Analysis of litter size and weight in mice differing in Ped gene phenotype and the Q region of the H-2 complex

A gene has been described, Ped (Preimplantation embryo development), that influences the rate of cleavage of preimplantation mouse embryos. Previous studies of linkage of Ped gene phenotype (fast or slow embryo development) and H-2 haplotype (H-2b or H-2k) in backcross embryos from the C57BL/10Sn and B10.BR congenic strains have shown that the Ped gene is located in the H-2 complex, the major histocompatibility complex (MHC) of the mouse. The present study was undertaken to localize the Ped gene to a particular subregion of the H-2 complex. Analysis of the B6.K1 and B6.K2 congenic strains, which differ at only the Q subregion of the MHC, was undertaken to test whether the Ped gene is located in the Q subregion of the MHC. Qa-2 antigen expression was used as a marker for the Q subregion and fast or slow development was used to assess Ped gene phenotype in backcross embryos generated from the mating of (B6.K1 x B6.K2)F1 and B6.K1 mice. The results showed linkage of Ped gene phenotype and Qa-2 antigen expression, which strongly supports the idea that the Ped gene is located in the Q subregion of the MHC. In a further set of experiments, litter size and weight were investigated in the B6.K1 and B6.K2 mice. Pure line and reciprocal crosses were made using sires and dams of both the B6.K1 and B6.K2 genotypes. Traits measured on pups included birth weight, weaning weight and weight per day of age from birth to weaning. Litter traits measured were number born and weaned and survivability. Sex effects existed for weaning weight and weight gain per day of age. Males gained more and were heavier than female pups (P less than 0.05). Pups whose sire or dam were B6.K2 were significantly (P less than 0.05) heavier at birth than those pups whose sire or dam were B6.K1, and B6.K2 homozygous pups were significantly (P less than 0.001) heavier than all others. Litters whose dams were B6.K2 had significantly more pups at birth (P less than 0.05) than those litters whose dams were B6.K1. Results suggest that pups that are heterozygous or homozygous B6.K2 are more apt to be heavier at birth and be in larger litters suggesting that genes in the Q region of the H-2 complex are advantageous to reproductive performance.     

Influence of the Preimplantation Embryo Development (Ped) Gene on Embryonic Platelet-Activating Factor (PAF) Levels

Purpose : A major gene responsible for the control of preimplantation cleavage rate is the Ped gene, the product of which is the Qa-2 protein. Fast, but not slow developing mouse embryos express the Qa-2 protein. Platelet-activating factor (PAF) is a novel and potent signaling phospholipid that has unique pleiotropic properties in addition to platelet activation. PAF plays a significant role in virtually every reproductive event, including ovulation, fertilization, implantation, and parturition. The role of the Ped gene in PAF production by preimplantation embryos is yet to be established. The presence of this gene provides embryos with a reproductive advantage over those that are Ped negative, and may also serve as a regulator of PAF synthesis. The study hypothesis is that the amount of PAF produced is dependent upon the presence or absence of the Ped gene. Methods : B6.K1 (Ped negative) and B6.K2 (Ped positive) mouse embryo-conditioned culture media were assayed for PAF content by a PAF-specific radioimmunoassay. Results : There was a significant (p < 0.001) difference in blastocyst development rates between the Ped+ B6.K2 (61.0%) and the Ped− B6.K1 (25.3%) embryo culture groups. There was a significant difference (p < 0.05) in PAF production between the Ped+ B6.K2 (4.70±0.46 pmol per embryo) embryo culture group and the Ped− B6.K1 (10.02±3.49 pmol per embryo) embryo group. The B6.K1 (Ped−) embryo group produced >2× more PAF than did the B6.K2 (Ped+) group. Conclusions : The Ped gene plays a role in PAF production and release in preimplantation stage embryos. The use of two mouse identical strains, except for the Ped gene, show that its presence is associated with an increase in developmental potential. Embryos where the Ped gene was absent produced significantly higher levels of PAF, which may aid in their survival.     

Preimplantation genetic diagnosis

Preimplantation genetic diagnosis is essentially an alternative to prenatal diagnosis, in which genetic testing is performed on embryos before a clinical pregnancy is established. Preimplantation genetic diagnosis has been applied to patients carrying chromosomal rearrangements, such as translocations, in which it has been proven to decrease the number of spontaneous abortions and prevent the birth of children affected with chromosome imbalance. Preimplantation genetic diagnosis techniques have also been applied to increase implantation rates, reduce the incidence of spontaneous abortion and prevent trisomic offspring in women of advanced maternal age undergoing fertility treatment. A third group of patients receiving preimplantation genetic diagnosis are those at risk of transmitting a single gene disorder to their children. The number of monogenic disorders that have been diagnosed in preimplantation embryos has increased each year. Recent protocols have tended to be more complex and more reliable than previous methods, making greater use of multiplex polymerase chain reaction. As well as an expansion in the variety of disorders for which preimplantation genetic diagnosis is offered, new indications have been reported including the use of human leukocyte antigen histocompatibility typing and the application of preimplantation genetic diagnosis to late onset diseases.     

Selection for faster development does not bias sex ratios resulting from blastocyst embryo transfer

Concerns of possible male biased sex ratios with IVF exist due to reports of faster preimplantation development for male relative to female embryos in a variety of mammals, including humans. This study reports the first direct test of the hypothesis that selection for faster preimplantation development to the blastocyst stage increases the likelihood of a male birth. Gender outcomes of all live births resulting from blastocyst stage embryo transfer within a large assisted reproduction practice were reviewed, along with associated embryological records. The developmental stage of transferred embryos relative to the average developmental stage of all viable embryos in each cohort was related to gender outcomes by logistic regression analysis. Gender data were available for 1,184 children born from 815 pregnancies resulting from blastocyst stage embryo transfer during the study period. Data on preimplantation embryonic development were available for 737 cycles resulting in the birth of 1,075 children. There was no relationship between selection for developmental rate and the gender of resulting children. Numbers of male and female children were nearly identical within this patient population (593 male versus 591 female). These results indicate that offspring gender is unrelated to selection for developmental rate among blastocyst transfer cycles.     

Seven years of experience of preimplantation HLA typing: a clinical overview of 327 cycles

Preimplantation human leukocyte antigen (HLA) typing allows the birth of healthy children who are potential donors of stem cells for their affected siblings. This technique can be used for acquired diseases such as leukaemia or can be used for single-gene disorders such as thalassaemia. This retrospective study presents clinical data obtained from 171 couples who had undergone 327 preimplantation HLA typing cycles: 262 cycles for HLA typing in combination with mutation analysis and 65 cycles for the sole purpose of HLA typing. Of the diagnosed embryos 17.6% were found to be HLA matched. Embryo transfer was performed in 212 cycles, 34.9% clinical pregnancy rate per transfer was achieved and 59 healthy and HLA-compatible children were born. Twenty-one sick children have been cured through haemopoietic stem cell transplantation. The effect of maternal age and ovarian reserve on reproductive outcome was assessed retrospectively. The data demonstrated that, once a mutation-free and HLA-compatible embryo was found, clinical pregnancy rate did not differ statistically significantly despite the presence of some cycle-related limitations such as advanced maternal age and/or diminished ovarian reserve. Preimplantation HLA typing is an effective therapeutic tool for curing an affected sibling even for poor-prognosis patients.Preimplantation human leukocyte antigent (HLA) typing allows the birth of healthy children who are potential donors of stem cells for their affected siblings. This technique can be used for acquired diseases such as leukaemia or can be used for single-gene disorders such as thalassaemia. This study presents clinical data obtained from 171 couples who underwent 327 preimplantation HLA-typing cycles. Of these, 262 cycles were performed for HLA typing in combination with mutation analysis and 65 cycles were performed for the sole purpose of HLA typing. A total of 17.6% of the diagnosed embryos were found to be HLA matched. Embryo transfer was performed in 212 cycles. The clinical pregnancy rate per transfer was 34.9% and 59 healthy and HLA compatible children were born. Twenty-one sick children have been cured through haemopoietic stem cell transplantation. The effect of maternal age and ovarian reserve on reproductive outcome was assessed retrospectively. The data demonstrated that, once a mutation-free and HLA-compatible embryo was found, clinical pregnancy rates did not differ statistically significantly by the presence of some cycle-related limitations such as advanced maternal age and/or diminished ovarian reserve. Preimplantation HLA typing is an effective therapeutic tool for curing an affected sibling even for poor-prognosis patients.     

Inhibition of preimplantation mouse embryo development by isoflurane.

OBJECTIVE: Preimplantation mouse embryos were exposed to a commonly used inhalational anesthetic agent, isoflurane, to determine its effects on embryo development. STUDY DESIGN: Two-cell embryos were exposed at various intervals (5 to 6 hours, 3 to 4 hours, and 0 to 1 hour) before the onset of their first cleavage in vitro. In addition, the effects of 5% isoflurane on four-cell embryos exposed about 2 hours after the first cleavage and morula stage embryos also were examined. RESULTS: Development to the blastocyst stage was inhibited by 3% and 5% isoflurane (p less than 0.005) but not by 1.5% isoflurane when two-cell embryos were exposed 3 to 4 hours or 0 to 1 hour before the onset of cleavage. Most of the affected embryos completed cell division and came to a halt at the three- to four-cell stage. The development of embryos exposed to isoflurane at the four-cell or morula stage was unaffected. CONCLUSIONS: Isoflurane adversely affects subsequent preimplantation development when two-cell mouse embryos are exposed just before the onset of their first cleavage in vitro.     

Effects of platelet activating factor on mouse embryo implantation in vitro

Purpose Our purpose was to assess the role of platelet activating factor (PAF) in embryo implantation, we examined the effects of PAF and a PAF antagonist on the in vitro implantation of mouse embryos, using an in vitro embryo culture system in the absence of the endometrium.Methods BDF1 mouse pronuclear-stage embryos were cultured until they developed to the two-cell, the four- to eight-cell, or the morula stage in the absence of PAF or its antagonist CV6209. The medium was then changed and supplemented with PAF or CV6209 at various concentrations. We also examined the reversible effects of PAF addition to the media containing the PAF antagonist.Results The addition of PAF to the culture from the two-cell stage significantly (P <0.05) increased the rates of embryo implantation in vitro (control, 69.8%; 10 ^–10 MPAF, 90.1%; 10 ^–9 MPAF, 95.5%). Similarly, the addition of PAF to the cultures from the four- to eight-cell and morula stage also significantly (P <0.05) increased their rates of implantation in vitro. In contrast, the addition of CV6209 to the culture significantly (P <0.01) decreased the rates of embryo implantation in vitro. CV6209 also decreased the rate of blastocyst formation. The degree of inhibition by CV6209 decreased with the advancing stage of embryos. The addition of PAF to media containing CV6209 reversed the inhibition and restored the implantation rate in vitro.Conclusions These results suggest that PAF may act directly on the mouse embryo and favor its implantation like an autocrine activating factor, irrespective of the presence or absence of the endometrium.     

Birth of a healthy infant after preimplantation genetic diagnosis by sequential blastomere and trophectoderm biopsy for β-thalassemia and HLA genotyping

Background

Preimplantation genetic diagnosis (PGD) is a widely used technique for couples at genetic risk and involves the diagnosis and transfer of unaffected embryos generated through in vitro fertilization (IVF) techniques.

Study design

For those couples who are at risk of transmitting a genetic disease to their offspring, preimplantation embryos can be selected according to their genetic status as well as human leukocyte antigen (HLA) compatibility with the affected child. Stem cells from the resulting baby's umbilical cord blood can be used for transplantation to the affected sibling without graft rejection.

Results

Here we report successful hematopoietic stem cell transplantation (HSCT) after the birth of a healthy infant, who was born after successful PGD testing with both cleavage stage and blastocyst stage biopsy for the purpose of diagnosis of β-thalassemia and HLA compatibility.

Conclusion

The specific feature of this work is not only to have the first successful HSCT achieved in Bulgaria after using preimplantation HLA typing technique, it also demonstrates how to accomplish this success via cross-border collaboration of different units, which makes the application of these sophisticated methods possible in hospitals not having the necessary equipments and expertise.     

Preimplantation HLA typing with aneuploidy testing

Preimplantation HLA typing has been introduced for the treatment of affected siblings, requiring HLA-identical stem cell transplantation. This was applied either in combination with preimplantation genetic diagnosis (PGD) to ensure that the preselected HLA-matched embryos were also free of the genetic disorder, or without PGD, with the only purpose of selecting and transferring the HLA-matched embryos. Because patients requesting preimplantation HLA typing are usually of advanced reproductive age, aneuploidy testing allows not only the avoidance of the birth of children with chromosomal disorders, but also improvement of the reproductive outcome, which is still not sufficiently high in preimplantation HLA typing at the present time. This study presents the results of the first experience of preimplantation HLA typing combined with aneuploidy testing, demonstrating feasibility and impact of aneuploidy testing on the accuracy and outcome of preimplantation HLA typing. Of a total of 138 cycles performed, 87 were combined with PGD and 52 without testing for the causative gene, of which aneuploidy testing was performed in 27 cycles, allowing the preselection and transfer of only those HLA-matched embryos that were also euploid. Although the euploid HLA-identical embryos were available for transfer in only half of these cycles, pregnancy and birth of unaffected HLA-identical children were observed in approximately half of these cycles, suggesting the potential usefulness of incorporating aneuploidy testing into preimplantation HLA typing.     

Effect of chromosomal translocations on the development of preimplantation human embryos in vitro

OBJECTIVE: To determine the reliability of a new technique for single human blastomere karyotyping during clinical cases for preimplantation genetic diagnosis of translocations. DESIGN: Controlled clinical study. SETTING: Preimplantation genetic diagnosis and IVF program. PATIENT(S): Nineteen preimplantation genetic diagnosis cases with 11 types of translocations (10 reciprocal and one Robertsonian) involving chromosomes 1, 5, 7, 8, 9, 11, 12, 13, 14, 15, 16, 18, 20, 21, and 22. INTERVENTION(S): Blastomere biopsy followed by blastomere nucleus conversion into metaphase chromosomes. Fluorescent in situ hybridization (whole chromosome painting) was used for the detection of chromosomally unbalanced preimplantation human embryos.Main Outcome Measure(s): Percentage of informative metaphase plates and effect of unbalanced translocations on preimplantation embryo development. RESULT(S): Informative metaphases were obtained for 84% of the blastomeres. Analysis of preimplantation development of the resulting embryos showed that an unbalanced chromosomal complement does not affect embryo ability to reach the blastocyst stage in vitro. CONCLUSION(S): For the translocations tested, there is no evident selection against chromosomally unbalanced embryos at the preimplantation stage of embryo development.     

Twin birth after preimplantation diagnosis for cystic fibrosis]

OBJECTIVE: Cystic fibrosis is a common autosomal recessive disease most often caused by a deletion (delta F508) in the CFTR gene. It is the most common indication for preimplantaion genetic diagnosis which allows genetic analysis of embryos obtained after in vitro fertilization and transfer of unaffected embryos into the patient's uterus. PATIENTS AND METHODS: We report the first preimplantation genetic diagnosis performed in Strasbourg for a couple at risk of having a child affected by severe cystic fibrosis due to a homozygous delta F508 mutation. Three days after fertilisation, embryos obtained after intra-cytoplasmic testiculare sperm injection were biopsied and analysed. PCR amplification of the genomic fragment containing the delta F508 locus allowed detection of the delta F508 mutation and transfer only of the unaffected embryos. RESULTS: Three embryos were transferred after this preimplantation genetic diagnosis. A twin pregnancy was obtained and the babies born from this cycle are both exempt from the mutation. CONCLUSIONS: Preimplantation genetic diagnosis for the cystic fibrosis delta F508 mutation is now available in our centre. In this report, we could resolve both the problem of infertility and the risk of transmission of a severe form of cystic fibrosis. Preimplantation genetic diagnosis is also available for other mutations involved in cystic fibrosis and also for other genetic diseases.     

The role of tapasin in MHC class I protein trafficking in embryos and T cells

Preimplantation mouse embryos express both classical (class Ia) and nonclassical (class Ib) MHC class I proteins, and yet are not rejected by the maternal immune system. Although the function of the embryonic MHC class Ia proteins is unknown, one MHC class Ib protein, Qa-2, the product of the preimplantation embryo development (Ped) gene, actually enhances reproductive success. Similar in structure to MHC class Ia proteins, Qa-2 protein is a trimer of the alpha (heavy) chain, β₂ microglobulin and a bound peptide. Studies on the folding, assembly and trafficking of MHC class Ia molecules to the cell surface have revealed this process to be dependent on multiple protein chaperone molecules, but information on the role of chaperone molecules in Qa-2 expression is incomplete. Here, we report the detection of mRNA for four chaperone molecules (TAP1, TAP2, calnexin and tapasin) in preimplantation embryos. We then focused on the role of the MHC-dedicated chaperone, tapasin, on Qa-2 protein expression. First, we demonstrated that tapasin protein is expressed by preimplantation embryos. Then, we used tapasin knockout mice to evaluate the role of tapasin in Qa-2 protein expression on both T cells and preimplantation embryos. We report here that optimal cell surface expression of Qa-2 is dependent on tapasin in both T cells and preimplantation embryos. Identification of the molecules involved in regulation of MHC class I protein expression in early embryos is an important first step in gaining insight into mechanisms of escape of embryos from destruction by the maternal immune system.     

Effects of Preconceptual Caffeine Exposure on Pregnancy and Progeny Viability

Objective: A previous study demonstrated for the first time that a drug such as caffeine, administered prior to ovulation and genomic activation, causes a quantitative difference in growth-promoting energy utilization in a proportion of 5-day-old blastocysts. The objective of the present study was to investigate whether developmental changes induced by caffeine administered throughout the estrus cycle prior to fertilization are sustained throughout pregnancy and after birth.

Methods: Caffeine was administered to rats throughout the estrus cycle prior to fertilization, with control and experimental groups subdivided into preimplantation and postimplantation categories. Preimplantation fertilization rate was assessed on day 4 of pregnancy by a pregnancy-induced elevation in maternal plasma progesterone concentration, or by flushing each uterine horn on day 5 of pregnancy to determine the presence or absence of a litter. Postimplantation fetuses were collected on gestational day 12 or allowed to go to term.

Results: Preconceptual caffeine exposure significantly reduced maternal fertility by the failure of a proportion of the litters to implant, rather than curtailing preimplantation development or postimplantation losses. Postnatal mortality between weeks 0 and 1 was elevated and the weekly incremental growth rate of the pups from week 3 through week 7 was significantly reduced in the preconceptually caffeine-treated offspring. Experimental females reached puberty at the same age as the controls but at a significantly lower body weight. Gestation length, birthweight, litter size, sex ratio, and anogenital distance (a measure of prenatal androgenization) were not affected by preconceptual caffeine treatment.

Conclusions: It was concluded that the reduced fertility rate in preconceptually caffeine-exposed rats was due to the failure of litters to implant rather than to a reduced fertilization rate, which was normal. It was further concluded that the growth rate over the neonatal and prepubertal periods of surviving pups in the caffeine-treated group was subnormal.     

Insulin-like growth factors (IGFs), IGF-binding proteins (IGFBPs), and IGFBP-3 protease activity in the peritoneal fluid of patients with and without endometriosis

OBJECTIVE: To evaluate embryotropic action of hemoglobin (Hb) and ethylenediaminetetraacetic acid (EDTA) on preimplantation embryo development. DESIGN: In vitro model study using mouse embryos. SETTING: University affiliated hospital, Pochon CHA University. ANIMALS: Four-week-old block strain ICR mice naturally mated after superovulation. INTERVENTION(S): One-cell embryos were cultured in serum-free, modified preimplantation-1 medium, to which 1 microg/ml Hb and/or 0.1 mM EDTA were added. MAIN OUTCOME MEASURE(S): Preimplantation development and blastomere number. RESULT(S): More (P<.05) 1-cell embryos developed to the 4-cell (52% vs. 67%-84%), 8-cell (48% vs. 65%-81%), and blastocyst (40% vs. 61%-79%) stages after the addition of hemoglobin (Hb) and/or EDTA than after no addition. Highest proportion of embryos developed to each stage after the combined addition of Hb+EDTA. EDTA specifically stimulated the development before the 8-cell stage, which was as similar as Hb+EDTA. On the contrary, higher ratio of morula to blastocyst transformation was obtained after the addition of Hb or Hb+EDTA than after no addition (0.76 vs. 0.96-0.98). Significant increases in the cell number of blastocysts (46.5-47.2 vs. 53.2 cells), inner cell mass (ICM) cells (16.7-17.5 vs. 21 cells), and the ratio of ICM cells to trophoblasts (0.3-0.37 to 0.39) were found after the combined addition of Hb+EDTA, compared with no addition or with the addition of EDTA or Hb alone. CONCLUSIONS: Hb and EDTA have stage-specific effects on supporting preimplantation embryo development; Hb promotes both the development before the 8-cell stage and the morula to blastocyst transformation, whereas EDTA mainly promotes the development to the 8-cell stage. The combined exposure of embryos to Hb and EDTA improves not only preimplantation development but also the growth and quality of blastocysts.     

The role for preimplantation genetic diagnosis in balanced translocation carriers

OBJECTIVE: Preimplantation genetic diagnosis is an established technique that provides an alternative to prenatal diagnosis for patients who are at risk of transmitting a serious genetic disorder to their offspring. Preimplantation genetic diagnosis has been used for couples who have been at risk for having offspring with single gene or X-linked disorders and for screening for common age-related aneuploidy and in couples who themselves carry balanced chromosomal rearrangements. The aim of this study was to summarize our experience using preimplantation genetic diagnosis after the identification of a parental balanced translocation, specifically as it relates to the number of embryos that are suitable for transfer after preimplantation genetic diagnosis for a known translocation and aneuploidy screening. STUDY DESIGN: This is a retrospective review of data from a single center that involved 6 couples that initiated the process of preimplantation genetic diagnosis for translocation and aneuploidy screening by fluorescent in situ hybridization. RESULTS: A total of 65 embryos were obtained, of which 56 embryos (86%) were suitable for fluorescent in situ hybridization analysis. After fluorescent in situ hybridization, 1 embryo was diagnosed as normal or balanced (1.7%). Forty-three embryos (76.8%) were unbalanced for the translocation; 8 embryos (14.3%) were aneuploid, and 4 embryos (7.1%) were uninformative. There were no clinical pregnancies. CONCLUSION: In our experience, there are very few embryos that are available for transfer from these patients after translocation and aneuploidy screening because of multiple unbalanced segregation products and a high rate of aneuploidy. Factors that contributed to this may be related to which parent carries the translocation, methods that were used for in vitro fertilization, and advanced maternal age. Although preimplantation genetic diagnosis for translocation carriers theoretically can enhance the pregnancy rate for a couple, there are limitations. This information should be shared with couples who are contemplating preimplantation genetic diagnosis for translocation, and the options of sperm or egg donor should be considered.     

Präimplantationsdiagnostik in Deutschland nach dem Urteil des Bundesgerichtshofes 2010

For couples with severe genetically determined diseases preimplantation genetic diagnosis offers the option to diagnose affected embryos before pregnancy has been established and thus to exclude such embryos from transfer to the womb. In this way a possibly stressful termination of pregnancy can be avoided. This article gives a review of the state of affairs regarding international preimplantation genetic diagnosis usage with commonly occurring indication as well as pregnancy and birth rates. It can be seen that the indication for preimplantation genetic diagnosis (PGD) is not liberally applied. Furthermore, the legal situation in Germany will be explained. After confrontational opinions with respect to the permissibility of PID collided over many years and physicians did not use PGD because they were afraid of prosecution, the judgment of the Federal High Court has now provided security for physicians and affected couples. PID has become possible in Germany! Preimplantation genetic aneuploidy screening (PGS) for diagnosis of chromosome abnormalities cannot be used for improvement of the pregnancy rate and reduction of the abortion rate in older patients. Therefore, PGS on embryos or oocytes (polar body analysis) should not be offered outside clinical studies.     

Birth weight and sex ratio after transfer at the blastocyst stage in humans.

OBJECTIVE: To analyze the birth weights and sex ratio of infants born after blastocyst transfer. DESIGN: Retrospective analysis. SETTING: Three infertility clinics. PATIENT(S): Patients admitted for IVF with blastocyst transfer. INTERVENTION: None. MAIN OUTCOME MEASURE(S): Birth weights and sex ratio of infants born after blastocyst transfer. RESULT(S): Statistically significantly more male infants were born after transfer of fresh blastocysts, either cocultured or cultured in sequential media. No specific differences in birth weight were observed between infants born after blastocyst transfer and those born after spontaneous conception. CONCLUSION(S): More male infants than female infants were born after blastocyst transfer when transfers were performed as soon as the blastocyst stage was reached and male embryos had a faster cleavage rate.     

Effect of glutamine on preimplantation mouse embryo development in vitro

OBJECTIVES: The purpose of this research was to study the independent effect of the amino acid glutamine on preimplantation mouse embryo development in vitro. STUDY DESIGN: Two-cell stage mouse embryos were cultured in human tubal fluid medium in the presence and absence of 1 mmol/L of glutamine. Outcomes for morphology and cleavage rates were compared with Fisher's and Mann-Whitney's tests, respectively. RESULTS: Glutamine increased the proportion of embryos that developed to the blastocyst stage (86.4%) compared with those cultured without glutamine (59.1%) (P =.052). The percentages of embryos developing to the morula or hatching blastocyst stages were comparable in the 2 groups. Blastocyst total cell numbers were significantly higher in the glutamine group (34+/-1.7 vs 18.5+/-3.5, respectively; values are mean+/-SEM, P =.044). CONCLUSION: The amino acid glutamine independently improves preimplantation mouse embryo development in vitro. Further studies are needed to examine the applicability of these results to humans.