Identification of chromosomal errors in human preimplantation embryos with oligonucleotide DNA microarray.

A previous study comparing the performance of different platforms for DNA microarray found that the oligonucleotide (oligo) microarray platform containing 385K isothermal probes had the best performance when evaluating dosage sensitivity, precision, specificity, sensitivity and copy number variations border definition. Although oligo microarray platform has been used in some research fields and clinics, it has not been used for aneuploidy screening in human embryos. The present study was designed to use this new microarray platform for preimplantation genetic screening in the human. A total of 383 blastocysts from 72 infertility patients with either advanced maternal age or with previous miscarriage were analyzed after biopsy and microarray. Euploid blastocysts were transferred to patients and clinical pregnancy and implantation rates were measured. Chromosomes in some aneuploid blastocysts were further analyzed by fluorescence in-situ hybridization (FISH) to evaluate accuracy of the results. We found that most (58.1%) of the blastocysts had chromosomal abnormalities that included single or multiple gains and/or losses of chromosome(s), partial chromosome deletions and/or duplications in both euploid and aneuploid embryos. Transfer of normal euploid blastocysts in 34 cycles resulted in 58.8% clinical pregnancy and 54.4% implantation rates. Examination of abnormal blastocysts by FISH showed that all embryos had matching results comparing microarray and FISH analysis. The present study indicates that oligo microarray conducted with a higher resolution and a greater number of probes is able to detect not only aneuploidy, but also minor chromosomal abnormalities, such as partial chromosome deletion and/or duplication in human embryos. Preimplantation genetic screening of the aneuploidy by DNA microarray is an advanced technology used to select embryos for transfer and improved embryo implantation can be obtained after transfer of the screened normal embryos.

DNA microarray reveals that high proportions of human blastocysts from women of advanced maternal age are aneuploid and mosaic.

Trophectoderm (TE) biopsy and DNA microarray have become the new technologies for preimplantation genetic diagnosis in humans. In this study, we comprehensively examined aneuploid formation in human blastocysts produced in vitro with microarray and investigated the clinical outcome after transfer of euploid embryos. Biopsied cells from either TE or inner cell mass (ICM) were processed for microarray to examine the errors in 23 pairs of chromosomes and the consistency between TE and ICM. It was found that 56.6% of blastocysts were aneuploid. Further analysis indicated that 62.3% of aneuploid blastocysts had single and 37.7% had multiple chromosomal abnormalities. Chromosome errors could occur in any chromosome, but errors in chromosome 21 accounted for the most (11.3%) among the 23 pairs of chromosomes. Transfer of array-screened blastocysts produced high pregnancy (70.2%) and implantation (63.5%) rates. Microarray of TE and ICM cells in the same blastocysts revealed that high proportions of aneuploid blastocysts (69.2%) were mosaic, including aneuploid TE and euploid ICM, inconsistent anomalies between ICM and TE, or euploid TE cells and aneuploid ICM in the same blastocyst. These results indicate that high proportions of human blastocysts produced in vitro from women of advanced maternal age are aneuploid and mosaic. Errors can occur in any of the 23 pairs of chromosomes in human blastocysts. Biopsy from TE in blastocysts does not exactly predict the chromosomal information in ICM if the embryos are aneuploid. Some mosaic blastocysts have euploid ICM, which may indicate important differentiate mechanism(s) of human preimplantation embryos.

Testosterone interacts with the feedback mechanisms engaged by Tyr985 of the leptin receptor and diet-induced obesity.

Inhibitory signaling through Tyr985 of the leptin receptor contributes to the attenuation of anorectic leptin action in obese animals. Leptin receptor (LEPR-B) Tyr985Leu homozygote mutant mice (termed l/l) were previously generated to study Tyr985's contributions to inhibition of LEPR-B signaling; young female l/l mice display a lean, leptin-sensitive phenotype, while young male l/l are not significantly different from wild-type. We report here that testosterone (but not estrogen) determines the sex-specificity of the l/l phenotype. This provides additional insight into the cellular mechanism by which gonadal hormones determine central sensitivity to leptin, and may help elucidate the long-noted sex differences in leptin sensitivity. Additionally, we observed that Tyr985 signaling protects against a diet-dependent switch that exacerbates obesity with high fat feeding, such that the enhanced leptin sensitivity of l/l mice on a normal diet leads to increased adiposity in the face of chronic high-fat diet.

IRS2 signaling in LepR-b neurons suppresses FoxO1 to control energy balance independently of leptin action.

Irs2-mediated insulin/IGF1 signaling in the CNS modulates energy balance and glucose homeostasis; however, the site for Irs2 function is unknown. The hormone leptin mediates energy balance by acting on leptin receptor (LepR-b)-expressing neurons. To determine whether LepR-b neurons mediate the metabolic actions of Irs2 in the brain, we utilized Lepr(cre) together with Irs2(L/L) to ablate Irs2 expression in LepR-b neurons (Lepr(ΔIrs2)). Lepr(ΔIrs2) mice developed obesity, glucose intolerance, and insulin resistance. Leptin action was not altered in young Lepr(ΔIrs2) mice, although insulin-stimulated FoxO1 nuclear exclusion was reduced in Lepr(ΔIrs2) mice. Indeed, deletion of Foxo1 from LepR-b neurons in Lepr(ΔIrs2) mice normalized energy balance, glucose homeostasis, and arcuate nucleus gene expression. Thus, Irs2 signaling in LepR-b neurons plays a crucial role in metabolic sensing and regulation. While not required for leptin action, Irs2 suppresses FoxO1 signaling in LepR-b neurons to promote energy balance and metabolism.

Elevated progesterone-to-estradiol ratio versus serum progesterone alone for predicting poor cycle outcome with in vitro fertilization.

OBJECTIVE: To determine whether a progesterone-to-estradiol (P/E2) ratio on day of human chorionic gonadotropin (hCG) administration would be a better predictor of in vitro fertilization (IVF) outcome than serum P alone. STUDY DESIGN: All 348 fresh IVF cycles performed in 2002 and 2003 at a university hospital center were reviewed for all cycle parameters as related to the peak P and peak P/E2 ration on day hCG administration. RESULTS: Out of the 348 cycles performed, 199 cycles resulted in clinical pregnancies. The mean P level (1.4 ng/mL) was equivalent in both conception and nonconception cycles. A P/E2 ratio > 1.0, however, was associated with a highly significant reduction in clinical pregnancy rate (38.2% vs. 62.6%, p< 0.01) and live birth rate (35.4% vs. 49.1%, p = 0.02). CONCLUSION: Cycles with elevated P/E2 ratios are associated with lower clinical pregnancy and live birth rates, which decrease further as the P/E2 ratio rises. P/E2 ratio improves the prediction of IVF outcome when compared to serum P levels alone.

A rare vulvar manifestation of neurofibromatosis 1 in a teen.

Neurofibromatosis 1 is an autosomal dominant disorder with cutaneous findings that include multiple café-au-lait spots, axillary/inguinal freckling, dermal, and plexiform neurofibromas. Skin manifestations, including involvement of the vulva, are often the most troubling physical finding to patients. Hormonal and growth factor changes during puberty have been implicated in neurofibroma growth. In the case presented here, an exceedingly rare isolated vulvar neurofibroma without clitoral involvement became enlarged and symptomatic, requiring excisional surgery after puberty. The diffuse involvement of these tumors makes complete resection very difficult and recurrence is common.

Detection of novel copy number variants in uterine leiomyomas using high-resolution SNP arrays.

Uterine leiomyomas (ULs) are benign monoclonal tumors originating from myometrial tissue in the uterus. Genetic pathways that lead to myometrial transformation into leiomyomas are largely unknown. Approximately 40% of ULs are karyotypically abnormal by G-banding; however, the remaining 60% of leiomyomas do not contain cytogenetically visible genomic rearrangements. Recent technological advances such as array based comparative genomic hybridization (array CGH) and dense single nucleotide polymorphism (SNP) arrays have enabled genome-wide scanning for genomic rearrangements missed by karyotype banding analysis. In the current study, we employed a high resolution SNP microarray on 16 randomly selected ULs and normal myometrium samples to detect submicroscopic (<5 Mb) chromosomal aberrations. The SNP array identified gene dosage changes in 56% of the fibroids (9/16), 25% of which (4/16) had aberrations >5 Mb, whereas 31% of which (5/16) contained only submicroscopic copy number changes (<5 Mb). We corroborated 3/5 submicroscopic changes using quantitative PCR, meaning that ultimately, 19% of our samples (3/16) were found to contain only submicroscopic changes. Novel submicroscopic aberrations on chromosomal segments 1q42.13, 11q13.1 and 13q12.13 and large, previously unreported deletions on 15q11.2-q23, 17p-q21.31 and 22q12.2-q12.3 were identified. Previously reported deletions on 1p, 3q, 7q, 13, and chromosome 14q were also noted. RHOU, MAP3K11 and WASF3 gene copy numbers were changed in the subset of leiomyomas with submicroscopic aberrations, and these genes have previously been implicated in tumorigenesis. Our findings support the hypothesis that a significant fraction of ULs without visible cytogenetic changes harbor submicroscopic genomic rearrangements which may in turn contribute to transformation of normal myometrial tissue into leiomyomas.

Comparison of FSH flare with and without pretreatment with oral contraceptive pills in poor responders undergoing in vitro fertilization.

OBJECTIVE: To compare FSH, LH, estrogen, and P flare response following 1 mg lupron injection in poor responders with or without pretreatment with oral contraceptive pills (OCPs). DESIGN: Prospective study. SETTING: University hospital. PATIENT(S): Poor responders undergoing IVF flare protocol from October 2002 to November 2003. INTERVENTION(S): Patients were divided into group A, who received OCPs before IVF cycle (n = 12), and group B, who did not (n = 7). One milligram Lupron was injected SC after measuring day 2 serum FSH, LH, estrogen, and P. After 24 hours, serum hormones were measured before lupron administration. MAIN OUTCOME MEASURE(S): Serum FSH, LH, estrogen, and P before and after 1 mg lupron RESULT(S): Basal FSH was similar in both groups (8.6 +/- 4.5 vs. 9.6 +/- 2.9 mIU/mL). Group A patients had significantly lower day 2 FSH (3.6 +/- 3.6 vs. 10.1 +/- 4.2 mIU/mL; P<.05). After lupron, although both groups had a significant rise in FSH and LH, mean LH rise in group B was 39.5 +/- 31 mIU/mL versus 11.3 +/- 4.6 mIU/mL in group A (P<.05). CONCLUSION(S): Pretreatment with OCPs in GnRH agonist flare protocol suppresses pre-Lupron FSH but does not blunt FSH flare. It blunts LH flare, which may be beneficial.