Allelic variants of the estrogen receptor genes and frailty phenotype in postmenopausal women.

OBJECTIVE: The prevalence of frailty has been related to menopause. Our main objective was to investigate whether single nucleotide polymorphisms (SNPs) of the estrogen receptor (ER) ERα and ERß genes were related to the frailty phenotype in a population of community-dwelling postmenopausal women. METHODS: A cross-sectional study was performed in which we selected five SNPs, three in the ERα gene and two in the ERß. Linear regression was used to estimate the percentage of phenotypic variance after adjusting for confounding variables. RESULTS: A total of 470 women (mean ± standard deviation age 63.83 ± 8.16 years) were included, of whom 137 women were frail. The SNP rs3798577 of the ERα gene was the only variant associated with frailty, but this significance faded in the multivariant analysis. Body mass index (p = 0.012), number of comorbidities (0 vs. ≥2, p = 0.002) and two reproductive variables, number of miscarriages (none vs. ≥2, p = 0.036) and of childbirths (one vs. ≥3, p = 0.008), were independently related to frailty. CONCLUSION: The five SNPs of the ERα and ERß genes tested were not correlated with frailty. Other SNPs of the ER warrant analysis to clarify whether variance in the gene response affects frailty status.

A translational approach from an animal model identifies CD80 as a candidate gene for the study of bone phenotypes in postmenopausal women.

This study represented a translational study that first compared gene expression of B cells of BM from ovariectomized and control mice, and then analyzed some of the differentially expressed genes in women. Results showed novel genetic associations with bone phenotypes and points to the CD80 gene as relevant in postmenopausal bone loss. INTRODUCTION: Osteoporosis is a multifactorial disease with a strong genetic component. However, to date, research into osteoporosis has only been able to explain a small part of its heritability. Moreover, several components of the immune system are involved in the regulation of bone metabolism. Among them, B cells occupy a prominent place. METHODS: The study consisted of two stages. In the first, gene expression in bone marrow B cells is compared between ovariectomized and SHAM control mice using microarrays. In the second, we studied the association of polymorphisms in some differentially expressed genes (DEG) in a cohort of postmenopausal women. RESULTS: The present study has found 2791 DEG (false discovery rate (FDR) <5%), of which 1569 genes were upregulated (56.2%) and 1122 genes (43.8%) were downregulated. Among the most altered pathways were inflammation, interleukin signaling, B cell activation, TGF-beta signaling, oxidative stress response, and Wnt-signaling. Sixteen DEG were validated by MALDI-TOF mass spectrometry or qPCR. The translational stage of the study genotyped nine single nucleotide polymorphisms (SNPs) of DEG or related and detected association with bone mineral density (BMD) (nominal P values), while adjusting for confounders, for SNPs in the CD80, CD86, and HDAC5 genes. In the logistic regression analysis adjusted for confounders, in addition to the SNPs in the aforementioned genes, the SNPs in the MMP9 and SOX4 genes were associated with an increased risk of osteoporosis. Finally, two SNPs (in the CD80 and SOX6 genes) were associated with an increased risk of bone fragility fracture (FF). However, after Bonferroni correction for multiple testing, only the association between CD80 with BMD and risk of osteoporosis remained significant. CONCLUSION: These results show that the use of animal models is an appropriate method for identifying genes associated with human bone phenotypes.

Endometrial response to concurrent treatment with vaginal progesterone and transdermal estradiol.

ABSTRACT Objective To describe the effect of the intermittent administration of vaginal progesterone and a low-dose estradiol patch on endometrial stability, as assessed by the rate of amenorrhea and endometrial stimulation. Methods This was an open study in which 64 moderately symptomatic, postmenopausal women were treated in the outpatient clinic of our University Hospital for different intervals up to 1 year. The treatment consisted of a combination of patches delivering 25 µg/day estradiol and intravaginal pills containing 100 mg of micronized progesterone. Patches and pills were administered concomitantly in a twice-a-week protocol. The endometrial response was assessed by endovaginal ultrasound completed with suction biopsy when required. Results Both cumulative amenorrhea and no-bleeding rates increased progressively and reached 88.9% and 100.0%, respectively, by the 12th month. Isolated or repetitive episodes of bleeding, bleeding and spotting, or only spotting were reported by three, four, and 12 women, respectively. Endometrial thickness remained unaltered. Endometrium was atrophic in the seven women in whom a biopsy was performed. Conclusion The substantially reduced progestogen load determined by this combination achieved an acceptable incidence of spotting or bleeding when associated with a low estrogenic dose. There was no apparent endometrial stimulation. Additional studies are required to confirm this observation.

Gene-gene interaction between CD40 and CD40L reduces bone mineral density and increases osteoporosis risk in women.

SUMMARY: We have analysed the association of single-nucleotide polymorphisms (SNPs) in CD40 and CD40L genes with bone mineral density (BMD) in our women. Results showed that women with TT genotype for rs1883832 (CD40) and for rs1126535 (CD40L) SNPs displayed reduced BMD and increased risk for osteopenia/osteoporosis. Our data notwithstanding, the results need to be replicated. INTRODUCTION: Recent data have revealed that the CD40/CD40L system can be implicated in bone metabolism regulation. Moreover, we previously demonstrated that rs1883832 in the CD40 gene was significantly associated with BMD and osteoporosis risk. The objective of the present work was to determine whether polymorphisms in CD40 and CD40L genes are associated with BMD and osteoporosis risk. METHODS: We conducted an association study of BMD values with SNPs in CD40 and CD40L genes in a population of 811 women of which 693 and 711 had femoral neck (FN) and lumbar spine (LS) densitometric studies, respectively. RESULTS: Women with the TT genotype for rs1883832 (CD40) showed a reduction in FN-BMD (P = 0.005) and LS-BMD (P = 0.020) when compared with women with the CC/CT genotype. Moreover, we found that rs1126535 (CD40L) was significantly associated with LS-BMD so that women with the TT genotype displayed lower BMD (P = 0.014) than did women with the CC/CT genotype. Interestingly, we have found a strong interaction between polymorphisms in these genes. Thus, women with the TT genotype for both rs1883832 and rs1126535 SNPs (TT + TT women) showed a lower age-adjusted BMD (Z-score) for FN (P = 0.0007) and LS (0.007) after adjusting by years since menopause, body mass index, smoking and menopausal status, densitometer type, hormone replacement therapy (HRT) use and HRT duration and after making the Bonferroni adjustment for multiple comparisons than did the remaining women. Logistic regression analysis adjusted by these covariates showed that TT + TT women had increased risk for FN (odds ratio (OR) = 2.76; P = 0.006) and LS (OR = 2.39; P = 0.020) osteopenia or osteoporosis than did the other women. CONCLUSIONS: Our results suggest that interaction between genetic variants in the CD40 and CD40L genes exerts a role on BMD regulation. Further studies, which we welcome, are needed to replicate these data in other populations.

Progestogens reduce thromboxane production by cultured human endothelial cells.

OBJECTIVES: Progestogens have been poorly studied concerning their roles in endothelial physiology. Prostanoids are vasoactive compounds, such as thromboxane A2, a potent vasoconstrictor, and prostacyclin, a vasodilator. We examined the effects of two progestogens used clinically, progesterone and medroxyprogesterone acetate, on thromboxane A2 production by cultured human umbilical vein endothelial cells (HUVEC) and investigated the role of progesterone receptors and the enzymes involved in production of thromboxane A2 and prostacyclin. METHODS: Cells were exposed to 1-100 nmol/l of either progesterone or medroxyprogesterone acetate, and thromboxane A2 production was measured in culture medium by enzyme immunoassay. Gene expression of prostacyclin synthase and thromboxane synthase was analyzed by quantitative real-time polymerase chain reaction. Expression of prostacyclin synthase protein was analyzed by Western blot. RESULTS: Both progestogens decreased thromboxane A2 release after 24 h. Protein and gene expression of prostacyclin synthase were increased after exposure to both progestogens, without changes in thromboxane synthase expression. These effects induced by progestogens were mediated through progesterone receptors, since they were decreased in the presence of the progesterone receptor antagonist RU486. The cyclo-oxygenase-1 selective inhibitor reduced thromboxane release. CONCLUSION: Progesterone and medroxyprogesterone acetate decreased HUVEC thromboxane release in a progesterone receptor-dependent manner, without changes in thromboxane synthase expression and enhanced prostacyclin synthase gene and protein expression.

Sudden cardiac death in a 20-year-old male swimmer.

In young adults, hypertrophic obstructive cardiomyopathy (HOCM) is an acknowledged risk factor for sudden cardiac death (SCD) in an otherwise healthy and active patient. While the incidence of SCD in young people is not high enough for extensive, wide-scale examinations, the potential for prevention of some deaths via pre-exercise imaging may be beneficial in certain patient populations, such as those with a family history of SCD or professional athletes. We present the case of a healthy 20-year-old man with no past medical history who died while swimming in a river, likely secondary to cardiac arrest in the setting of HOCM.

Accidental fire fatality with sustained combustion.

This is a case of a 59-year-old man found with extensive second to fourth degree thermal burns found lying on the ground several feet from a vehicle used as his domicile. Autopsy revealed extensive loss of soft tissue and fragmentation of bone, mostly to the trunk region, with partial sparing of the upper trunk, head, and extremities.The decedent had a history of acute and chronic substance abuse and it was reported that he was participating in a methadone program. Toxicology reports of autopsy blood obtained from the heart indicated methadone levels of 0.4 mg/L at the time of his death.There was also a trace amount of cocaine present and there was no ethanol detected. Sustained human combustion, or the "wick effect," is concisely defined as the partial destruction of a body by fire, where the victim's clothing absorbs liquefied fatty tissue and acts like a wick of a candle by perpetuating a flame that slowly destroys the body with heat. There are few nonexperimental cases describing this process in the world literature.

The new frontier of bone formation: a breakthrough in postmenopausal osteoporosis?

OBJECTIVE: Osteoporosis is a chronic disease that accelerates after menopause in many women. Most of the pharmacologic attempts to control the disease, such as hormone therapy, have emphasized the constraint of bone resorption. Since recent years have witnessed important advances in the field of bone formation, this review aims to update the present knowledge on the mechanisms affecting osteoblastogenesis and on the therapeutic results achieved by recently approved drugs. METHOD: We sought peer-reviewed, full-length basic and clinical articles published between 1995 and May 2008 using a PubMed search strategy, with the terms osteoporosis and osteoblast, osteoporosis and strontium ranelate, and osteoporosis and parathyroid hormone (PTH). This search was further supplemented by a hand-search of reference lists of selected review papers. After crossing-cleaning the reference lists, some 800 articles were selected. Articles on regulators of osteoblast differentiation and function, together with well-designed clinical studies, were surveyed. RESULTS: A complex network of systemic and local factors regulates osteoblastogenesis. Advances in fracture protection have been published in clinical studies with PTH. Some investigators claim an anabolic effect for strontium ranelate, which also confers protection against fracture. CONCLUSION: The control of bone formation offers new clinical potential. Stimulation of bone formation by PTH has translated into fracture protection. The action of strontium ranelate has been claimed to be mediated by some level of bone formation, but this hypothesis still needs clarification.

[Role of probiotics in Obstetrics and Gynecology]. / Papel de los probióticos en Obstetricia y Ginecología.

The human microbiota is estimated to be 2.5-3.0 kg. Bacterial colonization starts during delivery, due to fetal contact with vaginal and intestinal maternal microorganisms. The oligosaccharides in human breast milk stimulate the growth of bacteria, which provide the optimal environment for intestinal mucosal immunity development. Additionally, breast milk has its own microbiota and it is altered in mastitis. The vagina is another important microenvironment. Vaginal dysbiosis leads to bacterial vaginosis and vaginal candidiasis, both of them very frequent in reproductive life. The probiotics are a potential and encouraging treatment for all microbiota alterations. Nevertheless, additional studies are required to confirm the benefits of probiotics.

La microbiota de cada individuo se estima en 2,5-3 kg de peso. La colonización bacteriana comienza en el momento del parto, por contacto del feto con la microbiota vaginal e intestinal de la madre. Los oligosacáridos de la leche materna estimulan el crecimiento de bacterias, que proporcionan el ambiente adecuado para el desarrollo de la inmunidad de la mucosa intestinal. Además, la leche materna es portadora de su propia microbiota, la cual se altera en las mastitis. La vagina constituye otro importante microambiente. Las disbiosis en esta zona conducen a la aparición de vaginosis bacteriana y candidiasis, siendo ambas una patología muy frecuente en la mujer en edad fértil. Los probióticos se presentan como tratamiento potencial y alentador de toda la patología asociada a alteraciones de la microbiota. Son necesarios más estudios que confirmen el beneficio de los probióticos en este campo.

Embryo biopsy strategies for preimplantation diagnosis.

OBJECTIVE: To analyze different biopsy methods, embryo stages, and cellular masses that can be removed for preimplantation diagnosis of genetic diseases to find optimal biopsy conditions compatible with the subsequent development of the conceptus, the acquisition of intact viable blastomeres, and the reliability of the genetic analysis. DATA IDENTIFICATION: The most important published studies have been identified through a computerized bibliographical search (MEDLINE; Dialog, Palo Alto, CA). STUDY SELECTION: Studies reporting different embryo biopsy methods practiced at different stages have been selected. RESULTS: The analysis carried out in the current review shows, at the present time, the following: [1] the displacement and push methods may be more suitable than the stitch and pull and aspiration (puncturing the zona pellucida) approaches at cleavages stages; [2] the aspiration and stitch and pull procedures may assure higher success rates than the herniation procedure at the blastocyst stage; [3] the mechanical division method and the use of acid Tyrode's solution would not be advisable before the eight-cell stage; [4] human embryos at the two-cell and blastocyst stages may not be suitable for preimplantation diagnosis because of an excessive reduction of cellular mass at the two-cell stage and a low or zero pregnancy rate after transfer at the blastocyst stage; and [5] biopsy of a quarter of the embryonic cellular mass on day 2 after insemination may increase biochemical pregnancies if the cleavage rate is not preserved. CONCLUSIONS: At the present time, biopsy of a quarter of the embryo on day 3 after insemination may be the most feasible approach for preimplantation diagnosis.

Consequences of high ovarian response to gonadotropins: a cytogenetic analysis of unfertilized human oocytes.

Several reports have shown decreased fertilization and implantation rates in high-responder patients to gonadotropins. In this cytogenetic study of unfertilized human oocytes, we have investigated possible reasons for this clinical evidence. Three groups of patients were established according to the number of oocytes retrieved: group 1 (n = 21), 1 to 5 oocytes; group 2 (n = 32), 6 to 10 oocytes; and group 3 (n = 35), greater than or equal to 11 oocytes. Patients in group 3 had lower estradiol levels per follicle developed, lower follicular volume, higher incidence of diploid oocytes, and higher rate of oocytes with an additional set of prematurely condensed sperm chromosomes of the presynthetic gap than groups 1 and 2. These results suggest that women with greater than or equal to 11 oocytes retrieved have a relative higher cytoplasmic immaturity that could explain the lower fertilization and implantation rates found in other studies.

Oocyte maturation in humans: the role of gonadotropins and growth factors.

OBJECTIVE: To determine the effect of FSH/LH in vivo and epidermal growth factor (EGF) and insulin-like growth factor I (IGF-I) in vitro on human oocyte maturation. DESIGN: Oocyte-cumulus complexes were harvested from three different groups of patients: [1] unstimulated ovaries from women undergoing surgery; [2] multifollicular development achieved with a combination of FSH and LH in the absence of an ovulatory dose of hCG; and [3] oocyte-cumulus complexes retrieved after appropriate ovarian stimulation with FSH/LH and hCG for IVF purposes. SETTING, PATIENTS: In vitro fertilization program and patients undergoing surgery for benign disorders at the Instituto Valenciano de Infertilidad, Valencia, Spain. INTERVENTIONS: Oocyte-cumulus complexes from unstimulated ovaries collected at surgery by follicular puncture and washing. Oocyte-cumulus complexes from stimulated cycles obtained by ultrasound-guided transvaginal aspiration. Oocyte-cumulus complexes cultured in vitro in the absence or presence of different concentrations of EGF and IGF-I. MAIN OUTCOME MEASURE: Germinal vesicle breakdown and metaphase-II stage after 24 and 48 hours. RESULTS: Comparison of the spontaneous resumption of meiosis and metaphase II oocytes among groups showed significant differences between unstimulated and stimulated ovaries after 24 and 48 hours in culture. Administration of hCG accelerated the percentage of maturation by 24 hours. Further incubation of unstimulated oocyte-cumulus complexes with EGF and IGF-I significantly increased the percentage of metaphase-II oocytes after 24 and 48 hours in culture. CONCLUSIONS: Epidermal growth factor and IGF-I are able to augment spontaneous maturaion in immature human oocytes. Because spontaneous maturation is mainly observed when follicles have been exposed to pharmacological doses of hMG, it is suggested that increasing FSH levels within the follicle is coincident with the generation of a positive signal necessary to complete oocyte maturation in humans. This signal may be linked to the dynamics of growth factors within the follicle itself.

Chromosome anomalies in human oocytes in vitro.

OBJECTIVE: To determine possible effects on chromosomal status of human oocytes in vitro. DESIGN: A prospective cytogenetic study of human oocytes fixed at 4, 20 (noninseminated oocytes), or approximately 52 hours after pick-up (inseminated-unfertilized oocytes). SETTING: Primary treatment of infertility in an institutional practice. PATIENTS, PARTICIPANTS: Eighty-eight consecutive in vitro fertilization (IVF) patients (262 inseminated-unfertilized oocytes) and 47 IVF patients with greater than 8 oocytes retrieved (95 noninseminated oocytes). INTERVENTIONS: Ultrasound-guided transvaginal follicular aspiration. MAIN OUTCOME MEASURE(S): Planned before data collection began. RESULTS: The incidence of chromosome anomalies did not differ between groups. However, a negative correlation between oocytes apparently without chromosomes and oocytes with fragmented chromosomes was found. CONCLUSIONS: Incubation of oocyte in vitro does not affect the chromosome complement. The mechanisms leading to oocytes apparently without chromosomes and with fragmented chromosomes are self-excluding.

The heparin-glutathione test: an alternative to the hypo-osmotic swelling test to select viable sperm for intracytoplasmic sperm injection.

OBJECTIVE: To evaluate the heparin-glutathione test (HEGLUT) for the selection of viable sperm for intracytoplasmic sperm injection (ICSI). DESIGN: A prospective study. SETTING: Department of Pediatrics, Obstetrics and Gynecology, University of Valencia and Instituto Valenciano de Infertilidad. PATIENT(S): Semen samples from healthy donors and patients with infertility. INTERVENTION(S): Sperm samples were kept in culture for different periods in Ham's F-10 medium supplemented or not supplemented with heparin, reduced glutathione (GSH), or a heparin-GSH mixture. Control and heparin-GSH-treated spermatozoa were injected into hamster oocytes. The HEGLUT and ICSI were performed. MAIN OUTCOME MEASURE(S): Sperm nuclear decondensation, progressive and nonprogressive motility, and male pronucleus formation. RESULT(S): The maximum proportion of sperm nuclear decondensation (28.7%+/-2.1% versus 2.6%+/-0.5% in the control group) was reached after 60 minutes of incubation in the presence of a heparin-GSH mixture. Differences in the percentages of progressive and nonprogressive motility among treatments and times of incubation, although statistically significant, were biologically negligible. No statistically significant differences were observed in the rate of sperm head decondensation (8.2% [4/49] versus 11.1% [6/54]) and male pronucleus formation (18.4% [9/49] versus 22.2% [12/541) after the injection of control and treated spermatozoa into hamster oocytes. CONCLUSION(S): The HEGLUT may offer an alternative to the hypo-osmotic swelling test for the selection of viable sperm for ICSI.

Two-year prospective, randomized trial comparing an innovative twice-a-week progestin regimen with a continuous combined regimen as postmenopausal hormone therapy.

OBJECTIVE: To compare compliance, symptom control, bleeding patterns, endometrial response, and lipid changes in postmenopausal women treated with transdermal E2 and a regimen of either intermittent or continuous dosing of progestin. DESIGN: Randomized, prospective study. SETTING: Menopausal Outpatient Clinic at an academic tertiary care hospital. PATIENT(S): One hundred women who had reached menopause naturally and had been amenorrheic for at least 1 year. Fifty women were randomly assigned to receive each regimen. INTERVENTION(S): All patients received 50-microg E2 patches and medroxyprogesterone acetate, either 5 mg twice per week or 2.5 mg daily. The bleeding pattern was registered in diary cards. Endometrial status was assessed by vaginal ultrasound and endometrial biopsy. Lipid levels were measured by ELISA. MAIN OUTCOME MEASURE(S): The number of patients who dropped out of the study and the number of days each patient reported spotting or bleeding were recorded. Endometrial thickness was measured and histologic examination of the endometrial tissue was performed. Plasma levels of total cholesterol, triglycerides, high-density lipoprotein cholesterol, and low-density lipoprotein cholesterol were measured. RESULT(S): Fourteen women dropped out of the intermittent dosing group and 13 dropped out of the continuous dosing group. Irregular spotting or bleeding occurred at a similar rate in both groups. Biopsies performed at the end of the study showed adequate endometrial stabilization in both groups. No significant changes in lipid levels were detected with either regimen. CONCLUSION(S): Both regimens were similarly effective. The high rate of atrophic endometria suggests the possibility of further reduction of the progestin dose.

Effect of menopause and different combined estradiol-progestin regimens on basal and growth hormone-releasing hormone-stimulated serum growth hormone, insulin-like growth factor-1, insulin-like growth factor binding protein (IGFBP)-1, and IGFBP-3 levels.

OBJECTIVE: To determine the effects of menopause and three different formulations of E2 plus medroxyprogesterone acetate on serum concentrations of basal and growth hormone-releasing hormone (GHRH)-stimulated growth hormone (GH), insulin-like growth factor-1 (IGF-1), insulin-like growth factor binding protein (IGFBP)-1, IGFBP-3, insulin, and C peptide. DESIGN: Prospective, controlled trial. SETTING: Menopausal outpatient clinic at an academic tertiary care hospital. PATIENT(S): Nineteen postmenopausal women with different menopausal ages. Seventeen premenopausal women were included as controls. INTERVENTION(S): Oral estrogen (E2 valerate, 2 mg/d) or transdermal estrogen (50-microg or 100-microg E2 patch) was administered for 8 weeks. Medroxyprogesterone acetate (5 mg/d) was administered during weeks 3, 4, 7, and 8 of each protocol. Blood samples were collected before treatment and after the completion of each protocol from postmenopausal women, and on cycle days 6-8 from premenopausal women. MAIN OUTCOME MEASURE(S): Levels of GH, IGF-1, IGFBP-1, IGFBP-3, insulin, and C peptide. RESULT(S): Basal GH levels were negatively correlated with age in premenopausal women but not in postmenopausal women. The area under the GHRH-induced GH curve decreased in older postmenopausal women after the oral estrogen protocol. Levels of IGF-1 diminished after the oral E2 protocol in postmenopausal women. CONCLUSION(S): The administration of oral, but not transdermal, E2 plus medroxyprogesterone acetate at the usual clinical doses used in postmenopausal women decreased IGF-1 levels and the response of GH to GHRH in older women. No substantial changes were detected in IGFBP-1, IGFBP-3, insulin, or C peptide levels.

Human embryo biopsy on the 2nd day after insemination for preimplantation diagnosis: removal of a quarter of embryo retards cleavage.

OBJECTIVE: To assess any reduction in viability and development in vitro after biopsy of a quarter of the cells of human embryos on day 2 after insemination. DESIGN: A prospective study in which normally fertilized surplus embryos of good morphology with two to eight cells approximately 48 hours after insemination were randomly allocated to a control or biopsied group, respectively. SETTING: In vitro fertilization (IVF) unit and laboratories of the Hammersmith Hospital, Institute of Obstetrics and Gynaecology, London University. PATIENTS, PARTICIPANTS: One hundred twenty-nine embryos from 28 infertile IVF patients. INTERVENTIONS: Follicular aspiration by ultrasound-guided transvaginal puncture and embryo biopsy by micromanipulative procedures. MAIN OUTCOME MEASURE(S): Pyruvate uptake and cell number at the blastocyst stage. RESULTS: Embryo biopsy did not have an adverse effect on either the proportion developing to the blastocyst stage (50% [32 of 64] and 47.7% [31 of 65] for the control and biopsied groups, respectively) or embryo viability, measured indirectly through pyruvate uptake. However, the proportion of embryos that reached the morula stage after day 4 (retarded embryos) was significantly higher (44%, 11 of 25 versus 8.7%, 2 of 23) in the biopsied group. The total number of cells (29.6 +/- 3.1 versus 62.4 +/- 4.7), numbers of inner cell mass (7.7 +/- 2.2 versus 24.5 +/- 1.4) and trophectoderm (24.0 +/- 5.2 versus 45.0 +/- 6.4) cells, and the inner cell mass:trophectoderm ratio (34.7 +/- 7.9 versus 59.5 +/- 11.7) were strikingly reduced at the blastocyst stage in the biopsied group. This reduction was greater in embryos that reached the morula stage after day 4. CONCLUSIONS: More investigation is needed to assess whether the detrimental effects observed were because of the biopsy method used in this study or to a high sensitivity of human embryos at early stages to manipulation in vitro.

Failed in vitro fertilization of human oocytes: a cytogenetic analysis.

OBJECTIVE: To investigate possible causes of in vitro fertilization (IVF) failure. DESIGN: A retrospective cytogenetic study of human oocytes divided into four groups or, alternatively, into two groups according to fertilization rates and whether the patients became pregnant or not. Two additional groups included oocytes in which there was no or only partial fertilization. SETTING: Primary treatment of infertility in an institutional practice. PATIENTS, PARTICIPANTS: Two hundred fifty-three inseminated-unfertilized oocytes from 87 women entering the IVF program because of tubal, unknown, and male infertility. Immunological infertility was excluded. INTERVENTIONS: Ultrasound-guided transvaginal follicular aspiration. MAIN OUTCOME MEASURE(S): Planned after data collection. RESULTS: The rate of chromosome anomalies did not show any significant difference among the four groups established according to the fertilization rate and between pregnant and nonpregnant patients. Independently, our data identified male factor as responsible for 41%, chromosome anomalies 19.3%, oocyte immaturity 11.8%, and unknown etiology 41% of fertilization failures (based on analysis of 161 oocytes). CONCLUSIONS: Fertilization rate and pregnancy outcome after IVF are not related to the incidence of oocyte chromosome anomalies.

Effects of aging on the human ovary: the secretion of immunoreactive alpha-inhibin and progesterone.

OBJECTIVE: To investigate the changes induced by age in the function and secretory pattern of the human ovary. Immunoreactive alpha-inhibin, E2, and P secretion in vivo and in vitro have been compared in two different populations. DESIGN: Prospective study. Women undergoing IVF-ET were divided into two groups according to age: group 1 (32.0 +/- 0.7 years; mean +/- SEM) and group 2 (40.3 +/- 0.3 years). SETTING: In vitro fertilization program at the Instituto Valenciano de Infertilidad. PATIENTS: A total of 33 infertile women with regular menses, undergoing IVF-ET. INTERVENTIONS: Follicle aspiration performed by transvaginal ultrasound. Four follicles per patient were aspirated in individual plastic tubes. Granulosa-luteal cells isolated with Percoll columns and cultured in vitro up to 4 days in the presence of hCG. MAIN OUTCOME MEASURES: In vitro fertilization parameters, serum levels of E2, immunoreactive alpha-inhibin, and P, as well as the secretion of immunoreactive alpha-inhibin and P by the cultured granulosa-luteal cells. RESULTS: Serum immunoreactive alpha-inhibin levels the day of ovum pick-up were significantly lower in group 2 compared with group 1. Incubation of cells for 96 hours showed a significantly higher ability to accumulate immunoreactive alpha-inhibin in group 1 than 2. Human chorionic gonadotropin stimulated immunoreactive alpha-inhibin production after 96 hours. Cells from younger women displayed a significantly higher ability to secrete P than cells from older women. Human chorionic gonadotropin was able to significantly stimulate P production in group 1. CONCLUSIONS: These results confirm previous observations showing a reduced production of immunoreactive alpha-inhibin and steroids of ovaries from older women and suggest that a reduced cellular function, rather than a decrease in the follicular population, is the main mechanism by which these changes are produced.

The effect of oral hormone replacement therapy on lipoprotein profile, resistance of LDL to oxidation and LDL particle size.

OBJECTIVES: To disclose if oral estradiol (E(2)), alone or in combination with natural progesterone (P) or medroxyprogesterone acetate (MPA), may modify the oxidizability of low density lipoprotein (LDL), and if the effect is achieved at physiological dosages. LDL oxidizability was assessed by the resistance to oxidation by copper and by the particle size profile, since small particles have increased oxidation susceptibility. METHODS: Thirty-three women received two consecutive, two-month length doses of 1 and 2 mg/day of oral E(2). They were then randomly assigned to a fourteen-day treatment of 2 mg/day E(2) plus either 300 mg/day P or 5 mg/day MPA. A parallel group of experiments was performed on a pool of baseline plasma, where hormones were added at the desired concentration. Lipoprotein levels, resistance of LDL to oxidation, and LDL particle diameter, were measured at baseline and after each treatment. RESULTS: Estradiol reduced LDL levels and increased high density lipoprotein (HDL) and triglycerides. P abolished these changes, whereas MPA only reversed the increase of HDL. Estradiol protected LDL from oxidation in a dose-dependent manner, although only at pharmacological concentrations (1 microM or higher). Both P and MPA were inert at either physiological or pharmacological concentrations. The size of the LDL particles remained unaffected except under MPA, in which it was reduced. CONCLUSIONS: Estradiol has a protective effect against LDL oxidation, although only at pharmacological dosages. P and MPA did not limit the E(2) action. The size of the LDL particles remained unaltered after each E(2) dose, but MPA, and not P, was associated with a diminution.