Biomedical subjects

E W Carney

Publications and source records attributed to E W Carney.

At least 37 records · Page 2Linked to original sources

PubMed1999

Ethylene glycol developmental toxicity: unraveling the roles of glycolic acid and metabolic acidosis.

This study sought to determine the relative roles of glycolic acid (GA), a toxicologically important metabolite of ethylene glycol (EG), and metabolic acidosis in causing developmental toxicity in Sprague-Dawley rats. To tease apart these two interrelated factors, we developed an experimental approach in which high blood glycolate levels could be achieved, in either the presence or absence of metabolic acidosis. Initially, rats previously implanted with a carotid artery cannula were given, on gestation day (gd) 10, 40.3 mmol/kg (2500 mg/kg) of EG via gavage, 8.5 mmol/kg (650 mg/kg) of GA via gavage, 8.5 mmol/kg (833 mg/kg) of sodium glycolate (NaG; pH 7.4) via subcutaneous (sc) injection, or distilled water via gavage (control). Peak serum glycolate was nearly identical (8.4-8.8 mM) in the EG, GA, and NaG groups and, as expected, EG and GA caused a metabolic acidosis, but acid base balance was normal with NaG. Subsequently, these treatments were given on gd 6-15 to groups of 25 time-mated rats, followed by fetal evaluation on gd 21. EG and GA decreased fetal body weights and caused a similar spectrum of developmental effects, including numerous axial skeleton malformations. NaG treatment also caused slight decreases in fetal body weight, increases in skeletal variations, and totally malformed fetuses. These results indicate that glycolate, in the absence of metabolic acidosis, can cause the most sensitive of EG's developmental effects, whereas metabolic acidosis appears to interact with glycolate at very high doses to markedly enhance teratogenesis. These results support previous studies, which indicated that glycolate is the proximate developmental toxicant for EG, and that GA toxicokinetic parameters can be used to define a quantitative, physiologically based threshold for EG-induced developmental effects.

Acidosis↗

PubMed1999

Assessment of adult and neonatal reproductive parameters in Sprague-Dawley rats exposed to propylene glycol monomethyl ether vapors for two generations.

This study evaluated propylene glycol monomethyl ether (PGME) in a rat 2-generation reproduction study, which included non-traditional study end points, such as sperm count and motility, developmental landmarks, estrous cyclicity, and weanling organ weights. Groups of 30 male and 30 female Sprague-Dawley rats (6-weeks-old) were exposed to 0, 300, 1000, or 3000 ppm of PGME vapors via inhalation for 6 hours/day, 5 days/week prior to mating, and 6 hours/day, 7 days/week during mating, gestation, and lactation, for 2 generations. These concentrations corresponded to estimated oral equivalent doses of 0, 396, 1325, or 3974 mg/kg/day. At 3000 ppm, toxicity in the P1 and P2 adults was marked, as evidenced by sedation during and after exposure, and mean body weights which were as much as 21% lower than controls. This marked parental toxicity was accompanied by lengthened estrous cycles, decreased fertility, decreased ovary weights, and histologic ovarian atrophy in maternal rats. In the offspring from these dams, decreased body weights, reduced survival and litter size, slight delays in puberty onset, and histologic changes in liver and thymus in the F1 and F2 offspring were observed. The nature of the reproductive/neonatal effects and their close individual animal correlation with decreased maternal body weights suggested that these effects were secondary to general toxicity and/or nutritional stress. No such reproductive/neonatal effects were observed at 1000 ppm, a concentration which caused less marked, but significant body weight effects without sedation. There were no treatment-related effects of any kind noted at 300 ppm of PGME. Therefore, the no-observable-effect level (NOEL) for reproductive/neonatal effects was 1000 ppm, and that for parental toxicity was 300 ppm.

Administration, Inhalation↗

PubMed1999

Evaluation of the EDSTAC female pubertal assay in CD rats using 17beta-estradiol, steroid biosynthesis inhibitors, and a thyroid inhibitor.

The Endocrine Disrupter Screening and Testing Advisory Committee has recommended the female pubertal onset assay as a Tier I test to detect potential endocrine-disrupting chemicals (EDs). We evaluated this assay's ability to detect EDs acting through various mechanisms. In two similar experiments, weanling female rats were dosed for 20 days by gavage with vehicle (0.5% methocel) or the following test compounds (mg/kg/day): 17beta-estradiol (E2; 0.1, 2, or 4), ketoconazole (KETO; 24, 50, or 100), finasteride (FIN; 20), testolactone (TL; 220), fadrozole (FAD; 0.6, 1.2, or 6.0) or 6-propylthiouracil (PTU; 240). In vehicle-treated females, mean age at pubertal onset, as evidenced by vaginal opening (VO), varied interexperimentally from 32.3+/-1.6 days to 33.5+/-1.8 days. At 0.1 mg/kg E2, age at VO was reduced slightly to 31.0+/-1.6 days, but not significantly (alpha=0.05). Higher E2 doses (2.0 and 4.0) reduced age at VO to 28 days. KETO delayed VO, but this delay was significant only at 100 mg/kg (39.7+/-2.4 days). FIN and TL had no effect on age at pubertal onset; however, FAD significantly delayed VO. PTU delayed VO to 34.2+/-1.1 days and altered thyroid weight, histology, and hormone levels. With each compound, significant changes in age at VO were accompanied by decreased uterine or ovarian weights. Thus, although this assay did not detect TL or lower doses of E2 (0.1 mg/kg) or KETO (< or = 50 mg/kg), it was capable of detecting EDs operating through a variety of mechanisms.

5-alpha Reductase Inhibitors↗

PubMed1998

Dominant lethal test with rats exposed to 1,3-dichloropropene by inhalation.

The potential of 1,3-dichloropropene (1,3-DCP) to induce dominant lethal mutations in the germ cells of male CD rats following inhalation exposure was investigated. Groups of 11-week-old males (30 animals/group) were exposed to 1,3-DCP vapors by inhalation at targeted concentrations of 0 (negative control), 10, 60, and 150 ppm for 10 weeks (6 hr/day, 7 days/week). An additional group of 30 males (designated the pairfed group) was kept on dietary restriction for 10 weeks. This group served as a control for any effects of decreased feed consumption and the associated body weight loss on the dominant lethal indices in the males exposed to 1,3-DCP. At the termination of the exposures, each male was cohoused with naive adult virgin CD females for two consecutive mating trials (1 week/trial, 2 females/male). Females were necropsied 13 days after the conclusion of each weekly mating trial and the number of corpora lutea, live implantations, and resorptions were determined. There were no statistically significant increases in either the pre- or postimplantation embryonic/fetal loss in females mated with 1,3-DCP-exposed males relative to controls at any weekly mating period. Based on these results, it can be concluded that 1,3-DCP is not mutagenic to the male germ cells of CD rats at exposure levels < or = 150 ppm, the highest concentration tested.

Allyl Compounds↗

PubMed1996

Identification of proximate toxicant for ethylene glycol developmental toxicity using rat whole embryo culture.

The effects of ethylene glycol (EG) and its metabolite, glycolic acid (GA), were compared by culturing day 10.5 rat conceptuses for 46 h in media containing 0.5, 2.5, 12.5, 25 or 50 mM EG or GA. EG up to 50 mM was essentially without effect, whereas > or = 12.5 mM GA inhibited embryo growth and development. Craniofacial dysmorphogenesis was observed in 70% of the 12.5 mM GA embryos (0% in controls). To determine if GA toxicity in vitro was an indirect effect of medium acidification, embryos were cultured in 12.5 mM GA (pH 6.7), 12.5 mM sodium glycolate (pH 7.4), or in control medium (pH 7.4 or 6.7). The percentage of dysmorphic embryos was 67% for the 12.5 mM GA (pH 6.7) group, 58% for the sodium glycolate (pH 7.4) group, 8% in the pH 6.7 controls, and 0% in the pH 7.4 controls. These results suggest that GA, not parent EG, is the active toxicant for EG-induced developmental toxicity and that acidification of culture medium pH plays only a minor role in GA's effects in vitro. The identification of GA as the active toxicant is important for the risk assessment of EG because GA exhibits dose-rate-dependent, nonlinear kinetics in vivo.

Animals↗

PubMed1994

Localization of two angiogenic growth factors (PDECGF and VEGF) in human placentae throughout gestation.

The spatiotemporal distribution of two angiogenic growth factors, platelet-derived endothelial cell growth factor (PDECGF) and vascular endothelial growth factor (VEGF) were determined using immunohistochemistry on sections of human placentae from each trimester of pregnancy. In the first trimester PDECGF was detected in trophoblast and in a band in the centre of the villous core. During gestation staining spread throughout the stroma but began to weaken in trophoblast until, by term, it was found only in stroma and in some endothelial cells. VEGF was detected exclusively in cytotrophoblast during the first trimester and then in syncytiotrophoblast throughout the remainder of pregnancy. Western blot analysis revealed that PDECGF antisera bound to three bands approximately 27, 47 and 94 kDa. The lowest band was not detected in platelet lysate and may represent an alternatively processed form of this peptide in placenta. VEGF antisera bound strongly to bands approximately 36, 46, 54, 56 and 64 kDa. The intensity of most bands increased between the first and second trimesters, consistent with an increased level of angiogenesis as the placenta develops. The presence of both factors in trophoblast in early pregnancy may be indicative of the trophoblast playing an active role in influencing the development of the villous vascular network.

Blotting, Western↗

PubMed1994

Female reproductive and developmental toxicology: overview and current approaches.

In recent years, concern about possible female reproductive and developmental toxicity due to environmental contaminants, such as PCBs, has been growing. Because this area of toxicology had not been emphasized prior to this time, there are many gaps in current knowledge about female developmental and reproductive toxicology and only a limited number of validated tests to assay effects of toxicants on various parts of the reproductive and developmental cycle. This article reviews the current state of knowledge on this topic and also explores a variety of techniques for assessing female reproductive and developmental toxicity. These include an assay of the state of intercellular communication among the embryo, fetus and placenta; protocols for assessing toxicity in early pregnancy; and techniques for evaluating the role of glutathione in protecting the conceptus from xenobiotics.

Embryonic and Fetal Development↗

PubMed1993

Progressive expression of trophoblast-specific genes during formation of mouse trophoblast giant cells in vitro.

The expression of a battery of trophoblast-specific mRNAs was studied during trophectoderm development in vivo and in vitro to assess the use of these mRNAs as markers of trophoblast differentiation and to examine lineage relationships between various trophectoderm derivatives. In situ hybridization of sectioned day 6.5-18.5 mouse embryos localized mRNAs for mouse placental lactogens I and II and mouse proliferin (PLF) to trophoblast giant cells and proliferin-related protein mRNA to the spongiotrophoblast and giant cell layers. A fifth marker, cDNA 4311, was found only in spongiotrophoblast. Day 3.5 blastocyst outgrowths and day 7.5 diploid extraembryonic ectoderm (EX) and ectoplacental cone (EPC) were then cultured to produce polyploid giant cells in vitro. Cultures were processed for in situ hybridization after 2, 4, or 6 days. EX and EPC both formed secondary giant cells, which expressed all markers in the same sequence as was observed in vivo, and primary giant cells in blastocyst outgrowths expressed the early giant cell markers PLF and PL-I on days 4 and 6 of culture. EPC progressed through the sequence 2 days ahead of EX, indicating commitment of EPC to giant cell formation. These results suggest that EX, EPC, and primary and secondary giant cells all share in a common pathway of differentiation and that the highly ordered sequence of gene expression characteristic of this pathway occurs similarly in vivo and in vitro.

Animals↗

PubMed1992

Increasing carbon dioxide from five percent to ten percent improves rabbit blastocyst development from cultured zygotes.

One-cell rabbit zygotes were cultured at 39 degrees C in basal synthetic medium II (BSM-II) with 5%, 10%, or 15% CO2 and humidified air to determine the effect of CO2 concentration on development in vitro. After 4 days in culture, 37% of the embryos grown in 10% or 15% CO2 had reached the hatching blastocyst stage, but only 10% of the embryos were hatching when cultured under 5% CO2 (P = 0.01). Over all blastocysts, cell numbers were 207, 246, and 205 for the 5%, 10%, and 15% CO2 treatments, respectively. In a second experiment to determine if there was a beneficial effect, particularly at the blastocyst stage, of a higher concentration of CO2, embryos were cultured 4 days in either 5% or 10% CO2 or for 2 days in 5% CO2 followed by 2 days in 10% CO2. The numbers of blastomeres per embryo and embryo diameter were greater (P < 0.05) in embryos cultured continuously in 10% CO2 or in 10% CO2 only during days 3 and 4 of culture than in embryos cultured continuously in 5% CO2. In a third experiment, one-cell rabbit zygotes were cultured with 5% or 10% CO2 in a defined, protein-free medium consisting of 1:1 RPMI 1640 and Dulbecco's modified Eagle's medium. The proportion of embryos hatching and cell counts were significantly greater (P < 0.01) when cultured in the presence of 10% CO2. These data indicate that a 10% CO2 atmosphere exerts a beneficial effect on the development of zygotes into expanding and hatching rabbit blastocysts in vitro.

Animals↗

PubMed1992

The role of nutrients, peptide growth factors and co-culture cells in development of preimplantation embryos in vitro.

Our knowledge of the control of preimplantation embryo development and growth is deficient in many aspects as is evidenced by the great difficulty there is in growing embryos of many species in vitro while maintaining viability. This review discusses recent findings on the roles of nutrients, peptide growth factors and co-culture cells in embryo growth and development in vitro.

Journal Article↗

PubMed1991

Improved development of rabbit one-cell embryos to the hatching blastocyst stage by culture in a defined, protein-free culture medium.

In Exp. 1, Medium 199 and Medium RD (RPMI-1640 and Dulbecco's MEM, 1:1 v/v) were compared in a 2 x 2 factorial design by supplementing each with 15 mg bovine serum albumin (BSA)/ml of 1 mg polyvinyl alcohol (PVA)/ml. All media contained 5 micrograms insulin/ml, 5 micrograms transferrin/ml, 5 ng selenium/ml (ITS), and 10 ng epidermal growth factor (EGF)/ml. One-cell embryos were cultured at 39 degrees C with 5% CO2 in air for 65 h and then stained with Hoechst 33342 to determine blastomere number. Embryos in Medium 199 developed poorly (P less than 0.001) when PVA was used instead of BSA (30 vs 76 cells/embryo), but developed rapidly in Medium RD with PVA or BSA (118 and 121 cells). Similar results were obtained in Exp. 2 in BSA- and PVA-free medium. In Exp. 3, the development of 1-cell embryos after 65 h in unsupplemented (protein-free) Medium RD (68% blastocysts, 117 cells) did not differ (P greater than 0.37) from that obtained using Medium RD with insulin, ITS or EGF alone. Culture in protein-free Medium RD for 96 h resulted in 82% of the 1-cell embryos forming blastocysts and 40% hatching through the zona pellucida. In a preliminary test of viability, 1-cell embryos cultured in this medium for 48 or 65 h and transferred to synchronous recipients resulted in 5/18 (28%) and 3/24 (12%) Day-15 viable fetuses. Cell counts of approximately 120 per blastocyst after culturing 1-cell embryos for 65 h in Medium RD indicated that cell division was more rapid than that obtained with all other media tested previously in this laboratory. This is the first report of rabbit embryo development from the 1-cell to the hatching blastocyst stage in a defined protein-free culture medium.

Animals↗

PubMed1990

Co-culture of rabbit 2-cell embryos with rabbit oviduct epithelial cells and other somatic cells.

Rabbit 2-cell embryos were co-cultured in Basel Synthetic Medium II + 10% fetal bovine serum with one of the following: primary cultures of rabbit oviduct epithelial cells (ROEC), a rabbit kidney epithelioid cell line (RK13), a rabbit epidermal epithelioid cell line (Sf1), or a rabbit skin fibroblast-like cell line (RAB9). Embryos cultured in medium alone served as controls. After 4 d of culture at 39 degrees C in 5% CO2 in air, 77-93% of the rabbit embryos which were co-cultured with somatic cells had reached the blastocyst stage, and 60-76% were hatching through their zonae pellucidae. These percentages, however, were not significantly different (P greater than .05) from those of embryos in medium alone, of which 90% had reached the blastocyst stage and 83% were hatching. Mean intrazonal embryo diameters also did not differ significantly among treatments (239-302 microns). Bovine 1-8-cell embryos were also co-cultured with ROEC. This stimulated 60% of these embryos to develop beyond the so-called "16-cell block" in vitro, whereas 0% of the embryos cultured in medium alone developed past this block. Evaluation of the ROEC cultures by light microscopy, immunocytochemistry, and gel electrophoretic analysis of conditioned medium, together with the positive results with bovine embryos, indicate that the ROEC culture partially simulates oviductal conditions in vivo. Therefore, our results suggest that oviduct epithelial cells may play a less pivotal role in regulating early development in the rabbit than in the cow.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1990

Co-culture of rabbit one-cell embryos with rabbit oviduct epithelial cells.

Rabbit 1-cell embryos were co-cultured with rabbit oviduct epithelial cells (ROEC) to determine if ROEC can enhance embryo development in vitro. Primary ROEC were cultured in serum-free media at 39 degrees C in a 5% CO2:95% air environment. In experiment 1, 1-cell embryos were co-cultured in Ham's F10 with freshly collected or 4-d-old cultures of ROEC seeded in plastic culture wells or on collagen membranes. One-cell embryos cultured without ROEC served as controls. After 65 h in culture, embryos were stained with Hoechst 33342 to determine the number of cells per embryo. Cell numbers were higher (P less than 0.035) in all co-culture treatments when compared to controls. Optimal development was obtained by co-culture with 4-d-old ROEC grown on plastic (P less than 0.003). In experiment 2, Ham's F10, Medium 199, and CZB with glucose medium were compared for their ability to support embryo development in the presence or absence of 4-d-old ROEC grown on plastic. Cell number and the percentage of embryos becoming blastocysts were significantly (P less than 0.002) higher for embryos cultured in Medium 199 compared to the other media tested. In Medium 199, co-culture with ROEC resulted in only a slight, nonsignificant increase in cell number over culture in Medium 199 alone (110 vs. 96 cells). However, the percentage of embryos reaching the blastocyst stage when co-cultured in Medium 199 with ROEC (49%) was nearly twice (P = 0.01) that of embryos in Medium 199 without ROEC (26%). In experiment 3, transfer of embryos cultured in Medium 199 with or without ROEC of 24 or 48 h resulted in no significant differences in posttransfer development. These data indicate a beneficial effect of ROEC on blastogenesis and a salvage effect of ROEC on cell proliferation in embryos grown in a less supportive medium such as Ham's F10.

Animals↗

PubMed1990

Bovine 1-2-cell embryo development using a simple medium in three oviduct epithelial cell coculture systems.

These studies were designed to develop a coculture system using a simple medium to promote development of 1-cell bovine embryos through the 8-16-cell stage to morula and blastocyst stages. Monolayers for coculture were prepared from bovine oviduct epithelial cells (BOEC). In vivo-fertilized 1-2-cell embryos and ova (384) were surgically collected from superovulated cows. In Experiment 1, embryos cocultured in a simple glucose-free and serum-free medium (CZB) developed with superior scores of embryo quality than embryos cocultured in Ham's F-10 with serum, and a greater percentage developed past 8-16 cells than embryos cocultured in CMRL-1066 with serum (p less than 0.05). In Experiment 2, embryos cocultured with fresh BOEC monolayers averaged more (p less than 0.05) cells than did embryos in coculture with frozen-thawed BOEC monolayers or in BOEC-conditioned medium. Without glucose in the simple medium for the first 48 h of culture, more embryos blastulated (p less than 0.01) by Day 5.5 of culture (Day 6.5 of donor's estrous cycle) than embryos in the same medium with glucose present throughout. In Experiment 3, more embryos tended to hatch in BOEC coculture (p less than 0.10) than in conditioned medium. These results show that a chemically simple medium with fresh BOEC monolayers can provide a significant benefit for coculture of early bovine embryos.

Animals↗

PubMed1990

Effects of superovulation, embryo recovery, culture system and embryo transfer on development of rabbit embryos in vivo and in vitro.

Uninterrupted development of rabbit embryos in vivo was studied in 7 superovulated and 7 normally ovulating (GnRH-treated) does, while another 7 does were superovulated and 1-cell embryos were collected from them at 19 h after LH to compare development in vivo and in vitro. Embryos from the last group were either cultured in the presence or absence of rabbit oviduct epithelial cells for 65 h in Medium 199, or were immediately transferred to recipients. At 84 h after LH or GnRH, blastomere number, embryo volume and stage of development were assessed for all embryos. Intrazonal embryo volumes were significantly reduced in embryos recovered from superovulated donors. Superovulation also had a negative effect on embryo cell numbers. However, this reduction was more severe in embryos remaining in vivo in superovulated donors until 84 h after LH than it was in embryos transferred to nonsuperovulated recipients at the 1-cell stage (19 h after LH). The embryo recovery procedure apparently caused little harm to the embryos, except that the mucin layer on flushed and immediately transferred embryos was significantly thinner than that of embryos residing continuously in vivo. Co-culture with rabbit oviduct epithelial cells resulted in improved development in vitro, but this development was still significantly retarded compared with embryos developing in vivo.

Animals↗

PubMed1988

Factors limiting reproductive efficiency in selected laboratory animals.

The mouse, rat, hamster, and rabbit have had remarkable success in surviving in the wild, which attests to their high reproductive capability. In fact, Parkes refers to mice conceiving at each postpartum estrus having the potential for 13 litters per year. Paufler et al. used GnRH to repeatedly cause ovulation in 27 rabbits a few days after parturition, resulting in an average pregnancy rate of 71.5%, 7.0 young born, 5.8 young weaned, and 50 young per doe per year. All four species produce an excess of spermatozoa relative to the requirements for fertilization. The rabbit is suitable for semen collection, artificial insemination, and most of the techniques one might wish to model for reproductive studies in both males and females. A major limitation to reproduction in these species is that reproductive capacity fails long before the finite oocyte population formed prenatally is depleted. The uterus of the aged female appears to be the major cause of reproductive failure, as fertilized eggs replaced in such a uterus usually soon deteriorate. With in vitro techniques, much is still to be learned about harvesting, maturing and fertilizing oocytes and identifying those most likely to result in formation of a healthy neonate.

Animals↗

PubMed1987

Stimulatory and inhibitory effects of amino acids on the development of hamster eight-cell embryos in vitro.

Amino acids were added to a simple medium (TALP) in an effort to enhance the development of hamster eight-cell embryos in vitro. The addition of 1.0 mM glutamine to TALP medium markedly increased the percentage of embryos reaching the blastocyst stage. The addition of groups of 4 or 20 amino acids to TALP + glutamine medium produced differential effects, some groups inhibiting development while others stimulated it. The beneficial effect of glutamine was on the eight-cell to morula transition as well as on blastocyst formation, while the effects of the other 20 amino acids studied were only on blastocyst development. This study represents further progress made in identifying the culture requirements for growth of hamster preimplantation embryos in vitro.

Amino Acids↗

PubMed1987

Regulation of hamster embryo development in vitro by carbon dioxide.

We have examined the effect of collecting and culturing hamster eight-cell embryos in media containing high levels of bicarbonate and/or CO2 on development in vitro. An approximate doubling in the percentage of embryos developing to the blastocyst stage was observed upon raising the concentration of CO2 in the gas phase from 5% to 10% CO2. Development to the blastocyst stage was not affected by the bicarbonate concentration (6-50 mM), nor by the pH of the medium (6.5-7.4). However, escape of embryos from their zonae pellucidae was pH-dependent (optimum pH 7.1-7.4). We hypothesized that the beneficial effect of high concentrations of CO2 on blastocyst development was due to the action of CO2 as a weak acid in regulating intracellular pH (pHi). To test this hypothesis, eight-cell embryos were cultured under 5% CO2 in media containing various concentrations of organic weak acids (lactic or acetic acids, or the non-metabolizable compound 2,4-dimethyloxazolidine-dione). Embryos cultured in standard medium (TALP) under 5% and 10% CO2 served as low and high controls, respectively. At optimum concentrations, all of the media containing weak acids supported embryo development significantly better than 5% CO2-equilibrated low control medium, and gave a response similar to that obtained with high control medium equilibrated in 10% CO2. These studies demonstrate that culture in a 10% CO2 environment has a marked stimulatory effect on in vitro development of hamster eight-cell embryos and suggest that this effect is due to maintenance of pHi.

Animals↗