Comparative analysis of pregnancy proteins and regulation of the functional activity of T-suppressors in pregnancy

The relationship between the pregnancy specific proteins (SP1, SP3) and the functional activity of ConA-induced T-suppressors has been studied during normal gestation. There were a positive correlation between the activity of T-suppressors and the level of SP1 during 8-28 weeks of gestation; between the activity of T-suppressors and the level of SP3 during 8-36 weeks, especially during 29-36 weeks of gestation. It was shown that a progressive accumulation of these proteins apparently necessary for the formation a suppression's dominant in the condition of the immunoregulatory branches during gestation. These findings suggest that SP1 and SP3 as a natural endogenous immunoregulators may play an important role in reproductive strategy of human gestation.     

Distribution of endoglin in early human development reveals high levels on endocardial cushion tissue mesenchyme during valve formation

Endoglin is a component of the receptor complex for transforming growth factor (TGF)-β1 and TGF-β3. We analysed its expression by immunohistochemistry in human embryos at 4–8 weeks of gestation and in hearts ranging from 4–13 weeks old. We compared endoglin distribution with that of TGF-β receptors type I (TβR-I), type II (TβR-II) and betaglycan. Endoglin was found on endothelial cells in all tissues examined, consistent with its expression in adult blood vessels. TβR-I, TβR-II and betaglycan were observed on most cell types and had an overall similar pattern of distribution. Endoglin was detected on the endocardium as early as 4 weeks, but was absent from myocardium. It was present at high levels on the endocardial cushion tissue mesenchyme from 5–8 weeks’ gestation, during heart septation and valve formation, and subsequently decreased as the valves matured. Endoglin expression in heart extracts was confirmed by Western blot analysis. TβR-I, TβR-II and betaglycan were mostly found on cardiac myocytes, but were detectable at low levels on endocardium. They were expressed transiently on cushion mesenchyme, albeit at much lower levels than endoglin. All four components of the TGF-β receptor complex were detected by RT-PCR in embryonic heart. Thus transient up-regulation of the components of the TGF-β receptor complex, and particulartly of endoglin, is associated with heart septation and valve formation during early human development.     

The dynamics of the T-lymphocyte subpopulations and the peroxidase activity of the neutrophils in children immunized with a live mumps vaccine

The amount of T-helpers and T-suppressors and the T-helper/T-suppressor ratio were determined in 30 practically healthy children aged 1.5-2 years, immunized with live parotitis vaccine prepared from strain -3. One immunization dose of the vaccine contained 7,950 HADU50. In all children peroxidase activity in the cytoplasm of neutrophils was studied. This investigation revealed that in all examined children the formation of immune response to immunization with live parotitis vaccine was accompanied by the development of an imbalance of immunoregulatory T-cell subpopulations and by functional changes in neutrophils, characterized by the inhibition of peroxidase activity. The changes revealed in this study were most pronounced in children with a high level of antibody formation.     

Glycosphingolipid Glycosyltransferases in Human Fetal Brain

The developmental pattern of gangliosides in human fetal brain should reflect the activities of the respective glycosyltransferases. LA2-synthase activity, along with that of GM3-, GD3-, GM2-, and GM1-synthases, was determined in human fetal brain at 10-22 weeks of gestation. LA2-synthase is the pivotal enzyme in lacto series ganglioside formation. LA2-synthase activity decreased during the study period, mirroring a similar temporal decline in levels of the lacto series gangliosides, particularly 3'-isoLM1. The developmental profiles of the ganglio series glycosyltransferase activities demonstrate distinct changes that correspond to the ganglioside pattern between fetal weeks 10 and 22. In particular, the marked increase in GM2-synthase activity at 20 and 22 weeks of gestation and the decline in GD3-synthase activity after 15 weeks could explain the prominent expression of the a series gangliosides in this period of rapid neuronal outgrowth. However, a similar decline (two- to 2.5-fold) in GM3-synthase activity suggests a more likely conclusion, namely, that the two sialyltransferase activities are derived mainly from astroglial cells, which show a marked proliferation during the 10-15th fetal weeks. The data do not negate the hypothesis that GM3- and GD3-synthase are the critical enzymes in the regulation of ganglioside biosynthesis but do indicate a need to reevaluate the significance of GM2-synthase in expression of the a series gangliosides.     

Surfactant protein of molecular weight 28,000-36,000 in cultured human fetal lung: cellular localization and effect of dexamethasone

We have examined the effect of explant culture and hormones on the major surfactant associated protein of Mr 28,000-36,000 (SP 28-36) in human fetal lung. Explants of 16- to 23-week gestation lung were maintained for up to 5 days in culture. Polyclonal antibodies raised to SP 28-36 purified from alveolar proteinosis lung lavage were used in immunofluorescence experiments (n = 11). There was no specific fluorescence seen in frozen sections of preculture tissue. In explants cultured without serum or hormones, fluorescence was seen in most epithelial cells lining potential airspaces. In cultures treated with 10 nM dexamethasone and 2 nM T3 much brighter fluorescence was seen in virtually all epithelial cells. Immunofluorescence studies on cell monolayers prepared from explants confirmed that SP 28-36 is found in the cytoplasm of type II cells but not in fibroblasts. The pattern of fluorescence was consistent with the presence of SP 28-36 on rough endoplasmic reticulum. SP 28-36 mRNA was measured in isolated cell populations using a 32P-labeled cDNA probe. mRNA levels were manyfold higher in type II cell preparations (purity 78-92%) than in fibroblasts (purity 81-97%). A competitive enzyme linked assay was developed to quantify SP 28-36. The SP 28-36 content of five lungs before culture (17-23 weeks) was less than 0.02 microgram/mg DNA. During explant culture without hormones the SP 28-36 content increased exponentially. Exposure to dexamethasone accelerated the increase in SP 28-36 content. T3, alone or in the presence of dexamethasone, did not influence SP 28-36 content. We conclude that SP 28-36 content is very low in human fetal lung before 24 weeks gestation. Explant culture and treatment with dexamethasone synchronize development of type II cells from epithelial precursors, and induce synthesis of SP 28-36 in type II cells. These findings provide evidence of concomitant regulation by glucocorticoids of the phospholipid synthetic enzymes and the major protein of pulmonary surfactant.     

A shift in sensory processing that enables the developing human brain to discriminate touch from pain

When and how infants begin to discriminate noxious from innocuous stimuli is a fundamental question in neuroscience [1]. However, little is known about the development of the necessary cortical somatosensory functional prerequisites in the intact human brain. Recent studies of developing brain networks have emphasized the importance of transient spontaneous and evoked neuronal bursting activity in the formation of functional circuits [2, 3]. These neuronal bursts are present during development and precede the onset of sensory functions [4, 5]. Their disappearance and the emergence of more adult-like activity are therefore thought to signal the maturation of functional brain circuitry [2, 4]. Here we show the changing patterns of neuronal activity that underlie the onset of nociception and touch discrimination in the preterm infant. We have conducted noninvasive electroencephalogram (EEG) recording of the brain neuronal activity in response to time-locked touches and clinically essential noxious lances of the heel in infants aged 28-45?weeks gestation. We show a transition in brain response following tactile and noxious stimulation from nonspecific, evenly dispersed neuronal bursts to modality-specific, localized, evoked potentials. The results suggest that specific neural circuits necessary for discrimination between touch and nociception emerge from 35-37?weeks gestation in the human brain.     

Simultaneous observations of iodothyronine content in the thyroid gland, serum and thyroxine 5'- and 5-monodeiodinase activity in liver, during the neonatal period of the pig

Serum T4 and rT3 were high at about 4-12 h after birth, then they decreased to a nadir on day 3 (rT3) and day 7 (T4). Serum T3 concentration fell immediately after birth but then increased to a relatively stable level during the next 2-6 weeks, then fell after weaning. Reciprocal concentration profiles of T4, T3 and rT3 in the thyroid were found. The thyroidal iodothyronine content increased significantly after weaning. In the liver, 5'-monodeiodinating activity, low after birth, rose until day 3 and then decreased concomitantly with T3 in serum. The 5-monodeiodinating activity, high at birth, fell to a nadir at about 3 weeks. No changes in 5- and 5'-deiodinase activity after 3 weeks were observed. Opposite to the variations in absolute content, the iodothyronine relative proportion in thyroid tissue was practically unchanged until weaning time (6 weeks), when they rose. Serum T3/T4 and rT3/T4 ratios increased with age until weaning. The post-weaned pigs had T3/T4 and rT3/T4 ratios about two times smaller than 6-weeks-old pigs. Serum rT3/T3, high after birth, decreased with age. Summarizing, the results indicate that neither changes in the thyroid iodothyronine content nor in the liver T4-monodeiodinating activity can solely account for variations in serum TH during the early neonatal period in the pig. It is suggested that the rapid variations in serum TH levels can reflect changes in the thyroidal secretory activity in preferential T3 secretion and/or blood disappearance rates.     

Hormonal profile and glucose tolerance in late pregnancy and postpartum

Oral glucose tolerance, plasma insulin and basal levels of glucagon, hGH, hPRL, hPL, TSH, T4, T3, thyroxine-binding globulin (TBG), cortisol, corticosteroid-binding globulin (CBG) and estriol were measured in 23 normal pregnant women in late gestation (31 +/- 0.4 weeks of pregnancy). Twelve of these subjects could be re-examined 14 +/- 2 weeks postpartum. Blood glucose was lower basal and after glucose load (100 g) in the pregnant group. Fasting plasma insulin and glucose-induced insulin release were higher in pregnancy. The insulinogenic index and the beta cell response were significantly greater antepartum, while peripheral insulin activity was unchanged. The insulin:glucagon ratio as well as TSH and hGH showed no significant differences between ante- and postpartum values. However, T4, T3, TBG, cortisol, CBG, estriol, hPRL and hPL were significantly higher during gestation than after delivery. T4:TBG and T3:TBG ratios were much lower antepartum, while the cortisol:CBG ratio was comparable ante- and postpartum. To our knowledge this is the first report in which such an extensive hormonal and metabolic analysis was performed in the same women ante- and postpartum. It could be shown that glucose tolerance is not worsened during pregnancy in healthy subjects. The higher gestational insulin values are discussed with respect to the various significant hormonal changes.     

CD4+CD8- thymocytes from neonatal mice induce IgM production in SCID mice.

Defective recombination of both the TCR and Ig genes results in the absence of mature lymphocytes in mice with the scid mutation. We have shown previously that the transfer of neonatal, but not adult, thymocytes results in high levels of Ig production in 100% of C.B-17-scid (SCID) mice, in contrast to the 10 to 25% of SCID mice spontaneously producing low levels of oligoclonal Ig. In this report we demonstrate that neonatal CD4+8- thymocytes were able to induce this response; the CD4+8+ and CD4-8+ subpopulations were totally inactive and CD4-8- T cells had only limited activity several weeks after transfer. The stimulation of IgM production in SCID mice was detectable by 1 wk posttransfer of CD4+8- thymocytes or splenic T cells, and could be achieved with as few as 300 cells. The ability of neonatal CD4+8- thymocytes to induce Ig diminished gradually to insignificant levels at 3 wk postbirth; this loss of function was not associated with differential survival of neonatal T cells. Neonatal CD4+8- thymocytes from C.B-17 and other H-2d strains rescued Ig production, whereas cells from H-2b, H-2a, and H-2k strains were much less effective. These results suggest that a CD4+8- subpopulation found in both neonatal thymus and peripheral lymphoid tissues is able to induce the expansion or differentiation of the small numbers of functional B lymphocytes in SCID mice, and that the inducing T cell disappears shortly after birth, perhaps during the acquisition of self-tolerance.     

Maturation of the pituitary-thyroid axis during the perinatal period.

To clarify the maturation process of the pituitary-thyroid axis during the perinatal period, thyrotropin (TSH) response to thyrotropin releasing hormone (TRH) and serum thyroid hormone levels were examined in 26 healthy infants of 30 to 40 weeks gestation. A TRH stimulation test was performed on 10 to 20 postnatal days. Basal concentrations of serum thyroxine (T4), free thyroxine (free T4) and triiodothyronine (T3) were positively correlated to gestational age and birth weight (p less than 0.001-0.01). Seven infants of 30 to 35 gestational weeks demonstrated an exaggerated TSH response to TRH (49.7 +/- 6.7 microU/ml versus 22.1 +/- 4.8 microU/ml, p less than 0.001), which was gradually reduced with gestational age and normalized after 37 weeks gestation. A similar decrease in TSH responsiveness to TRH was also observed longitudinally in all of 5 high responders repeatedly examined. There was a negative correlation between basal or peak TSH concentrations and postconceptional age in high responders (r = -0.59 p less than 0.05, r = -0.66 p less than 0.01), whereas in the normal responders TSH response, remained at a constant level during 31 to 43 postconceptional weeks. On the other hand, there was no correlation between basal or peak TSH levels and serum thyroid hormones. These results indicate that (1) maturation of the pituitary-thyroid axis is intrinsically controlled by gestational age rather than by serum thyroid hormone levels, (2) hypersecretion of TSH in preterm infants induces a progressive increase in serum thyroid hormones, and (3) although there is individual variation in the maturation process, the feedback regulation of the pituitary-thyroid axis matures by approximately the 37th gestational week.     

The effect of kemantane on T-helpers and T-suppressors

The study of Kemantan on functionally alternative humoral immunity regulator cells: T-helpers and antigen-specific T-suppressors, including their induction, accumulation and functioning, was studied. Kemantan in doses of 0.2-200 mg/kg, introduced to the donors of T-helpers 2 days before they were taken, stimulated their activity 1.5- to 2-fold (with p less than 0.05). Kemantan had no influence on the functional activity of T-suppressors, as well as on their induction and accumulation.     

Sucrose feeding effects inhibition of gamma-glutamyltranspeptidase in the liver of the rat: possible mediation by thyroid hormone

The effect of sucrose on Fischer 344 rat liver gamma-glutamyltranspeptidase (gammaGT) was studied: in adults fed sucrose for 3 weeks; and rats exposed to sucrose from the 18th day of gestation to the 40th day after birth. Rats fed regular rodent chow served as controls. Sucrose caused mild lipemia; and in the liver an increase in size and fat build-up without damage. In adult sucrose-fed rats, compared to controls, plasma glucose levels were increased: 1.12-, 1.40- and 1.13-fold after 24, 48h and 3 week consumption of sucrose, respectively. Insulin levels were unaltered for the first week of sucrose consumption but increased from control levels: 16% at 1 week, and 2.0-fold at 3 weeks. The T3 levels were comparable to control levels 24h after the sucrose was started and were increased: 1.22-, 1.13- and 1.12-fold at 48h, 1 and 3 weeks, respectively. The T4 levels were comparable at all time points between sucrose-fed and control rats. Liver gammaGT activity exhibited a steady decrease from control levels: after 24, 48h, 1 and 3 weeks of sucrose feeding the decrease was 5, 8, 21 and 37%, respectively in homogenates; and 10, 17, 24 and 41%, respectively in plasma membranes. Perinatal sucrose exposure effected in 40-day-old rats, compared controls: a 1.09-fold increase in plasma glucose; no change in plasma insulin; an increase of 1.15- and 1.39-fold in plasma levels of total and free T3, respectively; a decrease of 20 and 14% in plasma levels of total and free T4, respectively. gammaGT activity was decreased in liver plasma membranes isolated from sucrose-exposed rats relative to those of control: 80% in the male; 82% in the female. Relative specific activities of gammaGT were the same in both males: 15.4 and 16.1 in control and sucrose-exposed male rats, respectively; and females: 14.1 and 15.4 in control and sucrose-exposed female rats, respectively. gammaGT was 2-fold higher in the livers of female relative to male rats in sucrose-exposed and control groups. Kidney gammaGT activities were the same in control and sucrose-exposed rats. The involvement of T3 in the sucrose-induced decrease in liver gammaGT is discussed.     

Controlling effect of T lymphocyte suppressors on the proliferation of cells in various tissues

The changes in mitotic index of mouse bone marrow, spleen, liver, kidneys, small intestine, cornea were studied during alterations in the number of T-suppressors. It was found that mitotic activity in all tissues investigated enhanced significantly after a decrease in the number of T-suppressors caused by single injection of an antiserum against T-suppressors. On the contrary, the mitotic index diminished significantly after the transfer of tolerant spleen lymphoid cell suspension enriched by T-suppressors to normal syngeneic mice. These data indicate that T-suppressors are responsible for the control over cell proliferation in nonlymphoid tissues.     

Genome‐wide association studies reveal genetic loci associated with plasma cholinesterase activity in ducks

Plasma cholinesterase (PCHE) activity is an important auxiliary test in human clinical medicine. It can distinguish liver diseases from non‐liver diseases and help detect organophosphorus poisoning. Animal experiments have confirmed that PCHE activity is associated with obesity and hypertension and changes with physiological changes in an animal's body. The objective of this study was to locate the genetic loci responsible for PCHE activity variation in ducks. PCHE activity of Pekin duck × mallard F2 ducks at 3 and 8 weeks of age were analyzed, and genome‐wide association studies were conducted. A region of about 1.5 Mb (21.8–23.3 Mb) on duck chromosome 9 was found to be associated with PCHE activity at both 3 and 8 weeks of age. The top SNP, g.22643979C>T in the butyrylcholinesterase (BCHE) gene, was most highly associated with PCHE activity at 3 weeks (?logP = 21.45) and 8 weeks (?logP = 27.60) of age. For the top SNP, the strong associations of CC and CT genotypes with low PCHE activity and the TT genotype with high PCHE activity indicates the dominant inheritance of low PCHE activity. Problems with block inheritance or linkage exist in this region. This study supports that BCHE is a functional gene for determining PCHE levels in ducks and that the genetic variations around this gene can cause phenotypic variations of PCHE activity.     

Modification of testicular and thyroid function by chronic exposure to short photoperiod: a comparison in four rodent species

Exposure of male Syrian hamsters (Mesocricetus auratus) for 10 weeks to short photoperiod (SP) providing 10 hr light: 14 hr darkness (10:14 LD) produced a significant reduction in the weights of the reproductive organs, plasma thyroxine (T4) levels and free T4 index (FT4I) compared to the values of animals exposed to long photoperiod (LP, 14:10 LD). C57bl male house mice (Mus musculus) kept in SP (10:14 LD) had reproductive organ weights equivalent to those of mice kept in long days (14:10 LD) and lower T3 uptake (T3U) values. Male gerbils (Meriones unguiculatus) exposed to 13 weeks of SP (10:14 LD) had lower body weights, testes and seminal vesicle weights and higher T3U values compared to LP (14:10 LD) controls. However, no effect was seen on plasma T4 and triiodothyronine (T3) values nor the FT4I and free T3 index (FT3I). White-footed male mice (Peromyscus leucopus) exposed to SP (8:16 LD) had significantly lower testes and seminal vesicle weights while plasma T4 and T3 levels were unaffected. Snell strain house mice (Mus musculus) exposed to SP (8:16 LD) had normal reproductive organ weights compared to the values of LP-exposed (16:8 LD) control animals. However, there was a significant depression in T3 and in the FT3I in the SP animals.     

Expression of a peptide binding to receptor for activated C-kinase (RACK1) inhibits phorbol myristoyl acetate-stimulated phospholipase D activity in C3H/10T1/2 cells: dissociation of phospholipase D-mediated phosphatidylcholine breakdown from its synthesis

The C3H/10T1/2 Cl8 HAbetaC2-1 cells used in this study express a peptide with a sequence shown to bind receptor for activated C-kinase (RACK1) and inhibit cPKC-mediated cell functions. Phorbol myristoyl acetate (PMA) strongly stimulated phosphatidylcholine (PtdCho)-specific phospholipase D (PLD) activity in the C3H/10T1/2 Cl8 parental cell line, but not in Cl8 HAbetaC2-1 cells, indicating that full PLD activity in PMA-treated Cl8 cells is dependent on a functional interaction of alpha/betaPKC with RACK1. In contrast, the PMA-stimulated uptake of choline and its subsequent incorporation into PtdCho, were not inhibited in Cl8 HAbetaC2-1 cells as compared to Cl8 cells, indicating a RACK1-independent but PKC-mediated process. Increased incorporation of labelled choline into PtdCho upon PMA treatment was not associated with changes of either CDP-choline: 1,2-diacylglycerol cholinephosphotransferase activity or the CTP:phosphocholine cytidylyltransferase distribution between cytosol and membrane fractions in Cl8 and Cl8 HAbetaC2-1 cells. The major effect of PMA on the PtdCho synthesis in C3H/10T1/2 fibroblasts was to increase the cellular uptake of choline. As a supporting experiment, we inhibited PMA-stimulated PtdH formation by PLD, and also putatively PtdH-derived DAG, in Cl8 cells with 1-butanol. Butanol did not influence the incorporation of [(14)C]choline into PtdCho. The present study shows: (1) PMA-stimulated PLD activity is dependent on a functional interaction between alpha/betaPKC and RACK1 in C3H/10T1/2 Cl8 fibroblasts; and (2) inhibition of PLD activity and PtdH formation did not reduce the cellular uptake and incorporation of labelled choline into PtdCho, indicating that these processes are not directly regulated by PtdCho-PLD activity in PMA-treated C3H/10T1/2 Cl8 fibroblasts.     

Ontogeny of human hepatic cytochromes P450

Significant changes in drug-metabolizing enzyme (DME) expression occur during ontogeny. Such changes can have a profound effect on therapeutic efficacy in the fetus and child, as well as the risk for adverse drug reactions. To gain a better understanding of DME ontogeny, enzyme contents for six key cytochromes P450 were measured in 240 human liver samples representing ages from 8 weeks gestation to 18 years. Where possible, both quantitative western blotting and activity assays with probe substrates were performed. Although oversimplified, the DME can be grouped into one of three categories. As typified by CYP3A7, some enzymes are expressed at their highest level during the first trimester and either remain at high concentrations or decrease during gestation and are silenced or expressed at low levels within 1-2 years after birth. These data cause one to query whether these enzymes have an important endogenous function. Representatives of a second group, CYP3A5 and CYP2C19, are expressed at relatively constant levels throughout gestation. Postnatal increases in CYP2C19 are observed within the first year, but not for CYP3A5. CYP2C9, 2E1, and 3A4 are more typical of a third group of enzymes that are not expressed or are expressed at low levels in the fetus with the onset of expression generally in either the second or third trimester. Substantial increases in expression are observed within the first 1-2 years after birth; however, considerable interindividual variability is observed in the immediate postnatal (1-6 months) onset or increase in expression of these enzymes, often resulting in a window of hypervariability.     

Crosstalk between PKCzeta and the IL4/Stat6 pathway during T-cell-mediated hepatitis

PKCzeta is required for nuclear factor kappa-B (NF-kappaB) activation in several cell systems. NF-kappaB is a suppressor of liver apoptosis during development and in concanavalin A (ConA)-induced T-cell-mediated hepatitis. Here we show that PKCzeta-/- mice display inhibited ConA-induced NF-kappaB activation and reduced damage in liver. As the IL-4/Stat6 pathway is necessary for ConA-induced hepatitis, we addressed here the potential role of PKCzeta in this cascade. Interestingly, the loss of PKCzeta severely attenuated serum IL-5 and liver eotaxin-1 levels, two critical mediators of liver damage. Stat6 tyrosine phosphorylation and Jak1 activation were ablated in the liver of ConA-injected PKCzeta-/- mice and in IL-4-stimulated PKCzeta-/- fibroblasts. PKCzeta interacts with and phosphorylates Jak1 and PKCzeta activity is required for Jak1 function. In contrast, Par-4-/- mice have increased sensitivity to ConA-induced liver damage and IL-4 signaling. This unveils a novel and critical involvement of PKCzeta in the IL-4/Stat6 signaling pathway in vitro and in vivo.