Tube Defects In Mouse Embryos Research Articles

RNA-Seq analysis reveals candidate genes that may explain neural tube defects in mouse embryos lacking SR-BI

RNA-Seq analysis reveals candidate genes that may explain neural tube defects in mouse embryos lacking SR-BI

Homocysteine is embryotoxic but does not cause neural tube defects in mouse embryos.

The observation of elevated maternal plasma homocysteine concentration in pregnancies affected by neural tube defects (NTD) suggests that folate metabolism may be disturbed in NTD cases. In addition, studies on the chick embryo suggest that hyperhomocysteinaemia may contribute directly to the development of NTD. In order to test the hypothesis that homocysteine may cause NTD, we cultured mouse embryos in the presence of homocysteine thiolactone during the period of cranial neural tube closure. At doses of 0.5 mM or above, exposure to homocysteine thiolactone caused growth retardation, blisters and abnormalities of somite development. Despite the teratogenic effects of homocysteine we did not detect any increase in the incidence of neural tube defects. Neither was there an effect on the incorporation of thymidine into DNA, a potential marker of alterations in the folate or homocysteine/methionine cycles. These observations suggest that homocysteine is unlikely to be a direct cause of NTD in humans. Rather, the elevated levels of homocysteine in human NTD pregnancies may reflect a disturbance in folate-related metabolism.

Perturbations in choline metabolism cause neural tube defects in mouse embryos in vitro.

A role for choline during early stages of mammalian embryogenesis has not been established, although recent studies show that inhibitors of choline uptake and metabolism, 2-dimethylaminoethanol (DMAE), and 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine (ET-18-OCH3), produce neural tube defects in mouse embryos grown in vitro. To determine potential mechanisms responsible for these abnormalities, choline metabolism in the presence or absence of these inhibitors was evaluated in cultured, neurulating mouse embryos by using chromatographic techniques. Results showed that 90%-95% of 14C-choline was incorporated into phosphocholine and phosphatidylcholine (PtdCho), which was metabolized to sphingomyelin. Choline was oxidized to betaine, and betaine homocysteine methyltransferase was expressed. Acetylcholine was synthesized in yolk sacs, but 70 kDa choline acetyltransferase was undetectable by immunoblot. DMAE reduced embryonic choline uptake and inhibited phosphocholine, PtdCho, phosphatidylethanolamine (PtdEtn), and sphingomyelin synthesis. ET-18-OCH3 also inhibited PtdCho synthesis. In embryos and yolk sacs incubated with 3H-ethanolamine, 95% of recovered label was PtdEtn, but PtdEtn was not converted to PtdCho, which suggested that phosphatidylethanolamine methyltransferase (PeMT) activity was absent. In ET-18-OCH3 treated yolk sacs, PtdEtn was increased, but PtdCho was still not generated through PeMT. Results suggest that endogenous PtdCho synthesis is important during neurulation and that perturbed choline metabolism contributes to neural tube defects produced by DMAE and ET-18-OCH3.

Perturbations in choline metabolism cause neural tube defects in mouse embryos in vitro.

Perturbations in choline metabolism cause neural tube defects in mouse embryos in vitro.

Antisense modulation of 5,10-methylenetetrahydrofolate reductase expression produces neural tube defects in mouse embryos

The role of folate metabolism in producing neural tube defects (NTDs) in humans is unknown. In the current study, antisense oligodeoxyribonucleotide technology was utilized to disrupt normal expression of the gene for 5,10-methylenetetrahydrofolate reductase (MTHFR) in organogenesis-stage mouse embryos. Two different antisense probes were microinjected into the amniotic sac of gestation day (GD) 8 mouse embryos with PBS or scrambled sense oligodeoxyribonucleotides injected into control embryos. Concentration-dependent increases in the frequencies of embryos with NTDs were observed for both antisense sequences. The level of mRNA for MTHFR was decreased in embryos treated with the higher concentration of one antisense sequence, indicating that the sequence is able to decrease gene expression. 5-methyltetrahydrofolate, the product of the MTHFR reaction, was able to decrease the incidence of antisense-induced NTDs, but co-injection with L-methionine did not. These results suggest that reduced expression of MTHFR may play a role in producing NTDs.

Neurulation and neural tube closure defects.

AbstractThe purpose of this chapter is to summarize some technical approaches and aspects of interpretation that are important when studying the process of neurulation and the origin of neural tube defects in mouse embryos. The techniques are not exclusive to studies of neurulation, so few protocols are described in practical detail. Instead, we emphasize the overall type of approach that appears most effective and give tips on the best application of standard techniques such as those described elsewhere in this and other volumes.KeywordsNeural TubeMouse EmbryoNeural Tube DefectBody AxisNeural Tube ClosureThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Cell proliferation, migration and CAM-dependent neurite outgrowth as developmental in vitro endpoints for screening teratogenic potential: Application to valproic acid and related analogues of varying potency

The in vivo teratogenic potential of valproic acid (VPA) and related teratogenic and non-teratogenic analogues has been correlated with their effects on specific in vitro endpoints of cell proliferation, migration and CAM-dependent neurite outgrowth, as these events are common to crucial epochs of development. The (±)-2- n-propyl-4-pentynoic acid [(±)-4-yn-VPA] and S-2- n-propyl-4-pentynoic acid [ S(-)-4-yn-VPA] analogues increased the incidence of neural tube defects in mouse embryos exposed to a single dose, whereas the E-2- n-propyl-2-pentenoic acid ( E-2-en-VPA) analogue and R-2- n-propyl-4-pentynoic acid [ R( + )-4-yn-VPA] enantiomer were without effect. VPA and related analogues tested exerted comparable G1 phase antiproliferative effects in C6 glioma and limb bud cells in a dose range of 0–3 mM; however, their relative potency did not correlate with in vivo teratogenicity. In contrast, VPA and all teratogenic analogues, at 3 mM, inhibited neuronal cell aggregation and limb bud chondrocyte differentiation in a manner that exhibited a reasonable correlation with their in vivo teratogenicity. The teratogenic S(−)-4-yn-VPA and non-teratogenic R( + )-4-yn-VPA enantiomers exhibited a differential inhibition of primary neurone outgrowth of neuntes stimulated by cell adhesion molecules [L1 and N-cadherin (NCAD)]. Half-maximal inhibition was observed at approximately 150 μM for the teratogenic S(−)-4-yn-VPA enantiomer, but not the non-teratogenic R( + )-4-yn-VPA form. These results suggest that in vitro perturbations of differentiation are likely to provide the greatest discriminatory power for in vivo teratogenicity.

Common hierarchies of susceptibility to the induction of neural tube defects in mouse embryos by valproic acid and its 4-propyl-4-pentenoic acid metabolite.

The teratogenic effects of valproic acid and its 4-propyl-4-pentenoic acid (4-en) metabolite were investigated in three inbred mouse strains that were known to possess differing sensitivity to heat-induced neural tube defects. In the heat-resistant DBA/2J strain, administration of either valproic acid or the metabolite during the critical period of neural tube development failed to produce any abnormal offspring. Similar treatment in the moderately heat-sensitive LM/Bc strain resulted in up to 19.8% exencephalic fetuses. The highly heat-sensitive SWV strain was also very susceptible to the induction of neural tube defects by either valproic acid or its 4-en metabolite. When administered on gestational day 8 plus 12 hours, the parent compound produced 35% exencephalic fetuses, while the metabolite had a response frequency of 32.4%. Thus, the hierarchy of susceptibility for the induction of neural tube defects in these inbred mouse strains was exactly the same whether the teratogen was a physical agent such as hyperthermia or a chemical compound such as valproic acid. If such diverse agents as these should interact to produce malformations, then it is possible that a wide variety of other agents might interact in a similar manner to produce neural tube defects.

Induction of neural tube defects and skeletal malformations in mice following brief hyperthermia in utero.

Hyperthermia was induced in ICR mice on day 8.5 of gestation by immersing them in hot water. Control mice were immersed in water at 38 degrees C for 15 min. In dams exposed to 42 degrees C for 12.5-15 min or to 43 degrees C for 7.5-10 min, externally malformed fetuses increased significantly and in a dose-related manner. Anterior neural tube defects (exencephaly, anencephaly, encephalocele, and cranial neural tube defect with facial cleft) were induced most frequently. Embryonic and fetal death and skeletal malformations also increased following heat stress. At intervals after heat exposure, sections of embryonic prosencephalon were prepared for light microscopy. At 2 h after heat stress, mitotic figures disappeared almost completely in embryonic tissues. Mitotic activity was inhibited for at least 3 h. At 6 h, there was a burst of new mitotic activity. Some damaged cells became pyknotic and abnormal cells were encountered in the neuroepithelial tissue after 3-12 h. Thus, neural tube defects in mouse embryos following maternal hyperthermia may result from a temporary cessation of cell proliferation and partial necrosis of the embryonic neuroepithelium.