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PubMed1991

Mouse model for the treatment of immune pregnancy loss.

Spontaneous abortions can be associated with preimplantation embryo loss, implantation problems and a variety of postimplantation pregnancy failures. The long list of possible causes for the postimplantation pregnancy loss includes, among others, genetic abnormalities in fetus, anatomical abnormalities of the uterus, endocrinological insufficiency, and microbiological problems. However, more than 50% of recurrent miscarriages still have no recognized causes. The concept that many such abortions may be immunologically mediated has gained increasing support over the years. Moreover, immunization of such women with husband's or third party leukocytes has resulted in more than 70% of subsequent pregnancies resulting in live births. Since neither the mechanisms leading to pregnancy loss nor the success of immunotherapy are clear, the set-up of animal models for recurrent abortions would be of supreme significance. Our recent data show that immunopotentiation of maternal immune system by Complete Freund Adjuvant significantly improves pregnancy rate in CBA x DBA/2 mouse combination with high percentage of fetal resorptions. This effect is followed by decrease of IL 2 production in spleen; increase of MAC 1-positive cells at placenta; amplification of suppressive activity of local and systemic lymphocytes and by reverse of embryotoxic effect of maternal serum. Data obtained in this model seems to be valuable in substantiation of rationale for nonspecific immunotherapy of human abortions.

Abortion, Spontaneous↗
PubMed2002

Specificity of E2F1, E2F2, and E2F3 in mediating phenotypes induced by loss of Rb.

The Rb/E2F pathway plays a critical role in the control ofcellular proliferation. Here, we report that E2F1, E2F2, and E2F3 make major individual contributions toward the in vivo phenotypic consequences of Rb deficiency. In the developing lens of Rb(-/-) embryos, loss of E2F1, E2F2, or E2F3 reduces the unscheduled proliferation of fiber cells, with the loss of E2F3 having the most pronounced effect. In Rb-deficient retinas, all three E2Fs contribute equally to the ectopic proliferation of postmitotic neuronal cells. In contrast, E2F1 is unique in mediating apoptosis in both Rb(-/-) lenses and retinas. In the central nervous system, loss of E2F1 or E2F3 can almost completely eliminate the ectopic DNA replication and apoptosis observed in Rb(-/-) embryos, and loss of E2F2 partially reduces the unscheduled DNA replication and has no effect on apoptosis. These results provide clear evidence for functional specificity among E2Fs in the control of Rb-dependent proliferation and apoptosis in a tissue-specific manner.

Animals↗
PubMed1980

[Nature of cytogenetic and morphologic disorders in rat oogenesis and embryogenesis induced by chloridin and 2,4,5-trichlorophenoxyacetic acid during the preovulatory phase of meiosis].

The effect of pyrimethamine (chloridine) and 2,4,5-trichlorophenoxyacetic acid (2,4,5-T) on embryo survival, cleavage rate and frequency of chromosome aberrations when injected into female rats just before ovulation was studied. Pyrimethamine induced 70% of embryonic death at a dose of 5 mg/kg and 27%--at a dose of 2 mg/kg. 200 mg/kg of 2,4,5-T had no embryotoxic effect, while 400 mg/kg increased the embryo loss up to 48%. Pyrimethamine (5 mg/kg) and 2,4,5-T (400 mg/kg), injected intragastrically just before ovulation, significantly affected the cleavage rate in the four day rat embryos. The frequency of chromosome aberrations in ovulated oocytes at metaphase, II subjected to pyrimethamine and 2,4,5-T, reached 6--8% and exceeded the control value in 5--10 times.

2,4,5-Trichlorophenoxyacetic Acid↗
PubMed1982

Early embryonic mortality in women.

Measurements of human chorionic gonadotropin (hCG) have been used to assess early embryo loss in women. Urine samples obtained from a control group of sterilized women with normal ovulatory menstrual cycles enabled a concentration limit of 56 IU/l to be determined so that any nontrophoblastic hCG or other cross-reacting compounds could be accounted for. One hundred ninety-eight ovulatory cycles were collected from a normal population attempting to conceive. Fecundability was 22% to 27% for this population. The risk of pregnancy in exposed ovulatory cycles was 59.6%; however, 61.9% of conceptuses will be lost prior to 12 weeks. Most of these losses (91.7%) occur subclinically, without the knowledge of the mother.

Adult↗
PubMed2004

Mastitis and fertility in cattle - possible involvement of inflammation or immune activation in embryonic mortality.

Causes for pre-implantation embryo loss, which can be as high as 50% or more of fertilized embryos, are multifactorial and largely undescribed. Studies in cattle using mastitis as a model indicate that one cause of early embryonic loss is infectious disease or activation of immune responses at sites outside the reproductive tract. Infection of the mammary gland in dairy cattle is associated with a reduction in pregnancy rate (proportion of inseminated cows that become pregnant) and an increase in the number of inseminations required to establish pregnancy. Also, intravenous challenge with bacterial peptidoglycan and polysaccharide at approximately days 3-5 after breeding reduced subsequent pregnancy rate in sheep that had been previously immunized against the same material. The mechanism by which extrauterine activation of immune and inflammatory responses leads to embryonic loss is not clear although cytokines probably play a crucial role. Effects could be exerted at the level of the hypothalamic-pituitary axis, ovary, reproductive tract or embryo. Interferon (IFN)-alpha, for example, which can reduce pregnancy rate in cattle when injected around 13-19 days after breeding, increases body temperature, inhibits secretion of luteinizing hormone, and reduces circulating concentrations of progesterone. Other cytokines or products of cytokine activation could cause embryonic loss by causing hyperthermia (as elevated temperature blocks oocyte function and embryonic development), exerting toxic effects on the corpus luteum [for example, IFN-gamma, tumor necrosis factor-alpha (TNF-alpha) and prostaglandin F(2alpha)], stimulating endometrial prostaglandin synthesis [TNF-alpha and interleukin(IL)-1beta], reducing endometrial cell proliferation (IL-1beta), and interfering with oocyte maturation and embryonic development (TNF-alpha, nitric oxide, and prostaglandin F(2alpha)). Although largely neglected by reproductive immunologists, study of the involvement of the immune system in pre-implantation embryonic loss is likely to lead to new methods for enhancing fertility.

Animals↗
PubMed1979

A possible influence of the bull on the incidence of embryonic mortality in cattle.

Milk progesterone profiles were used to study the high incidence of embryonic mortality between 30 to 80 days after insemination in 91 cows in a single herd. The progesterone levels dropped to basal values at 46.7 +/- 1.37 days after service in 22 per cent of apparently pregnant cows. Embryo loss occurred significantly more frequently following services by one bull (Z). The mortality rates were 14/32 (44 per cent) in cows conceiving to Z compared with 8/73 (11 per cent) in cows conceiving to other bulls. There was no significant effect of the age of the cow or yield on the incidence of embryonic mortality.

Animals↗
PubMed1990

Effects of a monoclonal antibody against progesterone, on embryo transport, development and implantation in laboratory mice.

A monoclonal antibody against progesterone (11P27) given on Day 2 of pregnancy interrupted pregnancy in BALB/c mice but not in BCF1 mice. The reason for this strain difference remains unclear, although it may involve discrimination by the recipient's immune system. The effects in BALB/c mice were reversed by progestin treatment. A dose of 5 nmol, which completely blocked implantation, had no significant effects on embryo transport and development on Day 4. A dose of 10 nmol did not increase the proportion of abnormal embryos, even though it accelerated tubal transport and increased embryo loss. No tubal retention was evident; most remaining embryos reached the uterus at the normal time. The transformation of morulae to blastocysts was only slightly delayed in 11P27-treated mice, and transfer experiments showed no decrease in embryo viability. The antibody appeared to act by blocking actions of endogenous progesterone on the uterus: uteri of 11P27-treated mice failed to develop a decidual cell reaction to intrauterine oil, and embryos from untreated donors failed to implant in 11P27-treated recipients. Antagonism of progesterone by antibody treatment prevented implantation in BALB/c mice apparently by actions on the uterus rather than on the embryo.

Animals↗
PubMed2004

p66shc, but not p53, is involved in early arrest of in vitro-produced bovine embryos.

High embryo loss occurs in the first week of bovine embryo development, with a high percentage of embryonic arrest. We hypothesized that arrested embryos enter a 'senescence-like state' and that both the cell cycle regulatory protein p53 and the stress-related protein p66(shc), which are involved in the onset of senescence in somatic cells, are responsible for this early embryonic arrest. In our in vitro production system, 13.5 +/- 0.5% of embryos arrest at the 2-4-cell stage. First cleavage occurs between 26 and 48 h post insemination (hpi), with early cleaving embryos showing only 0.6 +/- 0.3% arrest, with later cleaving embryos exhibiting up to 14.2 +/- 0.9% arrest. We compared 2-4-cell embryos collected at 28 hpi with those arrested at the 2-4-cell stage collected at day 8 post insemination. Quantification by real-time PCR and by semi-quantitative immunofluorescence showed significantly higher p66(shc) mRNA and protein levels in both arrested and late cleaving embryos versus 28 hpi embryos. By comparison, no significant changes in p53 mRNA, protein and phosphorylation levels were detected. Taken together, these results demonstrate that embryonic developmental potential is related to the time of first cleavage and that p66(shc), but not p53, is up-regulated in early arrested in vitro-produced bovine embryos.

Adaptor Proteins, Signal Transducing↗
PubMed2006

Embryo survival in dairy cows managed under pastoral conditions.

Efficient pasture-based milk production systems require a compact calving pattern aligned to the onset of the grazing season, a 365-day calving interval and low culling rates for infertility. Achievement of these targets requires high herd reproductive performance. While high genetic merit Holstein cows produce more milk in grass-based systems their fertility is compromised. Management of the modern high genetic merit Holstein dairy cow presents a major challenge in pasture-based systems of production. It appears that the extent of early embryo loss is greater (up to 20% points greater) in the modern high-producing dairy cow and that a much higher proportion of the embryos die before day 7 following insemination in contrast to heifers and lower yielding cows. About 7-8% of pregnancies are lost between days 30 and 90 of gestation with no evidence that loss rate is related to cow genetic merit, parity or level of production. Systemic concentrations of progesterone during both the cycle preceding and following insemination affect embryo survival rate with evidence that too low or indeed too high a concentration of progesterone been negatively associated with embryo survival rate. Peripheral concentrations of both progesterone and oestradiol are lowered by increased plane of feed intake due to increased metabolic clearance rate of the steroids, which is related to liver blood flow. It appears that high producing dairy cows have an increased risk of embryo death as a result of lowered peripheral concentrations of progesterone as a consequence of increased hepatic metabolism of progesterone. Uterine expression of mRNA for progesterone receptor, oestradiol receptor and retinol binding protein mRNA appears to be sensitive to changes in peripheral concentrations of progesterone during the first week after AI. It would appear that energy balance and dry matter intake during the 4 weeks, immediately after calving are critically important in determining conception rate when cows are inseminated at 70-100 days post-calving. Concentrate supplementation of cows at pasture during the breeding period has minimal affects on conception rates though sudden reduction in dietary intake should be avoided. For pasture-based systems of milk production more balanced breeding strategies, with greater emphasis on fertility and feed intake must be developed.

Animal Husbandry↗
PubMed2002

The effects of IgG purified from women with SLE and associated pregnancy loss on rat embryos in culture.

PROBLEM: Recurrent fetal loss occurs in approximately 1% of women. Autoimmune causes have been suggested as a factor in some of these cases. High rates of intrauterine fetal growth retardation and increased incidence of prematurity is associated with systemic lupus erythematosus (SLE) and the anti-phospholipid syndrome (APS). Autoantibodies from sera of SLE/APS patients affect reproductive outcome in pregnant mice, as was studied in vivo, where injection of immunoglobulin (Ig)G purified from patients with APS to mice caused fetal resorptions and growth retardation. METHODS: In order to investigate the direct effect of IgG purified from women with SLE or APS on the growth and viability of embryos, we cultured 11.5-day old-rat embryos in their yolk sacs in the presence of IgG purified from SLE and APS patients. RESULTS: IgG purified from SLE and recurrent pregnancy loss (RPL) patients affected directly the embryo and yolk sac reducing their growth. The purified IgG positive for anti-cardiolipin/anti-DNA antibodies reduced yolk sac and embryonic growth more than sera negative for these antibodies. CONCLUSION: Various antiphospholipid antibodies affect differently the growth and development of the embryo and the placenta.

Animals↗
PubMed2003

Involvement of fatty acid pathways and cortical interaction of the pronuclear complex in Caenorhabditis elegans embryonic polarity.

BACKGROUND: Cell polarity is essential for many decisions made during development. While investigation of polarity-specific factors has yielded great insights into the polarization process, little is known on how these polarity-specific factors link to the basic cellular mechanisms that function in non-polarity aspects of the cell. To better understand the mechanisms that establish embryonic polarity, we investigated genes required for polarity in the one-cell C. elegans embryo that are also required for other non-polarity functions. This has led to the identification of the Pod-class of mutants that are characterized by osmosensitive embryos and defects in anterior-posterior polarity. RESULTS: Mutation in either of two loci of this class, emb-8 and pod-2, disrupts embryonic polarization and results in osmotically-sensitive embryos. Loss of emb-8, a previously uncharacterized polarity gene, causes mislocalization of PAR-3 and PAR-2 that molecularly mark the anterior and posterior cortices. emb-8 encodes NADPH-cytochrome P450 reductase, a protein supplying electrons to cytochrome P450-family enzymes, some of which catalyze fatty acid modifications. Cloning of the previously characterized polarity gene pod-2 reveals it encodes acetyl-CoA carboxylase, an enzyme that catalyzes the first step in de novo fatty acid synthesis. Depletion of fatty acid synthase, the next enzyme in the biosynthetic pathway, by RNA-interference (RNAi) also causes similar loss of one-cell polarity. Furthermore, pod-2 polarity defects can be rescued by addition of exogenous fatty acids. By following the behavior of the pronucleus in emb-8 and pod-2 mutant embryos, we demonstrate that loss of polarity correlates with impaired interaction between the pronucleus-centrosome complex and the posterior cortex. CONCLUSIONS: The characterization of emb-8 and pod-2 mutant embryos suggests that the pronucleus-centrosome complex interaction with the cortex plays a direct role in establishing polarity and that fatty acid pathways are important for this polarizing event.

Acetyl-CoA Carboxylase↗
PubMed1988

Implantation potential of each pre-embryo in multiple pregnancies obtained by in vitro fertilization seems to be different.

Endometrial receptivity and pre-embryo quality are the main factors determining embryo implantation in in vitro fertilization. If one embryo implants normally and goes to term, the endometrium can be assumed to be normal. Eighty-one patients (March 1981 to 1987) had transfers of multiple pre-embryos, resulting in multiple pregnancies with at least one reaching term. The number of gestational sacs at 8 to 12 weeks of pregnancy, at 20 weeks, and the number of term pregnancies were studied longitudinally. Twelve patients (14.8%) had 2 pre-embryos transferred, 19 (23.5%) had 3, 21 (25.9%) had 4, 17 (21.0%) had 5, 9 (11.1%) had 6, and 3 (3.7%) had 7. At 20 weeks 18 patients (22.2%) had a single viable fetus, 56 (69.1%) had twins, 6 (7.4%) had triplets, and 1 (1.2%) had quadruplets. A total of 325 pre-embryos were transferred; 178 gestational sacs were observed at 8 to 12 weeks, but only 153 showed fetal heartbeat. Total embryo loss at 8 to 12 weeks was 53.0%; when the "index" embryo was excluded, it reached 61%. Regardless of the number of pre-embryos transferred, only 2.2 embryos on average were able to establish a normal pregnancy. (A)synchronism of the pre-embryos did not affect outcome. Results suggest that with normal endometrial receptivity, the implantation potential of each pre-embryo is different.

Embryo Transfer↗
PubMed2002

Plasma progesterone profiles and factors affecting embryo-fetal mortality following embryo transfer in dairy cattle.

The relationship between plasma progesterone (P4) levels and embryo survival, and the value of P4 profiles for the selection of cattle embryo transfer recipients is still a matter of controversy. This study reports a comparison between lactating cows and heifers (n = 407) from a single dairy herd, after transfer of either fresh or frozen-thawed good quality embryos, of their ability to sustain embryo-fetal development to term. Plasma P4 concentrations on the day of estrus (Day 0 = D0), Day 4, Day 7 and on Day 21 were measured and related to embryo survival. Plasma P4 levels on Days 0, 4 and 7 were similar in recipients later found pregnant or open. Plasma P4 levels on Day 7 were significantly higher (P < 0.01) in heifers than in cows, but they were similar in pregnant and nonpregnant heifers and in pregnant and nonpregnant cows. Pregnancy rates for fresh and frozen-thawed embryos were higher in heifers than in cows, but the differences did not reach significance. However, the overall late embryonic mortality was significantly higher (P < 0.01) and the calving rate for frozen-thawed embryos was significantly lower (P < 0.05) in cows than in heifers. As expected, plasma P4 on Day 21 was significantly higher (P < 0.001) in pregnant than in nonpregnant recipients, but there was no difference between pregnant cows and pregnant heifers. Plasma P4 levels on Day 7 of recipients presumed pregnant on Day 21 and later found pregnant or nonpregnant were similar, but plasma P4 levels on Day 21 were significantly higher (P < 0.001) in pregnant than in nonpregnant recipients. The results of this study suggest that plasma P4 levels until the day of transfer, except for the rejection of recipients with abnormal luteal function, are of limited practical use for embryo transfer recipient selection. However, in lactating cows low plasma P4 values on Day 7 might negatively affect embryo survival, while in heifers this effect is not noticeable. Lactating cows are more prone to embryo loss than heifers, especially in the case of frozen-thawed embryos; this is associated with a lower competence of the corpus luteum at Day 7.

Animals↗
PubMed2005

[Gamete and embryo protection against oxidative stress during medically assisted reproduction].

So-called oxidative stress is due to the generation of reactive oxygen species (ROS), the most important of which are the superoxide anion (O2-*), hydrogen peroxide, and the hydroxyl radical OH*. ROS can damage gametes and embryos, through DNA (and RNA) fragmentation and membrane lipid peroxidation, potentially resulting in miscarriage. ROS are produced both physiologically (by oxidative metabolism) and in response to environmental factors such as oxygen, light, and traces of divalent cations. The body possesses several defense mechanisms against ROS, including antioxidant enzymes such as superoxide dismutase (SOD), catalase, glutathione peroxidase, and also small molecules like vitamins (A, C and E), glutathione and pyruvate. These different systems generally act in concert. Embryonic cells also possess DNA repair machinery. As oxidative stress is one of the main sources of embryo loss in vitro, it is important to protect both gametes and embryos against these insults during medically assisted reproduction.

Antioxidants↗
PubMed2000

Production of interferon by red deer (Cervus elaphus) conceptuses and the effects of roIFN-tau on the timing of luteolysis and the success of asynchronous embryo transfer.

The role of interferon in early pregnancy in red deer was investigated by (a) measuring production of interferon by the conceptus, (b) testing the anti-luteolytic effect of recombinant interferon-tau in non-pregnant hinds, and (c) treatment of hinds with interferon after asynchronous embryo transfer. Blastocysts were collected from 34 hinds by uterine flushing 14 (n = 2), 16 (n = 2), 18 (n = 8), 20 (n = 13) or 22 (n = 9) days after synchronization of oestrus with progesterone withdrawal. Interferon anti-viral activity was detectable in uterine flushings from day 16 to day 22, and increased with duration of gestation (P < 0.01) and developmental stage (P < 0.01). When interferon-tau was administered daily between day 14 and day 20 to non-pregnant hinds to mimic natural blastocyst production, luteolysis was delayed by a dose of 0.2 mg day(-1) (27.3 +/- 1.3 days after synchronization, n = 4 versus 21 +/- 0 days in control hinds, n = 3; P < 0.05). Interferon-tau was administered to hinds after asynchronous embryo transfer to determine whether it protects the conceptus against early pregnancy loss. Embryos (n = 24) collected on day 6 from naturally mated, superovulated donors (n = 15) were transferred into synchronized recipients on day 10 or day 11. Interferon-tau treatment (0.2 mg daily from day 14 to 20) increased calving rate from 0 to 64% in all recipients (0/11 versus 7/11, P < 0.005), and from 0 to 67% in day 10 recipients (0/8 versus 6/9, P < 0.01). The increased success rate of asynchronous embryo transfer after interferon-tau treatment in cervids may be of benefit where mismatched embryo-maternal signalling leads to failure in the establishment of pregnancy.

Animals↗
PubMed2005

Drosophila WntD is a target and an inhibitor of the Dorsal/Twist/Snail network in the gastrulating embryo.

The maternal Toll signaling pathway sets up a nuclear gradient of the transcription factor Dorsal in the early Drosophila embryo. Dorsal activates twist and snail, and the Dorsal/Twist/Snail network activates and represses other zygotic genes to form the correct expression patterns along the dorsoventral axis. An essential function of this patterning is to promote ventral cell invagination during mesoderm formation, but how the downstream genes regulate ventral invagination is not known. We show here that wntD is a novel member of the Wnt family. The expression of wntD is activated by Dorsal and Twist, but the expression is much reduced in the ventral cells through repression by Snail. Overexpression of WntD in the early embryo inhibits ventral invagination, suggesting that the de-repressed WntD in snail mutant embryos may contribute to inhibiting ventral invagination. The overexpressed WntD inhibits invagination by antagonizing Dorsal nuclear localization, as well as twist and snail expression. Consistent with the early expression of WntD at the poles in wild-type embryos, loss of WntD leads to posterior expansion of nuclear Dorsal and snail expression, demonstrating that physiological levels of WntD can also attenuate Dorsal nuclear localization. We also show that the de-repressed WntD in snail mutant embryos contributes to the premature loss of snail expression, probably by inhibiting Dorsal. Thus, these results together demonstrate that WntD is regulated by the Dorsal/Twist/Snail network, and is an inhibitor of Dorsal nuclear localization and function.

Amino Acid Sequence↗
PubMed2002

A transgenic Lef1/beta-catenin-dependent reporter is expressed in spatially restricted domains throughout zebrafish development.

The Wnt/beta-catenin signaling pathway plays multiple roles during embryonic development, only a few of which have been extensively characterized. Although domains of Wnt expression have been identified throughout embryogenesis, anatomical and molecular characterization of responding cells has been mostly unexplored. We have generated a transgenic zebrafish line that expresses a destabilized green fluorescent protein (GFP) variant under the control of a beta-catenin responsive promoter. Early zygotic expression of this transgene (TOPdGFP) mirrors known domains of Wnt signaling in the embryo. Loss of Lef1 activity results in decreased reporter expression and posterior defects, while loss of Tcf3 (Headless, Hdl) activity does not alter reporter expression, even though it results in loss of forebrain structures. In addition, ectopic Wnt1 expression can activate the reporter. In older embryos, we identify a number of transgene-expressing cell populations as novel sites of beta-catenin signaling. We conclude that our TOP-dGFP reporter line faithfully illustrates domains of beta-catenin activity and enables the identification of responsive cell populations.

Animals↗
PubMed2002

Donor oocyte cytoplasmic transfer did not enhance implantation of embryos of women with poor ovarian reserve.

PURPOSE: To determine whether donor oocyte cytoplasm transferred into the oocytes of women < or = 40 years or with diminished ovarian reserve would enhance embryo quality, implantation, or pregnancy rates. METHODS: Study subjects included women > or = 40 years (15) or with abnormal FSH levels (3). Healthy volunteers (18) produced oocytes for cryopreservation. Donor oocytes were thawed and cytoplasm from surviving oocytes was injected with a single sperm into the cytoplasm of recipient oocytes. Outcome measures included embryo quality scores, implantation, and pregnancy rates. RESULTS: Eighteen donors produced 213 oocytes for cryopreservation and 39/171 (22.8%) survived thawing. Eighteen recipients initiated 25 IVF cycles with embryo transfer in 20 cycles after cytoplasmic transfer (CT). Four cycles resulted in three biochemical losses and one aneuploid clinical loss. Embryo quality did not improve with CT compared to pre-CT IVF cycles in six recipients. CONCLUSIONS: CT with cryopreserved donor oocyte cytoplasm did not enhance success in women with advanced reproductive age or low ovarian reserve.

Adult↗