Effect of acute maternal starvation on tyrosine metabolism and protein synthesis in fetal sheep

To determine the effects of acute maternal starvation on intrauterine growth, tyrosine concentration and specific activity values in plasma, intracellular free and protein bound pools were determined in catheterized ovine fetuses following an 8 h continuous infusion of L-[2,3,5,6 3H] or L-[U-14C] tyrosine into the ewe and fetus respectively at 115-125 days of gestation. From the kinetic data the rates of whole body and tissue fractional protein synthesis were calculated. Although placental protein synthesis was not significantly changed as a result of acute maternal starvation, fetal whole body protein synthesis was reduced from 63 g/d/kg in the fed to 25 g/d/kg in the starved condition. There was also a 10 fold reduction in the net placental transfer of tyrosine to the fetus in the starved ewes. In addition, a three fold increase was observed in the quantity of tyrosine used for oxidation by the fetuses of starved ewes, changing from 5.2% of tyrosine net utilization in the fed to 13.7% in the starved condition. Significant reductions in tissue fractional protein synthesis rates were also seen in the liver, brain, lung kidney and GIT tissues from 78, 37, 65, 45 and 71%/d respectively in the fed to 12, 10, 23, 22 and 35%/d in the fetuses of starved ewes. The data indicate that during acute maternal starvation the sheep fetus utilizes more tyrosine for oxidation and less for anabolic purposes which is reflected in a decrease both in whole body and tissue fractional rates of protein synthesis.     

Progesterone secretion and fetal development during prolonged starvation in the pig.

Roles of ovarian progesterone secretion and maternal nutrition in fetal development were investigated in a species that normally experiences considerable embryonic and fetal mortality. Pregnancies were maintained in 81% of Yorkshire pigs during prolonged starvation (e.g., 40 days; 0 kcal/day, water only) in either the middle third (days 30-70) or last third (days 70-110) of gestation compared with 100% in full-fed controls (7,028 kcal/day). In spite of severe maternal deprivation, fetal survival rates averaged 65% in starved dams and 63% in controls; mean number of living fetuses was 9.9 in starved and 9.6 in control dams. Fetal growth was reduced by maternal starvation during the middle third, but not the last third of pregnancy. Placental insufficiency was the primary cause of reduced fetal growth and resulted in abortion in a few of the dams. Progesterone in peripheral serum of dams starved either during middle or late pregnancy was maintained at levels similar (P greater than 0.05) to those in controls. Abortion occurred in starved dams only when serum progesterone concentrations dropped to less than 10 ng/ml within 3 days before loss of conceptuses.     

Cell number and size in selected organs of fetuses of rats malnourished and exposed to nitrofen

The effects of maternal exposure to nitrofen or protein-energy malnutrition on the number and sizes of cells in selected organs of the fetal rat have been studied. Pregnant rats were fed either an adequate (CON) or protein-energy deficient diet (PEM) throughout gestation. Each diet group was divided into two subgroups. One subgroup was gavaged with 25 mg nitrofen/kg body weight on gestational days 7-21 and the other, with corn oil carrier only. Fetal liver, kidneys, intestine, heart, lung, and brain were weighed and assayed for DNA, RNA, and protein. Maternal protein deficiency resulted in a reduction in organ weight and total DNA, RNA, and protein in all six organs. Maternal nitrofen exposure resulted in reduced weight and reduced protein in all organs except the brain. Total DNA and RNA were reduced in intestine, heart, and lung, and total RNA was also reduced in the liver following maternal nitrofen exposure. An interaction between diet and toxin affected lung weight, DNA, RNA, and protein, intestinal total protein, and heart DNA. Protein/DNA ratios were reduced in liver, intestine, and brain in the group fed the inadequate diet and in intestine only following nitrofen exposure. The deficient diet resulted in increased RNA/DNA ratio in the fetal liver and heart and a decreased ratio in the kidney and brain. Nitrofen exposure resulted in a lower RNA/DNA ratio in the liver. The data indicate that maternal protein-energy malnutrition results in smaller organs in the fetuses with fewer cells and containing less protein and RNA.(ABSTRACT TRUNCATED AT 250 WORDS)     

Maternal starvation and progesterone secretion, litter size, and growth in the pig.

The relationships of maternal nutrition and progesterone secretion to prenatal fetal loss and neonatal survival and growth were investigated in domesticated pigs that normally experience 40% fetal loss. Yorkshire pigs were subjected to prolonged starvation (40 days; 0 kcal/day; water only) in either the middle third or last third of pregnancy and then gradually realimented to a full diet and allowed to advance to parturition; controls received a full diet (7,028 kcal/day) throughout gestation. Pregnancies were maintained in 74% of the pigs starved during either middle or late gestation compared to 100% in controls. In dams starved from days 30--70, progesterone levels remained similar to controls, but were maintained at lower levels during realimentation between days 72 and 99. Progesterone levels in dams starved days 50--90 and controls were similar from days 50 to 130. At parturition, litter size in starved dams (9.4) was similar to full-diet controls (8.0) although birth weight and neonatal growth of piglets were reduced markedly from dams starved in middle or late pregnancy. These results indicate that severe maternal nutrient deprivation during the middle or last third of gestation has little effect on ovarian progesterone secretion and is not a major limitation to fetal survival in this litter-bearing species. Maternal nutrient deprivation imposes a significant detrimental effect, however, on birth weight and neonatal growth to 80 days of age.     

Comparison of DNA alkylation, fragmentation, and repair in maternal and fetal tissues of pregnant rats treated with a single dose of ethyl methanesulfonate, ethyl-N-nitrosourea, N-nitrosodiethylamine, and methyl-N-nitrosourea

The occurrence and persistence of DNA damage, as detected by the alkaline elution technique, have been studied in some tissues of both fetal and adult Sprague-Dawley rats (18th day of gestation) after administration of a single equimolar dose (0.5 mmol/kg) of ethyl methanesulfonate (EMS), N-ethyl-N-nitrosourea (ENU), N-nitrosodiethylamine (NDEA), and N-methyl-N-nitrosourea (MNU). EMS, ENU, and MNU, injected intravenously, produced a statistically significant increase of DNA elution rate, which is considered indicative of DNA fragmentation, in both maternal and fetal liver, kidney, and brain. NDEA, introduced by gastric gavage, induced DNA breaks in both liver and kidney of dams, but only in the liver of fetuses. The frequency of DNA lesions was found to vary with the four alkylating agents and in the three organs tested, to exhibit a different time course, and usually to be higher in maternal than in fetal tissues. Results provided by the concomitant determination of DNA binding levels demonstrated a satisfactory correlation with the amounts of DNA fragmentation. In contrast, the values of both these parameters did not show any positive correlation with the different susceptibility of the three organs to tumor induction. In conclusion, these findings suggest that when a compound is not available in radiolabeled form, measurement of DNA fragmentation may represent a useful alternative to the determination of DNA binding level in order to obtain information on the distribution of its reactive species in maternal and fetal tissues.     

Gene expression profile differences in left and right liver lobes from mid-gestation fetal baboons: a cautionary tale

Interpretation of gene array data presents many potential pitfalls in adult tissues. Gene array techniques applied to fetal tissues present additional confounding pitfalls. The left lobe of the fetal liver is supplied with blood containing more oxygen than the right lobe. Since synthetic activity and cell function are oxygen dependent, we hypothesized major differences in mRNA expression between the fetal right and left liver lobes. Our aim was to demonstrate the need to evaluate RNA samples from both lobes. We performed whole genome expression profiling on left and right liver lobe RNA from six 90-day gestation baboon fetuses (term 180 days). Comparing right with left, we found 875 differentially expressed genes – 312 genes were up-regulated and 563 down-regulated. Pathways for damaged DNA binding, endonuclease activity, interleukin binding and receptor activity were up-regulated in right lobe; ontological pathways related to cell signalling, cell organization, cell biogenesis, development, intracellular transport, phospholipid metabolism, protein biosynthesis, protein localization, protein metabolism, translational regulation and vesicle mediated transport were down-regulated in right lobe. Molecular pathway analysis showed down-regulation of pathways related to heat shock protein binding, ion channel and transporter activities, oxygen binding and transporter activities, translation initiation and translation regulator activities. Genes involved in amino acid biosynthesis, lipid biosynthesis and oxygen transport were also differentially expressed. This is the first demonstration of RNA differences between the two lobes of the fetal liver. The data support the argument that a complete interpretation of gene expression in the developing liver requires data from both lobes.     

Ontogeny of serine hydroxymethyltransferase isoenzymes in fetal sheep liver, kidney, and placenta

Serine is an amino acid that is not transported from the placenta to the ovine fetus. Thus, fetal plasma serine levels may be controlled by flux through their relevant biosynthetic pathways. This study was designed to determine, in fetal sheep tissues, the ontogeny of the three key enzymes in the biosynthetic pathway for serine, the cytosolic (c) and mitochondrial (m) isoforms of serine hydroxymethyltransferase (SHMT), phosphoglycerate dehydrogenase (PGD), and phosphoserine aminotransferase (PSAT). PGD and PSAT activity did not vary during gestation in either liver (PSAT, 9.4 +/- 1.3 nmol/min/mg cytosolic protein; and PGD, 76 +/- 10 mU/mg protein) or placenta (PGD, 8.0 +/- 3.6 mU/mg protein). In the liver, cSHMT activity was low early in gestation (0.6 +/- 0.5 nmol/min/mg protein at 45 days), rose in the last one-third of gestation, and peaked in the newborn period (25 +/- 3 nmol/min/mg protein at 1 week of age). Hepatic cSHMT RNA levels parallel the activity pattern. Mitochondrial SHMT was stable throughout gestation and with low constant mSHMT RNA levels. In contrast, the kidney and placenta had high mSHMT and steady low cSHMT activity throughout gestation. These data support the possible role of SHMT in the fetal control of plasma serine levels. While cSHMT may contribute to fetal hepatic serine production, its activity pattern does not support a primary role in the control of fetal hepatic serine biosynthesis. In the placenta, mSHMT may be important for glycine production from serine.     

Protein synthesis in the gastrointestinal tissues of the ovine fetus

An isotope dilution procedure employing an 8h continuous infusion of L-[2,3,5,6-3H] tyrosine was used to determine fractional protein synthetic rates in the gastrointestinal tissues of ovine fetuses. The infusions were made into the inferior vena cava of the fetuses at 120-130 days of gestation. Immediately following the termination of the infusion the ewes were sacrificed and fetal tissues were collected, frozen in liquid nitrogen and stored at -20 C. The specific activity of the intracellular free and protein bound amino acid pools was determined from which the fractional synthetic rates (%/day) were calculated. These rates were as follows: reticulo-rumen, 49%; omasum, 10%; abomasum, 14%; proximal duodenum, 93%; and distal colon, 15%. The contribution of duodenum to the whole body protein synthesis was 10.5% while the contribution of the entire GIT (13.9%) was very close to that of liver (14.4%). The specific activity of tyrosine in the amniotic fluid and fetal ruminal fluid averaged 22% and 36% respectively of the specific activity in the plasma. The high turnover of tissue proteins in the fetal gastrointestinal tract as well as the presence of labelled tyrosine in the ruminal fluid indicate the functional importance of gastrointestinal activity in utero preparing the gastrointestinal tract for post-natal life.     

Circulating thymic hormone levels in severe combined immunodeficiency.

Twenty-three patients with severe combined immunodeficiency disease were studied for circulating thymic hormone levels (facteur thymique serique, FTS), 21 prior to treatment by transplantation of bone marrow, thymus or fetal liver. Thirteen showed undetectable FTS activity. Only two had normal levels of this hormone. In serial determinations of FTS activity prior to and after transplantation, patients given bone marrow transplants developed sustained increments of serum FTS activity early in the course of their immunological reconstitution. However, patients given transplants of fetal liver alone or fetal liver plus thymus from fetuses of less than 12 weeks gestation generally did not show an increment of FTS activity during the period of observation. Transplantation of irradiated thymus derived from fetuses of more than 14 weeks gestation produced sustained increases of thymic hormone activity. These observations suggest that a cell of haematopoietic origin provides a stimulus necessary for differentiation or maturation of thymic secretory activity and that this cell(s) is present in post-natal marrow, but is either undeveloped or immature in the early fetal liver or fails to migrate to the thymus of an allogeneic host.     

Cellular development of liver and skeletal muscles and body composition of pigs from gestationally starved sows

Liver and skeletal muscle development and changes in body composition were studied in pigs from cross-bred sows subjected to starvation (allowed water and trace mineral salt) during the third trimester of gestation. Two groups of sows were taken off feed on days 93 (21-day) and 107 (7-day) of gestation respectively; a third group fed 1.82 kg of diet per day served as control. The pigs sacrificed at birth, were used to determine liver and skeletal muscle DNA, RNA and protein and body composition. There were no significant differences (P greater than .05) in body weight between the treated groups and the control. Liver weight was depressed in the progeny of 21-day and 7-day starvation groups (P less than .05). Liver cellular DNA was decreased (P less than .05) in the treated animals, RNA and protein content remained unchanged. The skeletal muscles studied responded differently to the treatment imposed; semitendinosus muscle weight, muscle DNA and RNA did not differ among treatments; whereas, gastrocnemius muscle weight, muscle DNA and RNA were significantly lower in the progeny of treated sows (P less than .05, P less than .01 and P less than .05 respectively). Muscle and liver protein content, RNA/DNA, and protein/DNA ratios were not affected by treatments. Body composition analysis showed no differences in per cent dry matter, lipid, ash and protein content. The results suggest that liver and gastrocnemius muscles were adversely affected by prenatal nutritional deficiency imposed while semitendinosus muscles remained unchanged. The differential response of skeletal muscles to prenatal nutritional deficiency indicated a need to study more than a single muscle in similar experiments designed to investigate muscle response. The lack of differences in body composition analysis showed that improved survival of newborn pigs previously reported can be achieved without changes in fetal chemical body composition.     

Dietary regulation of liver and muscle transport of amino acid.

Distribution ratios (intracellular concn/extracellular concn) of alpha-amino isobutyric acid (AIB) after intravenous injection were determined in fed, 12-h, 1-day, and 5-day starved rats. Progressive increases (over fourfold) in the distribution ratios of AIB in the liver and progressive decreases (over threefold) in the gastrocnemius muscle occurred within these periods. On full day of protein deprivation was without effect on AIB distribution ratios, but after 5 days it produced an increased distribution ratio of AIB in the liver (twofold), without affecting that of the muscle. A sudden increase in hepatic glucose output, induced by phlorizin, was followed by an increase in the liver distribution ratio of AIB. In starvation the increase in plasma concentration of glucagon and decrease in insulin level preceded the changes in AIB distribution ratios; in protein deprivation there was no change in plasma concentrations of these hormones. It is concluded that caloric restriction profoundly affects amino acid transport by the liver and by the skeletal muscle. These transport changes would enhance the availability of substrates for increased gluconeogenesis during starvation.     

Effect of protein deprivation and parenteral nitrogen feeding on tissue nucleic acids in albino rats with toxic hepatitis

The concentration of nucleic acids was studied in the tissues (skeletal and heart muscles, liver) of rats with toxic liver damage caused by CCl₄, associated with protein deprivation and parenteral administration of the amino acid mixture moriamines S-2 and improved casein hydrolysate. In protein-deprived albino rats with toxic hepatitis the RNA and DNA concentration is skeletal and heart muscle was considerably increased while, at the same time, their concentration in the liver was reduced. The RNA/DNA ratio under these circumstances was altered. The RNA content in the hepatocytes was reduced through a decrease in the nuclear fraction of RNA firmly bound with chromatin. Parenteral nitrogen feeding restored the normal nucleic acid concentration in all the tissues studied.Experimental Department, L'vov Institute of Hematology and Blood Transfusion. (Presented by Academician of the Academy of Medical Sciences of the USSR A. M. Chernukh.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 86, No. 8, pp. 161–164, August, 1978.     

Maternal nutrient deprivation induces sex-specific changes in thyroid hormone receptor and deiodinase expression in the fetal guinea pig brain

Thyroid hormone deprivation during fetal life has been implicated in neurodevelopmental morbidity. In humans, poor growth in utero is also associated with fetal hypothyroxinaemia. In guinea pigs, a short period (48 h) of maternal nutrient deprivation at gestational day (gd) 50 results in fetuses with hypothyroxinaemia and increased brain/body weight ratios. Thyroid hormone action is mediated by nuclear thyroid hormone receptors (TRs) and is dependent upon the prereceptor regulation of supply of triiodothyronine (T₃) by deiodinase enzymes. Examination of fetal guinea pig brains using in situ hybridization demonstrated widespread expression of mRNAs encoding TRα1, α2 and β1, with regional colocalization of deiodinase type 2 (D2) mRNA in the developing forebrain, limbic structures, brainstem and cerebellum at gd52. With maternal nutrient deprivation, TRα1 and β1 mRNA expression was significantly increased in the male, but decreased in the female fetal hippocampus and cerebellum and other areas showing high TR expression under euthyroid conditions. Maternal nutrient deprivation resulted in elevated D2 mRNA expression in males and females. Deiodinase type 3 (D3) mRNA expression was confined to the shell of the nucleus accumbens, the posterior amygdalohippocampal area, brainstem and cerebellum, and did not change with maternal nutrient deprivation. In conclusion, maternal nutrient deprivation resulted in sex-specific changes in TR mRNA expression and a generalized increase in D2 mRNAs within the fetal brain. These changes may represent a protective mechanism to maintain appropriate thyroid hormone action in the face of fetal hypothyroxinaemia in order to optimize brain development.     

The maturation of the rabbit fetal lung following maternal administration of pilocarpine

Pilocarpine HCl, a parasympathomimetic drug, was administered to pregnant white rabbits in a daily subcutaneous injection of 5 mg/kg on days 24 through 27 of gestation. Fetuses from these animals and from salineinjected controls were obtained by caesarean section at day 28 of gestation. Light microscopic examination revealed thinner alveolar septa in the lungs of pilocarpine-treated fetuses and, morphometrically, a significant increase in the number of mature type II cells, both per unit area and per 1,000 lung cells of any kind. Examination by electron microscopy revealed that the alveolar epithelium of pilocarpine-treated fetuses demonstrated morphologic correlates of increased maturation. These included thinning of type I cells to form blood-air barriers and substantial reductions in the glycogen content of both epithelial cell types. Type II cells of pilocarpine-treated fetuses contained (as indicated by morphometric analysis) more and larger lamellar inclusion bodies, as well as more multivesicular bodies than those of controls. Biochemical determination indicated that the glycogen content of fetal lung, but not liver, was reduced significantly in the pilocarpine-treated group. The findings of this study indicate that maternal administration of pilocarpine results in increased maturation of the fetal alveolar epithelium, thus providing a basis for the autonomic manipulation of fetal lung maturation.     

Protective effects of garlic juice against embryotoxicity of methylmercuric chloride administered to pregnant Fischer 344 rats

In order to investigate the beneficial effects of 0.5 or 1.0 g/kg Korean garlic juice against the embryotoxicity of 20 mg/kg methylmercury chloride (MMC, CH3HgCl), pregnant Fisher 344 rats were simultaneously orally administered on day 7 of gestation. On day 20 of gestation the dams were laparotomized under ether anesthesia, and the fetuses were removed and examined for toxicity of methylmercury. Garlic juice depressed the toxicity in terms of some parameters. In the case of simultaneous treatment with 0.1 g/kg garlic juice and MMC, rates of increase were 17.5% in maternal body weight, 13.2% and 41.9% in fetal and litters' weight respectively, and 37.0% in fetal survival rate. Decreasing rates were 10.0% in maternal death rate, and 6.9% and 31.3% in pre- and post-implantation loss respectively. Decreasing rates of mercury levels in dams were 67.2% in liver, 57.6% in brain, 47.2% in kidney, 42.1% in spleen and 40.9% in blood. As well, decreasing rates of mercury level in fetuses were 54.9% in all body burden, 55.9% in liver, 46.7% in kidney and 37% in brain, respectively. The number of fetal ossification centers were reduced by 23.8% to 58.0% following simultaneous treatment with 1.0 g/kg garlic juice. These findings indicated that garlic juice effectively inhibited the embryotoxicity of methylmercury in pregnant Fischer 344 rats.     

ACTH-producing cells of 21-day-old rat fetuses after maternal dexamethasone exposure

Adrenocorticotrophic hormone (ACTH) is essential for developmental maturation of numerous organ systems during the fetal period and for adaptation to environmental challenges. Immunocytochemical and stereological methods were used in the present study to examine the effects of dexamethasone (Dx) administration during pregnancy on fetal rat pituitary ACTH-producing cells. Doses of 0.5, 0.5 and 1.0 mg Dx/kg body weight/day were given to the dams on 3 consecutive days starting on day 16 of gestation. Morphometric analysis of the ACTH-producing cells of fetuses at 21 days of gestation revealed significant inhibition by 24% and 27%, respectively, of cell volume and cell number after maternal Dx administration, whereas the volume of cell nuclei and volume density of ACTH-stained cells were insignificantly decreased. Immunocytochemical analysis showed reduced numbers, sizes and immunopositivity of ACTH cells of 21-day-old fetuses from Dx-treated dams as compared with the control group. Maternal Dx treatment in the period of intense differentiation of the hypothalamo-hypophyseal-adrenal system had an inhibitory effect on fetal function and proliferative activity of ACTH-producing cells at 21 days of gestation. Thus, inhibition of activity of fetal ACTH-producing cells may lead to adrenal suppression, modified activity of the hypothalamo-pituitary-adrenal axis and reduced body weight possibly causing lasting functional abnormalities.     

不同妊娠状态下孕妇外周血中游离胎儿DNA定量分析

目的对不同妊娠状态下孕妇外周血中游离胎儿DNA(f DNA)定量分析,确定其平均浓度及临床参考值范围,初步探讨在不同妊娠状态下母血中f DNA的浓度变化,为临床应用提供科学依据。方法从孕妇外周血浆中提取fDNA,用实时荧光定量聚合酶链反应(FQ-PCR)方法检测其中Y性别决定区的SRY基因。结果在正常早期的孕妇组38例血浆标本中有32例检测到SRY基因,其平均浓度149.25拷贝数/ml,参考值范围为33.28~265.22拷贝数/ml;在正常晚期的孕妇组32例血浆标本中全部检测到SRY基因,其平均浓度为212.14拷贝数/ml,参考值范围为142.76~281.52拷贝数/ml;在晚期患有子痫前期的孕妇30例血浆标本中全部检测到SRY基因,其平均浓度为678.70拷贝数/ml,参考值范围为595.01~726.40拷贝数/ml。实验数据用单因素方差分析,组间差异显著性检验用LSD-t检验。妊娠晚期孕妇血浆中f DNA的含量较妊娠早期升高,约为1.4倍,有统计学意义(P<0.01);晚期患子痫前期的孕妇血浆f DNA的水平是同期正常对照组的3.9倍,有统计学意义(P<0.01)。结论1.用FQ-PCR法最早在孕48天孕妇外周血中即可检测到fDNA。2.随着妊娠的进展孕妇血浆中f DNA的含量升高。3.晚期患子痫前期孕妇其血浆f DNA的水平是同期正常对照组的3.9倍,有统计学意义(P<0.01)。4.f DNA在进行无创伤性产前诊断中有重要价值。