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Embryo loss, blastomere development and chromosome constitution after human chorionic gonadotropin-induced ovulation in mice and rats with regular cycles.

A dose of 7 IU human chorionic gonadotropin (hCG) given 14 h before the expected LH peak on proestrus significantly increased embryonic mortality in Swiss random-bred female mice to 55% of the number of corpora lutea. The use of luteinizing hormone-releasing hormone in a similar injection protocol did not induce embryonic death. The effect found in Swiss random-bred mice resembles that of a dose of 20 IU hCG in the rat. Afternoon-day-4 mouse embryos contained 39.1 +/- 12.6 nuclei after hCG-induced ovulation compared to 46.2 +/- 16.6 nuclei after spontaneous ovulation. For early-day-5 embryos of the rat, these figures were 34.2 +/- 10.1 and 31.7 +/- 8.4, respectively (mating was early on day 1). Numerical chromosome errors were estimated in secondary oocytes of the mouse and early-day-5 embryos of the rat. Compared with data from the literature, hCG seems to induce some extra meiotic nondisjunction in the rat only. Combining all genetic and physiological data, the loss of fecundity after hCG-induced ovulation is a maternal effect.

Animals

Pre- and post-implantation losses of embryos in aging female IVCS mice.

Mated female mice of the IVCS strain, aged 90 (control group), 180, 210, 240, 270, 300, 330, 360 and 420 days old, were studied for pre- and post-implantation loss of embryos. The percentage of pre-implantation loss in mice aged 90 to 210 days was 1.7/11.8 (14.4%) to 2.7/11.7 (23.1%). In mice aged 240 to 300 days it increased significantly as compared to the controls (46.5-90.2% versus 14.4%). It reached 100% in 300-days-old mice. The post-implantation loss of embryos and/or fetuses in mice aged 90 to 240 days was 1.0/10.2 (9.8%) to 2.5/9.0 (27.8%). In mice aged 270 to 300 days it increased significantly compared to the controls (100% versus 9.8%). The decrease in reproductive activity appeared first in a decrease in litter size, followed by a decrease in the number of blastocyst and implantation sites, and finally by anovulation during the process of aging in IVCS mice.

Aging

Early pregnancy factor (EPF) as a marker for the diagnosis of subclinical embryonic loss.

The validation of EPF as a possible correlate of early fertilization has made it possible to study and detect fertilization of the ovum in normal fertile women (during the luteal phase) and also in women with infertility, where the fertilization of the ovum may not be affected but there may be impairment in early embryonic development which results in early embryo loss or subclinical embryo loss. Our results have suggested that using EPF as a marker, we could detect subclinical embryonic loss in 57.8% of the infertile women where more than one menstrual cycle was studied and the blood was collected 4-7 days after ovulation. After the missed period, 80% of the patients who were negative for EPF but positive for hCG had spontaneous abortions. It would be interesting to study how EPF behaves as a marker, to detect subclinical embryonic loss in diverse pathological situations such as recurrent abortions, parental age and translocation carrier parents.

Adult

Effect of early ovariectomy and steroid hormone replacement of embryo transport, development and implantation in mice.

Bilateral ovariectomy on Day 1 of pregnancy increased abnormal embryo numbers on Day 4 and delayed passage of embryos to the uterus. Progestins given on Day 1 reversed these effects; given on Day 3 they reduced numbers of abnormal embryos, but did not restore normal transport. Oestrogen given alone after ovariectomy increased embryo loss, but restored preimplantation embryo development to normal when given on Day 3 after progestins on Day 1. The results suggested that both oestrogen and progesterone were necessary for normal preimplantation embryo development in vivo. However, although Day-1 progestins produced the greatest improvement in embryo transport and preimplantation development, they supported only low implantation rates compared with progestins starting on Day 3, and no progestin treatment returned implantation rates to normal. Sham ovariectomy on Day 1 also reduced implantation rate, suggesting that surgical stress of Day-1 ovariectomy had major adverse effects on embryo viability. This view was supported by experiments involving unilateral ovariectomy, which produced abnormalities in embryo transport, development and implantation, but only on the operated side. Furthermore, the major abnormality induced in embryo development by unilateral and bilateral ovariectomy, viz embryonic autolysis, was not increased in experiments in which pregnancy was blocked by non-surgical antagonism of progesterone. It is concluded that abnormalities in embryo development induced by early ovariectomy are not caused by a deficit of endogenous hormones, but result largely from effects of surgical trauma on oviduct function which can be reversed by treatment with exogenous hormones.

Animals

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

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

Manipulation of reproduction in sheep.

Active immunization of cyclic ewes against androstenedione (using melatonin, Fecundin) has been shown to increase reproductive output by about 30 extra lambs per 100 ewes put to the ram in UK sheep flocks. However, this technique has not been widely adopted within the industry. One possible reason for this is that it results in an unacceptable incidence of triplet births, even though the litter size distribution is similar to that obtained for untreated flocks with the same overall mean lambing percentage. The use of progestagen sponges plus pregnant mares' serum gonadotrophin (PMSG) for the induction of out-of-season breeding in ewes is associated with a number of recognized shortcomings. A possible reason for the variability in conception rates is the high incidence of complete embryo loss or fertilization failure in ewes that are induced to superovulate after treatment with the doses of PMSG required to promote a high oestrous response. Recent studies have indicated that slow release implants of melatonin (Regulin) can advance the onset of the breeding season of commercial sheep flocks, but only by about 4 weeks. This means that the optimum treatment window varies according to the natural breeding season characteristics for the breed. Nevertheless, this treatment appears to overcome a number of the shortcomings of progestagen sponges plus PMSG. It also produces a modest increase in litter size per ewe pregnant (about 15 extra lambs per 100 ewes) without the wide variation in birth types and birth weights associated with PMSG. The development of a laparoscopic procedure that improves conception rates after intrauterine insemination of frozen semen allows comparisons of genetic merit across flocks. This should promote far greater rates of genetic improvement for traits of high economic importance than those achieved by within-flock selection and in the last two years has allowed the development of sire-referencing schemes within the major terminal sire breeds. Laparoscopic procedures for the collection and transfer of embryos in sheep now have the potential to overcome many of the limitations of more invasive surgical approaches, although the unpredictability of superovulatory responses remains a particular obstacle to full exploitation of multiple ovulation embryo transfer (MOET) for genetic improvement through the female line.

Animals

Early detection of pathologic pregnancy by transvaginal sonography.

Vaginal ultrasonography, by virtue of its excellent resolution, has resulted in changing our concepts about early pregnancy both in normal and abnormal gestations. The purpose of this report is to examine the clinical application of this technique in abnormal early pregnancy. Pathologic pregnancy ultimately will prove to be either (1) an ectopic pregnancy or (2) an intrauterine gestation that may be destined to failure. (A third group, fetuses with anomalies, will be dealt with elsewhere). Vaginal sonography has enhanced very early pathologic diagnosis in pregnancy because of its ability to image things previously not seen with such clarity, and because the scanning process can be carried out at the time of pelvic examination when the clinical indication is first encountered.

Abortion, Missed

The embryotoxicity of ytterbium chloride in golden hamsters.

Two experiments were performed to assess the embryotoxicity of intravenously administered ytterbium chloride in golden hamsters: (1) 50, 75, 100, or 150 mg/kg ytterbium chloride were injected early on the 8th gestation day; (2) 100 mg/kg ytterbium chloride were administered at 4 different times during the critical period of organogenesis in this species (7th day, 4 PM; 8th day, 8 AM or 4 PM; 9th day, 8 AM). The pregnant females were killed on the 12th, 14th, or 15th gestation day and the frequency of resorptions was determined, and surviving offspring were studied for the presence of external and internal malformations as well as skeletal damage. Relatively few externally visible malformations and no significant internal malformations were produced. Damage to the skeletal system included fused and supernumerary ribs, and many different sites of poor ossification. Maximal susceptibility to these effects occurred on the 8th day.

Abnormalities, Drug-Induced

The effect of intrauterine position on the radiosensitivity of rat embryos.

Rats were exposed to gamma rays or helium ions on one of days 4-9 of gestation. Embryonic survival was recorded as a function of intrauterine position at autopsy on day 20 of gestation. Embryos located at the ovarian and cervical ends of the uterus experienced higher rates of mortality than did their littermates located at the middle of the uterine horn. This effect was observed in litters exposed to both radiation modalities on all days studied. The influence of intrauterine position on embryonic survival was directly proportional to radiation dose and to the number of fetuses occupying the uterus horn. Under the least advantageous conditions (i.e., a crowded uterine horn exposed to a moderately high radiation dose), the cervical embryo's probability of survival was less than half that of the litter as a whole. A disproportionately high rate of embryonic mortality at the cervical position was also observed in litters irradiated under hypoxic conditions, suggesting that the non-random distribution of radiation effect was not the result of variations in oxygen concentration within the uterus. In contrast, there was no indication that intrauterine position influenced the distribution of gross morphologic abnormalities in irradiated litters.

Animals

Mechanism of Shoutai Wan against recurrent spontaneous abortion: regulation of decidual vascular remodeling via ERβ-ANGPT2 signaling axis.

Shoutai Wan (STW), a classic traditional Chinese medicine formula used to tonify the kidney and prevent miscarriage, has been widely applied in the clinical management of recurrent spontaneous abortion (RSA). Increasing clinical evidence supports its efficacy in reducing miscarriage rates and improving pregnancy outcomes. However, the molecular basis by which STW alleviates defective decidual vascular remodeling in unexplained RSA remains insufficiently understood. Clinically, decidual ERβ and ANGPT2 expression, as well as serum estradiol, ANGPT2 and VEGFA levels were significantly decreased in RSA patients, accompanied by reduced decidual microvascular density. Furthermore, ERβ expression was positively correlated with ANGPT2 and microvascular density. In vivo, STW dose-dependently reduced embryo loss in RSA mice, repaired the damaged decidual-placental interface structure, and improved vascular maturation, structural stability and endothelial-pericyte ultrastructural connections. Mechanistically, STW upregulated ERβ expression. We demonstrated that ERβ binds to the ANGPT2 promoter, suggesting transcriptional upregulation of ANGPT2, thereby activating Tie2 and the downstream PI3K/AKT pathway and increasing NO and VEGFA secretion. In vitro, hypoxia inhibited ERβ nuclear translocation and ANGPT2 secretion in mDSCs, while STW-containing serum reversed these abnormalities. ERβ knockdown impaired the pro-angiogenic capacity of mDSCs, which was partially rescued by exogenous ANGPT2 supplementation. Network pharmacology predicted that STW targets were mainly enriched in PI3K-Akt, estrogen, VEGF and angiogenesis-related pathways. Transcriptomic GSEA further revealed that the gene signatures of angiogenesis and PI3K-Akt signaling were markedly suppressed in the RSA model, and STW treatment significantly normalized these transcriptional signatures.

Female

Regulatory interactions and role in cell type specification of the Malpighian tubules by the cut, Krüppel, and caudal genes of Drosophila.

Krüppel and caudal genes are both required for normal segmentation of the embryo, and the developmental regulatory gene cut is necessary for the normal specification of external sensory organs. These three genes are also expressed in the Malpighian tubules before and during differentiation. Two of the genes, Krüppel and cut, are known to be required for development of the tubules. We report that the absence of maternal and zygotic caudal function reduces their normal growth and elongation. Normal Krüppel function, which is known to be required for caudal expression, is also required for cut expression, while cut and caudal are expressed independently of each other. Cell type transformations of Malpighian tubules were studied by examining the effects of mutations on the expression of markers specific to Malpighian tubules, hindgut, or midgut of normal embryos. Loss of Krüppel activity confers hindgut characteristics on those cells that normally form the Malpighian tubules with all markers tested. Loss of cut function alters the expression of some markers but not others. The pathway of tissue specific gene regulation, apparently, branches beyond Krüppel to form at least a cut and a caudal branch.

Animals

Embryonic susceptibility of Microtus ochrogaster (common prairie vole) to phenyl mercuric acetate.

Pregnant M. ochrogaster received single intraperitoneal (i.p.) doses of 0.06, 0.125, 0.25, 0.5, 1,2, or 5 mg phenyl mercuric acetate (PMA/kg of body weight on day 8, 9, or 10 of gestation or 0.5 mg PMA/kg on day 7, 8, 9, 10, 11, or 12 of gestation. No toxicity was exhibited, and no abnormal fetuses were observed in any group. An embryocidal effect that depended on dose and stage of development was determined by increased numbers of resorption sites; resorption sites were dose-dependent, and the embryocidal effect decreased as the embryo matured. A dose of 0.06 mg PMA/kg of body weight produced no resorptions when given on day 8 or day 10 of gestation; 0.125 mg/kg produced no resorptions if given on day 10; 0.5 mg/kg resulted in live fetuses and resorptions in the same uterus when given on days 7 through 11; the same dose on day 12 produced all live fetuses; and 1, 2, or 5 mg PMA/kg given on day 8, 9, or 10 of gestation caused total embryo resorption. The i.p. LD50 of PMA in Microtus ochrogaster adult females was 10 mg/kg of body weight.

Animals

Incidence and significance of unequal gestational sac diameter or embryo crown-rump length in twin pregnancy.

The incidence of differences in gestational sac diameter and crown-rump length, measured at the time of the first ultrasound, in which at least one gestational sac or crown-rump length could be visualized, were analysed retrospectively in 260 twin pregnancies in which one or both fetuses were delivered at term. The difference in gestational sac diameter averaged 1.2 +/- 0.1 mm for pregnancies which ended in twin births, compared to 2.0 +/- 0.3 mm when pregnancy ended in single births (P less than 0.02). The difference in crown-rump length averaged 2.4 +/- 0.6 mm for pregnancies which ended in a single birth, compared to 0.9 +/- 0.1 mm for twin births (P = 0.02). Disparities of greater than or equal to 3 mm in gestational sac diameter (P less than 0.05) or crown-rump length (P less than 0.001) were associated with an embryo loss rate greater than or equal to 50%. The disparity in gestational sac diameter (P less than 0.04) and crown-rump length (P less than 0.01) was smaller in pregnancies resulting from assisted reproductive technologies, compared with pregnancies resulting from coitus or insemination. Differences in gestational sac diameter and crown-rump length in early pregnancy were unrelated to differences in birth weight, length or sex.

Extraembryonic Membranes