A positive umbilical venous-arterial difference of leptin level and its rapid decline after birth

OBJECTIVE: We investigated the site of leptin production in the fetoplacental circulation. STUDY DESIGN: We simultaneously determined plasma leptin levels in cord vessels and maternal peripheral veins of 38 healthy pregnant women. We also compared plasma leptin levels in 20 neonates at birth and on the fifth day after birth. RESULTS: Leptin levels in cord vessels were significantly (p < 0.001) lower than those in maternal veins (mean 29.5 ng/ml). Leptin levels in umbilical arteries (mean 9.8 ng/ml) were significantly (p < 0.01) lower than those in umbilical veins (mean 12.9 ng/ml). Leptin levels in neonatal veins on the fifth day (mean 3.0 ng/ml) were markedly (p < 0.001) lower than those in umbilical arteries of the same neonates (mean 10.9 ng/ml). CONCLUSION: The higher leptin levels in umbilical veins than in umbilical arteries and the marked decrease during the neonatal period suggest that the placenta is one of the major sources of leptin in the fetal circulation.     

Population pharmacokinetic modeling in very premature infants receiving midazolam during mechanical ventilation: midazolam neonatal pharmacokinetics

BACKGROUND: Midazolam is used widely as a sedative to facilitate mechanical ventilation. This prospective study investigated the population pharmacokinetics of midazolam in very premature infants. METHODS: Midazolam (100 microg/kg) was administered as a rapid intravenous bolus dose every 4-6 h to 60 very premature neonates with a mean (range) gestational age of 27 weeks (24-31 weeks), a birth weight of 965 g (523-1,470 g), and an age of 4.5 days (2-15 days). A median (range) of four (one to four) blood samples, 0.2 ml each, were drawn at random times after the first dose or during continuous treatment, and concentrations of midazolam in serum were assayed by high-performance liquid chromatography. A population analysis was conducted using a two-compartment pharmacokinetic model using the NONMEM program. RESULTS: Average parameter values (interpatient percent coefficient of variation) for infants with birth weights 1,000 g or less were total systemic clearance (Cl(T)) = 0.783 ml/min (83%), intercompartmental clearance (Cl(Q)) = 6.53 ml/min (116%), volume of distribution of the central compartment (V1) = 473 ml (70%), and volume of distribution of the peripheral compartment (V2) = 513 ml (146%). For infants with birth weights more than 1,000 g they were as follows: Cl(T) = 1.24 ml/min (78%), Cl(Q) = 9.82 ml/min (98%), V1 = 823 ml (43%), and V2 = 1,040 ml (193%). The intrapatient variability (percent coefficient of variation) in the data was 4.5% at the mean concentration midazolam in serum of 121 ng/mL CONCLUSIONS: Serum concentration-time data were used in modeling the population pharmacokinetics of midazolam in very premature, ventilated neonates. Clearance of midazolam was markedly decreased compared with previous data from term infants and older patients. Infants weighing less than 1,000 g at birth had significantly lower clearance than those weighing more than 1,000 g.     

Wasps are the cause of an increasing mastitis problem in dairy cattle in Israel

The purpose of this study was to estimate if the erythropoietin (EPO) concentration in cord arterial blood can be an indicator of a fetal risk. We studied EPO concentration measured by enzyme immonoassay in ten patient groups: (1) control group with healthy newborns (n = 72); (2) neonates born by elective caesarean section (n = 16); (3) newborns with acidosis at birth (n = 12); (4) newborns with 1-min-Apgar < 7 (n = 8); (5) preterm neonates (n = 25); (6) newborns with gestational age > or = 242 weeks (n = 19); (7) neonates born to mothers with hypertension (n = 16); (8) newborns with signs of fetal distress in CTG (n = 29); (9) neonates born to mothers with diabetes (n = 19), divided into two subgroups: diabetes White A-D (n = 8) and gestational diabetes (n = 11); (10) neonates born to mothers with diabetes White A-D and with acidosis at birth (n = 7). The geometric mean was 26.4 mU/ml in the control group. EPO levels was found significantly increased (p < 0.01) in the following groups: (3) newborns with acidosis (52 mU/ml); (6) newborns with gestational age > or = 242 weeks (63.5 mU/ml); (8) newborns with signs of fetal distress in CTG (47.1 mU/ml); (9) neonates born to mothers with diabetes White A-D (47.7 mU/ml); (10) neonates born to mothers with diabetes White A-D and with acidosis at birth (> 64 mU/ml). We came to the conclusion that the cord arterial EPO concentration indicates a chronic fetal hypoxia and a longer duration of hypoxia before birth.     

Pediatric annular pancreas: twenty years' experience

Characterization of pediatric annular pancreas is provided by this analysis of 24 cases (22 neonates, 1 infant, 1 child). Salient observations include: (1) Presentation is affected by the degree of duodenal obstruction at birth and by coexistent anomalies. (2) Polyhydramnios usually accompanies complete high intestinal obstruction by annular pancreas. (3) Primary biliary interruption was not encountered and jaundice was not unusually prevalent. (4) There is a high incidence of associated anomalies. (5) Duodenal bypass by duodenoenterostomy was employed with excellent results.     

Down-regulation of human FEN-1 gene expression during differentiation of promyelocytic leukemia cells

The presence of aflatoxin in corn and corn dust during relatively normal years and the increased risk of Aspergillus flavus infestation during drought conditions suggest that airborne agricultural exposures should be of considerable concern. Liquid extraction, thin layer chromatography, and high pressure liquid chromatography were used for the analysis of aflatoxin B1 in grain dust and bulk corn samples. A total of 24 samples of airborne dust were collected from 8 farms during harvest, 22 samples from 9 farms during animal feeding, and 14 sets of Andersen samples from 11 farms during bin cleaning. A total of 14 samples of settled dust and 18 samples of bulk corn were also collected and analyzed. The airborne concentration of aflatoxin B1 found in dust collected during harvest and grain unloading ranged from 0.04 to 92 ng/m3. Higher levels of aflatoxin B1 were found in the airborne dust samples collected from enclosed animal feeding buildings (5-421 ng/m3) and during bin cleaning (124-4849 ng/m3). Aflatoxin B1 up to 5100 ng/g were detected in settled dust collected from an enclosed animal feeding building; however, no apparent correlation was found between the airborne concentration of aflatoxin B1 and its concentration in settled dust or bulk corn. The data demonstrate that farmers and farm workers may be exposed to potentially hazardous concentrations of aflatoxin B1, particularly during bin cleaning and animal feeding in enclosed buildings.     

Secretion and Sex differences in cord serum growth hormone levels of the human fetus (author's transl)

Cord venous growth hormone (GH) and somatostatin levels were measured by radioimmunoassay in 86 infants and 11 anencephalic infants. 1) Cord GH levels were 69.2 +/- 20.2 ng/ml (N = 6, gestational weeks (GW) : 21-30W, mean +/- S.D.), 31.5 +/- 24.3 ng/ml (N = 11, GW : 31-36W), and 17.6 +/- 8.9 ng/ml (N = 71, GW: 37-41). In anencephalic infants, cord GH levels were significantly lower (2.3 +/- 1.4 ng/ml) than those in preterm and fullterm infants. 2) In fullterm infants, cord GH levels of small infants (19.2 +/- 9.2 ng/ml, N = 33, birth weight less than 3,250g) were significantly higher than that of large infants (15.1 +/- 8.1 ng/ml, N = 38, birth weight greater than of equal too 3,250g). On the other hands, cord somatostatin levels were relatively lower in small infants than in large infants. 3) There was a significant difference between male cord GH levels (20.6 +/- 8.9 ng/ml, N = 35) and female cord GH levels (14.6 +/- 8.0 ng/ml, N = 36). These results suggest that fetal GH secretion is suppressed by GH inhibiting factor (somatotropin release inhibiting factor) from mid gestation toward term and there is a sex difference of fetal GH secretion in fullterm fetus.     

The effect of traumatic brain injury on children with learning disability

In Argentina, there is rather little information about the natural occurrence of mycotoxins in feedstuffs. The aim of this work was to determine the fungal flora and natural incidence of aflatoxin B1 (AFB1), zearalenone (ZEA) and deoxynivalenol (DON) in poultry feeds from 5 factories of Río Cuarto, Córdoba. Three hundred samples were taken from May 1995 to May 1996. Fungal counts of poultry feeds ranged 10(4) to 10(6) CFU g-1. The lowest counts were obtained on the first months from the sampling (May to September 1995) with mean values significantly different from those found at the last of the sampling (October 1995 to April 1996). The most prevalent species isolated of poultry feed samples belonged to the genera Penicillium that was present in 98% of the samples, Fusarium (87%) and Aspergillus (52%). Fusarium species isolated were: F moniliforme in 73% of the samples, F subglutinans (35%), F graminearum (20%) and within Aspergillus species: A. parasiticus (33%) and A. flavus (8%) were identified. In poultry feeds aflatoxin B1 (AFB1) was the most significant mycotoxin with levels ranging from 17 to 197 ng/g. For deoxynivalenol (DON) the levels ranged from 240 to 410 ng/g. Only three out of 300 samples were contaminated with zearalenone (ZEA) in concentrations of 30, 120 and 280 ng/g. These are preliminary data on this subject in our region.     

Immunochemical approaches to AGE-structures: characterization of anti-AGE antibodies

An extractionless method for determining aflatoxin M1 (AFM1), a major metabolite of aflatoxin B1 (AFB1), in human urine was developed. The biological fluid is injected directly into the chromatographic system after simple dilution and centrifugation. A pre-column, packed with a cation-exchange phase and coupled on-line to a column-switching liquid chromatography (LC) system, is used for sample pre-treatment and concentration. The analytes are non-selectively desorbed with the LC eluent and cleaned by means of a column-switching procedure. Pre-treatment and analysis were performed within 40 min. Average AFMI recovery reached 97% in the 10-100 ng/l range of urine. The detection limit of AFM1 in urine and milk was 2.5 ng/l for 1 ml of injected sample. A comparison with an immunoaffinity column clean-up and LC method was performed. The method was applied to determine AFM1 in the urine of AFB1 gavaged rats, and in the urine of both potentially exposed and supposedly unexposed workers. The method was also extended to milk.     

Gilbert syndrome and glucose-6-phosphate dehydrogenase deficiency: a dose-dependent genetic interaction crucial to neonatal hyperbilirubinemia

Severe jaundice leading to kernicterus or death in the newborn is the most devastating consequence of glucose-6-phosphate dehydrogenase (EC 1.1.1.49; G-6-PD) deficiency. We asked whether the TA repeat promoter polymorphism in the gene for uridinediphosphoglucuronate glucuronosyltransferase 1 (EC 2.4.1.17; UDPGT1), associated with benign jaundice in adults (Gilbert syndrome), increases the incidence of neonatal hyperbilirubinemia in G-6-PD deficiency. DNA from term neonates was analyzed for UDPGT1 polymorphism (normal homozygotes, heterozygotes, variant homozygotes), and for G-6-PD Mediterranean deficiency. The variant UDPGT1 promoter allele frequency was similar in G-6-PD-deficient and normal neonates. Thirty (22.9%) G-6-PD deficient neonates developed serum total bilirubin >/= 257 micromol/liter, vs. 22 (9.2%) normals (P = 0.0005). Of those with the normal homozygous UDPGT1 genotype, the incidence of hyperbilirubinemia was similar in G-6-PD-deficients and controls (9.7% and 9.9%). In contrast, in the G-6-PD-deficient neonates, those with the heterozygous or homozygous variant UDPGT1 genotype had a higher incidence of hyperbilirubinemia than corresponding controls (heterozygotes: 31.6% vs. 6.7%, P < 0.0001; variant homozygotes: 50% vs. 14.7%, P = 0.02). Among G-6-PD-deficient infants the incidence of hyperbilirubinemia was greater in those with the heterozygous (31.6%, P = 0.006) or variant homozygous (50%, P = 0.003) UDPGT1 genotype than in normal homozygotes. In contrast, among those normal for G-6-PD, the UDPGT1 polymorphism had no significant effect (heterozygotes: 6.7%; variant homozygotes: 14.7%). Thus, neither G-6-PD deficiency nor the variant UDPGT1 promoter, alone, increased the incidence of hyperbilirubinemia, but both in combination did. This gene interaction may serve as a paradigm of the interaction of benign genetic polymorphisms in the causation of disease.     

Does excessive adenosine 5''-triphosphate formation in cells lead to malignancy? A hypothesis on cancer

PROBLEM: Neonatal outcome of in vitro fertilization (IVF) pregnancies has been described by different authors, but several issues have yet to be resolved. The aim of the present study was to evaluate neonates conceived in vitro and to direct special attention to neonatal morbidity and prevalence of minor abnormalities. The information that has been accumulated so far is scant. METHOD: The first 100 babies conceived in vitro, and subsequently born in our institute, were investigated and compared with the general, spontaneously conceived newborn population. All infants were examined by a senior neonatologist, and the data that were recorded included gestational age at delivery, birth weight, gender, major malformations, minor congenital abnormalities, neonatal mortality, and neonatal morbidity (including asphyxia, jaundice, meconium aspiration, hypoglycemia, and hypocalcemia). RESULTS AND CONCLUSIONS: The data indicate that the IVF neonates assessed had a higher rate of low birth weight (37%), twinning (30%), and preterm birth (20%) in comparison with the general reference population (p < 0.05). However, no differences were encountered either in the rate of small for gestational age infants or the incidence of major malformations and minor abnormalities between these groups of newborns. The overall neonatal morbidity in IVF babies was found to exceed that of the general population. Nevertheless, at birth, there were no clinical pathognomonic signs typical of IVF babies, although more detailed metabolic, endocrine, and neurobehavioral studies are still required to confirm that newborns conceived in vitro do not differ from those conceived spontaneously in any of these respects.     

Survey of the occurrence of aflatoxin M1 in dairy products marketed in Italy

During 1995, 159 samples of milk, 97 samples of dry milk for infant formula, and 114 samples of yogurt were randomly collected in supermarkets and drug stores in four large Italian cities and checked for aflatoxin M1 (AFM1) by immunoaffinity column extraction and HPLC. AFM1 was detected in 136 (86%) of the milk samples (in amounts ranging from < 1 ng/liter to 108.5 ng/liter; mean level: 10.19 ng/liter), in 81 (84%) of the dry milk samples (in amounts ranging from < 1 ng/liter to 101.3 ng/kg; mean level: 21.77 ng/kg), and in 91 (80%) of the yogurt samples (in amounts ranging from < 1 ng/liter to 496.5 ng/liter; mean level: 18.08 ng/liter). Altogether, only two samples of milk, two samples of yogurt, and one sample of dry milk had levels of AFM1 exceeding the Swiss legal limits, which are the most restrictive in the world. AFM1 contamination levels in milk and yogurt samples collected in the period of November to April were ca. four times as high as those in samples collected in the period of May to October. It is concluded that during 1995, despite the widespread occurrence of AFM1, the mean contamination levels in dairy products sold in Italy were not a serious human health hazard.     

Mutagenesis of nitrite reductase from Pseudomonas aeruginosa: tyrosine-10 in the c heme domain is not involved in catalysis

It has been proposed that some neonates infected with hepatitis B virus (HBV), acquire their infections in utero as demonstrated by HBV seromarkers in venous blood samples at birth. In this study, paired blood samples from 13 HBsAg-positive, 19 HBsAg- and HBeAg-positive, 2 HBsAg-negative mothers and 34 of their neonates, were drawn 24-72 hours after birth and tested for HBV-DNA in their peripheral blood mononuclear cells (PBMC). The presence of HBV-DNA in PBMC was detected in 69.2% (9/13) of HBsAg-positive mothers, 94.7% (18/19) of HBsAg- and HBeAg-positive mothers, and in none of their neonates. The conclusion from these results is that the evidence for hepatitis B infections occurring in neonates of hepatitis B carrier mothers in utero is uncommon.     

Metoclopramide plasma concentration in neonates

During the course of a multiple-dose metoclopramide (M) oral treatment, M plasma concentrations were measured just before (Cmin) and 1 hour after the administration (C1h) at steady-state in 5 (3 premature and 2 term) neonates. Mean Cmin was equal to 91.6 +/- 45.5 ng/ml and higher than C1h (87.4 +/- 43.2 ng/ml), but not significantly. A significant negative correlation was found between Cmin plasma concentration and gestational age as well as with postconceptional age, suggesting that the lower the gestational and postconceptional age, the lower the metoclopramide dosage should be.     

Case-control study of pancreatic cancer in Serbia, Yugoslavia

To determine the cardiac output change in neonates who were under nasal continuous positive airway pressure (CPAP) therapy, eleven newborn neonates, who were admitted to our neonatal intensive care unit with respiratory diseases, were enrolled in the study. Cardiac output was measured by pulsed Doppler echocardiogram at various pressures of 0, 2, 4, 6, 8, and 10 cm H2O and revealed 301 +/- 47, 300 +/- 49, 289 +/- 55, 275 +/- 64, 269 +/- 59 and 242 +/- 50 ml/min/kg, respectively. Cardiac output depressed significantly between 0 cm H2O and 8 cm H2O (P = 0.025), and between 0 cm H2O and 10 cm H2O (P = 0.004). We conclude that cardiac output may be depressed in neonates who are under high levels of nasal CPAP therapy, and suggest that high levels of nasal CPAP therapy must be used with caution, especially when the therapy is applied to the low birth weight neonates.     

Determination of transforming growth factor-beta 1 (TGF-beta 1) and insulin-like growth factor (IGF-1) in bovine colostrum samples

The major growth factors in bovine colostrum are transforming growth factor-beta s (TGF-beta 1 and TGF-beta 2) and insulin-like growth factors (IGF-1 and IGF-2). Recently, TGF-beta 2 content of bovine colostrum was measured using a TGF-beta 2 specific ELISA (1) and now we have validated ELISAs for for bovine TGF-beta 1 and IGF-1. The concentrations of IGF-1 and TGF-beta 1 in the first milking after calving were 248-1850 ng/ml and 12.4-42.6 ng/ml, respectively, and they declined in correlation with total protein concentration to 27.0-101 ng/ml (IGF-1) and 0.80-3.49 ng/ml(TGF-beta 1) by the fifth milkings. The amount of TGF-beta 1 was on average 5.3 +/- 1.4% of that of TGF-beta 2 and there is a high correlation (r = 0.966) between the concentrations of these growth factors in the same samples. No free TGF-beta 1 form of could be detected.     

Cultured human airway smooth muscle cells stimulated by interleukin-1beta enhance eosinophil survival

Airway smooth muscle may be an important cellular source of proinflammatory mediators and cytokines and may participate directly in airway inflammation. In this study we have examined whether airway smooth muscle cells could contribute to mechanisms of eosinophil accumulation by prolonging their survival. To investigate this possibility, conditioned medium from human airway smooth muscle cells stimulated with interleukin (IL)-1beta was examined on the in vitro survival of highly purified human peripheral blood eosinophils. After 7 d, when cultured in control medium, less than 1 +/- 0.2% of the initial eosinophil population remained viable. In contrast, culture in medium conditioned for 96 h by human airway smooth muscle cells stimulated with IL-1beta (1 pg-100 ng/ml) resulted in a concentration-dependent increase in eosinophil survival. (The concentration that produced 50% of this effect was 0.03 ng/ml IL-1beta.) Maximum eosinophil survival occurred at 1 to 3 ng/ml IL-1beta. This effect was also time-dependent and was readily detected in airway smooth muscle cell-conditioned medium after just 3 h of stimulation with IL-1beta (1 ng/ml). It continued to increase before reaching a plateau around 24 h, with no decrease in activity for up to 120 h of stimulation. Conditioned medium from unstimulated airway smooth muscle cells did not enhance eosinophil survival. The survival-enhancing activity was completely inhibited (the concentration that inhibited 50% [IC50] was 6.9 microg/ml) by a polyclonal goat antihuman antibody to granulocyte-macrophage colony stimulating factor (GM-CSF) (0.3-100 microg/ml), but antibodies (10-100 microg/ml) to IL-3 and IL-5, and a normal goat immunoglobulin G control had no effect on the eosinophil survival-enhancing activity. GM-CSF levels in culture medium from smooth muscle cells were markedly increased by IL-1beta and were maximum at 30 ng/ml (0.037 ng/ml/10(6) cells versus 3.561 ng/ml/10(6) cells, unstimulated versus 30 ng/ml IL-1beta). The IL-1 receptor antagonist inhibited both the production of GM-CSF (IC50 19. 1 ng/ml) and the eosinophil survival-enhancing (IC50 53.7 ng/ml) activity stimulated by IL-1beta. Release of GM-CSF elicited by IL-1beta was inhibited by dexamethasone but not by indomethacin. These data indicate that cultured human airway smooth muscle cells stimulated with IL-1beta support eosinophil survival through production of GM-CSF and thus may contribute to the local control of inflammatory cell accumulation in the airways.     

Does mandating continuing education mandate competence?

Serial prostate-specific antigen (PSA) measurements (PSA velocity) as an additional instrument to detect prostatic cancer was introduced in 1992. It has previously been reported that PSA increase per year differed in the last 5 years prior to diagnosis in patients with benign prostatic hyperplasia (0.18 ng/ml/year), locally confined (0.75 ng/ml/year) and metastasized (4.4 ng/ml/year) cancer of the prostate (CaP) in contrast to healthy men (0.04 ng/ml/year). The ability of PSA velocity to detect organ-confined CaP in patients with intermediate PSA serum values depends therefore on a reliable and reproducible PSA result. The present study comprised 85 men with PSA values between 3 and 8 ng/ml (Abbott IMx). PSA measurements were repeated with Abbott IMx (n = 85 patients) and Hybritech Tandem-E (n = 59 patients) assays. The PSA serum values differed from one examination to the other from 0.02 to 2.74 ng/ml with the Abbott IMx. Standard deviation amounted to 0.35 ng/ml with the Abbott IMx PSA assay. Using the Hybritech Tandem-E assay, mean standard deviation was 1.15 ng/ml and therefore higher than with the Abbott IMx assay. The difference from one test to the other ranged from 0.05 to 4.05 ng/ml with the Hybritech Tandem-E. Using the Abbott IMx assay, 10.6% of all repeat measurements exceeded 1 ng/ml whereas in the Hybritech Tandem-E assay 62.7% of the second measurements differed > 1 ng/ml from the first PSA result. An increase in PSA serum values may therefore be due to intratest variation, physiological day-to-day variation as well as prostatic disease. It is important to notice that the intra-assay variation may be greater than the PSA increase per year in a patient with CaP. Therefore, PSA velocity seems to be of limited value.     

Reduced production of immunoreactive interleukin-1 by peripheral blood monocytes of patients with acute and chronic viral hepatitis

The in vitro production of interleukin-1 beta by peripheral blood monocytes derived from patients with various liver diseases was studied. An impaired production of immunoreactive interleukin-1 (IL-1) (mean +/- SEM) by monocytes stimulated with an optimal dose (100 ng/ml) of lipopolysaccharide was observed in patients with chronic hepatitis B (N = 13; 32 +/- 6 pg/ml) or chronic hepatitis C (N = 13; 61 +/- 12 pg/ml) as compared to those of healthy control individuals (N = 35; 166 +/- 24 pg/ml; P = 0.0003 and P = 0.015, respectively), whereas an unaltered IL-1 production was seen in patients with alcoholic cirrhosis (N = 23; 125 +/- 28 pg/ml) and primary biliary cirrhosis (N = 6; 111 +/- 33 pg/ml). Similar to the situation seen in chronic viral hepatitis, lipopolysaccharide-stimulated monocytes from patients with acute hepatitis also showed a decreased IL-1 production in the first week after onset of jaundice (N = 17; 55 +/- 20 pg/ml; P = 0.001) and a return to normal in the second and third week. An impaired production of IL-1 in chronic as well as acute viral hepatitis is a further example of the known disturbed immunoregulation in this disease.     

Robotic automated analysis of foods for aflatoxin

Immunoaffinity column-based sample preparation procedures for determination of aflatoxins B1, B2, G1, and G2 in several food matrixes and aflatoxin M1 in milk have been automated by using flexible automation, or robotics. Components used to assemble the system were purchased commercially or developed and built in-house. A liquid-level sensor developed in-house to assist elution of the immunoaffinity column is described. After immunoaffinity column cleanup, aflatoxins are separated by reversed-phase liquid chromatography and determined by fluorescence without derivatization. Mean recoveries of aflatoxins B1, B2, and G1 added to corn and nuts at 9-36 ng/g total aflatoxins were > 85% (coefficient of variation [CV] = 16%). Recoveries of aflatoxin G2 averaged 50% (CV = 28%). Recoveries of aflatoxin M1 added to milk at 0.12-0.50 ng/mL averaged 78% (CV = 19%). The ability of the automated system to reproduce its results is demonstrated by the fact that the CV of replicate assays is generally better than 10%. Comparability between the automated procedure and the AOAC official method is demonstrated.     

Increase by tri-iodothyronine of endothelin-1, fibronectin and von Willebrand factor in cultured endothelial cells

Hyperthyroidism is associated with elevated plasma levels of endothelium-derived proteins such as von Willebrand factor (vWF), fibronectin (FN) and endothelin-1 (ET-1). This study was designed to characterize the mechanisms involved in this phenomenon at the cellular level. vWF, FN and ET-1 secretion and mRNA expression were measured in human umbilical vein endothelial cells (HUVECs) exposed to tri-iodothyronine (T3) for 13 +/- 1 days, using ELISA, Western blot, RIA and Northern blot analysis respectively. Exposure of HUVECs to T3 significantly increased vWF secretion (50 ng T3/ml: 117 +/- 5%, P < 0.01; 100 ng T3/ml: 127 +/- 26%, P < 0.01) as well as vWF mRNA expression (50 ng/ml: 116 +/- 13%, P < 0.001; 100 ng/ml: 136 +/- 30%, P < 0.002) (results are means +/- S.D. analysed by the Wilcoxon signed rank test). FN secretion was significantly affected by 50 (145 +/- 42% of control, P < 0.05) and 100 (116.8 +/- 16% of control, P < 0.05) ng T3/ml, and FN mRNA expression by 50 ng T3/ml (123 +/- 20%, P < 0.05). Long-term incubation with T3 increased both ET-1 secretion (25 ng/ml: 124 +/- 25%, P < 0.001; 50 ng/ml: 165 +/- 53%, P < 0.05; 100 ng/ml: 116 +/- 17%, P < 0.05) and prepro-ET-1 mRNA expression (25 ng/ml: 112 +/- 16%, P < 0.05; 50 ng/ml: 134 +/- 43%, P < 0.02; 100 ng/ml: 120 +/- 20%, P < 0.02). Protein kinase C (PKC) isoforms epsilon and beta II were not significantly affected by T3, whereas PKC alpha was increased in whole cell lysates and in membrane fractions of cells incubated with 100 but not 50 ng T3/ml. Prepro-ET-1 mRNA stability, cell numbers and proliferation, measured by [3H]thymidine assays, remained unaffected in HUVECs after exposure to T3. These data indicate thyroid hormone-induced upregulation of mRNA expression and protein synthesis of vWF, FN and ET-1, by PKC alpha-, beta II- and epsilon-independent pathways, explaining, at least in part, increased plasma concentrations of endothelial proteins and peptides in the hyperthyroid state.