The Bioavailability of Substances Administered to Chick Embryos: The Maximum Effective Route of Administration

Toxicokinetic studies are of key importance in both the design and the interpretation of developmental toxicity studies. The aim of this study was to determine concentrations of test substances within the chick embryo following the administration schedule recommended in the chick embryotoxicity screening test (CHEST). The concentration-time relationships were investigated by using four labelled substances with various physicochemical and embryo-toxic properties ([14C] sodium acetate, [14C] palmitic acid, [3H] Cortisol and [3H] cytosine arabinoside). These labelled chemicals were mixed with cold substances and singly administered at two dose levels to chick embryos on days 2, 3 and 4 of incubation. Extrachorial and subgerminal routes were used on day 2, and extrachorial and intra-amniotic applications were chosen on days 3 and 4. The concentration of labelled chemical present within the embryo was assessed at predetermined intervals by scintillation fluorimetry (from 6 minutes to 96 hours after administration), and used for estimating the concentration curves. Regardless of the substance, dose and application route, the concentration curves exhibited a characteristic pattern, reaching their peaks within the first 6 hours, and dropping down to near zero 48–96 hours after administration. The decrease followed the first order law, demonstrating that, within the CHEST system, the avian embryo does not act as a closed system. With regard to the total amount of substance entering the embryo, extrachorial administration appeared to be superior to subgerminal administration on day 2. Intra-amniotic administration was superior to extrachorial administration on days 3 and 4. These differences were most pronounced after administration of lipid-soluble palmitic acid. The concentrations within embryonic tissues were directly dose-dependent. After consideration of all these findings, we concluded that the CHEST system probably has closer similarity to the toxicokinetics of exposure of mammalian embryos (i.e. reaching a peak and then a gradual decline over time) than any other in vitro test of developmental toxicity, where the chemical is simply added to culture media. Several practical recommendations for improving the CHEST system were derived.

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The Use of Biomarkers as Alternatives to Current Animal Tests on Food Chemicals

Alternatives to Laboratory Animals ◽

10.1177/026119299802600411 ◽

1998 ◽

Vol 26 (4) ◽

pp. 421-480

Author(s):

Krys Bottrill

Keyword(s):

Animal Studies ◽

Developmental Toxicity ◽

Reproductive Toxicity ◽

In Vitro Test ◽

Animal Testing ◽

Dietary Biomarkers ◽

Recent Developments ◽

Mechanistic Basis ◽

The Three Rs

Recent developments in biomarkers relating to the interrelationship of diet, disease and health were surveyed. Most emphasis was placed on biomarkers of deleterious effects, since these are of greatest relevance to the subject of this review. The area of greatest activity was found to be that relating to biomarkers of mutagenic, genotoxic and carcinogenic effects. This is also one of the major areas of concern in considerations of the beneficial and deleterious effects of dietary components, and also the area in which regulatory testing requires studies of the longest duration. A degree of progress has also been made in the identification and development of biomarkers relating to certain classes of target organ toxicity. Biomarkers for other types of toxicity, such as immunotoxicity, neurotoxicity, reproductive toxicity and developmental toxicity, are less developed, and further investigation in these areas is required before a comprehensive biomarker strategy can be established. A criticism that recurs constantly in the biomarker literature is the lack of standardisation in the methods used, and the lack of reference standards for the purposes of validation and quality control. It is encouraging to note the growing acknowledgement of the need for validation of biomarkers and biomarker assays. Some validation studies have already been initiated. This review puts forward proposals for criteria to be used in biomarker validation. More discussion on this subject is required. It is concluded that the use of biomarkers can, in some cases, facilitate the implementation of the Three Rs with respect to the testing of food chemicals and studies on the effects of diet on health. The greatest potential is seen to be in the refinement of animal testing, in which biomarkers could serve as early and sensitive endpoints, in order to reduce the duration of the studies and also reduce the number of animals required. Biomarkers could also contribute to establishing a mechanistic basis for in vitro test systems and to facilitating their validation and acceptance. Finally, the increased information that could result from the incorporation of biomarker determinations into population studies could reduce the need for supplementary animal studies. This review makes a number of recommendations concerning the prioritisation of future activities on dietary biomarkers in relation to the Three Rs. It is emphasised, however, that further discussions will be required among toxicologists, epidemiologists and others researching the relationship between diet and health.

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Metabolites Secreted by Bovine Embryos In Vitro Predict Pregnancies That the Recipient Plasma Metabolome Cannot, and Vice Versa

Metabolites ◽

10.3390/metabo11030162 ◽

2021 ◽

Vol 11 (3) ◽

pp. 162

Author(s):

Enrique Gomez ◽

Nuria Canela ◽

Pol Herrero ◽

Adrià Cereto ◽

Isabel Gimeno ◽

...

Keyword(s):

Mass Spectrometry ◽

Stearic Acid ◽

Palmitic Acid ◽

Hippuric Acid ◽

Culture Media ◽

Capric Acid ◽

Glyceryl Monostearate ◽

Invasive Approach ◽

Hydrocinnamic Acid

This work describes the use of mass spectrometry-based metabolomics as a non-invasive approach to accurately predict birth prior to embryo transfer (ET) starting from embryo culture media and plasma recipient. Metabolomics was used here as a predictive platform. Day-6 in vitro produced embryos developed singly in modified synthetic oviduct fluid culture medium (CM) drops for 24 h were vitrified as Day-7 blastocysts and transferred to recipients. Day-0 and Day-7 recipient plasma (N = 36 × 2) and CM (N = 36) were analyzed by gas chromatography coupled to the quadrupole time of flight mass spectrometry (GC-qTOF). Metabolites quantified in CM and plasma were analyzed as a function to predict pregnancy at Day-40, Day-62, and birth (univariate and multivariate statistics). Subsequently, a Boolean matrix (F1 score) was constructed with metabolite pairs (one from the embryo, and one from the recipient) to combine the predictive power of embryos and recipients. Validation was performed in independent cohorts of ETs analyzed. Embryos that did not reach birth released more stearic acid, capric acid, palmitic acid, and glyceryl monostearate in CM (i.e., (p < 0.05, FDR < 0.05, Receiver Operator Characteristic—area under curve (ROC-AUC)> 0.669). Within Holstein recipients, hydrocinnamic acid, alanine, and lysine predicted birth (ROC-AUC > 0.778). Asturiana de los Valles recipients that reached birth showed lower concentrations of 6-methyl-5-hepten-2-one, stearic acid, palmitic acid, and hippuric acid (ROC-AUC > 0.832). Embryonal capric acid and glyceryl-monostearate formed F1 scores generally >0.900, with metabolites found both to differ (e.g., hippuric acid, hydrocinnamic acid) or not (e.g., heptadecanoic acid, citric acid) with pregnancy in plasmas, as hypothesized. Efficient lipid metabolism in the embryo and the recipient can allow pregnancy to proceed. Changes in phenolics from plasma suggest that microbiota and liver metabolism influence the pregnancy establishment in cattle.

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Investigation of the developmental toxicity of cocaine in in vitro cultured chick embryos: Correlation of effects with intra-embryonic drug levels

Toxicology in Vitro ◽

10.1016/0887-2333(91)90004-w ◽

1991 ◽

Vol 5 (4) ◽

pp. 285-293 ◽

Cited By ~ 5

Author(s):

G.J. Burin ◽

L.K. Al-Ghaith ◽

K.G. Anitole ◽

M.K. Barber ◽

K.M. Brown

Keyword(s):

Developmental Toxicity ◽

Chick Embryos ◽

Drug Levels

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Hepatic induction in the avian embryo: Specificity of reactive endoderm and inductive mesoderm

Development ◽

10.1242/dev.63.1.111 ◽

1981 ◽

Vol 63 (1) ◽

pp. 111-125

Author(s):

Sumiko Fukuda-Taira

Keyword(s):

Tissue Specificity ◽

Normal Development ◽

Mouse Embryos ◽

Avian Embryo ◽

Chick Embryos ◽

Cardiac Region ◽

Myocardial Layers ◽

Cardiac Mesoderm

Mesoderm of precardiac and cardiac region (‘cardiac’ mesoderm) of chick, quail and mouse embryos could induce hepatic epithelium in the endoderm of the anterior half of young quail or chick embryos (anterior endoderm) in vitro as well as in vivo. No species specificity in the induction of hepatic epithelium by the ‘cardiac’ mesoderm could be observed. The hepatic induction, was controlled strictly by tissue specificity of both endoderm and mesoderm. Replacement of the ‘cardiac’ mesoderm or the anterior endoderm by noncardiac mesoderms or endoderms other than the anterior endoderm resulted in failure of hepatic induction. Only the anterior endoderm was found to have competence for hepatic induction, indicating that it was committed, in unknown ways, to react with ‘cardiac’ mesoderm, and can properly be called pre-hepatic endoderm. Comparison between the development of hepatic endoderm and the hepatic induction potency of ‘cardiac’ mesoderm, which was most intense during 1- to 1·5- incubation days and decreased gradually with the increase of the stage, suggests that in normal development the ‘cardiac’ mesoderm actually induces hepatic epithelium in the competent endoderm. Hepaticinduction potency remained up to 6 days, and was found in truncus arteriosus, ventricle and auricle areas and in endocardial and myocardial layers of the heart.

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From simple in vitro test to complex models: The example of developmental toxicity of organophosphorus compounds

Toxicology Letters ◽

10.1016/j.toxlet.2016.06.1174 ◽

2016 ◽

Vol 258 ◽

pp. S17

Author(s):

Eugenio Vilanova ◽

Andrea C. Romero ◽

David Pamies ◽

Carmen Estevan ◽

Miguel A. Sogorb

Keyword(s):

Organophosphorus Compounds ◽

Developmental Toxicity ◽

In Vitro Test ◽

Complex Models ◽

Vitro Test

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Total Replacement of the Yolk of Chick Embryos

Development ◽

10.1242/dev.5.2.210 ◽

1957 ◽

Vol 5 (2) ◽

pp. 210-214

Author(s):

C. R. Grau ◽

N. W. Klein ◽

T. L. Lau

Keyword(s):

Avian Embryo ◽

Chick Embryos ◽

Food Material ◽

Fundamental Study ◽

Essential Step ◽

Total Replacement ◽

Young Embryo ◽

Normal Food ◽

Embryonic Nutrition

Isolation of the avian embryo from its normal food material is an essential step in a fundamental study of embryonic nutrition. With present methods, however, it has not been possible to grow embryos in culture for much longer than one day. In 1932 Waddington described methods of growing explanted embryos on serum clots. Several years later Spratt (1947, 1952) devised simplified media for embryos cultured on agar with which he studied various aspects of morphogenesis and carbohydrate needs of the young embryo. New (1955) has recently reported another technique for growing explanted embryos which allows development to proceed to about 60 hours, 40 hours of which are in vitro. A number of workers have injected various nutrients, antimetabolites, drugs, poisons, and hormones, directly into the yolk sac or other areas of the egg (e.g. Landauer, 1954).

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Immunochemical study of hypoxanthine-dehydrogenase in different organs of the chick

Development ◽

10.1242/dev.27.2.261 ◽

1972 ◽

Vol 27 (2) ◽

pp. 261-275

Author(s):

Yvon Croisille

Keyword(s):

Molecular Weight ◽

Culture Media ◽

Chick Embryos ◽

Immunochemical Study ◽

Direct Association ◽

Embryonic Life ◽

Low Levels ◽

Starch Gels

Immunotitration of hypoxanthine-dehydrogenase confirms earlier findings according to which, in the liver, HXDH activity stays low throughout embryonic life and increases suddenly at the period of hatching. Whether from liver, kidney, intestine, pancreas or mesonephros, HXDH appears to have the same electrophoretic mobility (in agar, agarose and starch gels), the same molecular weight (estimated to be about 290000 by Sephadex G 200 filtration) and the same immunochemical properties (as tested with 24 anti-liver, 18 anti-kidney, and 4 anti-mesonephros sera). Various attempts to interfere with HXDH synthesis in vivo or in vitro (injection of substrate or liver extracts; explanation of fragments of mesonephros, metanephros or intestine from 9- to 12-day-old chick embryos on substrate-containing culture media; direct association of tissues containing high and low levels of enzyme) have so far failed.

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Structural integrity after cold storage of human epidermal model in hypothermosol: A correlative ultrastructural and fluorescent assay

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100169080 ◽

1994 ◽

Vol 52 ◽

pp. 270-271

Author(s):

Henry H. Eichelberger ◽

John G. Baust ◽

Robert G. Van Buskirk

Keyword(s):

Cold Storage ◽

Structural Integrity ◽

Culture Media ◽

Cell Structure ◽

Natural State ◽

Sodium Cacodylate ◽

Ruthenium Tetroxide ◽

Cacodylate Buffer ◽

Before And After

For research in cell differentiation and in vitro toxicology it is essential to provide a natural state of cell structure as a benchmark for interpreting results. Hypothermosol (Cryomedical Sciences, Rockville, MD) has proven useful in insuring the viability of synthetic human epidermis during cold-storage and in maintaining the epidermis’ ability to continue to differentiate following warming.Human epidermal equivalent, EpiDerm (MatTek Corporation, Ashland, MA) consisting of fully differentiated stratified human epidermal cells were grown on a microporous membrane. EpiDerm samples were fixed before and after cold-storage (4°C) for 5 days in Hypothermosol or skin culture media (MatTek Corporation) and allowed to recover for 7 days at 37°C. EpiDerm samples were fixed 1 hour in 2.5% glutaraldehyde in sodium cacodylate buffer (pH 7.2). A secondary fixation with 0.2% ruthenium tetroxide (Polysciences, Inc., Warrington, PA) in sodium cacodylate was carried out for 3 hours at 4°C. Other samples were similarly fixed, but with 1% Osmium tetroxide in place of ruthenium tetroxide. Samples were dehydrated through a graded acetone series, infiltrated with Spurrs resin (Polysciences Inc.) and polymerized at 70°C.

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Haemostatic Platelet Plug Formation in the Isolated Rabbit Mesenteric Preparation - An Analysis of Red Blood Cell Participation

Thrombosis and Haemostasis ◽

10.1055/s-0038-1650069 ◽

1980 ◽

Vol 44 (01) ◽

pp. 006-008 ◽

Cited By ~ 12

Author(s):

D Bergqvist ◽

K-E Arfors

Keyword(s):

Whole Blood ◽

Platelet Rich Plasma ◽

Adenosine Diphosphate ◽

In Vitro Test ◽

Pharmacological Agents ◽

Vitro Test ◽

Releasing Effect ◽

Plug Formation

SummaryIn a model using an isolated rabbit mesenteric preparation microvessels were transected and the time until haemostatic plugs formed was registered. Perfusion of platelet rich plasma gave no haemostasis whereas whole blood did. Addition of chlorpromazine or adenosine to the whole blood significantly prolonged the time for haemostasis, and addition of ADP to the platelet rich plasma significantly shortened it. It is concluded that red cells are necessary for a normal haemostasis in this model, probably by a combination of a haemodynamic and ADP releasing effect.The fundamental role of platelets in haemostatic plug formation is unquestionable but there are still problems concerning the stimulus for this process to start. Three platelet aggregating substances have been discussed – thrombin, adenosine diphosphate (ADP) and collagen. Evidence speaking in favour of thrombin is, however, very minimal, and the discussion has to be focused on collagen and ADP. In an in vitro system using polyethylene tubings we have shown that "haemostasis" can be obtained without the presence of collagen but against these results can be argued that it is only another in vitro test for platelet aggregation (1).To be able to induce haemostasis in this model, however, the presence of red blood cells is necessary. To further study this problem we have developed a model where haemostatic plug formation can be studied in the isolated rabbit mesentery and we have briefly reported on this (2).Thus, it is possible to perfuse the vessels with whole blood as well as with platelet rich plasma (PRP) and different pharmacological agents of importance.

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