Katsuto Eguchi

Determination of tryptophan and its metabolites in human plasma and serum by high-performance liquid chromatography with automated sample clean-up system

An automated high-performance liquid chromatographic method that incorporates direct injection of biological samples followed by chromatographic sample clean-up in a precolumn is described for the determination of tryptophan and its metabolites in human plasma and serum. The system gave reproducible data with a coefficient of variation of less than 3% with a sample size of 100 microliters of human plasma. The major tryptophan metabolites found in 100 microliters of human plasma were kynurenine, indolelactic acid, indoleacetic acid, indolepropionic acid, serotonin and 5-hydroxyindoleacetic acid. The level of tryptophan and kynurenine in individuals was constant in comparison with other metabolites. Analysis of samples from normal controls, diabetics, gravida and their foetuses showed a tendency for tryptophan metabolites to be low in maternal plasma.

Urinary and Plasma Epidermal Growth Factor Levels Are Decreased in Neonates with Intrauterine Growth Retardation and in Their Mothers

Epidermal growth factor (EGF) concentrations in urine and plasma samples collected from pregnant women and neonates were measured by RIA. The EGF concentration of the first voided urine was higher in appropriate-for-date (AFD) neonates (33.9 +/- 23.0 ng/mg creatinine) than in those with intrauterine growth retardation (IUGR; 23.5 +/- 7.7 ng/mg creatinine, p less than 0.05) and heavy-for-date (19.8 +/- 5.2 ng/mg creatinine, p less than 0.05) neonates. The urinary EGF concentration of pregnant women showed no marked changes throughout pregnancy. Urinary EGF concentrations of women with AFD fetuses (45.9 +/- 31.2 ng/mg creatinine) did not differ significantly from those of women with diabetes (39.9 +/- 26.8 ng/mg creatinine) or women with multiple fetuses (44.6 +/- 30.6 ng/mg creatinine). However, women with IUGR fetuses showed lower urinary EGF concentrations (13.8 +/- 7.4 ng/mg creatinine, p less than 0.05) than women with AFD fetuses. Maternal and fetal platelet-poor plasma EGF concentrations at delivery were lower in the IUGR group (mother: 2.62 +/- 0.38 ng/ml, fetus: 2.16 +/- 0.07 ng/ml, respectively, p less than 0.05 and p less than 0.005) than in the AFD group (mother: 3.34 +/- 0.64 ng/ml, fetus: 3.24 +/- 0.93 ng/ml). In the IUGR group, the EGF concentration in fetal blood was always lower than that in maternal blood (p less than 0.05), although the AFD groups showed no such difference. These data suggest that EGF levels are closely related to fetal growth.

Comparison of plasma concentrations of arginine vasopressin (AVP) and atrial natriuretic peptide (ANP) in normal and preeclamptic pregnancies.

The study was undertaken to measure both plasma arginine vasopressin (AVP) and atrial natriuretic peptide (ANP) concentrations in normal and preeclamptic pregnancies to elucidate the mechanisms of plasma volume regulation during pregnancy. The mean plasma AVP concentration in healthy pregnant women was much lower than nonpregnant controls; however, the concentration in preeclamptic patients was similar to that in the comparative period of normal pregnancy. Despite volume expansion, the mean plasma concentration of ANP in normal pregnancy showed no changes compared to nonpregnant controls. The concentration of plasma ANP in patients complicated by mild preeclampsia was higher than that in normal pregnant women. The concentration of plasma ANP in those with severe preeclampsia was much higher roughly in proportion to the severity of the disease. It is inferable that the lowered concentration of plasma AVP is involved in plasma volume expansion in normal pregnancy, largely but not in a depletion of plasma volume in preeclampsia. The elevated concentration of plasma ANP appears to be closely related to impaired plasma volume expansion in preeclampsia.

Developmental Changes of Glutamate Dehydrogenase Activity in Rat Liver Mitochondria and Its Enhancement by Branched-Chain Amino Acids

The developmental changes of glutamate dehydrogenase activity in the fetal and neonatal rat liver were investigated, as well as the effects of branched-chain amino acids on this enzyme. Hepatic glutamate dehydrogenase activity showed a marked increase at the end of the fetal period and peaked on the 5th day of neonate at approximately 3 times higher than the adult level. Glutamate dehydrogenase was activated by leucine, isoleucine, and valine in this order when they were added to isolated intact liver mitochondria in vitro. The enhancement of enzyme activity was more marked in fetal rats than in adults. In contrast, when branched-chain amino acids were added after disrupting the mitochondrial membrane by sonication, only leucine slightly activated glutamate dehydrogenase, while isoleucine and valine slightly inhibited its activity. Our findings suggest that glutamate may be actively synthesized in the developing rat liver mitochondria and then transaminated to other nonessential amino acids for protein synthesis, and that increased intramitochondrial branched-chain amino acid concentrations may enhance glutamate dehydrogenase activity. This anabolic metabolism will contribute to the fetal growth and development.

Effects of Epidermal Growth Factor on Ornithine Decarboxylase Activity and DNA Synthesis in Rats during the Perinatal Period

The effects of epidermal growth factor (EGF) on ornithine decarboxylase (ODC) activity and DNA synthesis were studied during the perinatal period in rats. EGF administration to neonatal rats increased ODC activity and DNA synthesis in neonatal rat liver, but not in brain. EGF administration to maternal rats increased ODC activity in maternal rat liver, but not in rat placenta and fetal rat liver. These data suggest that EGF has mitogenic effects on neonatal and maternal rat liver and play some important roles in fetal and neonatal growth.

Developmental Changes of Polyamine Biosynthesis in Rat Liver

To investigate the role of polyamine metabolism in fetal development, we determined the hepatic polyamine content and the activity of enzymes related to polyamine metabolism in fetal, newborn and adult rats. In the fetuses, spermidine was detected at the highest level, followed by spermine and then putrescine. Putrescine and spermidine levels were higher in the fetuses than in adult rats, while the spermine level was significantly lower in the fetuses. The activities of the enzymes investigated were also higher in the fetuses than in adult rats. These findings indicate that polyamine synthesis was enhanced in the fetal liver, and that degradation of spermidine via spermidine N1-acetyltransferase contributes to the increase in putrescine levels. Thus, our study suggests that an increase in putrescine is important for fetal growth and that putrescine per se has a significant role in fetal cellular proliferation.

Alterations of red blood cells' polyamines during pregnancy and neonatal period.

Red blood cell polyamines were measured using high-performance liquid chromatography. Spermidine and spermine continued to increase with pregnancy and retained a high level during 24-35 weeks of gestation, then decreased at term; putrescine, however, kept the low nonpregnant level during pregnancy and puerperium. At delivery, umbilical blood showed a significantly higher polyamine level than maternal blood (p less than 0.005 or p less than 0.001). In the neonatal period, each polyamine showed a slight increase on the 1st day after birth, then spermidine and spermine decreased day by day, but putrescine kept the same level. Alterations of polyamines seemed to have some relation to the erythropoiesis in bone marrow and rapid cell proliferations.