Cholinephosphotransferase activities in early rat embryos and their associated placentas

CDP-Choline:1,2-diglycerolcholinephosphotransferase (EC 2.7.8.2, cholinephosphotransferase) activities were determined in subcellular fractions prepared from rat embryos, placentas, or yolk sacs obtained on the fourteenth day of gestation. It was found that, in all of the tissues studied, cholinephosphotransferase activity (1) copurified with NADPH-cytochrome c reductase activity (EC 1.6.2.4), (2) was maximal around pH 8.0; (3) was stimulated by MgCl₂, exogenous diolein, and cytidine diphosphocholine (CDP-choline); and (4) was highest in homogenates of placentas, lowest in those of embryos, and intermediate in those of yolk sacs. These data substantiate, for the first time, that the early mammalian (rat) embryo, placenta, and yolk sac have the ability to synthesize phospholipids de novo.     

Cadmium-induced forelimb ectrodactyly: a proposed mechanism of teratogenesis

We have attempted to elucidate the mechanism of cadmium teratogenesis utilizing inbred mouse strains sensitive (C57BL/6J) or resistant (SWV) to the embryotoxic effect of this common heavy metal contaminant. Carbonic anhydrase activity of whole-embryo homogenates was moderately depressed in C57BL/6J mice compared to a slight and transient decrease in the resistant SWV mice. Embryonic erythrocytes were similarly examined, and the cadmium did not have any effect on carbonic anhydrase activity in either strain. Likewise, histochemical examination of carbonic anhydrase activity did not reveal any effect of cadmium in the embryos of their strain. Generally, the zinc concentration of embryos was not affected by cadmium administration. However, increased levels of zinc were observed in cadmium-exposed yolk sacs of both strains suggesting that cadmium produces an adverse effect on yolk sac function. Untreated C57BL/6J units (embryo plus surrounding extraembryonic membranes), embryos, and yolk sacs had much lower hemoglobin concentrations than those observed in untreated SWV units, embryos, and yolk sacs. Additionally, cadmium exposure significantly decreased C57BL/6J embryonic hemoglobin levels on gestation day 10 (PM) and increased C57BL/6J yolk sac hemoglobin levels on gestation days 10 (AM) and 10 (PM). No difference in hemoglobin concentration was observed between untreated and cadmium-treated SWV embryos or yolk sacs. We propose that cadmium induces forelimb ectrodactyly by creating an acidotic embryonic environment and that the primary site at which cadmium exerts its teratogenic effect might be the yolk sac.     

The pattern of apolipoprotein B100 containing lipoprotein subclasses produced by the isolated visceral rat yolk sac depends on developmental stage and fatty acid availability

Explants of visceral rat yolk sacs from gestational days 16, 18 and 22 were used for studying developmental changes of secretion and density distribution of lipoproteins, particularly of those containing apoB. Moreover, the influence of fatty acid supply on the amount and density distribution of secreted apolipoproteins was studied on day 18 of gestation. Active lipoprotein production was observed in yolk sacs taken on days 16 and 18 of gestation. It declined considerably on day 22 of gestation in parallel with the production of total protein, triacylglycerols and cholesterol. On all gestational days, apoB floated mainly in the LDL range (⩾ 70%) with differences in the distribution pattern of LDL subclasses. The lowest density of secreted LDL was found on day 18 of gestation (peak at d = 1.025 g/ml) followed by day 16 (peak at d = 1.035 g/ml) and day 22 of gestation (peak at d = 1.045 g/ml). ApoIV, apoE and apoAI floated exclusively in the HDL range with a peak at d = 1.089 g/ml independently of the gestational day. After incubation of yolk sacs from the 18th day of gestation with 0.4 mM or 0.8 mM oleate, the density of secreted apoB containing particles was decreased (peaks in the VLDL and IDL density range), whereas palmitate in the same concentrations caused a redistribution of secreted apoB toward higher densities (peaks at d ≥ g/ml). Taken together, the data provide evidence that the density of L.DL subclasses produced by isolated yalk sacs between days 16 and 22 of gestation depended on the gestational stage. Moreover, addition of unsaturated or saturated fatty acids to the organ culture differently affected the secretory rate and the density of lipoproteins delivered by yolk sacs on day 18 of gestation.     

Phosphatidylethanolamine synthesis in castor bean endosperm

Phosphatidylethanolamine synthesis by CDP-ethanolamine:1,2-diacylglycerol ethanolaminephosphotransferase (EC 2.7.8.1) from the endoplasmic reticulum of castor bean (Ricinus communis L. var. Hale) endosperm was characterized. The Michaelis-Menten constant of the enzyme for CDP-ethanolamine was approximately 8.0 micromolar. The pH optimum was 6.5 and a divalent cation was an absolute requirement for activity, with Mg²⁺ giving the greatest stimulation at 3 millimolar. Sulfhydryl reagents variously affected enzyme activity. No discernible differences were detected between the responses of the ethanolaminephosphotransferase and CDP-choline:1,2-diacylglycerol cholinephosphotransferase (EC 2.7.8.2) to a variety of treatments. CDP-choline and CDP-ethanolamine were competitive inhibitors of the ethanolaminephosphotransferase and cholinephosphotransferase reactions, respectively.     

Cadherin mRNAs during rat embryo development in vivo and in vitro.

Whole rat embryo cultures are being used in increasing numbers of laboratories to study the mechanisms by which teratogens disturb development. The development of early somite stage embryos in vitro is very similar morphologically to that in vivo, yet few biochemical comparisons have been made. The purpose of this study was to determine the steady-state mRNA concentrations of a family of Ca(2+)-dependent cell adhesion molecules, the cadherins, during rat embryonic development in vivo and in vitro. Embryos and yolk sacs were collected on days 10, 11, and 12 of gestation (in vivo); they were also obtained from day 10 embryos after growth in culture for 24 hr (day 11 in vitro) or 45 hr (day 12 in vitro). Total RNAs isolated from embryos and yolk sacs were studied by Northern blot analysis using specific cDNA probes for three cadherins, E-cadherin, N-cadherin, and P-cadherin. Although E-cadherin mRNA was detected in embryos, it was present at much higher concentrations in yolk sacs. In addition, multiple species of E-cadherin mRNA ranging from 3.0 to 13 kb were detected. Interestingly, the concentration of the major 4.5-kb E-cadherin mRNA species in yolk sac after 45 hr in culture was increased 2.8-fold over that on day 12 of gestation in vivo. Second, two species (4.3 and 3.5 kb) of N-cadherin mRNA were detected, almost exclusively in embryos. In yolk sac, N-cadherin mRNA was detected only after 45 hr in culture. Third, P-cadherin mRNA was detected as a single 3.5-kb species, mainly in embryos. P-cadherin mRNA concentrations in yolk sac after 45 hr in culture were 5.6-fold higher than in vivo. Thus, these results demonstrate that there is a differential distribution of cadherin mRNAs in rat embryos and yolk sacs. Further, there appear to be multiple species of mRNAs for E-cadherin and N-cadherin. Finally, while whole embryo culture in vitro did not significantly alter the steady-state concentrations of cadherin mRNAs in the embryo, these concentrations were dramatically increased in the yolk sac.     

Lysolecithin as regulator of de novo lecithin synthesis in rat liver microsomes.

1-Acyl-sn-glycero-3-phosphocholine (lysolecithin) was found to affect 1,2-diacyl-sn-glycerol:CDPcholine cholinephosphotransferase (CPT; EC 2.7.8.2) activity of rat liver microsomes in a concentration dependent, characteristic manner. Cholinephosphate transfer was activated at lysolecithin concentrations below 0.5 mM with a maximum stimulation occurring at 75–100 μM lysolecithin levels. At concentrations above 0.5 mM, CPT activity was inhibited by lysolecithin. It was shown that CPT inhibition by lysolecithin is competitive (Ki ≈ 0.6 mM) with respect to CDPcholine. The possible role of lysolecithin as regulator of de novo lecithin synthesis in vivo is outlined.     

Role of reactive oxygen species (ROS) in the diabetes-induced anomalies in rat embryos in vitro: reduction in antioxidant enzymes and low-molecular-weight antioxidants (LMWA) may be the causative factor for increased anomalies

A disturbed embryonic antioxidant defense mechanism may play a major role in diabetes-induced teratogenesis. We therefore studied the antioxidant capacity of 10.5-day-old rat embryos and their yolk sacs after culture for 28 hr in vitro under diabetic conditions (3 mg/ml glucose, 2 mg/ml beta-hydroxybutyrate (BHOB) and 10 microg/ml of acetoacetate), as compared with control embryos in vitro. We found a high rate of congenital anomalies, decreased growth and protein content, and a decrease in the activity of both superoxide dismutase (SOD) and catalase (CAT) under diabetic conditions, as compared with controls. The reducing power, which reflects the concentration and type of water-soluble and of lipid-soluble low-molecular-weight antioxidants (LMWA), was measured by cyclic voltammetry. Generally, LMWA were reduced in the embryos and yolk sacs under diabetic conditions. In the water-soluble fraction of control embryos and yolk sacs, two peak potentials were found, indicating two major groups of LMWA, while only one peak potential was found under diabetic conditions, indicating that an entire group of LMWA is missing. HPLC studies have demonstrated a decrease in vitamin C (water-soluble fraction) and in vitamin E (lipid-soluble fraction) under diabetic culture conditions, and an increase in uric acid. Generally, the concentration of LMWA was higher in the embryos than in the yolk sac. LMWA concentration, protein content, and antioxidant enzyme activity were lower in the malformed experimental embryos than in experimental embryos without anomalies. The addition of vitamins C and E to the diabetic culture medium abolished the deleterious effects of the diabetic serum on the embryos. The disturbed antioxidant defense mechanism under diabetic conditions may be explained, at least in part, by a direct effect of diabetic metabolic factors on the activity of antioxidant enzymes and on the concentration of reducing equivalents. This, in turn, may be embryotoxic.     

Cholinephosphotransferase activity during development of the sea urchin, Arbacia punctulata

Cholinephosphotransferase activity was examined during early development of Arbacia punctulata embryos. CMP was rapidly incorporated by enzyme preparation from Arbacia embryos into a compound identified as CDP-choline. Activity was dependent upon the presence of Mg²⁺, Mn²⁺, or Co²⁺, and was maximal in phosphate buffer, pH 7.0, containing 3 × 10^?2M MgCl₂ and 2 × 10^?5M CMP. Addition of ATP or egg lecithin had no effect on enzyme activity. The activity was localized in the 1000 g and 10,000 g pellets, with little or no activity in the microsomal fraction.Upon fertilization, cholinephosphotransferase activity decreased rapidly, detectable changes in activity being observed within 2 min after fertilization. After reaching a minimum activity 2 hr after fertilization, the enzyme activity increased up to the gastrula stage, then decreased once more. Possible interference by competing enzymes was examined and found to be negligible.     

Radioactive labeling of proteins in cultured postimplantation mouse embryos I. Influence of the embryo preparation method

Summary Conditions for optimum incorporation of radioactive amino acids into proteins of cultured postimplantation mouse embryos were investigated under the aspect of using these proteins for two-dimensional electrophoretic separations followed by fluorography. The aim was to obtain highly radioactive proteins under conditions as physiological as possible. Embryos at Days 10, 11, and 12 of gestation were prepared in different ways and incubated for 4 h in Tyrode’s solution containing [³H]amino acids (mixture) at a concentration of 27 μCi/ml medium. The preparations were: a) yolk sac opened, placenta and blood circulation intact; (b) yolk sac and amnion opened, placenta and blood circulation intact (Day 10 embryos only); c) placenta, yolk sac, and amnion removed (embryo “naked”); d) naked embryos cut randomly into pieces (Day 10 mebryos only). After incubation whole embryos or certain parts (tail, liver, rest body) were investigated by determining the radioactivity taken up by the protein. The results are given in dpm per mg protein per embryo. Radioactivity of proteins was about 3 times higher in naked mebryos than in embryos left in their yolk sacs. This was true for all three stages investigated. However, the degree of radioactivity in the various parts of naked embryos differed by a factor of 15, whereas radioactivity was evenly distributed in embryos incubated in their yolk sacs. Therefore, embryos prepared according to the first methods (see above) fulfilled the conditions required at the best. This work was supported by grants from the Deutsche Forschungsgemeinschaft awarded to the project K1 237/3-2 (Systematic Analysis of Cell Proteins).     

DIFFERENCES IN THE SUBCELLULAR AND SUBSYNAPTOSOMAL DISTRIBUTION OF THE PUTATIVE ENDOPLASMIC RETICULUM MARKERS, NADPH-CYTOCHROME c REDUCTASE, ESTRONE SULFATE SULFOHYDROLASE AND CDP-CHOLINE DIACYLGLYCEROL CHOLINEPHOSPHOTRANSFERASE IN RAT BRAIN

Abstract— A comprehensive study has been undertaken on the subcellular and subsynaptosomal distribution of a number of markers for subcellular organelles in preparations from rat brain. Although the activity of most enzymatic markers was decreased by freezing and storage at - 70^oC, no significant changes were noted in the distribution of these activities. This demonstrates that contamination of brain fractions by subcellular organelles can be accurately assessed after freezing and thawing. A marked discrepancy was noted between the distribution of three putative markers for endoplasmic reticulum. CDP-choline-diacylglycerol cholinephosphotransferase (EC 2.7.8.1) activity was mainly limited to the microsomal fraction and was present to a lesser extent in the synaptosomal fraction than the other putative markers for endoplasmic reticulum. Estrone sulfate sulfohydrolase (EC 3.1.6.2) activity demonstrated a bimodal distribution between the crude nuclear and microsomal fractions. However, considerable activity was associated with the synaptosomal fraction. NADPH-cytochrome c reductase (EC 2.3.1.15) activity sedimented in the microsomal and the synaptosomal fractions. Calculations based on the relative specific activities of the microsomal and synaptic plasma membrane fraction indicated that the contamination of the synaptic plasma membranes by endoplasmic reticulum was 44.5% (NADPH-cytochrome c reductase), 38.0% (estrone sulfatase) and 9.0% (cholinephosphotransferase). Since it is believed that virtually all of the synthesis of phosphatidylcholine by cholinephosphotransferase occurs in the neuronal and glial cell bodies, it was concluded that cholinephosphotransferase is a satisfactory marker for the endoplasmic reticulum derived from these sources. The results suggest that NADPH-cytochrome c reductase and estrone sulfatase may be present in the smooth endoplasmic reticulum system responsible for the fast transport of macromolecules along the axon to the nerve endings as well as in the endoplasmic reticulum of the cell bodies. The possible relation between that portion of the smooth endoplasmic reticulum involved in fast axonal transport and the GERL (Golgi, Endoplasmic Reticulum, Lysosomes) complex discovered by Novikoff and his coworkers (Novikoff , 1976) is discussed.     

Delta5-3beta-hydroxysteroid dehydrogenase and estradiol-17beta-hydroxysteroid dehydrogenase activity in preimplantation hamster embryos

Preimplantation golden hamster (Mesocricetus auratus) embryos were recovered on days 1 (= day of finding spermatozoa in the vagina) through 4 of pregnancy. Postimplantation embryos were studied in sectioned gestation sacs excised on days 5 and 6. Δ⁵-3β-Hydroxysteroid dehydrogenase (3β-HSD) activity in embryos was determined histochemically. There was no enzyme activity on days 1 and 2. Weak activity was first observed at 08:00–09:00 hr on day 3, the activity then increased, peaked at 01:00–03:00 hr on day 4, considerably declined by 08:00–09:00 hr (day 4), and was absent on days 5 and 6. These results suggest that the preimplantation embryos synthesize steroid hormones. It was previously hypothesized (Dickmann and Dey, 1973, Dickmann and Dey, 1974) that, hormones synthesized by the preimplantation rat embryo participate in the regulation of morula to blastocyst transformation and implantation of the blastocyst. This hypothesis is applicable to the hamster.In addition to 3βHSD, estradiol-17β-hydroxysteroid dehydrogenase activity was observed in day 3 embryos, suggesting that the embryo synthesizes estrogen.     

Contribution of calumin to embryogenesis through participation in the endoplasmic reticulum-associated degradation activity

Calumin is an endoplasmic reticulum (ER)-transmembrane protein, and little is known about its physiological roles. Here we showed that calumin homozygous mutant embryos die at embryonic days (E) 10.5−11.5. At mid-gestation, calumin was expressed predominantly in the yolk sac. Apoptosis was enhanced in calumin homozygous mutant yolk sacs at E9.5, pointing to a possible link to the embryonic lethality. Calumin co-immunoprecipitated with ERAD components such as p97, BIP, derlin-1, derlin-2 and VIMP, suggesting its involvement in ERAD. Indeed, calumin knockdown in HEK 293 cells resulted in ERAD being less efficient, as demonstrated by attenuation in both degradations of a misfolded α1-antitrypsin variant and the ER-to-cytosol dislocation of cholera toxin A1 subunit. In calumin homozygous mutant yolk sac endoderm cells, ER stress-associated alterations were observed, including lipid droplet accumulation, fragmentation of the ER and dissociation of ribosomes from the ER. In this context, the ER-overload response, assumed to be cytoprotective, was also triggered in the mutant endoderm cells, but seemed to fully counteract the excessive ER stress generated due to defective ERAD. Taken together, our findings suggested that calumin serves to maintain the yolk sac integrity through participation in the ERAD activity, contributing to embryonic development.     

Vitamins C and E improve rat embryonic antioxidant defense mechanism in diabetic culture medium.

BACKGROUND: Diabetes teratogenicity seems to be related to embryonic oxidative stress and the extent of the embryonic damage can apparently be reduced by antioxidants. We have studied the mechanism by which antioxidants, such as vitamins C and E, reduce diabetes-induced embryonic damage. We therefore compared the antioxidant capacity of 10.5-day-old rat embryos and their yolk sacs cultured for 28h in diabetic culture medium with or without vitamins C and E. METHODS: The embryos were cultured in 90% rat serum to which 2mg/ml glucose, 2mg/ml beta hydroxy butyrate (BHOB) and 10 microg/ml of acetoacetate were added. Rat embryos were also cultured in a diabetic medium with 25 microg/ml of vitamin E and 50 microg/ml of vitamin C. Control embryos were cultured in normal rat serum with or without vitamins C and E. RESULTS: Decreased activity of Cu/Zn superoxide dismutase (SOD) and of catalase (CAT) in the "diabetic" embryos and their yolk sacs, and reduced concentrations of low molecular weight antioxidant (LMWA) were found. Under these conditions we also found a decrease in vitamin C and vitamin E concentrations in the embryos, as measured by HPLC. In situ hybridization for SOD mRNA showed a marked reduction of SOD mRNA in the brain, spinal cord, heart and liver of embryos cultured in diabetic medium in comparison to controls. Following the addition of vitamins C and E to the diabetic culture medium, SOD and CAT activity, the concentrations of LMWA, the levels of vitamin C and E and the expression of SOD mRNA in the embryos and yolk sacs returned to normal. CONCLUSIONS: Diabetic metabolic factors seem to have a direct effect on embryonic SOD gene and perhaps genes of other antioxidant enzymes, reducing embryonic endogenous antioxidant defense mechanism. This in turn may cause a depletion of the LMWA, such as vitamins C and E. The addition of these vitamins normalizes the embryonic antioxidant defense mechanism, reducing the damage caused by the diabetic environment.     

Early development of marine catfishes (Ariidae): from mouth brooding to the release of juveniles in nursery habitats

The development and allometric growth patterns of the ariid catfishes Cathorops spixii and Cathorops agassizii were studied from neurula embryos to juveniles. The ontogenetic sequence revealed that prior to hatching, embryos of both species are well developed, and their axial and appendicular skeletons are well ossified. Embryos of both species grow slowly longitudinally, but positively allometric growth (growth coefficient, β₁ > 1) was observed in head width and eye diameter. It is hypothesized that these growth patterns might be related to functional priorities for the development of sensory organs, such as the inner ears (otoliths), the Weberian apparatus, eyes and nostrils, during the embryonic period. The first appearance of vertebrae and otoliths, as well as the ossification of otoliths and the Weberian apparatus, occur earlier in embryos of C. agassizii than in embryos of C. spixii. After hatching, mouth‐brooded free embryos of both species grow isometrically. Negatively allometric growth was observed in head width and eye diameter during the yolk‐sac period, which is expected given that the sensory organs are already formed. Free embryos of C. agassizii are distinguishable from those of C. spixii by their larger eyes, longer snouts, longer heads and heavier yolk sacs. The end of the yolk‐sac period is characterized by a direct change from free embryo to juvenile, without a true larval period. The juveniles of the two species can also be distinguished from each other by the larger eyes of C. agassizii compared with C. spixii, as in adult fishes.     

Mouse Embryonic Development in a Serum-free Whole Embryo Culture System

Mid-gestation stage mouse embryos were cultured utilizing a serum-free culture medium prepared from commercially available stem cell media supplements in an oxygenated rolling bottle culture system. Mouse embryos at E10.5 were carefully isolated from the uterus with intact yolk sac and in a process involving precise surgical maneuver the embryos were gently exteriorized from the yolk sac while maintaining the vascular continuity of the embryo with the yolk sac. Compared to embryos prepared with intact yolk sac or with the yolk sac removed, these embryos exhibited superior survival rate and developmental progression when cultured under similar conditions. We show that these mouse embryos, when cultured in a defined medium in an atmosphere of 95% O₂ / 5% CO₂ in a rolling bottle culture apparatus at 37 °​C for 16-40 hr, exhibit morphological growth and development comparable to the embryos developing in utero. We believe this method will be useful for investigators needing to utilize whole embryo culture to study signaling interactions important in embryonic organogenesis.     

Adsorptive pinocytosis of polycationic copolymers of vinylpyrrolidone with vinylamine by rat yolk sac and rat peritoneal macrophage

Polycationic copolymers of vinylpyrrolidone and vinylamine (10:0.77) were prepared, and ¹²⁵I-labelled with either Bolton-Hunter reagent or methyl 3,5-di-[¹²⁵I]iodohydroxybenzimidate. The rate of pinocytic capture of the copolymer was compared with that of ¹²⁵I-labelled polyvinylpyrrolidone, using rat visceral yolk sacs and rat macrophages cultured in vitro as test systems. Whereas polyvinylpyrrolidone was captured entirely by non-adsorptive pinocytosis, the cationic derivative was captured more efficiently, probably because it adsorbs to the cell surface. Copolymer of M_r 120 000 was internalized by macrophages somewhat more rapidly than copolymer of M_r 46 000, but was excluded from the yolk sac.     

Accumulation of calcium and phosphorus in pigeon (Columba livia) embryos

Summary Calcium and phosphorus were measured in the yolk and albumen of fertile pigeon (Columba livia) eggs incubated for 0–17 days, and in embryos and hatchlings. Shell provided most of the calcium for skeletal mineralization of the embryos, whereas phosphorus was derived from the yolk and albumen. Mobilization of calcium from the shell to the embryo commenced at approximately day 11 of incubation, accumulating both in the embryo and the yolk sac. There was 1.4 times more calcium in squab yolk sacs than that contained in newly laid egg yolks. The results suggest that whereas general patterns of calcium and phosphorus accumulation during embryogenesis in altricial birds closely resemble those of precocial birds, calcium mobilization from the shell begins later, proceeds at a slower rate and results in a less mineralized hatchling.CIDA/NSERC Visiting Research Associate Permanent address: Department of Animal Science, University of Peradeniya, Peradeniya, Sri Lanka     

Evidence that the rb Locus Alters the Starch Content of Developing Pea Embryos through an Effect on ADP Glucose Pyrophosphorylase

The aim of this work was to discover whether the rb locus of peas (Pisum sativum L.) affects seed starch content through action on an enzyme of starch synthesis in the developing embryo. The phenotypic effects of this locus are like those of the better characterised, unlinked r locus, which affects seed starch content through action on starch-branching enzyme. Embryos recessive at one or both of these loci (RRrbrb, rrRbRb, rrrbrb) have lower starch contents from an early stage of development than embryos dominant at these loci (RRRbRb). Maximum catalytic activities of enzymes of the pathway from sucrose to starch (sucrose synthase EC 2.4.1.13, UDP glucose pyrophosphorylase EC 2.7.7.9, ADP glucose pyrophosphorylase EC 2.7.7.27, ADP glucose-starch synthase EC 2.4.1.21, starch-branching enzyme EC 2.4.1.18) were compared in developing embryos of three lines of rbrb peas and four lines of RbRb peas. The only consistent difference between the two sorts of embryo was in the activity of ADP glucose pyrophosphorylase, which was at least tenfold lower in rbrb than in RbRb embryos. The activity in rbrb embryos was in most cases less than the estimated rate of starch synthesis of RRRbRb embryos. We conclude that the effect of the rb locus on the starch content of pea seeds is mediated through an alteration in the activity of ADP glucose pyrophosphorylase in the developing embryo.     

The effects of high-sucrose diets and of maternal diabetes on the ultrastructure of the visceral yolk sac endoderm in rat embryos developing in vivo and in vitro

The ultrastructure of the visceral yolk sac endoderm of in vivo developing 9- to 13-day-old embryos from 2 diabetic rat models (streptozotocin diabetes and Cohen--genetically determined--diabetes) and from nondiabetic rats fed high sucrose diets have been studied. This was compared to yolk sacs from 9.5-day-old embryos cultured for 48 h in sera from diabetic and nondiabetic rats fed a high-sucrose diet. Light-microscopic, TEM and SEM studies showed that the pathological cellular changes in the visceral yolk sac endoderm from diabetic rats were first observed on day 9 and were most severe among 11-day-old embryos. In vitro culture of control rat embryos in serum from experimental animals induced a reduction in the number of microvilli, of vacuolar intracellular inclusions and an increase in the number of degenerated endodermal cells. SEM studies showed that in addition to disappearance of microvilli, the majority of cells were collapsed and had degenerated cell membranes. Culture of embryos from diabetic animals in control serum only slightly reversed the pathological changes in the visceral yolk sac endoderm. A good correlation exists between the rate of embryonic malformations in diabetic rats and an index of endodermal-cell damage in the visceral yolk sac.