• Toxicological Research
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• v.26 no.4
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• pp.275-283
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• 2010

Placenta transfer study in non-human primate (NHP) is one of the crucial components in the assessment of developmental toxicity because of the similarity between NHP and humans. To establish the method to determine placenta transfer in non-human primate, toxicokinetics of valproic acid (VPA), a drug used to treat epilepsy in pregnant women, were determined in pregnant cynomolgus monkeys. After mating, pregnancy-proven females were daily administered with VPA at dose levels of 0, 20, 60 and 180 mg/kg by oral route during the organogenesis period from gestation day (GD) 20 to 50. Concentrations of VPA and its metabolite, 4-ene-VPA, in maternal plasma on GDs 20 and 50, and concentrations of VPA and 4-ene-VPA in placenta, amniotic fluid and fetus on GD 50 were analyzed using LC/MS/MS. Following single oral administration of VPA to pregnant monkeys, concentrations of VPA and 4-ene-VPA were generally quantifiable in the plasma from all treatment groups up to 4-24 hours post-dose, demonstrating that VPA was absorbed and the monkeys were systemically exposed to VPA and 4-ene-VPA. After repeated administration of VPA to the monkeys, VPA was detected in amniotic fluid, placenta and fetus from all treatment groups, demonstrating that VPA was transferred via placenta and the fetus was exposed to VPA, and the exposures were increased with increasing dose. Concentrations of 4-ene-VPA in amniotic fluid and fetus were below the limit of quantification, but small amount of 4-ene-VPA was detected in placenta. In conclusion, pregnant monkeys were exposed to VPA and 4-ene-VPA after oral administration of VPA at dose levels of 20, 60 and 180 mg/kg during the organogenesis period. VPA was transferred via placenta and the fetus was exposed to VPA with dose-dependent exposure. The metabolite, 4-ene VPA, was not detected in both amniotic fluid and fetus, but small amount of 4-ene-VPA was detected in placenta. These results demonstrated that proper procedures to investigate placenta transfer in NHP, such as mating and diagnosis of pregnancy via examining gestational sac with ultrasonography, collection of amniotic fluid, placenta and fetus after Caesarean section followed by adequate bioanalysis and toxicokinetic analysis, were established in this study using cynomolugus monkeys.

• Asian-Australasian Journal of Animal Sciences
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• v.30 no.8
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• pp.1124-1134
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• 2017

Objective: This study explored the physico-chemical properties of late-incubation egg amniotic fluid and a potential in ovo feed (IOF) supplement. Methods: Amniotic fluid was collected from broiler breeders (Ross 308, 51 weeks and Cobb 500, 35 weeks) on day 17 after incubation. A mixture of high-quality soy protein supplement - Hamlet Protein AviStart (HPA) was serially diluted in MilliQ water to obtain solutions ranging from 150 to 9.375 mg/mL. The mixtures were heat-treated (0, 30, 60 minutes) in a waterbath ($80^{\circ}C$) and then centrifuged to obtain supernatants. The amniotic fluid and HPA supernatants were analysed for their physico-chemical properties. Results: Only viscosity and $K^+$ were significantly (p<0.05) different in both strains. Of all essential amino acids, leucine and lysine were in the highest concentration in both strains. The osmolality, viscosity and $pCO_2$ of the supernatants decreased (p<0.05) with decreasing HPA concentration. Heat treatment significantly (p<0.05) affected osmolality, pH, and $pCO_2$, of the supernatants. The interactions between HPA concentration and heat treatment were significant with regards to osmolality (p<0.01), pH (p<0.01), $pCO_2$ (p<0.05), glucose (p<0.05), lactate (p<0.01) and acid-base status (p<0.01) of HPA solutions. The $Ca^{2+}$, $K^+$, glucose, and lactate increased with increasing concentration of HPA solution. The protein content of HPA solutions decreased (p<0.05) with reduced HPA solution concentrations. The supernatant from 150 mg/mL HPA solution was richest in glutamic acid, aspartic acid, arginine and lysine. Amino acids concentrations were reduced (p<0.05) with each serial dilution but increased with longer heating. Conclusion: The values obtained in the primary solution (highest concentration) are close to the profiles of high-protein ingredients. This supplement, as a solution, hence, may be suitable for use as an IOF supplement and should be tested for this potential.

• YAKHAK HOEJI
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• v.41 no.2
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• pp.212-219
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• 1997

Multiple forms of extracellular phospholipase $A_2$ have been detected in human amniotic fluid (HAF). When HAF was subjected to heparin-Sepharose column chromatography, phospholipase $A_2$ activity was detected in both heparin-non binding and binding fraction. The activity of heparin-non binding fraction was further purified by sequential uses of column chromatographies on butyl-Toy-opearl 650M and DEAE-Sephacel. DEAE-Sephacel fraction contained three different phospholipase $A_2$ activities (Peak I, II, III). The molecular weight of DEAE-Sephacel fraction phospholipase $A_2$ determined by SDS-PAGE were about 52KDa (Peak I). Peak II, III required micromolar $Ca^{2+}$ ion for its maximum activity, but Peak I enzyme showed calcium independent phospholipase $A_2$ activity and showed broad range of pH (6.0~10.0) optimum. All these enzymes were not recognized by a monoclonal antibody raised against phospholipase $A_2$ from human synovial fluid. These results suggest that HAF might contain multiple forms of extracellular phospholipase $A_2$, which may neither belong to the 14KDa group II phospholipase $A_2$ family nor cytosolic phospholipase $A_2$.

• Development and Reproduction
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• v.22 no.2
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• pp.199-203
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• 2018

Although trisomy 16 is commonly detected in spontaneous abortions and accounts for over 30% of cases of autosomal trisomy detected after spontaneous abortion, trisomy 16 mosaicism is rarely detected by amniocentesis in the second trimester. Here, we report a case of level III trisomy 16 mosaicism (47,XX,+16[8]/46,XX[31]) diagnosed by cytogenetic analysis of independently cultured amniotic fluid cells. The female baby was delivered at full term with low birth weight and intrauterine growth retardation, and interestingly, her karyotype was normal (46,XX). Given the difficulty in predicting the outcomes of fetuses with this mosaicism, it is recommended to inform the possibility of mosaicisms including this trisomy 16 mosaicism during prenatal genetic diagnosis and genetic counseling for parents.

• Development and Reproduction
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• v.20 no.1
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• pp.63-71
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• 2016

Human embryonic stem cells (hESCs) have been routinely cultured on mouse embryonic fibroblast feeder layers with a medium containing animal materials. For clinical application of hESCs, animal-derived products from the animal feeder cells, animal substrates such as gelatin or Matrigel and animal serum are strictly to be eliminated in the culture system. In this study, we performed that SNUhES32 and H1 were cultured on human amniotic fluid cells (hAFCs) with KO-SR XenoFree and a humanized substrate. All of hESCs were relatively well propagated on hAFCs feeders with xeno-free conditions and they expressed pluripotent stem cell markers, alkaline phosphatase, SSEA-4, TRA1-60, TRA1-81, Oct-4, and Nanog like hESCs cultured on STO or human foreskin fibroblast feeders. In addition, we observed the expression of nonhuman N-glycolylneuraminic acid (Neu5GC) molecules by flow cytometry, which was xenotransplantation components of contamination in hESCs cultured on animal feeder conditions, was not detected in this xeno-free condition. In conclusion, SNUhES32 and H1 could be maintained on hAFCs for humanized culture conditions, therefore, we suggested that new xeno-free conditions for clinical grade hESCs culture will be useful data in future clinical studies.

• Journal of Microbiology and Biotechnology
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• v.20 no.6
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• pp.959-967
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• 2010

Down syndrome (DS) is an abnormality of the 21st chromosome that commonly occurs in children born to older women. Thus, amniotic fluid (AF) is usually collected from such women for prenatal diagnosis. This study analyzed human AF supernatants (AFS) using a mass spectrometric (MS) approach to search for candidate biomarkers of a DS pregnancy. The AFS were collected from older pregnant women at weeks 16-18 of their gestation by amniocentesis for cytogenetic analysis. The AFS from the pregnancies carrying DS (n=4) or chromosomally normal (n=6) fetuses, as revealed by the cytogenetic analysis, were then subjected to global protein profiling based on liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). Affinity chromatography was also applied prior to the LC-ESI-MS/MS to minimize the masking effect of highly abundant albumin and immunoglobulin and thereby increase the diversity of the identified proteins. As a result, at least 30 new AFS proteins were identified and 44 AFS proteins were found to be differentially expressed between the DS and normal cases, where 6 of the proteins were unique to the DS cases and 11 were unique to the chromosomally normal cases. In addition, in the DS cases, 19 AFS proteins were downregulated and 8 were upregulated to varying degrees. A Western blot analysis confirmed the LC-ESI-MS/MS data, indicating that the combined detection of apolipoprotein A-II (apoA-II) and alpha-fetoprotein (AFP) could be a potential tool for diagnosing DS cases.

• The Korean Journal of Physiology
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• v.18 no.1
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• pp.1-8
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• 1984

In order to determine the extent of the placental transfer of Lithium ion, pregnant rabbits at $27{\sim}29$ days of gestation, which has hemochorial placenta similar to the human placenta, received 2 mM/Kg of $Li^+$ in the form of LiCl intravenously. Maternal arterial blood, placental sinus blood, fetal blood, amniotic fluid and maternal urine were drawn two hours after the single dose of LiCl. Concentrations of $Li^+$, $Na^+$, $K^+$ and osmolarity were measured in plasma of collected bloods, amniotic fluid and urine. Followings are the results obtained. 1) Evident level of $Li^+$ was detected in fetal blood, although fetal plasma concentration of $Li^+$ found to be almost one third of maternal plasma. 2) Plasma concentration of $Li^+$ in placental sinus blood was higher than that in fetal plasma but lower than that in maternal plasma. It means that downward concentration gradient of $Li^+$ from mother to fetus was still remarkable two hours after the injection. 3) Significant level of $Li^+$ was also detected in amniotic fluid. It seemed likely that $Li^+$, at least in part, excreted by the fetal urinary tract. 4) There were no differences in $Na^+$ and osmolar concentration between fetal and maternal blood. 5) From above results, it was concluded that $Li^+$ may transfer across the placenta but limited passage capacity through placental barrier for $Li^+$ is significant, beacause net transfer assumed to be going on even at two hours, at which time maternal equlibrium has been reached.

• Journal of Genetic Medicine
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• v.12 no.1
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• pp.31-37
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• 2015

Purpose: We investigated the neurogenic potentials of amniotic fluid-derived stem cells (AFSCs) according to the expression levels of stem cell markers and ingredients in the neural induction media. Materials and Methods: Four samples of AFSCs with different levels of Oct-4 and c-kit expression were differentiated neurally, using three kinds of induction media containing retinoic acid (RA) and/or a mixture of 3-isobutyl-1-methylxanthine/indomethacin/insulin (neuromix), and examined by immunofluorescence and reverse transcription-polymerase chain reaction (RT-PCR) for their expression of neurospecific markers. Results: The cells in neuromix-containing media displayed small nuclei and long processes that were characteristic of neural cells. RT-PCR analysis revealed that the number of neural markers showing upregulation was greater in cells cultured in the neuromix-containing media than in those cultured in RA-only medium. Neurospecific gene expression was also higher in Oct-4 and c-kit double-positive cells than in c-kit-low or -negative cells. Conclusion: The stem cell marker c-kit (rather than Oct-4) and the ingredient neuromix (rather than RA) exert greater effects on neurogenesis of AFSCs.

• Clinical and Experimental Pediatrics
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• v.52 no.4
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• pp.429-434
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• 2009

Purpose : Severe aspiration of the amniotic fluid is known to cause fatal respiratory distress in neonates. We conducted this study to investigate the clinical findings of severe amniotic fluid aspiration pneumonia (AFAP) in neonates and the effect of pulmonary surfactant replacement therapy (SRT). Methods : Retrospective analysis of medical records was conducted on 28 patients who received ventilator care due to severe AFAP in a neonatal intensive care unit over a 7-year period (2000-2006). Patients whose amniotic fluid was contaminated with meconium were excluded. Results : A large number of cases were term infants (82.1%) and infants born by caesarean section (85.7%), and the 1- and 5-min Apgar scores of these patients were $6.5{\pm}1.2$ and $7.5{\pm}1.3$, respectively. Soon after birth, the amount of amniotic fluid sucked out from airway below the vocal cord was $16.0{\pm}10.1$ mL. All patients received SRT with a modified bovine-derived surfactant (120 mg/kg/dose), and one dose was administered in most cases (75%). Compared with pre-SRT, the oxygenation index ($8.0{\pm}9.6$ vs. $18.9{\pm}7.3$) according to ventilator care was a significant improvement at 12 h after SRT (P<0.001). Furthermore, most cases showed radiological improvement for aeration at 12 h post-treatment. Many cases (46.4%) had cardiorespiratory complications, but their final outcomes were excellent (survival rate, 96.4%). Conclusion : AFAP may be an important cause of serious respiratory distress in near-term and term infants, and SRT seems to be an effective adjuvant therapy in mechanically ventilated neonates with severe AFAP.

• Korean Journal of Veterinary Research
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• v.31 no.4
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• pp.411-418
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• 1991

This study was carried out to investigate the amino acid and protein concentrations in amniotic fluid and the potency of the teratogenic effect of ethylenethiourea(2-imidazolidinethione, ETU) in the fetuses due to different dose amounts of this compound. The S.P.F. Sprague-Dawley female rats(10 weeks) were used in this study and these animals were divided into four groups; control group(25pregnant female rats), group I (dosed ETU from day 7 to day 17 of gestation at 10mg/kg/day), group II (dosed ETU from day 7 to day 17 of gestation at 30mg/kg/day), group III (dosed ETU from day 7 to day 17 of gestation at 50mg/kg/ day). 250mg/100mg ETU in group I, 750mg/100ml ETU in group II and 1,250mg/100ml ETU in group III were administered 4ml/kg 13.W by oral route. The results obtained were summarized as follows; 1. The anomalies of the external examination werf meningocele in the head, kinky tail, clubfoot and sharp tail.(Meningocele, in group III, significantly increased from control value at p<0.001). 2. The skeletal variations and delayed ossification were Lumbar ribs, asymmetric sternebrae, asymmetric 13th rib and delayed ossification of skull. Asymmetric sternebrae(group III ) was significantly increased from control value at p<0.05 and delayed ossification of skull (group II and III ) were significantly increased from control value at p<0.05 and p<0.01, respectively. 3. The internal soft tissue anomalies were hydroencephaly of 3th lateral ventricle, dilatation of ureter, dilatation of renal pelvis and cleft palate. (Hydroencephaly, 28.1% in group I, 88.3% in group II and 100% in group III ). 4. Protein values in amniotic fluids are not significantly decreased in 10mg/kg group but significantly(p<0.05) decreased in 30mg/kg group and 50mg/kg group from control group. 5. In the levels of amino acid in amniotic fluids, the levels of glntamic acid, iso-lencine, leucine, tyrosine and phenylalanine of 10mg/kg group are significantly decreased from control group. In 50mg/kg group, except for glycine, valine and methionine, all amino acid levels are significantly(p<0.05) decreased from control group.