Comparison of Pregnancy Outcomes Between Single-Morula Embryo Transfer and Single-Blastocyst Transfer in Fresh IVF/ICSI Cycles.

BACKGROUND This study investigated the effectiveness and feasibility of day 4 (D4) morula embryo transfer (ET) in comparison with day 5 (D5) blastocyst ET, with regards to their clinical data, laboratory test results, and pregnancy outcomes. MATERIAL AND METHODS This retrospective cohort study enrolled 1070 patients, including 178 cases in group D4 and 892 cases in group D5. The endpoint was live birth rate after fresh embryo transfer. Furthermore, the clinical outcomes of D4 embryos with different morphology were compared and assigned to 3 groups: in group 1 (n=66) the embryos were compacted but not expanded, in group 2 (n=102) the embryos were compacted and expanded (early blastocyst), and in group 3 (n=10) the embryos were not compacted. RESULTS Groups D4 and D5 had comparable clinical pregnancy rates (53.37% vs. 59.97%) and live birth rates (43.25% vs 50.89%), and there were no significant differences between the 2 groups. In group 3, there was only 1 clinical pregnancy and no live birth. In comparison between group 1 and group 2, the clinical pregnancy rate of group 2 showed an upward trend (48.48% vs 60.78%), but there was no significant difference. There was also no statistically significant difference in the live birth rate between the 2 groups (42.42% vs 49.01%). CONCLUSIONS Transferring of compacted embryos or early blastocysts can result in high clinical pregnancy rates and live birth rates. In addition to the cleavage and blastocyst ET, morula ET may serve as an alternative option for the clinician.

Morula transfer achieves better clinical outcomes than post-thawed cleavage embryos after overnight culture in frozen embryo transfer (FET) cycles.

PURPOSE: This study aimed to investigate the clinical outcomes of morula stage transfer derived from post-thawed cleavage embryos undergoing overnight culture in frozen embryo transfer (FET) cycles. METHODS: We performed a retrospective study that included 392 FET cycles with 784 thawed embryos undergoing overnight culture between January 2014 and December 2018. Embryos were divided into three groups in terms of their status: 8-16 cells without morula (group I), one morula (group II), and two morulae (group III). The clinical outcomes of these cycles were then compared between the three groups. Logistic regression analysis was performed to control for confounders. RESULTS: Group III was associated with a significantly higher clinical pregnancy rate (odds ratio [OR] 2.35; 95% confidence interval [CI] 1.29-4.27; P = 0.005), implantation rate (OR 3.00; CI 1.75-5.16; P < 0.001), multiple pregnancy rate (OR 4.91; CI 2.11-11.40; P < 0.001), and live birth rate (OR 1.96; CI 1.10-3.49; P = 0.022) than group I. Group II had a higher live birth rate than group I after adjustment (OR 1.70; CI 1.04-2.79; P = 0.035). There was no difference in the rate of premature delivery when compared across the three groups after adjustment. CONCLUSION: The transfer of morula stage embryos following the overnight culture of post-thawed cleavage embryos led to an improvement in the clinical outcomes of FET cycles. It is important to reduce the number of morula embryos transferred in order to achieve a singleton pregnancy.

Developmental cell death programs license cytotoxic cells to eliminate histocompatible partners.

In a primitive chordate model of natural chimerism, one chimeric partner is often eliminated in a process of allogeneic resorption. Here, we identify the cellular framework underlying loss of tolerance to one partner within a natural Botryllus schlosseri chimera. We show that the principal cell type mediating chimeric partner elimination is a cytotoxic morula cell (MC). Proinflammatory, developmental cell death programs render MCs cytotoxic and, in collaboration with activated phagocytes, eliminate chimeric partners during the "takeover" phase of blastogenic development. Among these genes, the proinflammatory cytokine IL-17 enhances cytotoxicity in allorecognition assays. Cellular transfer of FACS-purified MCs from allogeneic donors into recipients shows that the resorption response can be adoptively acquired. Transfer of 1 × 10(5) allogeneic MCs eliminated 33 of 78 (42%) recipient primary buds and 20 of 76 (20.5%) adult parental adult organisms (zooids) by 14 d whereas transfer of allogeneic cell populations lacking MCs had only minimal effects on recipient colonies. Furthermore, reactivity of transferred cells coincided with the onset of developmental-regulated cell death programs and disproportionately affected developing tissues within a chimera. Among chimeric partner "losers," severe developmental defects were observed in asexually propagating tissues, reflecting a pathologic switch in gene expression in developmental programs. These studies provide evidence that elimination of one partner in a chimera is an immune cell-based rejection that operates within histocompatible pairs and that maximal allogeneic responses involve the coordination of both phagocytic programs and the "arming" of cytotoxic cells.

Production of chimeras derived from murine embryonic stem cells.

Embryonic stem cells are undifferentiated cells derived from early mouse embryos, which under appropriate culture conditions proliferate continuously in vitro. ES cells have been demonstrated to be pluripotent in vivo from their capacity to form teratocarcinomas and germ-line chimeric mice, dependent on the environment into which the stem cells are introduced. When ES cells are introduced under the kidney capsule, in vivo differentiation is chaotic with the teratocarcinoma composed of a wide variety of different cell types. If, however, the stem cells are returned into a preimplantation mouse embryo, in vivo differentiation proceeds in a normal and organized manner, and the ES cells colonize the three primary cell lineages of the developing embryo: the primitive ectoderm, endoderm, and mesoderm. This leads to the formation of chimeric offspring composed of cells of two different genetic constitutions: the host embryonic cells and those derived from the ES cells. The ES cells are capable of contributing to every tissue in the fetus, including the primordial germ cells. Furthermore, the trophectodermal and primitive endodermal derivatives in the extraembryonic tissues of the conceptus may also be colonized by ES cells. Recent studies have shown that murine ES cells are capable of supporting complete fetal development, following the aggregation of stem cells with tetraploid-cleavage-stage embryos.

Cryopreservation of mouse half-morulae and chimeric embryos by vitrification.

Mouse half-morulae were cryopreserved less than or equal to 1, 3, 6, and 12 hr after bisection by the vitrification method using 25% glycerol and 25% 1,2-propanediol as cryoprotectant. The developmental rates of the frozen-thawed half-embryos to blastocysts in vitro were 77.8% (63/81), 82.0% (41/50), 92.1% (117/127), and 0% (0/37), respectively. Sixty-one of the half-embryos that had been vitrified 6 hr after the bisection followed by transfer to five recipients resulted in a total of ten (16.4%) normal fetuses. Chimeric mouse embryos constructed by two half-morulae were also vitrified 6 and 16 hr after aggregation. Survivors were obtained from the former case: 40 (80.0%) of 50 frozen-thawed embryos developed in vitro to blastocysts, and, after transfer, six chimeric offspring were obtained from the 34 vitrified chimeric embryos. These results showed that mouse half-morulae and chimeric embryos could be cryopreserved by the vitrification method. It seems possible to manufacture a chimeric mouse embryo of defined genotypic composition that can be analyzed during its frozen state using the identical half-embryos of the components.

Cleavage capability of water buffalo follicular oocytes classified by cumulus cells and fertilized in vitro.

Water buffalo (Murrah) oocytes were collected from ovaries obtained from the slaughter house. They were classified according to the character of the cumulus cells under a stereomicroscope, and cultured in 25 mM Hepes buffered Tissue Culture Medium-199 (TCM-199) supplemented with 5% estrous water buffalo serum in an atmosphere containing 5% CO2 in air at 39 degrees C. After 20-24 hr of in vitro maturation, the oocytes were fertilized using capacitated sperm obtained from 4 different bulls. For cleavage the oocytes were cultured at 39 degrees C in TCM-199 supplemented with 1% estrous water buffalo serum and in an atmosphere containing 5% CO2 in air. The good oocytes, with compact and dense cumulus cells cleaved significantly higher (p less than 0.01, 67.3%), than those of fair. partially naked oocytes with thin cumulus layers (27.5%, 25/91) or small remnants of cumulus cells and poor naked oocytes (3/100). A substantial variation cumulus layers (27.5% 25/91) or small remnants of cumulus cells and poor naked oocytes (3/100). A substantial variation in fertilization and developmental rates (16.0% to 43.8%) was observed among 4 different bulls. Late non-surgically into 14 buffalo recipients on day 6 or 7 of their estrous cycle. One recipient was diagnosed to be pregnant by rectal palpation on day 60 and confirmed to be so on day 90 post-estrus.

Sexing of half-embryos produced by microsurgical bisection of mouse morulae and production of chimeric mouse of defined sex composition by aggregating two sexed half-embryos.

Using the halved morulae of mice obtained with microsurgical technique, the following two experiments were performed. 1) Sexing of half-embryos by chromosomal analysis and transfer of the half-embryos after determining the sex of the other monozygotic half. One half of the bisected embryo was cultured in Colcemid solution (0.04 micrograms/ml) to be ensured for chromosomal preparation. More than 50% (152/270) of the blastulated embryos from the halves could be sexed by direct sex chromosome analysis. Thirty-nine of the half-embryos of which the co-twin halves were sexed, were transplanted in to the uterine horns of 18 pseudopregnant mice, and twelve became pregnant. The autopsies of them on Day 18 to 20 of pregnancy, revealed the presence of 16 fetuses. The morphological sex of these fetuses thus obtained coincided completely with the previous judgement based on the chromosomal sexing. 2) Production of chimeras of defined sex composition by aggregating two half-morulae of defined sex. Out of 147 pairs of half-morulae of two different strains (ICR and C3H/He), which were replaced in pairs into empty zona pellucidae, 107 (72.8%) were aggregated successfully and developed in vitro into full expanding blastocysts of typical form. Among the 107 aggregate blastocysts, 31 were sexed for both component embryos by chromosomal analysis on the co-twin half-embryos. When these 31 blastocysts were transferred, 11 living offspring including 4 chimeras were obtained. Transfer of 12 male-male and 5 female-female aggregate blastocysts resulted in 8 males and 1 female, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

The development of sheep embryos when two are transferred into only one uterine horn or into each uterine horn.

In two series of experiments, two D6 morulae were surgically transferred either to each uterine horn of recipient ewes or to the same horn. When embryos were transferred to each side, the percentages of lambs born were not significantly different (68 and 62.5% respectively) whether the corpora lutea of recipient ewes were on each ovary or on the same ovary (in this case, embryos were transferred to the horn ipsilateral to the corpora lutea). When embryos were transferred to the same horn, the percentages of lambs born (71 and 92.6% respectively) were not significantly different whether the corpora lutea were on each side or on the same side. The percentages of lambs born were not significantly different whether the two morulae were transferred to each uterine horn or to the same horn.

Embryo transfer for conserving valuable genetic material from swine herds with pseudorabies.

Embryo transfer was used to conserve genetic material from 2 swine herds seropositive for pseudorabies virus (PRV). Embryos (n = 805) were recovered from 38 PRV-seropositive Duroc sows in Iowa and, after 4 to 10 hours' culture and shipment to Illinois, were transferred to 34 recipients from a herd seronegative for PRV. All recipients remained seronegative for PRV, and 22 of the recipients farrowed 208 pigs (189 alive) that also were seronegative for PRV. There was no evidence of PRV in the embryo recovery medium or in the uterine and oviductal cells recovered with the embryos. Transfer of morulae resulted in higher (P less than 0.02) farrowing rates than did transfer of 4- to 8-cell embryos, but litter size was not affected.

Factors influencing the results of transfers of rabbit embryos stored at -196 degrees C.

Rabbit embryos at the 8-cell and morula stages were frozen and stored at -196 degrees C for 2-200 days. After thawing the embryos were examined for their viability in vitro and in vivo. In vitro, 62.5% of frozen 8-cell embryos and 81.4% of frozen morulae developed to blastocysts. In the control group of unfrozen embryos, 93.2% 8-cell embryos and 92.4% morulae developed to the blastocyst stage. Culture permitted a more reliable elimination of the embryos damaged during freezing and thawing. Embryos were transferred into the reproductive tracts of the recipients either directly after thawing or after 24 h in culture. Synchronous transfers of frozen rabbit embryos were not successful. After asynchronous transfers of morulae and blastocysts into the oviducts, implantation was 31.8% and 42.9%, respectively. After transfer of blastocysts into the uterine horns of the recipients, 47.6% embryos implanted.