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61.
BackgroundPremature neonates might be exposed to toxic metals during their stay in the neonatal intensive care unit (NICU), which could adversely affect neurodevelopment; however, limited evidence is available. The present study was therefore designed to assess the exposure to mercury, lead, cadmium, arsenic, and manganese of preterm neonates who received total parenteral nutrition (TPN) and/or red blood cell (RBC) transfusions during their NICU stay and the risk of neurodevelopment delay at the age of 2 months.MethodsWe recruited 33 preterm neonates who required TPN during their NICU admission. Blood samples were collected for metal analysis at two different time points (admission and before discharge). Metals in the daily TPN received by preterm neonates were analyzed. Neurodevelopment was assessed using the Ages and Stages Questionnaire Edition 3 (ASQ-3).ResultsAll samples of TPN had metal contamination: 96% exceeded the critical arsenic limit (0.3 μg/kg body weight/day); daily manganese intake from TPN for preterm neonates exceeded the recommended dose (1 µg/kg body weight) as it was added intentionally to TPN solutions, raising potential safety concerns. All samples of RBC transfusions exceeded the estimated intravenous reference dose for lead (0.19 µg/kg body weight). Levels of mercury, lead and manganese in preterm neonates at discharge decreased 0.867 µg/L (95% CI, 0.76, 0.988), 0.831 (95%CI, 0.779, 0.886) and 0.847 µg/L (95% CI, 0.775, 0.926), respectively. A decrease in ASQ-3-problem solving scores was associated with higher levels of blood lead in preterm neonates taken at admission (ß = −0.405, 95%CI = −0.655, −0.014), and with plasma manganese (ß = −0.562, 95%CI = −0.995, −0.172). We also observed an association between decreased personal social domain scores with higher blood lead levels of preterm neonates before discharge (ß = −0.537, 95%CI = −0.905, −0.045).ConclusionOur findings provide evidence to suggest negative impacts on the neurodevelopment at 2 months of preterm infants exposed to certain metals, possibly related to TPN intake and/or blood transfusions received during their NICU stay. Preterm neonates may be exposed to levels of metals in utero. 相似文献
62.
The human platelet integrin GPIIb/IIIa (228 kDa), a Ca-dependent heterodimer formed by the IIb subunit (GPIIb, 136 kDa) and the 3 subunit (GPIIIa, 92 kDa), serves as the fibrinogen receptor at the surface of activated platelets. The degree of dissociation of the GPIIb/IIIa heterodimer (s°20
*, 8.9 S) into its constituent glycoproteins (GPIIb, 5.8 S; and GPIIIa, 3.9 S) has been assessed by analytical ultracentrifugation in Triton X100 buffers, and its Ca2+- and temperature-dependence correlated with Ca2+-binding to GPIIb/IIIa and its temperature dependence. At 21°C half-maximal dissociation of GPIIb/IIIa occurs at 5.5 ± 2.5 × 10–8 M Ca2+, very close to the dissociation constant of the high affinity Ca-binding site of GPIIb/IIIa (Kd1 8 ± 3 × 10–8 M) (Rivas and González-Rodríguez, 1991) and much lower than the Kd of the 3.4 medium affinity Ca-binding sites (Kd2 4 ± 1.5 × 10–5 M), which seems to demonstrate that the stability of the heterodimer in solution at room temperature is regulated by the degree of saturation of the high-affinity Ca-binding site. At 4°C, the stability of the heterodimer is apparently Ca2+-independent, while at room and physiological temperatures (15–37°C) the degree of dissociation of the heterodimer is regulated by the degree of dissociation of the high- and medium-affinity Ca-binding sites, respectively. On increasing the Ca2+ concentration up to 1 × 10–4 M after dissociation in Triton X100 solutions, the reconstitution of the GPIIb/IIIa heterodimer depends on the time and temperature at which the dissociated heterodimer was maintained, being almost complete within the first 5–10 min at 37°C and within the first 1–2 h at 21°C. After this time, a time- and temperature-dependent irreversible autoassociation of GPIIb (covalent) and GPIIIa (non-covalent) occurs, which hinders both the isolation of permanently stable monoamers of GPIIb and GPIIIa and the reconstitution of the GPIIb/IIIa heterodimer in Triton X100 solutions.
Abbreviations: GPIIb, GPIIIa, and GPIIb/IIIa, glycoprotein IIb, IIIa, and the heterodimer formed by them, respectively; s°20
*, the sedimentation coefficient of the glycoprotein-detergent complexes determined at 20°C, after extrapolation to zero-glycoprotein concentration
Offprint requests to: J. González-Rodríguez 相似文献
63.
Calmodulin copurifies with platelet plasma membranes isolated by glycerol-induced lysis and density gradient centrifugation. These membranes also bind 125I-labeled calmodulin in vitro in the presence of Ca2+. Binding is largely reduced by replacing Ca2+ by Mg2+ or by addition of an excess unlabeled calmodulin. The specific component of binding is saturable, with an apparent Kd of 27 nM and a maximum of 15.9 pmol binding sites per mg of membrane protein. This is equivalent to approx. 4100 binding sites per platelet. Binding was inhibited by addition of phenothiazines, a group of calmodulin antagonists. Half-maximal inhibition was attained with approx. 20 μM trifluoperazine or 50 μM chlorpromazine. In contrast, chlorpromazine-sulfoxide which is inactive towards calmodulin, did not affect the binding. Calmodulin binding polypeptides of the plasma membrane were identified by a gel-overlay technique. A major calmodulin-binding component of molecular weight 149 000 was detected. Binding to this band was Ca2+-dependent and inhibited by chlorpromazine. The molecular weight of this polypeptide is similar to that of glycoprotein I and also that of the red cell (Ca2+ + Mg2+)-stimulated ATPase, which is known to bind calmodulin. The possible role of calmodulin in platelet activation is analysed. 相似文献
64.
We investigated the restoration of [Ca2+]i in fura-2-loaded human platelets following discharge of internal Ca2+ stores in the absence of external Ca2+. After stimulation by thrombin [Ca2+]i returned from a peak level of 0.6 μM to resting levels within 4 min. When ionomycin discharged the internal stores the recovery was slower with [Ca2+]i still elevated at around 0.5 μM after 5 min. Thrombin added shortly after ionomycin could accelerate the recovery of [Ca2+]i and restore resting levels within 5 min, an effect that was mimicked by phorbol-12-myristate-13-acetate (PMA). Since the continued presence of ionomycin precluded reuptake into the internal stores we conclude that thrombin and PMA stimulate Ca2+ efflux, perhaps via protein kinase C actions on a plasma membrane Ca2+ pump. 相似文献
65.
Sergio Grinstein Wendy Furuya 《Biochimica et Biophysica Acta (BBA)/Molecular Cell Research》1984,803(4):221-228
The site and mechanism of accumulation of acridine derivatives into platelets and their isolated organelles were investigated. In addition, their suitability as indicators of cytoplasmic pH was analysed. Direct microscopic observation showed that quinacrine and 9-aminoacridine are concentrated inside organelles in platelets. Using fractionation studies, the acridine derivatives were found to accumulate particularly in dense and α-granules. Uptake into these organelles is driven by a pH differential across their membrane (acidic inside). Because of their cellular distribution, acridine derivatives were found to be poor indicators of cytoplasmic pH. In contrast, a poorly permeant dicarboxylated fluorescein derivative, generated in situ by cytosolic enzymes, is shown to be a more reliable probe of intracellular pH. The results are compared with previous reports of the use of 9-aminoacridine as a cytoplasmic pH probe in platelets and of quinacrine as a selective dense-granule marker. 相似文献
66.
Trifluoperazine, chlorpromazine and other drugs to inhibit calmodulin-dependent processes are also known to inhibit protein kinase C. The effect of these agents on secretion evoked by known activators of C-kinase has been studied in human platelets loaded with the fluorescent Ca indicator, quin2 and preincubated with aspirin. The secretory response stimulated by phorbol ester and exogenous diacyglycerol, at basal levels of cytoplasmic free Ca2+, [Ca2+]i, was suppressed by trifluoperazine, chlorpromazine and W-7, as was the secretion evoked by collagen that occurs without a change in [Ca2+]i, The response to thrombin, which is accompanied by elevated [Ca2+]i was barely affected. Modest elevation of [Ca2+]i by Ca ionophore was able to overcome the inhibitory effect of these drugs on the response to phorbol ester, diacylglycerol, and collagen. 相似文献
67.
Platelet glycerol lysis membranes and α-granule membranes were compared with respect to protein and lipid composition. Crossed immunoelectrophoresis using antibodies against whole platelets, and sodium dodecyl sulphate polyacrylamide gel electrophoresis, revealed the presence of the glycoproteins IIb and IIIa, myosin and an antigen termed G4 in both membrane fractions. The glycoproteins Ia, Ib and IIIb, in addition to β2-microglobulin and actin, appeared specific for the glycerol lysis membranes, whereas two antigens, termed G8 and G18, were observed only in the α-granule membranes. The localization of glycoprotein IIa was inconclusive. Comparison with the surface-located proteins revealed that the glycerol lysis membranes represented a reasonable approximation to a plasma membrane preparation. Radioactively labelled immunoprecipitates obtained after crossed immunoelectrophoresis of 125I-labelled platelets were cut out and applied to sodium dodecyl sulphate electrophoresis on polyacrylamide slab gels. Autoradiography of the dried gels revealed that antigen G4 represented a protein with an average molecular weight of 146 000 in its unreduced state and 132 000 in its reduced state. Antigen G18 represented a protein of molecular weight 130 000–135 000 in the reduced as well as unreduced state. Quantitation of protein and lipids showed that the α-granule membranes contained about one-third as much cholesterol and 2-times as much protein in relation to phospholipids as compared to the glycerol lysis membranes. No significant difference between the two membrane preparations was found as regards the composition of their phospholipids. 相似文献
68.
(1) Exposure of phospholipids at the outer surface of activated and control platelets was studied by incubation with a mixture of phospholipase A2 from Naja naja and bee venom, solely or in combination with sphingomyelinase from Staphylococcus aureus, using conditions under which cell lysis remained below 10%. (2) Incubation with phospholipase A2 alone revealed a markedly increased susceptibility of the phospholipids in platelets activated by a mixture of collagen plus thrombin, by the SH-oxydizing compound diamide, or by calcium ionophore A23187, as compared to control platelets or platelets activated separately by collagen or thrombin. (3) Collagen plus thrombin, diamide, and ionophore treated platelets revealed an increased exposure of phosphatidylserine at the outer surface accompanied by a decreased exposure of sphingomyelin, as could be concluded from incubations with a combination of phospholipase A2 and sphingomyelinase. These alterations were much less apparent in platelets activated either by thrombin or by collagen alone. (4) The increased exposure of phosphatidylserine in activated platelets is accompanied by an increased ability of the platelets to enhance the conversion of prothrombin to thrombin by coagulation factor Xa, in the presence of factor Va and calcium. (5) It is concluded that the altered orientation of the phospholipids in the plasma membrane of platelets activated by collagen plus thrombin, by diamide, or by calcium ionophore, is the result of a transbilayer movement. Moreover, the increased exposure of phosphatidylserine in platelets stimulated by the combined action of collagen and thrombin might be of considerable importance for the hemostatic process. 相似文献
69.
Christian J. Le Peuch Danielle A.M. Le Peuch Sidney Katz Jacques G. Demaille Maxwell T. Hincke R. Bredoux J. Enouf S. Levy-Toledano Jacques Caen 《生物化学与生物物理学报:生物膜》1983,731(3):456-464
Calcium-accumulating vesicles were isolated by differential centrifugation of sonicated platelets. Such vesicles exhibit a activity of about 10 nmol (min·mg)?1 and an ATP-dependent Ca2+ uptake of about 10 nmol (min·mg)?1. When incubated in the presence of Mg[γ-32P]ATP, the pump is phosphorylated and the acyl phosphate bond is sensitive to hydroxylamine. The [32P]phosphate-labeled Ca2+ pump exhibits a subunit molecular weight of 120 000 when analyzed by lithium dodecyl sulfate-polyacrylamide gel electrophoresis. Platelet calcium-accumulating vesicles contain a 23 kDa membrane protein that is phosphorylatable by the catalytic subunit of cAMP-dependent protein kinase but not by protein kinase C. This phosphate acceptor is not phosphorylated when the vesicles are incubated in the presence of either Ca2+ or Ca2+ plus calmodulin. The latter protein is bound to the vesicles and represents 0.5% of the proteins present in the membrane fraction. Binding of 125I-labeled calmodulin to this membrane fraction was of high affinity (16 nM), and the use of an overlay technique revealed four major calmodulin-binding proteins in the platelet cytosol (). Some minor calmodulin-binding proteins were enriched in the membrane fractions (). When the vesicles are phosphorylated in the presence of MgATP and of the catalytic subunit of cAMP-dependent protein kinase, the rate of Ca2+ uptake is essentially unaltered, while the Ca2+ capacity is diminished as a consequence of a doubling in the rate of Ca2+ efflux. Therefore, the inhibitory effect of cAMP on platelet function cannot be explained in such simple terms as an increased rate of Ca2+ removal from the cytosol. Calmodulin, on the other hand, was observed to have no effect on the initial rate of calcium efflux when added either in the absence or in the presence of the catalytic subunit of the cyclic AMP-dependent protein kinase, nor did the addition of 0.5 μM calmodulin result in increased levels of vesicle phosphorylation. 相似文献
70.
Valery N. Bochkov Tatyana A. Voino-Yasenetskaya Vsevolod A. Tkachuk 《生物化学与生物物理学报:疾病的分子基础》1991,1097(2):123-127
Low density lipoproteins activate phosphoinositide turnover, increase free cytoplasmic calcium concentration and stimulate phosphorylation of 20- and 47-kDa proteins in blood platelets. All these effects are substantially potentiated by epinephrine. 相似文献