J.R. Alegretti

Prospective randomized comparison of human oocyte cryopreservation with slow-rate freezing or vitrification

OBJECTIVE To compare cryopreservation of mature human oocytes with slow-rate freezing and vitrification and determine which is most efficient at establishing a pregnancy. DESIGN Prospective randomized. SETTING Academically affiliated, private fertility center. PATIENT(S) Consenting patients with concerns about embryo cryopreservation and more than nine mature oocytes at retrieval were randomized to slow-rate freezing or vitrification of supernumerary (more than nine) oocytes. INTERVENTION(S) Oocytes were frozen or vitrified, and upon request oocytes were thawed or warmed, respectively. MAIN OUTCOME MEASURE(S) Oocyte survival, fertilization, embryo development, and clinical pregnancy. RESULT(S) Patient use has resulted in 30 thaws and 48 warmings. Womens age at time of cryopreservation was similar. Oocyte survival was significantly higher following vitrification/warming (81%) compared with freezing/thawing (67%). Fertilization was more successful in oocytes vitrified/warmed compared with frozen/thawed. Fertilized oocytes from vitrification/warming had significantly better cleavage rates (84%) compared with freezing/thawing (71%) and resulted in embryos with significantly better morphology. Although similar numbers of embryos were transferred, embryos resulting from vitrified oocytes had significantly enhanced clinical (38%) pregnancy rates compared with embryos resulting from frozen oocyte (13%). Miscarriage and/or spontaneous abortion rates were similar. CONCLUSION(S) Our results suggest that vitrification/warming is currently the most efficient means of oocyte cryopreservation in relation to subsequent success in establishing pregnancy.

Extensive Excision of Deep Infiltrative Endometriosis before In Vitro Fertilization Significantly Improves Pregnancy Rates

STUDY OBJECTIVE We sought to compare the outcomes of in vitro fertilization (IVF) treatments in women with infertility-associated deep infiltrative endometriosis (DIE) who underwent extensive laparoscopic excision of endometriosis before IVF with those who underwent IVF only. DESIGN Prospective cohort study. SETTING Infertility clinic and private hospital in São Paulo, Brazil. PATIENTS A total of 179 infertile patients younger than 38 years had symptoms and/or signs of endometriosis and sonographic images suggestive of DIE. INTERVENTIONS After thorough counseling, 179 women were invited to participate in a prospective cohort study with 2 treatment options: IVF without undergoing laparoscopic surgery (group A, n = 105) and extensive laparoscopic excision of DIE before IVF (group B, n = 64). Ten women were lost to follow-up. The IVF outcomes were compared between the 2 groups. MEASUREMENTS AND MAIN RESULTS In group B, patients had 5 +/- 2 (mean +/- SD) DIE lesions excised during laparoscopy. Patient characteristics in groups A and B, respectively, were: age (32 +/- 3 vs 32 +/- 3 years, p = .94), infertility duration (29 +/- 20 vs 27 +/- 17 months, p = .45), day-3 serum follicle-stimulating hormone levels (5.6 +/- 2.5 vs 5.9 +/- 2.5 IU/L, p = .50), and previous IVF attempts (1 +/- 1 vs 2 +/- 1, p = .01). The IVF outcomes differed between groups A and B, respectively, with regard to total dose of recombinant follicle-stimulating hormone required to accomplish ovulation induction (2380 +/- 911 vs 2542 +/- 1012 IU, p = .01), number of oocytes retrieved (10 +/- 5 vs 9 +/- 5, p = .04), and pregnancy rates (24% vs 41%, p = .004), but not number of embryos transferred (3 +/- 1 vs 3 +/- 1, p = 1). The odds ratio of achieving a pregnancy were 2.45 times greater in group B than in group A. CONCLUSION Extensive laparoscopic excision of DIE significantly improved IVF pregnancy rates of women with infertility-associated DIE.

High implantation and pregnancy rates with transfer of human blastocysts developed in preimplantation stage one and blastocyst media

OBJECTIVE To evaluate the proficiency of preimplantation stage one (P1) and blastocyst media in supporting human blastocyst development and to document implantation and clinical pregnancy rates from the transfer of the normally developed blastocysts. DESIGN Retrospective clinical study. SETTING Private IVF unit of a university-affiliated center. PATIENT(S) Twenty-eight women aged 33.7 +/- 2.9 years who underwent IVF treatment for infertility. INTERVENTION(S) Bipronucleate oocytes obtained from IVF and intracytoplasmic sperm injection were cultured in vitro with P1 and blastocyst media for 96-120 hours. One to three embryos were transferred (2.1 +/- 0.2 for the patients who became pregnant and 1.5 +/- 0.3 for those who did not become pregnant). MAIN OUTCOME MEASURE(S) Total number and percentage of developed blastocysts, frequency of blastocysts of grades A and B, and implantation and pregnancy rates. RESULT(S) From 431 oocytes retrieved, 269 bipronucleate oocytes were cultured, producing 81 blastocysts that resulted in the transfer of 54 embryos in 27 procedures. Blastocysts developed in 39.7% +/- 5.5% of the pregnant group and 30.2% +/- 4.5% of the nonpregnant group. From 15 (15/27 = 55.6%) clinical pregnancies, 18 (18/54 = 33.3%) gestational sacs were visualized. The rate of implantation in the pregnant group was 58.1% (18/31). CONCLUSION(S) These results provide evidence for the benefits of extending human embryo culture with P1 and blastocyst media for all normally fertilized embryos in vitro.

Does slow embryo development predict a high aneuploidy rate on trophectoderm biopsy

The aneuploidy rates in expanded blastocysts biopsied on days 5 and 6 development were assessed in women undergoing IVF followed by array comparative genomic hybridization. This study included 1171 expanded blastocysts from 465 patients. Among the 465 patients, 215 and 141 underwent embryo biopsy on day 5 and day 6 (46.2% and 30.3%, respectively), and 109 underwent biopsy on both days 5 and 6 (23.4%). The cycles of 206 women were cancelled because only aneuploidy embryos were present (44.3%). The aneuploid embryos were classified according to the type as single, double or complex aneuploidy. No differences were observed in the distributions of these three categories according to the day of the biopsy. The aneuploidy rate was also evaluated according to maternal age, and was found to be higher in older patients; however, no differences in this rate were detected between embryos biopsied on days 5 and 6 according to maternal age. Biopsy was carried out when blastocysts reached the expanded stage. The embryos biopsied on day 6 had a higher rate of aneuploidy (69.9%) than those biopsied on day 5 (61.4%); however, the euploid embryos transferred had similar chances for successful and healthy gestation.

A Euploid Line of Human Embryonic Stem Cells Derived from a 43,XX,dup(9q),+12,-14,-15,-18,-21 Embryo

Aneuploid embryos diagnosed by FISH-based preimplantation genetic screening (PGS) have been shown to yield euploid lines of human embryonic stem cells (hESCs) with a relatively high frequency. Given that the diagnostic procedure is usually based on the analysis of 1–2 blastomeres of 5 to 10-cell cleavage-stage embryos, mosaicism has been a likely explanation for the phenomena. However, FISH-based PGS can have a significant rate of misdiagnosis, and therefore some of those lines may have been derived from euploid embryos misdiagnosed as aneuploid. More recently, coupling of trophectoderm (TE) biopsy at the blastocyst stage and array-CGH lead to a more informative form of PGS. Here we describe the establishment of a new line of hESCs from an embryo with a 43,XX,dup(9q),+12,-14,-15,-18,-21 chromosomal content based on array-CGH of TE biopsy. We show that, despite the complex chromosomal abnormality, the corresponding hESC line BR-6 is euploid (46,XX). Single nucleotide polymorphism analysis showed that the embryo´s missing chromosomes were not duplicated in BR-6, suggesting the existence of extensive mosaicism in the TE lineage.

How general semen quality influences the blastocyst formation rate: Analysis of 4205 IVF cycles

Objective To select embryos with higher implantation potential, the extended culture has been the most frequently applied strategy worldwide, and consequently leads to higher live birth rates per transfer. Sperm quality is a determining feature, and it may influence the outcomes of IVF from fertilization to embryo development. Therefore, we hypothesize that blastocyst formation may also be impaired by general semen quality. Methods We analyzed 4205 IVF cycles. Four study groups were designed according to semen quality: normal, mild alteration, severe alteration and epididymis. All cycles were intended to extend embryo culture until the blastocyst stage, and embryo development was evaluated. Results Regarding cleavage rate, the normal and mild alteration semen groups were equivalent, and the severe alteration and epididymis semen groups were equivalent to each other. The blastocyst formation rate decreased with semen quality. At least one blastocyst formed in 79.9% of cycles for the normal semen group, whereas the percentage of cycles with the formation of at least one blastocyst was slightly lower for the mild alteration (75.6%), severe alteration (76.4%) and epididymis (76.8%) semen groups. A multivariate logistic regression showed that for each additional cleaved embryo on day 3, the chance of having at least one blastocyst doubles. Additionally, the chance of having at least one blastocyst decreased when semen presented mild or severe alterations. Conclusion The general quality of sperm is a good predictor of blastocyst formation, significantly affecting the likelihood of having at least one blastocyst at the end of the cycle. Based on our findings, it is necessary to consider general semen quality and the number of cleaved embryos when forecasting the possibility of blastocyst formation and transfer in an extended culture system.