population arise, at least partly, from kariokinesis without cytokinesis.

High maternal serum chorionic gonadotropin level in Downs' syndrome pregnancies is caused by elevation of both subunits messenger ribonucleic acid level in trophoblasts.

A unique product of human placenta is CG. Its concentration in maternal blood rises exponentially until 9-10 weeks' gestation, thereafter, it decreases to about 20% of the maximum, remaining constant from 16-17 until 40 weeks. High second-trimester maternal blood level indicates an increased risk for Downs' Syndrome (DS). This study's aim was to determine whether changes occur in the genetic expression of CG subunits in cultured trisomy-21 trophoblasts compared with various gestational age controls. Second-trimester trisomy-21 trophoblasts secrete 10 times more CG than gestational age-matched controls during the first day in culture: 878 (range, 235-2230) IU/g vs. 87 (range, 20-150) IU/g (P < 0.05). This high secretion closely resembles quantities secreted by first-trimester normal trophoblasts: 7500 (range, 3,850-10,000) IU/g. Both subunits' messenger RNA content are substantially increased, CG beta much more than CG alpha, although these genes are not located on chromosome 21. We conclude that at least one cause of high second-trimester maternal blood CG in DS pregnancies is a rise in alpha and beta CG messenger RNA levels in the trophoblast. We propose that at 12-14 weeks, when rapid decrease in maternal blood CG levels can be found, higher than normal values may indicate an increased risk for DS.

Relationship between serum inhibin A and B and ovarian follicle development after a daily fixed dose administration of recombinant follicle-stimulating hormone.

The aim of this study was to investigate the relationship of serum inhibin A and inhibin B to ovarian follicular development in women undergoing pituitary down-regulation and ovarian stimulation with a fixed daily dose of recombinant human FSH in an in vitro fertilization program. Thirty-eight patients were treated randomly with either 100 or 200 IU/day recombinant human FSH (Puregon) for a period of 9-14 days. Serum FSH, inhibin A, inhibin B, 17beta-estradiol, and follicular size and number were determined before FSH treatment and every second day from days 4-6 throughout FSH treatment. Serum FSH increased in a dose-related manner to reach a maximum by days 4-6 and remained unchanged over the duration of treatment. Serum inhibin A and 17beta-estradiol also increased with increasing FSH dose and continued to rise throughout the FSH treatment period. By contrast, serum inhibin B was increased by days 4-6 at both doses of FSH to reach a maximum by days 7-8, remaining unchanged thereafter. Serum inhibin B and, to a lesser extent, inhibin A correlated significantly with the number of oocytes retrieved even when assessed early (days 4-6) in the treatment period (inhibin B vs. number of oocytes: r = 0.89; P < 0.001; inhibin A vs. number of oocytes: r = 0.61; P < 0.05). Serum inhibin A, inhibin B, and 17beta-estradiol were weakly correlated with the number of follicles less than 11 mm when assessed on a daily basis; stronger correlations were observed with the greater than 11-mm follicles during the late stages of treatment. It is concluded that serum inhibin B levels determined during the early stages (e.g. days 4-6) of fixed dose FSH treatment provide an early indicator of the number of recruited follicles that are destined to form mature oocytes. In this context, serum inhibin B may be of predictive value in monitoring ovarian hyperstimulation treatment for in vitro fertilization.

Imidazoline binding sites in human placenta: evidence for heterogeneity and a search for physiological function.

1. An alpha 2-adrenoceptor antagonist, idazoxan, that binds to both alpha 2-adrenoceptors and to imidazoline sites (IR), has been used to characterize human placental IR. Human placenta is shown to be the richest source of IR (1800 +/- 100 fmol mg-1 protein; Kd 38.9 +/- 3.4 nM). 2. Primary cells derived from human placenta and grown in monolayers, also displayed a high density of receptors (3209 +/- 136 fmol mg-1 in cytotrophoblasts and 3642 +/- 144 fmol mg-1 protein in syncytiotrophoblast enriched cell culture). 3. [3H]-idazoxan did not show binding characteristics of alpha 2-adrenoceptors in human placental membranes or human trophoblastic cells, thus making it a ligand of choice to study the imidazoline site. The tissue appeared to be lacking alpha 2-adrenoceptors in that other alpha 2-adrenoceptor ligands, [3H]-rauwolscine and [3H]-clonidine, do not bind to alpha 2-adrenoceptors in human placenta. 4. IRs are localized on the cell surface, as determined by the release of bound [3H]-idazoxan from cells, when washed with high ionic/acidic medium. 5. Imidazoline receptors of human placenta display high affinity for amiloride (72 +/- 27 nM). The high affinity was used as a criterion to classify IR to IRa subtype (placenta, rabbit kidney, rabbit liver and rabbit adipose cells) as opposed to the IRb subtype which display low affinity for amiloride (greater than 2 microM, in all the other tissues).6. Several novel ligands comprising a guanido functional group attached to an aromatic residue (e.g. benziliden-amino-guanidine (BAG), guanido pyrole) display pronounced selectivity for IR over the M2-adrenoceptors as the affinity of BAG is about 40 fold higher (Kd= 18.9 +/- 13.8 nM in human placenta), than the affinity for M2-adrenoceptors (Kd = 768 +/- 299 nM in human platelets). Imidazoline sites bind selectively BAG and other guanido ligands thus indicating a distinct structural requirement at its site of binding.7. K+ channel blockers and monovalent ions (e.g. Cs' and NH4+) interfere with idazoxan binding to IR, indicating a possible involvement of IR in K+ transport.

Interaction between choriocarcinoma cell line (JAr) and human cytotrophoblasts in vitro.

Cytotrophoblasts (from term placentae) and cells from the choriocarcinoma cell line JAr were cultivated either separately or in co-culture for 72 h. RNA was isolated from the cell cultures and Northern blots were developed using equal amounts of RNA. The RNA was hybridized with cDNA probes for CG alpha, CG beta and hPL. Corresponding m-RNAs were detected in the three RNAs except for hPL m-RNA which was absent from JAr cells RNA. The abundance of CG alpha and CG beta m-RNA in the RNA of the co-culture was higher than their accumulative abundances in the RNAs from cytotrophoblasts and JAr cells cultured alone and the abundance of hPL m-RNA in the RNA of the co-cultures was as high as that in the RNA from cytotrophoblasts cultured alone. On the basis of previous findings (Hochberg et al, 1991), it can be assumed that the cytotrophoblasts in the co-cultures are responsible for the increase in hormonal m-RNA production. It could be calculated that the abundances of the CG alpha, CG beta and hPL m-RNAs in the RNA which originated in the cytotrophoblast nuclei were 20, 100 and 10-fold higher respectively in the co-culture compared to those in the culture of cytotrophoblasts. This effect is limited to certain genes only as the concentration of the 92kD collagenase m-RNA and uPA (urokinase type plasminogen activator) m-RNA, which are both produced in cytotrophoblasts to a much higher extent than in JAr cells, and are not increased by cultivating the cytotrophoblasts with JAr cells in co-culture.(ABSTRACT TRUNCATED AT 250 WORDS)

Attenuation of ovarian response by low-dose ketoconazole during superovulation in patients with polycystic ovary syndrome.

OBJECTIVE: To investigate the clinical efficacy of mild inhibition of ovarian steroidogenesis by very low-dose ketoconazole during induction of ovulation in patients with polycystic ovary syndrome (PCOS). DESIGN: Prospective, randomized, cross-controlled study in consecutive cycles. SETTING: Large tertiary care center. PATIENT(S): Eighteen patients with PCOS undergoing hMG superovulation with or without ketoconazole. INTERVENTION(S): A fixed hMG dosage was initiated on cycle days 5-9 in both of the study cycles. Further hMG adjustment was done according to serum E2 levels and follicular measurements. Ketoconazole was administered in one of the cycles by two protocols. MAIN OUTCOME MEASURE(S): Serum E2 and P levels, lead follicles, pregnancy rate, and development of ovarian hyperstimulation syndrome. RESULT(S): Although higher daily hMG doses were needed in cycles with ketoconazole compared with cycles without the drug, the peak E2 levels were substantially lower in the ketoconazole cycles. Although the number of lead follicles did not differ between treatments, the addition of ketoconazole significantly reduced the number of hyperstimulated cycles. Consequently, the cancellation rate dropped dramatically, thus yielding a higher pregnancy rate per patient in the ketoconazole protocols. CONCLUSION(S): Use of a very low dose of ketoconazole during ovulation induction effectively attenuates ovarian steroidogenesis in patients with PCOS. This effect may serve as an adjunct to better control the ovarian response in women who are prone to hyperstimulated cycles.

Different influence of incongruent follicular development on in vitro fertilization-embryo transfer and gamete intrafallopian transfer pregnancy rates.

OBJECTIVE: To compare the influence of incongruent (asymmetric) follicular development on treatment outcome in IVF-ET and GIFT cycles. DESIGN: A retrospective comparative study. SETTING: Tertiary referral center for infertility. PATIENT(S): Five hundred forty-three consecutive assisted reproduction cycles (428 IVF-ET and 115 GIFT) in 422 infertile patients. INTERVENTION(S): Controlled ovarian hyperstimulation (COH) and IVF-ET or GIFT. MAIN OUTCOME MEASURE(S): The incongruity ratio as a parameter of the asymmetry in follicular development and pregnancy rate (PR). RESULT(S): For GIFT cycles, the PRs were 37.8% and 15.7% in cycles with congruent and incongruent follicular development, respectively. However, for IVF-ET cycles, the PR was not affected by incongruent follicular development: 28.2% and 29.0%, respectively. An inverse relationship was observed between the degree of incongruity and the estimated probability of pregnancy in GIFT cycles but not in IVF-ET cycles. Neither the side of the dominant ovary nor the degree of incongruity were consistent in consecutive cycles. CONCLUSION(S): Incongruent follicular development during COH has a significantly negative influence on the outcome of GIFT cycles but not on the outcome of IVF-ET cycles. The reason for this difference is not clear. We recommend considering IVF-ET instead of GIFT if incongruent follicular development occurs.

Effect of intramural, subserosal, and submucosal uterine fibroids on the outcome of assisted reproductive technology treatment.

OBJECTIVE: To investigate the effect of subserosal, intramural, and submucosal fibroids on the outcome of assisted reproductive technology (ART) treatment. DESIGN: A retrospective comparative study. SETTING: A tertiary referral center for infertility. PATIENT(S): Treatment outcome of 106 ART cycles in 88 patients with uterine fibroids (33 subserosal, 46 intramural without cavity distortion, and 9 submucosal) was compared with that of 318 ART cycles in age-matched patients without fibroids. INTERVENTION(S): Controlled ovarian hyperstimulation and ART. MAIN OUTCOME MEASURE(S): Findings on transvaginal uterine ultrasonography performed before the initiation of treatment and pregnancy and implantation rates. RESULT(S): The pregnancy rates per transfer were 34.1%, 16.4%, 10%, and 30.1% in the patients with subserosal fibroids, intramural fibroids, submucosal fibroids and no fibroids, respectively. The implantation rates were 15.1%, 6.4%, 4.3%, and 15.7%, respectively. Both rates were significantly lower in patients with intramural fibroids than in those with subserosal fibroids or no fibroids. CONCLUSION(S): Pregnancy and implantation rates were significantly lower in the groups of patients with intramural and submucosal fibroids, even when there was no deformation of the uterine cavity. Pregnancy and implantation rates were not influenced by the presence of subserosal fibroids. Surgical or medical treatment should be considered in infertile patients who have intramural and/or submucosal fibroids before resorting to ART treatment.

Elevated serum progesterone levels during pituitary suppression may signify adrenal hyperandrogenism.

OBJECTIVE: To investigate whether elevated serum P levels after pituitary down-regulation signify adrenal enzyme defects or hyperandrogenism. DESIGN: Prospective study. SETTING: Assisted reproduction unit in a university medical center. PATIENT(S): Two hundred twenty-seven IVF patients treated by the long down-regulation protocol. INTERVENTION(S): Oral dexamethasone (DEX) administration if P level exceeded 0.8 ng/mL (conversion factor to SI unit, 3.180) after pituitary suppression. MAIN OUTCOME MEASURE(S): Serum concentrations of P, E2, LH, DHEAS, and 17 alpha-hydroxyprogesterone and ACTH stimulation tests. RESULT(S): In eight patients (3.5%), serum P levels exceeded 0.8 ng/mL and E2 and LH levels confirmed pituitary down-regulation. Mean DHEAS levels in the patients in this group were significantly higher than in the other patients. All eight patients demonstrated a significant decrease in serum P level after DEX administration. In five patients the ACTH stimulation test suggested an adrenal defect. Five pregnancies were achieved after the addition of DEX to the treatment protocol. CONCLUSION(S): High serum P levels after pituitary down-regulation appear to be of adrenal origin and may be the first indication of an adrenal enzyme defect. Further investigation such as an ACTH stimulation test is recommended, followed by treatment with DEX if indicated.

Twin gestations and prophylactic hospitalization.

The study evaluates the benefit of elective hospitalization in preventing premature deliveries of twin gestations. Three groups of women with twin gestations, having no other complications of pregnancy which could cause premature delivery, were evaluated. The study group was comprised of 43 women who were electively hospitalized between 30-32 and 36 weeks of gestation. Control group 1 was comprised of 55 women who were not hospitalized but were instructed to rest at home. Control group 2 was comprised of 53 women who were not hospitalized and were not instructed to rest at home. Our results showed that elective hospitalization did not significantly affect the gestational duration or the prematurity rate. However the mean birthweight difference between the study group and the two control groups were 143 +/- 83 g and 205 +/- 84 g, respectively. This result was more significant in multiparous women. The slight increase in birthweight of the hospitalized women compared to the controls, does not seem to justify the cost of hospitalization.

Preimplantation genetic diagnosis for fetal neonatal alloimmune thrombocytopenia due to antihuman platelet antigen maternal antibodies.

OBJECTIVE: To develop a reliable preimplantation genetic diagnosis protocol for antihuman platelet antigen-1 incompatibility for a family in whom antenatal treatment was not possible because of the mother's hypersensitivity to intravenous immunoglobulin (IVIG). METHODS: Haplotypes were constructed from genomic DNA of the family members. A polymerase chain reaction protocol that included eight microsatellite polymorphic markers and the ITGB3-specific (T196C, rs5918) polymorphism were multiplexed to be used in a single cell protocol, and single blastomeres were analyzed. RESULTS: In one preimplantation genetic diagnosis cycle, out of 28 retrieved oocytes, 24 embryos fertilized and 12 underwent biopsy. Three embryos were found to be antihuman platelet antigen-1b/1b homozygotes and two were transferred. This cycle resulted in an uneventful pregnancy and birth of a healthy child. CONCLUSION: In cases in which there is antihuman platelet antigen incompatibility and IVIG cannot be administered, preimplantation genetic diagnosis is a reliable alternative to enable birth of unaffected children.

Successful polar body-based preimplantation genetic diagnosis for achondroplasia.

Achondroplasia, the most common form of dwarfism, is a candidate for preimplantation genetic diagnosis (PGD) because a single mutation accounts for almost all cases. Multiplex fluorescent assay including the common G380R mutation in the FGFR3 gene and eight close polymorphic markers was developed. First and second polar bodies (PB) were used for PGD analysis. An affected woman was treated with routine long-protocol ovarian stimulation and puncture. In the first PGD cycle, out of four fertilized oocytes, PB analysis revealed two mutant oocytes, one with total amplification failure of the maternal allele and one with inconclusive results. In the second PGD cycle, 14 oocytes were retrieved following a higher FSH dose and by performing oocyte retrieval and by placing the patient in the anti-Trendelenburg position using abdominal pressure to allow all follicles to be drained. Following PB analysis, two embryos containing the wild-type FGFR3 allele were transferred. This led to an uncomplicated pregnancy and delivery by Caesarean section at week 38 of a healthy boy, carrying the FGFR3 wild-type maternal allele. In conclusion, oocyte retrieval, while difficult in patients with achondroplasia, can be successfully performed. PB analysis is a reliable and sensitive method for PGD for maternal achondroplasia.

Advantages of multiple markers and polar body analysis in preimplantation genetic diagnosis for Alagille disease.

OBJECTIVE: The development of a preimplantation genetic diagnosis (PGD) protocol for Alagille syndrome (AGS), a rare autosomal dominant disorder with hepatic, cardiac and ophthalmologic involvement. METHODS: We developed a polar body (PB)-based multiplex fluorescent PCR reaction for a female affected with AGS. The protocol included analysis of the Jagged 1 (JAG1) familial mutation and five closely linked highly polymorphic markers (D20S162, D20S901, D20S894, and D20S186). RESULTS: In two cycles of PGD 9 of ten embryos were accurately diagnosed by assessment of first and second PBs, one embryo required additional blastomere biopsy. CONCLUSIONS: This protocol takes advantage of the larger window of opportunity for transfer and the increased accuracy of diagnosis afforded by the combination of PB biopsy and multiple marker analysis. Two cycles resulted in the transfer of two and three mutation-free embryos and a subsequent pregnancy as measured by the rising hCG levels.