Conventionally, in vitro–fertilized (IVF) bovine embryos are morphologically evaluated at the time of embryo transfer to select those that are likely to establish a pregnancy. This method is, however, subjective and results in unreliable selection. Here we describe a novel selection system for IVF bovine blastocysts for transfer that traces the development of individual embryos with time-lapse cinematography in our developed microwell culture dish and analyzes embryonic metabolism. The system can noninvasively identify prognostic factors that reflect not only blastocyst qualities detected with histological, cytogenetic, and molecular analysis but also viability after transfer. By assessing a combination of identified prognostic factors—(i) timing of the first cleavage; (ii) number of blastomeres at the end of the first cleavage; (iii) presence or absence of multiple fragments at the end of the first cleavage; (iv) number of blastomeres at the onset of lag-phase, which results in temporary developmental arrest during the fourth or fifth cell cycle; and (v) oxygen consumption at the blastocyst stage—pregnancy success could be accurately predicted (78.9%). The conventional method or individual prognostic factors could not accurately predict pregnancy. No newborn calves showed neonatal overgrowth or death. Our results demonstrate that these five predictors and our system could provide objective and reliable selection of healthy IVF bovine embryos.
Selection of optimal quality embryos for in vitro fertilization (IVF) transfer is critical to successful live birth outcomes. Currently, embryos are chosen based on subjective assessment of morphologic developmental maturity. A non-invasive means to quantitatively measure an embryo's developmental maturity would reduce the variability introduced by the current standard. We present a method that exploits the scaling electrical properties of pre-transfer embryos to quantitatively discern embryo developmental maturity using light-induced dielectrophoresis (DEP). We show that an embryo's DEP response is highly correlated with its developmental stage. Uniquely, this technique allows one to select, in sequence and under blinded conditions, the most developmentally mature embryos among a mixed cohort of morphologically indistinguishable embryos cultured in optimized and sub-optimal culture media. Following assay, embryos continue to develop normally in vitro. Light-induced dielectrophoresis provides a non-invasive, quantitative, and reproducible means to select embryos for applications including IVF transfer and embryonic stem cell harvest.
The assisted reproductive technologies (ARTs) have been used in the production of rhesus monkey offspring at the Oregon National Primate Research Center (ONPRC) and that experience is summarized here. Additionally these technologies serve as a source of oocytes/embryos for monozygotic twinning, embryonic stem (ES) cell derivation and cloning. High fertilization efficiencies were realized with conventional insemination or following the use of intracytoplasmic sperm injection (ICSI) and approximately 50% of the resulting embryos grew in vitro to blastocysts. Both fresh and frozen sperm were employed in fertilization by ICSI and the resulting embryos could be low temperature stored for subsequent thawing and transfer when a synchronized recipient female was available or after shipment to another facility. Following the transfer of up to 3 embryos, an overall pregnancy rate of 30% was achieved with increasing rates dependent upon the number of embryos transferred. Singleton pregnancy outcomes following the transfer of ART produced embryos were similar to those observed in a control group of animals in the timed mated breeding colony at ONPRC. ICSI produced embryos were used in efforts to create monozygotic twins by blastomere separation or blastocyst splitting. While pregnancies were achieved following the transfer of demi-embryos, only one was a twin and it was lost to spontaneous abortion. ICSI produced embryos have also served as the source of blastocysts for the derivation of embryonic stem cells. These pluripotent cells hold potential for cell based therapies and we consider the monkey an important translational model in which to evaluate safety, efficacy and feasibility of regenerative medicine approaches based on the transplantation of stem cell-derived progeny. Finally, efforts to produce genetically-identical monkeys by nuclear transfer have been briefly summarized.
Non-invasive gamete and embryo assessment is considered an important focus in assisted reproductive technologies (ART). Currently, the selection of embryos for transfer is based on morphological indices. Though successful, the field of ART would benefit from a non-invasive quantitative method of viability determination. Omics technologies, including transcriptomics, proteomics and metabolomics, have already begun providing evidence that viable gametes and embryos possess unique molecular profiles with potential biomarkers that can be utilized for developmental and/or viability selection. Unlike the human genome that is relatively fixed and steady throughout the human body, the human proteome, estimated at over a million proteins, is more complex, diverse and dynamic. It is the proteins themselves that contribute to the physiological homeostasis in any cell or tissue. Of particular interest in ART is the secretome, those proteins that are produced within the embryo and secreted into the surrounding environment. Defining the human embryonic secretome has the potential to expand our knowledge of embryonic cellular processes, including the complex dialogue between the developing embryo and its maternal environment, and may also assist in identifying those embryos with the highest implantation potential. Advances in proteomic technologies have allowed the non-invasive profiling of the human embryonic secretome with ongoing research focused on correlation with outcome. From a clinical perspective, embryo selection based on morphological assessment and non-invasive analysis of the human embryonic secretome may improve IVF success and lead to routine single embryo transfers.
proteomics; embryo; secretome; non-invasive assessment
Studies using animal models demonstrated the importance of autocrine/paracrine factors secreted by preimplantation embryos and reproductive tracts for embryonic development and implantation. Although in vitro fertilization-embryo transfer (IVF-ET) is an established procedure, there is no evidence that present culture conditions are optimal for human early embryonic development. In this study, key polypeptide ligands known to be important for early embryonic development in animal models were tested for their ability to improve human early embryo development and blastocyst outgrowth in vitro. We confirmed the expression of key ligand/receptor pairs in cleavage embryos derived from discarded human tri-pronuclear zygotes and in human endometrium. Combined treatment with key embryonic growth factors (brain-derived neurotrophic factor, colony-stimulating factor, epidermal growth factor, granulocyte macrophage colony-stimulating factor, insulin-like growth factor-1, glial cell-line derived neurotrophic factor, and artemin) in serum-free media promoted >2.5-fold the development of tri-pronuclear zygotes to blastocysts. For normally fertilized embryos, day 3 surplus embryos cultured individually with the key growth factors showed >3-fold increases in the development of 6–8 cell stage embryos to blastocysts and >7-fold increase in the proportion of high quality blastocysts based on Gardner’s criteria. Growth factor treatment also led to a 2-fold promotion of blastocyst outgrowth in vitro when day 7 surplus hatching blastocysts were used. When failed-to-be-fertilized oocytes were used to perform somatic cell nuclear transfer (SCNT) using fibroblasts as donor karyoplasts, inclusion of growth factors increased the progression of reconstructed SCNT embryos to >4-cell stage embryos. Growth factor supplementation of serum-free cultures could promote optimal early embryonic development and implantation in IVF-ET and SCNT procedures. This approach is valuable for infertility treatment and future derivation of patient-specific embryonic stem cells.
The recent development of genomic selection induces dramatic changes in the way genetic selection schemes are to be conducted. This review describes the new context and corresponding needs for genomic based selection schemes and how reproductive technologies can be used to meet those needs. Information brought by reproductive physiology will provide new markers and new improved phenotypes that will increase the efficiency of selection schemes for reproductive traits. In this context, the value of the reproductive techniques including assisted embryo based reproductive technologies (Multiple Ovaluation Embryo Transfer and Ovum pick up associated to in vitro Fertilization) is also revisited. The interest of embryo typing is discussed. The recent results obtained with this emerging technology which are compatible with the use of the last generation of chips for genotype analysis may lead to very promising applications for the breeding industry. The combined use of several embryo based reproductive technologies will probably be more important in the near future to satisfy the needs of genomic selection for increasing the number of candidates and to preserve at the same time genetic variability.
We report the derivation and characterization of two new human embryonic stem cells (hESC) lines (CU1 and CU2) from embryos with an irreversible loss of integrated organismic function. In addition, we analyzed retrospective data of morphological progression from embryonic day (ED) 5 to ED6 for 2480 embryos not suitable for clinical use to assess grading criteria indicative of loss of viability on ED5. Our analysis indicated that a large proportion of in vitro fertilization (IVF) embryos not suitable for clinical use could be used for hESC derivation. Based on these combined findings, we propose that criteria commonly used in IVF clinics to determine optimal embryos for uterine transfer can be employed to predict the potential for hESC derivation from poor quality embryos without the destruction of vital human embryos.
Objective: Embryonic chromosomal abnormality is one of the main reasons for invitro fertilization (IVF) failure. This study aimed at evaluating the value of Fluorescence in-situ Hybridization (FISH)-based Preimplantation Genetic Diagnosis (PGD) in screening for embryonic chromosomal abnormality to increase the successful rate of IVF. Method: Ten couples, four with high risk of chromosomal abnormality and six infertile couples, underwent FISH-based PGD during IVF procedure. At day 3, one or two blastomeres were aspirated from each embryo. Biopsied blastomeres were examined using FISH analysis to screen out embryos with chromosomal abnormalities. At day 4, embryos without detectable chromosomal abnormality were transferred to the mother bodies as in regular IVF. Results: Among 54 embryos screened using FISH-based PGD, 30 embryos were detected to have chromosomal abnormalities. The 24 healthy embryos were implanted, resulting in four clinical pregnancies, two of which led to successful normal birth of two healthy babies; one to ongoing pregnancy during the writing of this article; and one to ectopic pregnancy. Conclusion: FISH-based PGD is an effective method for detecting embryonic chromosomal abnormality, which is one of the common causes of spontaneous miscarriages and chromosomally unbalanced offsprings.
Preimplantation genetic diagnosis; Fluorescence in-situ Hybridization (FISH); Chromosome abnormality
In assisted reproductive technology (ART), the pregnancy and birth rates following in vitro fertilization (IVF) attempts are still low. Recently, apoptotic markers have been suggested as new criteria for oocyte and embryo quality selection. Many studies have provided evidence that poor oocyte and embryo quality can be associated with apoptosis. The aim of this review is to summarize our current knowledge on the apoptotic process in oocytes and embryos, and focus on the possibility for using apoptotic markers as a reliable and predictive marker to select competent oocytes and embryos during IVF. Moreover, it is currently accepted that IVF failures, linked to poor embryo quality, are, in part, associated with suboptimal in vitro culture conditions. Here, we also review the current state of knowledge concerning how the genetic control of apoptosis during folliculogenesis and pre- implantation embryonic development is affected by in vitro culture conditions during IVF. In the future, identification of apoptotic markers in ART for oocyte and embryo selection should result in the development of new agonistic or antagonistic molecules of apoptosis by medicinal chemistry.
Apoptosis; Biological Markers; Embryonic Development; Female; Fertilization in Vitro; Humans; Male; Oocytes; cytology; Signal Transduction; IVF; Apoptosis markers; Genetic control; Oocyte; Embryo; Microenvironment; Diagnosis
In IVF-ICSI cycles with single embryo transfer (SET), embryo selection for transfer is of crucial importance. The present study aimed to define which embryo parameters might be related to the implantation potential of advanced blastocysts.
Overall, in 203 cycles with SET, developmental characteristics of 93 implanted (group A) and 110 non-implanted (group B) advanced blastocysts of good quality were compared. The following developmental parameters were assessed in the two groups: normal fertilization, developmental stage on day 5, number of blastomeres on day 2 and on day 3, fragmentation rate on day 3, compaction on day 4 and cleavage pattern on day 2 and day 3.
Expanded blastocysts compared to full blastocysts have higher implantation potential (56.5% vs. 29.3%, p < 0.05). In group B, a higher proportion of advanced blastocysts showed between 10% and 50% anucleated fragments on day 3 than in group A (23.6 vs 11.8, P = 0.03). Advanced blastocysts with >10–50% fragments on day 3 showed a significant lower implantation (29.7%) than those with ≤ 10%fragments (49.4%, P = 0.03). All the other parameters analysed were comparable for the two groups.
Developmental stage on day 5 and fragmentation rate on day 3 were related to the implantation potential of advanced blastocysts and should also be taken into account in the selection of the best advanced blastocyst for transfer.
HLA-G is a major histocompatibility complex (MHC), class Ib molecule that is selectively expressed at the fetal–maternal interface. It is thought to play a role in protecting the fetus from the maternal immune response. Interestingly, the preimplantation embryo development (Ped) gene product Qa-2 is also a mouse MHC class Ib protein that affects cleavage and division of preimplantation mouse embryos and subsequent embryonic survival. Data from many human in vitro fertilization (IVF) clinics suggest that the mouse Ped phenomenon also exists in human because embryos fertilized at the same time have different cleavage rates and consequently different IVF outcomes. As HLA-G is expressed in human early embryos, it is highly regarded as the functional homologue of Qa-2. Whether HLA-G expression is correlated with the cleavage rate of human embryos has great potential clinical value.
In this study, 45 human early abnormal fertilized embryos (3 PN) from patients undergoing in vitro fertilization were used to test the effects of HLA-G knock-down via infection with adenovirus carrying its specific siRNA on the cleavage rate in a 2-day culture period. One-way ANOVA, Post hoc and Chi-square were used to compare groups. A p-value smaller than 0.05 was considered statistically significant.
Knocking-down HLA-G in human pre-implantation stage embryos resulted in a higher cell arrest rate and a slower cleavage rate.
The results from the present study suggested that HLA-G might play an important role in early human embryo development.
Adenovirus vector; Cleavage; HLA-G; In vitro fertilization; Mouse Qa-2 antigen; Preimplantation embryo; RNAi
Preimplantation genetic diagnosis (PGD) testing is the practice of obtaining a cellular biopsy sample from a developing human
oocyte or embryo, acquired via a cycle of in vitro fertilization (IVF); evaluating the genetic composition of this sample; and using
this information to determine which embryos will be optimal for subsequent uterine transfer. PGD has become an increasingly
useful adjunct to IVF procedures. The ability to provide couples who are known carriers of genetic abnormalities the opportunity
to deliver healthy babies has opened a new frontier in reproductive medicine. The purpose of the PGD is enables us to choose
which embryos will be implanted into the mother. In the present study 137 families who had undergone IVF at Habib Medical
Centre, were enrolled for the PGD analysis. The couple visited the clinic for the sex selection, recurrent fetal loss and with the
recurrent IVF failure. 802 embryos were tested by the biopsy method and 512 are found to be normal and 290 were abnormal
embryos. In this study only 24% of the embryos were transferred and the remaining was not transferred because of the
abnormalities or undesired sex of the embryos. The structural and numerical abnormalities were found to be 16.8%.
Preimplantation genetic diagnosis (PGD); invitro fertilization (IVF); Biopsy; Saudi Arabia
Assisted reproductive technology (ART) has been recognized for its success in treating infertility, a condition that affects 15 percent of couples in the United States. The most popular option is in vitro fertilization (IVF), which relies on embryo culture, selection, and transfer for implantation, with the ultimate aim of pregnancy. Previous embryo selection methods relied on morphological factors to select for greatest viability. At Yale’s Frontiers in Reproduction Conference on April 29, 2011, at the New Haven Lawn Club, Dr. Denny Sakkas of Yale’s Department of Obstetrics, Gynecology, and Reproductive Sciences presented a paradigm shift: using morphological factors along with metabolic, protein, and genetic markers in culture media to enhance embryo selection and IVF success rates.
in vitro fertilization; assisted reproductive technology; multiple pregnancies
With improved survival rates among cancer patients, fertility preservation is now being recognized as an issue of great importance. There are currently several methods of fertility preservation available in female cancer patients and the options and techniques via assisted reproduction and cryopreservation are increasing, but some are still experimental and continues to be evaluated. The established means of preserving fertility include embryo cryopreservation, gonadal shielding during radiation therapy, ovarian transposition, conservative gynecologic surgery such as radical trachelectomy, donor embryos/oocytes, gestational surrogacy, and adoption. The experimental methods include oocyte cryopreservation, ovarian cryopreservation and transplantation, in vitro maturation, and ovarian suppression. With advances in methods for the preservation of fertility, providing information about risk of infertility and possible options of fertility preservation to all young patients with cancer, and discussing future fertility with them should be also considered as one of the important parts of consultation at the time of cancer diagnosis.
A valuable tool for both research and industry, in vitro fertilization (IVF) has applications range from gamete selection and preservation of traits to cloning. Although IVF has achieved worldwide use, with approximately 339,685 bovine embryos transferred in 2010 alone, there are still continuing difficulties with efficiency. It is rare to have more than 40% of fertilized in vitro cattle oocytes reach blastocyst stage by day 8 of culture, and pregnancy rates are reported as less than 45% for in vitro produced embryos. To investigate potential influences in-vitro fertilization (IVF) has on embryonic development, this study compares in vivo- and in vitro-derived bovine blastocysts at a similar stage and quality grade (expanded, excellent quality) to determine the degree of transcriptomic variation beyond morphology using RNA-Seq.
A total of 26,906,451 and 38,184,547 fragments were sequenced for in vitro and in vivo embryo pools, respectively. We detected expression for a total of 17,634 genes, with 793 genes showing differential expression between the two embryo populations with false discovery rate (FDR) < 0.05. There were also 395 novel transcribed units found, of which 45 were differentially expressed (FDR < 0.05). In addition, 4,800 genes showed evidence of alternative splicing, with 873 genes displaying differential alternative splicing between the two pools (FDR < 0.05). Using GO enrichment analysis, multiple biological pathways were found to be significantly enriched for differentially expressed genes (FDR < 0.01), including cholesterol and sterol synthesis, system development, and cell differentiation.
Thus, our results support that IVF may influence at the transcriptomic level and that morphology is limited in full characterization of bovine preimplantation embryos.
In vivo embryo; In vitro fertilization; RNA-Seq; Transcriptome; Alternative splicing
Uses and potential uses of in vitro fertilization are: (1) a research tool for investigating biochemistry of fertilization, (2) an assay for fertilizing ability, (3) a potentially useful clinical approach for certain cases of infertility, and (4) a potentially useful means for improving animal breeding. In vitro fertilization methodology is sufficiently advanced for gametes of several mammalian species, especially mouse, rabbit, and rat, for use in evaluating effects imposed by toxic agents of environmental or genetic origin. Alteration of the normal events of fertilization and/or embryonic development following transfer of in vitro fertilized embryos into surrogate dams can serve as end point(s) in applying this means for toxicity testing. In vitro fertilization of mouse and rat ova has been explored as an alternative to in vivo fertilization in male contraceptive development studies. Original observations on toxicity of abnormal O2 concentrations for rabbit fertilization in vitro are reported here. Ova were fertilized under 0, 20, 48, and 95% O2, but toxicity from the extreme conditions, 0 and 95% was apparent from comparison of proportions of fertilized ova reaching the 4-cell stage during the 24-hr incubation. Toxicity was further evidenced by inability of embryos fertilized under 0 and 95% O2 tensions, in contrast to those fertilized under 20% O2, to sustain normal gestation following their transfer into recipient does. Recent success in the fertilization of cow ova in vitro in this laboratory provides encouragement to develop a useful means for testing normalcy of gametes in this species. Such studies might lead to useful screening procedures for avoidance of human infertility resulting from hazardous environmental conditions.
Interspecies somatic cell nuclear transfer (iSCNT) has been proposed as a tool to address basic developmental questions and to improve the feasibility of cell therapy. However, the low efficiency of iSCNT embryonic development is a crucial problem when compared to in vitro fertilization (IVF) and intraspecies SCNT. Thus, we examined the effect of donor cell species on the early development of SCNT embryos after reconstruction with bovine ooplasm.
No apparent difference in cleavage rate was found among IVF, monkey-bovine (MB)-iSCNT, and bovine-bovine (BB)-SCNT embryos. However, MB-iSCNT embryos failed to develop beyond the 8- or 16-cell stages and lacked expression of the genes involved in embryonic genome activation (EGA) at the 8-cell stage. From ultrastructural observations made during the peri-EGA period using transmission electron microscopy (TEM), we found that the nucleoli of MB-iSCNT embryos were morphologically abnormal or arrested at the primary stage of nucleologenesis. Consistent with the TEM analysis, nucleolar component proteins, such as upstream binding transcription factor, fibrillarin, nucleolin, and nucleophosmin, showed decreased expression and were structurally disorganized in MB-iSCNT embryos compared to IVF and BB-SCNT embryos, as revealed by real-time PCR and immunofluorescence confocal laser scanning microscopy, respectively.
The down-regulation of housekeeping and imprinting genes, abnormal nucleolar morphology, and aberrant patterns of nucleolar proteins during EGA resulted in developmental failure in MB-iSCNT embryos. These results provide insight into the unresolved problems of early embryonic development in iSCNT embryos.
Currently, the assessment of sperm function in a raw or processed semen sample is not able to reliably predict sperm ability to withstand freezing and thawing procedures and in vivo fertility and/or assisted reproductive biotechnologies (ART) outcome. The aim of the present study was to investigate which parameters among a battery of analyses could predict subsequent spermatozoa in vitro fertilization ability and hence blastocyst output in a goat model. Ejaculates were obtained by artificial vagina from 3 adult goats (Capra hircus) aged 2 years (A, B and C). In order to assess the predictive value of viability, computer assisted sperm analyzer (CASA) motility parameters and ATP intracellular concentration before and after thawing and of DNA integrity after thawing on subsequent embryo output after an in vitro fertility test, a logistic regression analysis was used. Individual differences in semen parameters were evident for semen viability after thawing and DNA integrity. Results of IVF test showed that spermatozoa collected from A and B lead to higher cleavage rates (0 < 0.01) and blastocysts output (p < 0.05) compared with C. Logistic regression analysis model explained a deviance of 72% (p < 0.0001), directly related with the mean percentage of rapid spermatozoa in fresh semen (p < 0.01), semen viability after thawing (p < 0.01), and with two of the three comet parameters considered, i.e tail DNA percentage and comet length (p < 0.0001). DNA integrity alone had a high predictive value on IVF outcome with frozen/thawed semen (deviance explained: 57%). The model proposed here represents one of the many possible ways to explain differences found in embryo output following IVF with different semen donors and may represent a useful tool to select the most suitable donors for semen cryopreservation.
Preimplantation genetic diagnosis (PGD) is considered highly successful in respect to its accuracy in detecting genetic anomalies but the effects of embryo biopsy on embryonic/fetal growth and development are less known, particularly in conjunction with in vitro fertilization (IVF). Here, we compared biopsied (B) and non-biopsied (NB) mouse embryos for their developmental competence. Embryos C57BL/6 (B6) and B6D2F2 (F2) generated by IVF were subjected to single blastomere biopsy at the 4-cell stage, and were either cultured for 120 h and subjected to differential inner cell mass (ICM) and trophoblast (T) staining, or were transferred into the uterine tubes of surrogate mothers after 72 h of culture, to examine their pre- and post-implantation development, respectively. Non-biopsied embryos from the same IVF cohorts served as controls. Embryo biopsy negatively affected preimplantation development to blastocyst in C57BL/6 (69 vs 79%, P<0.01) but not in B6D2F1 mice (89 vs 91%, P=NS). Although B6 embryos had lower total cell number than F2 (B6: 47 and 61 vs. F1: 53 and 70; B and NB, respectively, P<0.05) there were no differences between B and NB blastocysts in %ICM (B6: 19.8 vs 19.8; F2: 20.9 vs 20.4, P=NS) and ICM:T ratio (B6: 4.7 vs 4.7; F2: 4.4 vs. 4.7) in both mouse strains. Post-implantation development to live fetuses of B embryos as compared to NB counterparts was impaired in C57BL/6 (6 vs 18%, P<0.001) but not in B6D2F1 mice (26 vs 35%, P=NS). We conclude that blastomere biopsy impairs embryonic/fetal development in mice known to be sensitive to in vitro culture and manipulations. Such mice model infertile couples with poor quality gametes seeking help in assisted reproduction technologies (ART) clinics.
embryo manipulation; assisted reproductive technology; development; in vitro fertilization; embryo biopsy; C57BL/6
Infertility and subfertility are important and pervasive reproductive problems in both domestic animals and humans. The majority of embryonic loss occurs during the first three weeks of pregnancy in cattle and women due, in part, to inadequate endometrial receptivity for support of embryo implantation. To identify heifers of contrasting fertility, serial rounds of artificial insemination (AI) were conducted in 201 synchronized crossbred beef heifers. The heifers were then fertility classified based on number of pregnancies detected on day 35 in four AI opportunities. Heifers, classified as having high fertility, subfertility or infertility, were selected for further study. The fertility-classified heifers were superovulated and flushed, and the recovered embryos were graded and then transferred to synchronized recipients. Quantity of embryos recovered per flush, embryo quality, and subsequent recipient pregnancy rates did not differ by fertility classification. Two in vivo-produced bovine embryos (stage 4 or 5, grade 1 or 2) were then transferred into each heifer on day 7 post-estrus. Pregnancy rates were greater in high fertility than lower fertility heifers when heifers were used as embryo recipients. The reproductive tracts of the classified heifers were obtained on day 14 of the estrous cycle. No obvious morphological differences in reproductive tract structures and histology of the uterus were observed in the heifers. Microarray analysis revealed differences in the endometrial transcriptome based on fertility classification. A genome-wide association study, based on SNP genotyping, detected 7 moderate associations with fertility across 6 different chromosomes. Collectively, these studies support the idea that innate differences in uterine function underlie fertility and early pregnancy loss in ruminants. Cattle with defined early pregnancy success or loss is useful to elucidate the complex biological and genetic mechanisms governing endometrial receptivity and uterine competency for pregnancy.
To assess the effects of light from an integrated optical microscope and evaluate the safety of time-lapse observations using a built-in microscope incubator.
We prospectively compared the fertilization rate and embryonic morphology after intracytoplasmic sperm injection between embryos cultured with time-lapse observations every 15 min in an incubator with an integrated optical microscope and embryos with intermittent observations (once a day) in conventional incubators.
No significant differences were observed in the fertilization rate (57.5% vs. 57.5%) or the rate of excellent-good cleavage embryos (36.0% vs. 36.0%).
These results suggest that time-lapse observations using an incubator with an integrated optical microscope may therefore be safely utilized in clinical practice.
Built-in microscope incubator; Embryo grade; ICSI; Time-lapse observation
Assessment of embryo viability is a key component of in vitro fertilization (IVF) and currently relies largely on embryo morphology and cleavage rate. In this study, we used receiver operating characteristic (ROC) analysis to compare the Viability Score (generated by metabolomic profiling of spent embryo culture media using near infrared (NIR) spectroscopy) to morphologic grading for predicting pregnancy in women undergoing single embryo transfer (SET) on day 5.
A total of 198 spent embryo culture media samples were collected in four IVF centers located in the USA, Europe and Australia. First, 137 samples (training set) were analyzed by NIR to develop an algorithm that generates a Viability Score predictive of pregnancy for each sample. Next, 61 samples (validation set) were analyzed by observers blinded to embryo morphology and IVF outcome, using the Day 5 algorithm generated with the training set. Pregnancy was defined as fetal cardiac activity (FCA) at 12 weeks of gestation.
The Area Under the Curve (AUC) was greater for the metabolomic Viability Score compared to Morphology [Training set: 0.75 versus 0.55, p = 0.0011; Validation set: 0.68 versus 0.50, P = 0.021], and for a Composite score (obtained using a model combining Viability Score with morphologic grading), compared to morphology alone [0.74 versus 0.50, p = 0.004].
Our findings suggest that Viability Score alone or in combination with morphologic grading has the potential to be a better classifier for pregnancy outcome than morphology alone in women undergoing SET on day 5.
ROC analysis; Assisted reproductive technologies; ART; In vitro fertilization; IVF; Morphologic grade; Metabolomics; Viability Score
Smooth atraumatic embryo transfer is paramount for the success of in-vitro fertilization (IVF). In difficult cases, cervical canal manipulation may be required.
To see if surgical correction of the cervical canal or cervical canal refashioning could improve ease of embryo transfer.
Private infertility and IVF hospital.
MATERIALS AND METHODS:
Patients: 11 women with failed 1-3 IVF cycles with history of extremely difficult embryo transfers (ETs) despite undergoing cervical dilatation in the cycle prior to IVF.
Operative hysteroscopy using Versapoint for refashioning of the cervical canal.
MAIN OUTCOME MEASURES:
Ease of ET in the subsequent IVF cycle. Secondary outcome measure was to assess reproductive outcome.
Easy and atraumatic ET in the IVF cycle after procedure in 100% patients. PR was 46.5%.
Use of Versapoint for refashioning the cervical canal can improve the quality of ET and PR.
Cervical stenosis; embryo transfer; hysteroscopy; in vitro fertilization; Versapoint
Purpose: Case studies of four in vitro fertilization (IVF) cycles where embryo transport by commercial airline followed by biopsy and genetic analysis with subsequent culture to Day 5 and resulting ongoing pregnancy.
Method: Retrospective clinical case study of 4 patients requiring preimplantation genetic diagnosis (PGD) testing. Normally fertilized embryos were transported in a battery-powered portable incubator by commercial airline following evaluation for fertilization under controlled conditions from the Center for Assisted Reproduction, Bedford, Texas to the Reproductive Genetic Institute, Chicago, Illinois. Following Day 3 embryo biopsy and genetic analysis, embryos were transported back to the Center for Assisted Reproduction for Day 5 embryo transfer.
Results: Ongoing clinical pregnancy resulted for all patients receiving embryo transfer.
Conclusion: These results demonstrate the feasibility of embryo transport by air for centers that do not have the in-house capabilities to perform genetic analysis. With successful pregnancies obtained through extended culture to Day 5, embryos requiring genetic analysis can be successfully transported by air, tested, and returned to the initial facility for embryo transfer without time restriction.
clinical pregnancy; embryo transport; extended culture; PGD
Purpose: To examine the effect of intracytoplasmic sperminjection (ICSI) on embryo fragmentation and implantationrates in those embryos chosen for transfer compared toconventional in vitro fertilization (IVF).
Methods: We compared 253 infertility patients (71 ICSI and182 IVF) with respect to age, semen analysis, number ofembryos transferred, embryo fragmentation, implantationrate, and pregnancy rate. Embryo fragmentation wasdetermined by one observer at the same laboratory over the entirestudy period.
Results: A statistically significant difference was observedin mean embryo grade between IVF (2.2 ± 0.84) and ICSI(2.5 ± 0.77), P = 0.01. Additionally, the IVF patients hadsignificantly more nonfragmented (grade I) embryoscompared to the ICSI group, P < 0.01.
Conclusions: These data suggest that ICSI, irrespective ofsemen parameters, may increase embryo fragmentation andproduce fewer nonfragmented grade I embryos while maintaining implantation and pregnancy rates similar toconventional IVF.
embryo fragmentation; in vitro fertilization; intracytoplasmic sperm injection; pregnancy rate; sperm parameters