A cluster of protein kinases and phosphatases modulated in fetal Down syndrome (trisomy 21) brain.

Down syndrome (DS; trisomy 21) is the most frequent cause of mental retardation with major cognitive and behavioral deficits. Although a series of aberrant biochemical pathways has been reported, work on signaling proteins is limited. It was, therefore, the aim of the study to test a selection of protein kinases and phosphatases known to be essential for memory and learning mechanisms in fetal DS brain. 12 frontal cortices from DS brain were compared to 12 frontal cortices from controls obtained at legal abortions. Proteins were extracted from brains and western blotting with specific antibodies was carried out. Primary results were used for networking (IntAct Molecular Interaction Database) and individual predicted pathway components were subsequently quantified by western blotting. Levels of calcium-calmodulin kinase II alpha, transforming growth factor beta-activated kinase 1 as well as phosphatase and tensin homolog (PTEN) were reduced in cortex of DS subjects and network generation pointed to interaction between PTEN and the dendritic spine protein drebrin that was subsequently determined and reduced levels were observed. The findings of reduced levels of cognitive-function-related protein kinases and the phosphatase may be relevant for interpretation of previous work and may be useful for the design of future studies on signaling in DS brain. Moreover, decreased drebrin levels may point to dendritic spine abnormalities.

Effect of puerarin on matrix metalloproteinase-2 in human fetal scleral fibroblasts treated with low frequency electromagnetic fields.

OBJECTIVE: To study the effect of puerarin on matrix metalloproteinase-2 (MMP-2) gene and protein expression in human fetal scleral fibroblasts (HFSFs) exposed to extremely low frequency electromagnetic fields (ELF-EMF). METHODS: Cultured HFSFs were exposed to 0.2 mT ELF-EMF for 24 h. The experimental groups were divided into subgroups treated with 0, 0.1, 1 and 10 microM puerarin respectively. The expression of MMP-2 mRNA and protein were detected with real-time polymerase chain reaction and western-blot analysis respectively. RESULTS: MMP-2 mRNA and protein expression increased by 0.793 and 1.130 folds respectively under the exposure of ELF-EMFs at 0.2 mT flux density for 24 h. Puerarin at the concentration of 0.1 microM reversed this effect by 8.53% in mRNA and by 17.97% in protein expression (P < 0.05). The effect was more prominent at higher concentrations (1 and 10 microM, P < 0.01). CONCLUSION: Exposure to ELF-EMFs increased the expression of MMP-2 mRNA and protein in HFSF cells. Puerarin reversed the action to some extent in a specific concentration range. Our results implied that the puerarin might protect scleral tissue from increased expression induced by exposure to ELF-EMFs.

Genomic analysis of neuroendocrine development of fetal brain-pituitary-adrenal axis in late gestation.

The present study was performed to identify the changes in genomic expression of critical components of the hypothalamus-pituitary-adrenal (HPA) axis in the second half of gestation in fetal sheep. We isolated mRNA from pituitary, hypothalamus, hippocampus, and brain stem in fetal sheep at 80, 100, 120, 130, and 145 days of gestation and 1 and 7 days after delivery (n = 4-5/group). Using real-time RT-PCR, we measured mRNA expression levels of glucocorticoid receptor (GR), mineralocorticoid receptor (MR), serum- and glucocorticoid-induced kinase-1 (sgk1), proopiomelanocortin (POMC), CRF, and arginine vasopressin (AVP). Both MR and GR were highly expressed in pituitary and hippocampus; in all tissues GR was more highly expressed than MR. AVP was more highly expressed than CRF in hypothalamus. MR, GR, and sgk1 expression were increased postnatally in brain stem, and sgk1 expression was increased postnatally in hypothalamus. GR expression was reduced in pituitary in term fetuses compared with younger ages. Hypothalamic CRF expression was increased at the end of gestation compared with younger ages, and AVP expression was increased in newborn lambs. Pituitary POMC was increased at 100 days of gestation compared with 80 days; hypothalamic POMC was increased at 120 days. Overall, the results demonstrate the expression of both MR and GR in brain regions important for control of the HPA axis. Decreases in expression of GR in pituitary at the end of gestation might contribute to the decreased corticosteroid negative feedback sensitivity at term in this species.

Identification of a novel substrate for tyrosine kinase in human testes.

Identification of genes specifically expressed in adult and fetal testes is important in furthering our understanding of testis development and function. In this study, a novel human specific transcript, designated NYD-SP10, was identified by hybridization of adult and fetal human testis probes with a human cDNA microarray. NYD-SP10 (GenBank Accession no. AF332192) was expressed at a higher level in adult testes than in fetal testes. Bioinformatic analysis illustrated NYD-SP10 is a splice variant of regulatory factor X4, while it is lack of RFX DNA-binding domain which binds to the X box of MHC class II genes. Most telling of all, NYD-SP10 uniquely possesses a tyrosine kinase phosphorylation site. Expression profiles demonstrated NYD-SP10 is specifically and strongly expressed in human adult testes and ejaculated spermatozoa. Taken together, these results suggest that NYD-SP10 is a novel testis-specific gene and it is possibly a novel substrate for tyrosine kinase and involved in the function of human spermatozoa, especially during capacitation.

Enzymes involved in the formation and transformation of steroid hormones in the fetal and placental compartments.

Human fetal and placental compartments have all the enzymatic systems necessary to produce steroid hormones. However, their activities are different and complementary: the fetus is very active in converting acetate into cholesterol, in transforming pregnanes to androstanes, various hydroxylases, sulfotransferases, whilst all these transformations are absent or very limited in the placenta. This compartment can transform cholesterol to C21-steroids, convert 5-ene to 4-ene steroids, and has a high capacity to aromatize C19 precursors and to hydrolyse sulfates. Steroid hormone receptors are present at an early stage of gestation and are functional for important physiological activities. The production rate of some steroids increases drastically with fetal evolution (e.g. estriol increases 500-1000 times in relation to non-pregnant women). We can hypothesize that the control of active steroid hormones could be carried out by fetal and placental factors, which act by stimulating or inhibiting the enzymes involved in their formation and transformation during pregnancy evolution and, consequently, limiting the high levels of the biologically active hormone.

Dopamine synthesis by non-dopaminergic neurons in the arcuate nucleus of rat fetuses.

The aim of the present work was to verify the hypothesis that non-dopaminergic neurons expressing individual complementary dopamine synthesis enzymes can perform the co-located synthesis of dopamine. According to this hypothesis, neurons expressing tyrosine hydroxylase use L-tyrosine for the synthesis of L-dihydroxyphenylalanine (L-DOPA), which then enters neurons expressing aromatic amino acid decarboxylase, which converts L-DOPA to dopamine. Experiments were performed using the mediobasal hypothalamus of rat fetuses, which mostly contains single-enzyme neurons (>99%) and occasional double-enzyme neurons (<1%). Controls were obtained from the fetal substantia nigra, which is enriched with dopaminergic neurons. High-performance liquid chromatography was used to measure levels of dopamine and L-DOPA in cell extracts and the incubation medium after incubation in the presence and absence of exogenous L-tyrosine. Addition of L-tyrosine to the medium led to increases in the level of synthesis and release of L-DOPA in the mediobasal hypothalamus and substantia nigra. In addition, L-tyrosine increased dopamine synthesis in the substantia nigra and decreased dopamine synthesis in the mediobasal hypothalamus. This regional difference in levels of dopamine synthesis is probably due to inhibition of the uptake of L-DOPA from the intercellular medium by neurons in the mediobasal hypothalamus containing aromatic amino acid decarboxylase, due to the competitive binding of the L-DOPA transporter by L-tyrosine. Thus, these results provide the first evidence for the co-located synthesis of dopamine by non-dopaminergic neurons expressing single complementary enzymes involved in the synthesis of this neurotransmitter.

Molecular cloning, identification and characteristics of a novel isoform of carbamyl phosphate synthetase I in human testis.

A gene coding a novel isoform of carbamyl phosphate synthetase I (CPS1) was cloned from a human testicular library. As shown by cDNA microarray hybridization, this gene was expressed at a higher level in human adult testes than in fetal testes. The full length of its cDNA was 3831 bp, with a 3149 bp open reading frame, encoding a 1050-amino-acid protein. The cDNA sequence was deposited in the GenBank (AY317138). Sequence analysis showed that it was homologous to the human CPS1 gene. The putative protein contained functional domains composing the intact large subunit of carbamoyl phosphate synthetase, thus indicated it has the capability of arginine biosynthesis. A multiple tissue expression profile showed high expression of this gene in human testis, suggesting the novel alternative splicing form of CPS1 may be correlated with human spermatogenesis.

Tyrosine aminotransferase and gamma-glutamyl transferase activity in human fetal hepatocyte primary cultures under proliferative conditions.

The ontogeny of gamma-glutamyl transferase (GGTase; E.C.2.3.2.2) and tyrosine aminotransferase (TAT; E.C.2.6.1.5) activities in 14 to 36 weeks gestational and neonatal hepatocytes during development of human fetal liver was studied. Subsequently, 20-24 weeks gestational hepatocytes were cultured in media supplemented with epidermal growth factor (EGF) and insulin with or without glucagon and dexamethasone to investigate the proliferation and differentiation of fetal hepatocyte in vitro using GGTase and TAT as biochemical markers. During the development of the liver, the activity of GGTase increased continuously from the first trimester through the third trimester and decreased (p < 0.001) in neonates. A low basal level of TAT activity was seen only during the third trimester, which then increased significantly (p < 0.001) in neonates. Fetal hepatocytes, in the presence of EGF and insulin, undergo proliferation from the fourth to 10th day with an increase in cell number (p < 0.001) and concomitant increase (p < 0.001) in GGTase activity. As the cells attain confluence, enzyme activity decreased significantly (p < 0.001) from the 10th to 16th day. Maximal TAT activity (p < 0.001) was observed at 48 h of culture, which decreased, but not significantly, during cell proliferation and the enzyme activity was regained as the cultures attained confluence. Furthermore, TAT activity was induced synergistically (p<0.001) in the presence of glucagon and dexamethasone, while GGTase was inhibited (p<0.001). These results indicate that GGTase increases with proliferation, whereas TAT, once it has been expressed, is not suppressed during cell proliferation. In conclusion, human fetal hepatocytes undergo enzymic differentiation by 48 h of culture, and proliferate with an increase in GGTase in the presence of growth factors with maintenance of differentiated status up to the studied 16 days of culture.

Development of paraventricular nucleus (PVN) associated with immunoreactivity to tyrosine hydroxylase (TH) in the second half of gestation.

The aim of this study was to shed more light on the developmental characteristics of human paraventricular nucleus (PVN) and hypothalamus in general, using modern immunohistochemical techniques to detect the activity of tyrosine hydroxylase (TH) in the synthesis of catecholamine (CA). Fetal brains were examined at 12, 16, 20 and 23 weeks gestation. Immunohistochemical staining used for sections is a qualitative method for detection and distribution of the chosen protein. The amplification of positive signals was carried out using AVIDIN/BIOTIN technique. The first positive results were obtained at 16th week of gestation and immunoreactivity) grew with the advencement of gestation and that there was no homogenisation of immunoreactivity in some parts of the nucleus. The PVN showed to be an oval formation. nearly parallel with the lateral wall of the 3rd cerebral ventricle, where the upper apex of the nucleus is situated closer to the ependymal ventricular layer. There was an ascendant dorsal movement of the nucleus in the embryonic phase of hypothalamus development.

Antisense inhibition of epidermal growth factor receptor decreases expression of human surfactant protein A.

Epidermal growth factor (EGF) stimulates surfactant protein A (SP-A) synthesis in fetal lung tissue through ligand binding to the EGF receptor. We hypothesized that inhibition of EGF receptor messenger RNA (mRNA) would block SP-A expression in human fetal lung tissue during alveolar type II cell differentiation in vitro. Midtrimester human fetal lung explants were maintained in serum-free Waymouth's medium for 3 to 5 d in the presence or absence of an antisense 18-mer phosphorothioate oligonucleotide (ON) complementary to the initiation codon region of EGF receptor mRNA. Sense and scrambled ONs similarly modified were used as additional controls. The concentration of EGF receptor mRNA was semiquantitatively determined by reverse transcriptase/polymerase chain reaction (RT-PCR). We found a significant 3-fold decrease in EGF receptor mRNA levels in the antisense-treated groups compared with the control group with no effect in the sense condition. Immunohistochemical staining revealed a decrease in the amount of staining for EGF receptor protein in distal pulmonary epithelial cells in the antisense-treated groups compared with either control or sense conditions. Treatment with antisense EGF receptor ON decreased both SP-A mRNA and protein compared with controls with no effect in the sense condition. The ONs did not affect tissue viability as measured by the release of lactate dehydrogenase. We conclude that selective degradation of EGF receptor mRNA with antisense ON treatment results in a decrease in SP-A expression in human fetal lung. These findings support the critical importance of the EGF receptor for the regulation of SP-A gene expression during human alveolar type II cell differentiation.

Biochemical properties, tissue expression, and gene structure of a short chain dehydrogenase/ reductase able to catalyze cis-retinol oxidation.

We have identified a retinol dehydrogenase (cRDH) that catalyzes the oxidation of 9-cis- but not all-trans-retinol and proposed that this enzyme plays an important role in synthesis of the transcriptionally active retinoid, 9-cis-retinoic acid. There is little information regarding either the biochemical properties of cRDH or how its 9-cis-retinol substrate is formed. We now report studies of the properties and expression of human and mouse cRDH and of the characteristics and location of the murine cRDH gene. Additionally, we report mouse hepatic 9-cis-retinol concentrations and demonstrate that 9-cis-retinol is formed in a time- and protein-dependent manner upon incubation of all-trans -retinol with cell homogenate. Human and mouse cRDH display similar substrate specificities for cis-isomers of retinol and retinaldehyde. Moreover, human and mouse cRDH show marked sensitivity to inhibition by 13-cis-retinoic acid, with both being inhibited by approximately 50% by 0.15 microm 13-cis-retinoic acid (for substrate concentrations of 10 microm). Lesser inhibition is seen for 9-cis- or all-trans-retinoic acids. Immunoblot analysis using antiserum directed against human cRDH demonstrates cRDH expression in several tissues from first trimester human fetuses, indicating that cRDH is expressed early in embryogenesis. Adult mouse brain, liver, kidney, and to a lesser extent small intestine and placenta express cRDH. The murine cRDH gene consists of at least 5 exons and spans approximately 6 kb of genomic DNA. Backcross analysis mapped the mouse cRDH gene to the most distal region of chromosome 10. Taken together, these data extend our understanding of the properties of cRDH and provide additional support for our hypothesis that cRDH may play an important role in 9-cis-retinoic acid formation.

Expression of receptor tyrosine kinase RYK in developing rat central nervous system.

Receptor tyrosine kinase RYK is a mammalian homologue of Drosophila Lio, which is involved in learning and memory and in axon guidance. We cloned a rat ryk gene and characterized its expression pattern in the central nervous system. Northern blot analysis of the whole brain revealed that the RYK mRNA was abundant during the period from 13 to 18 embryonic days (E13-18) and it decreased by E20. In the postnatal brain, the RYK signal was higher in postnatal one week (P1W) cerebrum and in P2W cerebellum than in later stages. In situ hybridization revealed that RYK was expressed throughout the central nervous system, mainly in the ventricular zone on E11 and E13. On E18 and E20, the remarkable level of RYK mRNA was detected in the ventricular zone as well as in the cortical plate of the forebrain. These two regions overlapped the immunoreactive areas of nestin and MAP2, a neural stem cell marker and a mature neural marker, respectively. Moreover, the double-labeling analysis showed that the same cells expressed both RYK and nestin in the ventricular zone. In the postnatal brain, RYK was predominantly expressed in neurons of various regions. These observations suggest that RYK plays a contributory role as a multifunctional molecule in the differentiation and maturation of neuronal cells in the central nervous system.

Cultures of cells from fetal rat brain: methods to control composition, morphology, and biochemical activity.

Fetal tissue transplantation is a promising new approach for the treatment of neurodegenerative diseases, but the optimal conditions for preparing cells for transplantation have not been defined. The growth of a population of septal brain cells, primarily containing cholinergic neurons and glia, was characterized after seeding at densities from 5 x 10(4) to 6 x 10(5) cells/cm2, on polystyrene-, collagen-, laminin-, and fibronectin-coated surfaces, in the presence of serum and/or serum-free medium. Differentiated glial cells were selected by culture on fibronectin or laminin surfaces, in the presence of low amounts of serum (2.5% FBS) and G5, a soluble factor containing EGF and insulin. Differentiated neuronal cells were selected by culture on laminin, in the presence of low amounts of serum (2.5% FBS) and N2, a soluble factor containing supplemental hormones. In each case, a minimum seeding density of 1 x 10(5) cells/cm2 was required. Neuronal growth could be maintained long term (21 days) with high levels of neuronal activity (ChAT activity).

Expression of cytochrome P450RAI (CYP26) in human fetal hepatic and cephalic tissues.

PCR amplifications with two sets of degenerate primers that were targeted to CYP26-specific regions were performed with cDNAs from human fetal liver and brain as templates. PCR products were purified, cloned, sequenced and analyzed with the BLAST program. Our results revealed expression of CYP26 in both human fetal liver and brain. Furthermore, human fetal CYP26 cDNA exhibited 99.2%-100% nucleotide sequence identity to its adult counterpart. Novel isoforms, that would have indicated additional CYP26 genes, were not found. A Northern blot containing poly(A+)RNAs from 43 human adult and 7 human fetal tissues was tested for CYP26 expression. We were able to detect CYP26 message in most tissues but hybridization signals varied in intensity. Highest levels of transcription were in adult liver, heart, pituitary gland, adrenal gland, placenta and regions of the brain. CYP26 expression in fetal tissues was strongest in the brain and comparable with message levels in adult tissues.

Expression of 11 beta-hydroxylase (CYP11B1) and aldosterone synthase (CYP11B2) in the human fetal adrenal.

OBJECTIVE: To understand better the steroidogenic capacity of the human fetal adrenal (HFA), we evaluated the expression of 11 beta-hydroxylase (CYP11B1) and aldosterone synthase (CYP11B2) in the fetal zone and neocortex of the HFA using a specific RNase protection assay. METHODS: Adrenal glands were obtained at the time of elective termination of pregnancy. Whole adrenals (n = 7) were frozen in liquid nitrogen, and subsequently total RNA extraction was performed by tissue homogenization followed by guanidinium/chloroform purification. In addition, RNA was obtained from separated fetal zone (n = 4) and neocortex (n = 4) tissues obtained by dissection. RNase protection assays were then performed using radiolabeled complementary RNA probes generated by T7 RNA polymerase directed against transcripts for CYP11B1, CYP11B2, and actin, the latter of which was used as a control for RNA integrity. Transcripts also were examined using a reverse transcription polymerase chain reaction (RT-PCR) protocol specific for CYP11B1 or CYP11B2. RESULTS: The RNase protection assay was designed to distinguish specific bands that corresponded to CYP11B1 (232 bp), CYP11B2 (262 bp), and actin (221 bp). RNA isolated from whole HFA was observed to have high levels of CYP11B1 transcript, whereas CYP11B2 was not detected. Dissected neocortex and fetal zones were found to contain transcript for CYP11B1 using both the RNase protection assay and RT-PCR analysis. In contrast, using the RNase protection assay, CYP11B2 mRNA was not observed in the RNA from the fetal zone, but after prolonged exposure there was a band corresponding in size to CYP11B2 observed in RNA from the neocortex. Using the more sensitive RT-PCR method, transcript for CYP11B2 was found in both neocortex and fetal zone. CONCLUSION: The HFA expresses low levels of CYP11B2 in accordance with its low production of mineralocorticoid. The expression of CYP11B1 in the fetal zone is intriguing because this enzyme is not necessary for the production of C19 steroids. Definition of the molecular mechanisms controlling expression of the CYP11B genes will be necessary to determine why the HFA differentially expresses these isoenzymes.

Cloning and characterization of fetal liver phosphatase 1, a nuclear protein tyrosine phosphatase isolated from hematopoietic stem cells.

We report the isolation of cDNAs encoding protein tyrosine phosphatases (PTPs) from highly purified hematopoietic stem cell populations. One such cDNA encodes a novel PTP, designated fetal liver phosphatase 1 (FLP1), which consists of one PTP domain followed by a carboxy terminal domain of 160 amino acids. Northern blot and in situ hybridization analysis showed that expression of FLP1 mRNA is restricted to thymus in 15.5-day-old and 17.5-day-old mouse embryos and to kidney and hematopoietic tissues in adult mice. Furthermore, polymerase chain reaction-based analysis shows that FLP1 is expressed in hematopoietic stem cells as well as in more mature hematopoietic cells. Peptide antisera against FLP1 immunoprecipitated a 48-kD protein that is localized in the nuclei of Ba/F3 lymphoid cells. We have analyzed the effects of overexpressing either wild-type FLP1 or a functionally inactive mutant of FLP1 in hematopoietic cells. In the progenitor K562 cell line, cells ectopically expressing functional FLP1 differentiated normally to megakaryocytes after induction with tetradecanoyl phorbol acetate (TPA). In contrast, when K562 transfectants expressing an inactive mutant FLP1 protein were treated with TPA, the characteristic cell spreading and substrate adhesion that accompany megakaryocytic differentiation did not occur. We show that, in these cells, the induction of the differentiation marker alphaIIb beta3 is not affected. However, both constitutive and TPA-induced expression of alpha2 integrin, a late megakaryocytic marker, are inhibited. These results suggest that the expression of an inactive form of FLP1 affects late signaling events of K562 megakaryocytic differentiation.

Cloning and production of antisera to human placental 11 beta-hydroxysteroid dehydrogenase type 2.

By inactivating potent glucocorticoid hormones (cortisol and corticosterone), 11 beta-hydroxysteroid dehydrogenase type 2 (11 beta-HSD2) plays an important role in the placenta by controlling fetal exposure to maternal glucocorticoids, and in aldosterone target tissues by controlling ligand access to co-localized glucocorticoid and mineralocorticoid receptors. Amino acid sequence from homogeneous human placental 11 beta-HSD2 was used to isolate a 1897 bp cDNA encoding this enzyme (predicted M(r) 44126; predicted pI 9.9). Transfection into mammalian (CHO) cells produces 11 beta-HSD2 activity which is NAD(+)-dependent, is without reductase activity, avidly metabolizes glucocorticoids (Km values for corticosterone, cortisol and dexamethasone of 12.4 +/- 1.5, 43.9 +/- 8.5 and 119 +/- 15 nM respectively) and is inhibited by glycyrrhetinic acid and carbenoxolone (IC50 values 10-20 nM). Rabbit antisera recognizing 11 beta-HSD2 have been raised to an 11 beta-HSD2-(370--383)-peptide-carrier conjugate. Recombinant 11 beta-HSD2, like native human placental 11 beta-HSD2, is detectable with affinity labelling and anti-11 beta-HSD2 antisera, and appears to require little post-translational processing for activity. 11 beta-HSD2 mRNA (approximately 1.9 kb transcript) is expressed in placenta, aldosterone target tissues (kidney, parotid, colon and skin) and pancreas. In situ hybridization and immunohistochemistry localize abundant 11 beta-HSD2 expression to the distal nephron in human adult kidney and to the trophoblast in the placenta. 11 beta-HSD2 transcripts are expressed in fetal kidney (but not lung, liver or brain) at 21-26 weeks, suggesting that an 11 beta-HSD2 distribution resembling that in the adult is established by this stage in human development.

Promoting phorbol ester-TPA enhances migration of C3H 10T1/2 cells. The role of protein kinase C and its relation to carcinogenesis.

Treatment of 10T1/2 cells with promoting phorbol ester drastically enhanced migration of the studied fibroblasts in a serum-supplemented medium. The same cells when exposed to ionomycin or TPA in a serum-free medium did not show any migration. The addition of 1% of serum induced spontaneous and TPA stimulated migration. Also EGF and PDGF separately or together induced the migration of 10T1/2 cells. Parallel studies of protein kinase C documented low enzymatic activity after treatment with TPA, whereas transcripts of PKC were shown independently of TPA treatment.

Relative expression of aromatase cytochrome P450 in human fetal tissues as determined by competitive polymerase chain reaction amplification.

The aromatase enzyme complex is responsible for the conversion of C19 steroids to estrogens. Aromatase activities ranging from moderate to very low have been measured in human fetal tissues. The inability to demonstrate aromatase cytochrome P450 (P450AROM) messenger RNA (mRNA) in several fetal tissues by northern blotting has been attributed to low levels of specific message. In order to identify and compare P450AROM mRNA levels in fetal tissues, we developed a specific competitive polymerase chain reaction amplification technique. This reaction uses coamplification of a rat P450AROM complementary RNA to normalize differences in amplification efficiencies. Using this technique, P450AROM mRNA was identified in all fetal tissues studied including; liver, lung, brain, skin, intestine, kidney, spleen, and heart. Fetal liver contained far more P450AROM mRNA per total RNA than any other tissues studied. Fetal brain and intestine also tended to have slightly higher levels than other tissues.

Evaluation of biochemical evidence of congenital nutritional myopathy in two-week prepartum fetuses from selenium-deficient ewes.

Muscle damage attributable to selenium (Se)/vitamin E deficiencies is known to develop at birth or later in lambs. The purpose of this study was to determine whether and when muscle damage develops in utero. Thirty pregnant ewes maintained on Se-deficient forages from birth were allotted to 3 equal groups. Half of each group was given a single IM injection of 0.056 mg of Se/kg of body weight, 1 month before parturition. At 3 weeks before parturition, cesarean section-derived fetuses from Se-deficient ewes did not have evidence of muscle damage. At 2 weeks before parturition, fetuses from Se-deficient ewes had biochemical evidence of congenital nutritional myopathy, as evidenced by low blood Se concentration (P less than 0.05) and by increased plasma creatinine kinase (P less than 0.001) and lactate dehydrogenase (P less than 0.01) activities, compared with fetuses from Se-treated ewes. Thus, for optimal protection of fetuses and newborn lambs in Se-deficient areas, Se should be administered to ewes at least 1 month before parturition.