The regulative potential of the limb region in 11.5-day rat embryos following the amputation of the fore-limb bud

Summary The regulative potential of the fore-limb region following the removal of the limb bud was investigated in 11.5-day rat embryos. Fore-limb buds were amputated from a total of 54 embryos. Five embryos were immediately examined via scanning electron microscopy (SEM) to assess the quality of the operation and the reproducibility of the technique. In all cases, the forelimb bud and adjacent tissues extending down to the celomic cavity at the same level were completely removed. The remaining 49 operated embryos were cultured in vitro and examined at different time intervals. Gross inspection of embryos which had been cultured for 24 h, revealed that 24 out of 37 had developed a pair of limb bud-like protrusions at the operation site. Twelve formed only a single protrusion, while nothing was found in the remaining embryo. These protrusions were examined in greater detail under SEM and light microscopy. SEM observations showed that these protrusions were covered with an intact layer of ectoderm. In embryos with a pair of protrusions, these outgrowths developed opposite somites 7 to 13. The failure of either one of these two outgrowths to form, produced our second type of experimental embryo, those which had just a single protrusion. Histological examination revealed that an apical ectodermal ridge (AER) was discernible on the protrusions of 36% of the embryos. Finally, we have established how these protrusions were constructed from SEM observations of operated embryos cultured for 6 h and 10 h.     

Sensitivity of early mouse embryos to the antifolic drug pyrimethamine

The effect of the antifolic drug pyrimethamine (2,4-diamino-5-p-chlorophenyl-6-ethylpyrimidine) on cleavage of CBA mouse embryos was studied. Pyrimethamine does not affect the development of early mouse embryos in utero but considerably inhibits cleavage of embryos explanted into medium containing the drug. The sensitivity of mouse and rat embryos to pyrimethamine is compared and it is concluded that there are no interspecific differences in their response to the teratogen at the level of the embryonic cells themselves.Department of Embryology, Institute of Experimental Medicine, Academy of Medical Sciences of the USSR, Leningrad. (Presented by Academician of the Academy of Medical Sciences of the USSR A. N. Klimov.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 82, No. 10, pp. 1247–1250, October, 1976.     

Relations between Meckel's cartilage and the morphogenesis of the mandible in the human embryo

Summary Based on a series of human embryos classified in stages [28], the authors studied the evolution of Meckel's cartilage in its ventral portion and its relations to the morphogenesis of the mandible. Three stages appeared particularly important: stage 16, appearance of Meckel's cartilage; stage 20, beginning of membranous ossification; and stage 23, end of the embryonic period (8th week). The primitive bony nodule which develops from the embryonic mesenchyme appears as a double bony layer forming a groove containing the neurovascular bundle, into which the dental lamina is also invaginated.

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Relations entre le cartilage de Meckel et la morphogénèse de la mandibule chez l'embryon humain

Résumé A partir d'une série d'embryons humains classés en stades [28], les auteurs étudient l'évolution du cartilage de Meckel dans sa portion ventrale et ses relations avec la morphogénèse de la mandibule. Trois stades apparaissent comme particulièrement importants : Stade 16 : Apparition du cartilage de Meckel ; Stade 20 : Début de l'ossification membraneuse. Stade 23 : Fin de la période embryonnaire (8ème semaine). Le nodule osseux primitif, qui s'est formé aux dépens du mésenchyme embryonnaire, se présente sous l'aspect d'une double lame osseuse formant une gouttière, lit du paquet vasculo-nerveux, dans laquelle vient également s'invaginer la lame dentaire.

     

Fertilization and early embryology: Effects of persistent chlorinated hydrocarbons on fertility and embryonic development in the rabbit

The commercial polychlorinated biphenyl (PCB) formulation Aroclor1260 (4 mg/kg body weight), technical grade dichlorodiphenyltrichloroethane(DDT; 3 mg) and Lindane (-hexachlorocyclohexane; 0.8 mg) wereadministered orally, either separately or in combination, tosexually mature female rabbits three times per week for 12–15weeks. Oviductal and uterine luminal fluid, cleavage stage embryos(day 1 post coitum), blastocysts (day 6), fetuses, exocoelicfluid and placentae (day 11) were analysed, firstly for chlorinatedhydrocarbon residues, and secondly for embryonic and fetal development.The doses applied were well tolerated by the treated animals.PCB and DDT accumulated in uterine secretions (day 6) but notin oviductal luminal fluid (day 1). Both chlorinated hydrocarbonswere found in preimplantation blastocysts. Residues in day 11fetuses were 16- (DDT) or 18-fold (PCB) higher than in day 6blastocysts. Significant amounts were also detected in placentaltissue and in exocoelic fluid. A specific accumulation of thehighly chlorinated biphenyl congener no. 180 was noted in fetuses,placentae and exocoelic fluid. The clear accumulation of thechlorinated hydrocarbon compounds in luminal fluid and embryonictissue is contrasted by rather weak effects on fertility. Nostatistically significant differences between treated animalsand controls were observed for fertilization rate and pre- andpost-implantation (up to day 11 post coitum) losses. However,in females exposed to PCB, a 20% higher loss of blastocystswas noticed, as compared with controls (P > 0.05). This effectwas shown on day 6 of embryonic development and may be due tothe embryotoxic activities of PCB.     

Fertilization and early embryology: The effect of equilibration temperature and time on the outcome of ultrarapid freezing of 1-cell mouse embryos

Mouse 1-cell embryos were frozen ultrarapidly at a rate of 2500°C/minin solutions containing 0.25 M sucrose, 0.5% (w/v) bovine serumalbumin (BSA) and 3 or 4.5 M dimethyl sulphoxide (DMSO) or 3or 4.5 M 1, 2-propanediol (PROH) in HEPES-buffered modifiedEarle's medium. We investigated the effect of pre-freeze equilibrationfor 1, 3, 5 or 10 min at 22°C and for 1, 3, 5, 10, 15 or20 min at 4°C. After thawing in a 22°C water bath ata rate of 2500°C/min and dilution in 1 M sucrose in HEPES-bufferedmodified Earle's medium, embryos were cultured in vitro in bicarbonate-bufferedmodified Earle‘s medium with 0.5% (w/v) crystalline BSA.Embryo viability was expressed as the percentage of hatchingor hatched blastocysts resulting from the initial number offrozen-thawed 1-cell embryos. To determine the toxicity of thefreezing solutions, embryo viability was evaluated after equilibrationwithout freezing. Our results demonstrated that the concentration,the equilibration temperature and time are very important factorsin ultrarapid freezing of mouse 1-cell embryos. Optimal viabilitywas found when equilibration was done in 4.5 M DMSO for 3–5min at 22°C and in 4.5 M PROH for 3–5 min at 4°C.The results with regard to exposure to the freezing solutionsindicated that the loss of viability beyond an optimum is notdue solely to cryoprotectant toxicity, in particular not at4°C and not for DMSO. It is suggested that the temperatureand time of equilibration influence the degree of cryoprotectantpermeation and subsequent rehydration, which play a role indetermining freezing susceptibility in terms of ice formation.We conclude that both DMSO and, in contradiction to previousreports, PROH can be used perfectly adequately for ultrarapidfreezing on condition that concentration, and the temperatureand time of equilibration are controlled.     

扬子鳄胚胎视网膜发育的免疫组化研究

目的:检测6种激素在孵育第8天至孵出时扬子鳄胚胎视网膜的表达及变化。方法:免疫组织化学方法。结果:(1)生长抑素(SS)在孵育第8～30天有表达,第12天数量最多,30天以后消失;(2)5-羟色胺(5-HT)从孵育第24天开始表达,第30～40天数量最多,孵出时数量减少;(3)转化生长因子β_1(TGFβ_1)从孵育第40天开始表达,孵出时增多;(4)神经丝蛋白(NFP)阳性细胞和纤维在所检测的各个发育阶段均有表达,P53和P物质在各期胚胎视网膜均未检出阳性细胞。结论:视网膜的发育是个渐进的过程,不同激素在扬子鳄胚胎视网膜中的表达规律不同。     

Reproductive epigenetics

Epigenetics refers to covalent modifications of DNA and core histones that regulate gene activity without altering DNA sequence. To date, the best-characterized DNA modification associated with the modulation of gene activity is methylation of cytosine residues within CpG dinucleotides. Human disorders associated with epigenetic abnormalities include rare imprinting diseases, molar pregnancies, and childhood cancers. Germ cell development and early embryo development are critical times when epigenetic patterns are initiated or maintained. This review focuses on the epigenetic modification DNA methylation and discusses recent progress that has been made in understanding when and how epigenetic patterns are differentially established in the male and female germlines, the mouse, and human disorders associated with abnormalities in epigenetic programming in germ cells and early embryos, as well as genetic and other modulators (e.g. nutrition and drugs) of reproductive epigenetic events.     

Human embryonic stem cells: research,ethics and policy

The use of human embryos for research on embryonic stem (ES) cells is currently high on the ethical and political agenda in many countries. Despite the potential benefit of using human ES cells in the treatment of disease, their use remains controversial because of their derivation from early embryos. Here, we address some of the ethical issues surrounding the use of human embryos and human ES cells in the context of state-of-the-art research on the development of stem cell based transplantation therapy.