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Biomedical subjects

L Berry

Publications and source records attributed to L Berry.

At least 19 recordsLinked to original sources
PubMed2002

Blood-compatible biomaterials by surface coating with a novel antithrombin-heparin covalent complex.

Covalent antithrombin-heparin complex (ATH) was covalently grafted to a polycarbonate urethane (Corethane) endoluminal graft (a kind gift of Corvita Corporation) after being activated using 0.3% m/m NaOCl in 0.15 M phosphate pH 6.0. ATH graft density (1.98 x 10(-7) mol/m2) was 6 times the maximum amount of unfractionated heparin (UFH) that could be bound to polycarbonate urethane surfaces. Surface-bound ATH could be stored in sterile 0.15 M NaCl at 4 degrees C for at least 2 months with good antithrombotic activity before being implanted into rabbits. Analysis of ATH-coated tubing showed that it contained significant direct thrombin inhibitory activity. In vivo testing in a rabbit model was compared to non-activated non-coated surfaces, activated-non-coated surfaces, hirudin-coated surfaces and antithrombin (AT)-coated surfaces. The weight of the clot generated in the ATH-coated graft tubing was significantly less than the weight of the clot generated within the hirudin-coated graft (p = 0.03 with a 1-tailed Student's t test). The anticoagulant nature of ATH grafts in vivo was shown to be due to bound ATH because boththe AT-coated surfaces and non-coated but activated surfaces showed similar thromboresistant efficacy to that of untreated material (ANOVA; p < 0.05). Apart from the direct antithrombin activity that contributed to much of the prolonged patency in vivo, surface-bound ATH likely catalyzed AT inhibition of thrombin, as evidenced by a significant number of 125I-AT binding sites (> or = 1.5 x 10(-8) mol/m2). Thus, ATH appears to be a good candidate for coating cardiovascular devices, such as endoluminal grafts, with high levels of substitution and significant long-term blood-compatibility.

Animals↗
PubMed2001

In vitro and in vivo antimalarial activity of ferrochloroquine, a ferrocenyl analogue of chloroquine against chloroquine-resistant malaria parasites.

Previous studies have shown that ferrochloroquine (FQ) exhibited an antimalarial activity against Plasmodium spp. The present work confirmed this activity, described the curative effect on P. vinckei and investigated the FQ toxicity in vitro and in vivo. The in vitro and in vivo growth inhibition of P. falciparum and P. berghei N, respectively, showed that FQ antimalarial activity was 1.5-10 times more potent than chloroquine. FQ completely inhibited the in vivo development of both chloroquine-susceptible and resistant P. vinckei strains and protected mice from lethal infection at a dose of 8.4 mg kg(-1) day(-1) given for 4 days subcutaneously or orally. This curative effect was 5-20 times more potent than chloroquine, according to the strains' resistance to chloroquine. At this curative dose, no clinical changes were observed in mice up to 14 days after the last administration. Nevertheless, the acute toxicity and lethality of ferrochloroquine seemed to be dependent on gastric surfeit. The FQ security index determined in vitro confirmed that it might be a promising compound.

Administration, Oral↗
PubMed2001

Plasmodium falciparum signal sequences: simply sequences or special signals?

The malaria parasite, Plasmodium falciparum, synthesises and exports several proteins inducing morphological and biochemical modifications of erythrocytes during the erythrocytic cycle. The protein trafficking machinery of the parasite is similar to that of other eukaryotic cells in several ways. However, some unusual features are also observed. The secretion of various polypeptides was inhibited when P. falciparum-infected erythrocytes were incubated with Brefeldin A. Immunoelectron microscopy studies revealed substantial morphological changes in the endoplasmic reticulum following exposure of parasitised erythrocytes to the drug. Immunofluorescence studies of Brefeldin A-treated parasites suggest that polypeptide sorting to different intracellular destinations begins at the endoplasmic reticulum. The parasite also secretes polypeptides by a Brefeldin A-insensitive route that bypasses the classical endoplasmic reticulum-Golgi complex pathway.

Animals↗
PubMed2000

Pfsbp1, a Maurer's cleft Plasmodium falciparum protein, is associated with the erythrocyte skeleton.

Antibodies from hyperimmune monkey sera, selected by absorption to Plasmodium falciparum-infected erythrocytes, and elution at acidic pH, allowed us to characterize a novel parasite protein, Pfsbp1 (P. falciparum skeleton binding protein 1). Pfsbp1 is an integral membrane protein of parasite-induced membranous structures associated with the erythrocyte plasma membrane and referred to as Maurer's clefts. The carboxy-terminal domain of Pfsbp1, exposed within the cytoplasm of the host cell, interacts with a 35 kDa erythrocyte skeletal protein and might participate in the binding of the Maurer's clefts to the erythrocyte submembrane skeleton. Antibodies to the carboxy- and amino-terminal domains of Pfsbp1 labelled similar vesicular structures in the cytoplasm of Plasmodium chabaudi and Plasmodium berghei-infected murine erythrocytes, suggesting that the protein is conserved among malaria species, consistent with an important role of Maurer's cleft-like structures in the intraerythrocytic development of malaria parasites.

Amino Acid Sequence↗
PubMed2000

Novel ferrocenic artemisinin derivatives: synthesis, in vitro antimalarial activity and affinity of binding with ferroprotoporphyrin IX.

Following our search for novel compounds with high antimalarial activity, a series of artemisinin (QHS) derivatives containing a ferrocenic nucleus was prepared and tested in vitro against Plasmodium falciparum strains. Two new metallocenic derivatives (1 and 3) were found as potent as QHS. All compounds showed a capacity to bind with ferroprotoporphyrin IX. A decrease in the Soret band absorbance of ferroprotoporphyrin IX, resulting from the addition of different drugs concentrations, was shown. The association stoichiometry of compounds to ferroprotoporphyrin IX appears to be 1:2 at equilibrium, with an intermediate 1:1 complexation. These results appear to strengthen the role of adducts between artemisinin derivatives and heme in generation of artemisinin radicals. Such interaction of artemisinin ferrocenyl derivatives with ferroprotoporphyrin IX and its biological significance could form a basis in future drug development.

Animals↗
PubMed2000

Influence of exogenous factor VIIa on thrombin generation in cord plasma of full-term and pre-term newborns.

Factor (F) VIIa has been used to treat adults and children with a variety of bleeding disorders. The results from these studies cannot be extrapolated to newborns because their hemostatic system differs significantly from adults, which may influence the effects of FVIIa on thrombin (IIa) generation. We compared the effects of FVIIa concentrates on IIa generation in plasmas from adults, full-term newborns and pre-term newborns. Defibrinated plasma (using arvin) from adults, or umbilical cords from full-term or pre-term deliveries was supplemented with FVIIa (Novo Nordisk, Bagsvaerd, Denmark), mixed with dilute thromboplastin reagent, and the resultant reaction mixture subsampled periodically into ethylenediamine tetraacetic acid, followed by measurement of total IIa activity (S-2238). Thrombin-alpha2 macroglobulin complexes, determined as residual activity after neutralization with heparin and antithrombin, were subtracted from total IIa to give free IIa. Prothrombin (FII) and inhibitor complexes were measured by enzyme-linked immunosorbent assays. Addition of FVIIa caused a reduction in the lag phase for the appearance of free IIa and consumption of FII, which was more pronounced in newborn plasma. There was no increase in peak IIa levels regardless of the amount of FVIIa added. Final inhibitor complex concentrations were increased in plasmas from adults compared with newborns, likely reflecting higher plasma concentrations of FII in adults. Generation of IIa was more rapid in pre-term plasma compared with that in adult and full-term cord plasmas due to increased endogenous tissue factor (TF). In summary, FVIIa enhanced IIa generation in plasma from different age groups, with the effect being more pronounced in plasma from pre-term newborns, possibly due to increased levels of plasma TF.

Adult↗
PubMed1999

The transport of the histidine-rich protein I from Plasmodium falciparum is insensitive to brefeldin A.

During its intraerythrocytic development, Plasmodium falciparum synthesizes several proteins that are exported beyond its membrane. Some of these secreted antigens are involved in the formation of protuberances or knobs, a major virulence factor, at the erythrocyte membrane. Various secreted malarial polypeptides, the transport of which is sensitive to brefeldin A, are translocated in vitro into dog pancreatic microsomes. We present evidence that the histidine-rich protein I (PfHRPI) is secreted by the parasite via a novel pathway, independent of the ER/Golgi apparatus. The secretion of PfHRPI was not blocked by incubation of parasite cultures at 15 degrees C and 20 degrees C or 37 degrees C in the presence of brefeldin A. PfHRPI was not translocated into microsomes in an in vitro translation-translocation cell-free system. Unlike other polypeptides from eukaryotic cells that bypass the ER/Golgi pathway and do not have a signal peptide, PfHRPI has an atypical signal sequence consisting of 21 amino acids, including eight positively charged residues followed by 11 hydrophobic residues. We also found that the unusually charged PfHRPI signal sequence diverts Exp-1, which is usually exported, away from the translocation machinery of microsomal membranes.

Animals↗
PubMed1998

Investigation of the anticoagulant mechanisms of a covalent antithrombin-heparin complex.

Recently, we developed a covalent antithrombin-heparin complex (ATH) as a possible treatment for respiratory distress syndrome. ATH reacted rapidly with thrombin and efficiently catalyzed the inhibition of either thrombin or factor Xa by exogenous antithrombin. In order to investigate mechanisms for the conjugate's unusual anticoagulant properties, changes in fluorescence due to covalent linkage or addition of exogenous antithrombin were studied in relation to reaction with thrombin derivatives or factor Xa. The emission spectrum of ATH was similar to that of antithrombin plus heparin mixtures. ATH quickly inhibited thrombin or factor Xa activities, as measured by a fluorogenic substrate. Fluorescein-labeled heparin was displaced from either thrombin or active site blocked thrombin by ATH, indicating that thrombin must bind to the conjugate's heparin moiety. Interaction of thrombin with ATH's heparin component was confirmed by a slow reaction rate of conjugate with a thrombin mutant that has weak heparin binding. Total intrinsic fluorescence increased when exogenous antithrombin was added to ATH, indicating that the catalytic mechanism may occur through a second inhibitor binding site. Thus, ATH reacts directly with thrombin through a bridge mechanism and probably catalyzes the reaction of thrombin with antithrombin by a second binding sequence on its heparin chain.

Anticoagulants↗
PubMed1998

Covalent heparin cofactor II-heparin and heparin cofactor II-dermatan sulfate complexes. Characterization of novel anticoagulants.

Heparin cofactor II is a naturally occurring anticoagulant that acts by specifically inhibiting thrombin and is facilitated by the binding of glycosaminoglycans such as heparin and dermatan sulfate. In vivo, heparin cofactor II-glycosaminoglycan complexes dissociate, leaving the inhibitor less active in its ability to function as a component of the anticoagulation pathway. We have produced permanently activated heparin cofactor II molecules by covalent linkage to either heparin or dermatan sulfate. Covalent heparin cofactor II-heparin and heparin cofactor II-dermatan sulfate complexes had catalytic antithrombin activities similar to those of the corresponding starting heparin and dermatan sulfate (86% and 110% of standard heparin and dermatan sulfate activity, respectively). Both heparin cofactor II-heparin and heparin cofactor II-dermatan sulfate had fast bimolecular rate constants of 1.4 x 10(7) M-1 s-1 and 1.3 x 10(7) M-1 s-1, respectively, for reaction with thrombin. The intravenous half-life of the covalent complexes in rabbits was significantly longer than that of free heparin or dermatan sulfate (4.4, 3.4, 0.33, and 0.50 h for heparin cofactor II-heparin, heparin cofactor II-dermatan sulfate, heparin, and dermatan sulfate, respectively). Given their unique properties, these conjugates may have a clinical application for long term, selective inhibition of thrombin.

Animals↗
PubMed1998

Anticoagulant dermatan sulfate proteoglycan (decorin) in the term human placenta.

Normal pregnancy is characterized by increased in vivo thrombin generation. A greater proportion of endogenously generated thrombin is complexed to heparin cofactor II in plasma from pregnant women compared to plasma from nonpregnant ones. The increase in thrombin-heparin cofactor II complexes suggests that the site of the additional thrombin generation is relatively rich in dermatan sulfate. We postulated that the site of thrombin generation may be the placenta since endogenous thrombin generation returns rapidly to normal after delivery. This report describes the isolation and characterization of placental dermatan sulfate proteoglycan from villous tissue of the term human placenta. Placental dermatan sulfate was isolated by guanidine HCI extraction and anion exchange chromatography. The isolated material was found to have anticoagulant activity with a relative activity of approximately 40% of that of a porcine mucosal dermatan sulfate which is undergoing clinical trial as an antithrombotic agent. The dermatan sulfate was present as a proteoglycan with a molecular mass of 90-150 kD. Upon degradation with chondroitin ABC lyase, the core protein was demonstrated to be a doublet with molecular masses of 42 and 44 kD. This core protein reacted with antiserum against the core protein of decorin on Western analysis. The role of this placental dermatan sulfate in local regulation of thrombin in the placenta warrants further study.

Alcian Blue↗
PubMed1998

Bioactive and immunoreactive FSH concentrations in ewe and ram lambs over the first year of life.

Several studies suggest that the concentration of immunoreactive (I) FSH measured in peripheral plasma by radioimmunoassay does not always reflect the level of bioactive (B) hormone capable of eliciting a biological response (e.g. oestradiol synthesis by Sertoli cells in vitro). The aim of this study was to measure both B-FSH and I-FSH concentrations in male and female sheep during the first year of life, and to relate this to pubertal development. The hypothesis being tested was that B-FSH is present in both male and female sheep during the prepubertal period and that discrete changes in B-FSH are associated with the onset of puberty. Eight ewe lambs and eight rams lambs were blood sampled fortnightly form 2 to 52 weeks of age. All samples were assayed for B-FSH content. Pubertal development was monitored in ewe lambs from behavioural oestrus and from plasma progesterone concentrations, and in ram lambs from penile and testicular development and from plasma testosterone concentrations. Mean I-FSH concentrations varied significantly with time after birth, in both females and males (P < 0.01). In contrast, B-FSH was found to vary with time in females only (P < 0.01). Around the expected time of puberty in ram lambs (i.e. at 30-40 weeks of age), and thereafter, I-FSH concentrations were undetectable (< 0.2 ng ml-1), whereas the B-FSH concentrations were measurable at concentrations up to twice the assay detection limit (0.8 ng ml-1) until 38 weeks of age. In ewe lambs, but not ram lambs, there was a significant linear relationship between B-FSH and I-FSH values (R = 0.595; P < 0.005). When standardised about the time of puberty, B-FSH (P < 0.05) but not I-FSH was significantly higher in ewe lambs that failed to reach puberty. No differences for either B-FSH or I-FSH between pubertal and non-pubertal ram lambs were noted. In summary, B-FSH was soften measurable in plasma throughout prepubertal development in sheep and the concentrations often differed from those of I-FSH, especially in ram lambs. However, there appeared to be no discrete change in B-FSH that could be directly related to specific pubertal events. It is concluded that although FSH may be a prerequisite for prepubertal testicular development and/or ovarian follicular growth, it is not a critical factor in determining whether puberty is attained during the first year of life in this seasonally breeding species.

Aging↗
PubMed1998

Polypeptide-polysaccharide conjugates produced by spontaneous non-enzymatic glycation.

A fundamental dogma has developed over the past 20 years that non-enzymatic glycation involving saccharide chains of greater than 3 to 4 residues is an extremely unlikely reaction. Our investigations using glycosaminoglycans have shown that, given sufficient time, polypeptide-polysaccharide conjugates form via the Schiff base-Amadori rearrangement mechanism. Further, even though these straight chain polysaccharides are relatively charged and sterically hindered, spontaneous glycation can also occur in vivo. A complete reinvestigation of all aldose terminating polysaccharides is required to elucidate this new class of macromolecules, which is likely to contain unusual polypeptide-polysaccharide combinations and functions.

Animals↗
PubMed1998

Effect of a novel covalent antithrombin-heparin complex on thrombin generation on fetal distal lung epithelium.

Respiratory distress syndrome is characterized by fibrin deposition in the lung. Fibrin adversely affects surfactant function and stimulates proliferation of fibroblasts. There is evidence that these properties may be important to the development of bronchopulmonary dysplasia. Despite successful initial treatment of neonatal respiratory distress syndrome with surfactant, the incidence of bronchopulmonary dysplasia has not decreased. In previous studies, it has been demonstrated that rat fetal distal lung epithelium (FDLE) possesses both procoagulant and anticoagulant properties. In this report, we have demonstrated (using factor VII-deficient plasma) that tissue factor is expressed on the FDLE surface and promotes thrombin generation. To regulate thrombin within this procoagulant environment, we have developed a novel anticoagulant, antithrombin-heparin covalent complex (ATH) that can be retained within the lung after intrapulmonary instillation. We have demonstrated that ATH was superior to antithrombin plus standard heparin in suppressing thrombin generation (P < 0.001) and prothrombin consumption (P < 0.01) in recalcified defibrinated plasma on the surface of FDLE. Further studies with ATH in vivo need to be performed.

Adult↗
PubMed1998

Influence of mechanical stretch on thrombin regulation by fetal mixed lung cells.

Respiratory distress syndrome (RDS) is characterized by intrapulmonary fibrin deposition, which can adversely affect surfactant function, and stimulate fibroblast proliferation, which may contribute to the development of bronchopulmonary dysplasia (BPD). We speculated that the premature lung may have impaired regulation of thrombin, thus making preterm infants susceptible to fibrin formation within the lung. Therefore, we studied the effect of stretch, which simulates fetal breathing movements (FBMs), on the generation and inhibition of a key hemostatic enzyme-thrombin-by rat fetal mixed lung cells (FMLCs). Our results showed that stretch induced glycosaminoglycan production with increased antithrombin activity due to an increase in the concentration of active chondroitin sulfate. Stretch downregulated secretion of tissue factor procoagulant activity, which may lead to decreased thrombin generation on the surface of FMLCs. Overall, stretch enhanced the local control of thrombin by FMLCs. These results suggest that premature infants, who will have experienced less FBM, may have impaired thrombin regulation. Impaired thrombin regulation likely contributes to increased fibrin deposition and, potentially, the development of BPD.

Animals↗
PubMed1998

Morphologic and biochemical features affecting the antithrombotic properties of the inferior vena cava of rabbit pups and adult rabbits.

The incidence of venous thromboembolic disease is reduced in children compared with adults. Thromboprotective mechanisms, some of which have already been identified in plasma, must be present in children. Blood vessel walls have important antithrombotic properties that maintain blood fluidity. This is in part due to proteoglycan (PG)-related glycosaminoglycan (GAG) molecules within vessel walls. PGs are macromolecules with covalently attached GAG chains, either chondroitin, dermatan, heparan, or keratan sulfate. The influence of age on the concentration and anticoagulant activities of PGs and GAGs, within vein walls before puberty has not been previously investigated. We hypothesized that developmental differences in vein walls may contribute to the reduced risk of thrombosis in children. We used a rabbit model to examine morphologic and biochemical features of inferior venae cavae (IVCs). We assessed IVC wall morphology, PG distribution, GAG mass, and GAG antithrombin activity. Morphologically, there were only minor differences between pups and adult rabbits' IVCs. However, there was a significant increase in GAGs by mass in IVCs from pups compared with adult rabbits (p = 0.012). In addition the total antithrombin activity (p = 0.04), and especially that of heparan sulfate (p = 0.01) was significantly increased in pups compared with adult rabbits. These results demonstrate important differences in the antithrombotic properties of IVC walls in pups and adult rabbits. In summary, developmental differences in vein wall PG content and activity exist which may contribute to the reduced risk of venous thromboembolism in children. Further characterization of these differences is required.

Age Factors↗
PubMed1998

Morphological and biochemical features affecting the antithrombotic properties of the aorta in adult rabbits and rabbit pups.

We hypothesised that there are important physiologic differences in arterial wall structure and function with respect to antithrombotic activity in the very young (pre-puberty) compared to adults. Electron microscopy, gel electrophoresis, and activity assays were used to examine differences in aorta structure and function comparing prepubertal rabbits (pups) to adult rabbits. Differences in endothelial function, extracellular matrix structure, proteoglycan (PG) distribution and glycosaminoglycan (GAG) content and function were shown. In both intima and media, total PG, chondroitin sulfate (CS) PG and heparan sulfate (HS) PG content were significantly increased in pups compared to adult rabbits. These findings corresponded to increased concentrations by mass analyses of CS GAG and DS GAG in aortas from pups. There was also a significant increase in antithrombin activity in pups due to HS GAG. In conclusion, differences in both structure and antithrombin activity of aortas from pups compared to adult rabbits suggest that young arteries may have greater antithrombotic potential that is, at least in part, related to increased HS GAG.

Aging↗
PubMed1998

Enhanced thrombin regulation during warfarin therapy in children compared to adults.

The intensity of warfarin therapy for prevention of primary and secondary thromboembolic complications in paediatric patients, is extrapolated from guidelines for adults, which may not be optimal. Therefore, we assessed thrombin regulation ex vivo and in vitro in plasmas from 40 children (1 to 18 years old with a median age of 13 years) and 27 adults receiving warfarin with an international normalized ratio of 2 to 3 (child: 2.5 +/- 0.15; adult: 2.4 +/- 0.14). Ex vivo concentrations of prothrombin fragment 1.2 were significantly lower in children (0.30 +/- 0.03 nM) compared to adults (0.45 +/- 0.04 nM; p <0.01). Thrombin generation in defibrinated plasmas (Arvin) was decreased and delayed for children compared to adults when activated by either activated partial thromboplastin time (child = 32 +/- 1.7, adult = 45 +/- 1.9 microM x s) or prothrombin time (child = 35 +/- 0.7, adult = 46 +/- 1.0 microM x s) reagents (p <0.01 for both). Although plasma concentrations of factors (F) II, FVII, FIX, F X, protein C and protein S were similar, more of the thrombin generated was complexed to alpha2 macroglobulin (alpha2M) at times close to peak thrombin activity (60 s) in plasma from children (general linear analysis of variance; p <0.03). Thus, increased alpha2M levels may enhance thrombin regulation in paediatric compared to adult patients receiving warfarin, suggesting that clinical trials in children, using less intense warfarin treatment, may be required to determine optimum therapy.

Adolescent↗