Tissue factor pathway inhibitor activity in factor VII deficiency.
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Anticoagulants↗
Biomedical subjects
R Paolini
Publications and source records attributed to R Paolini.
Kinase activation through the high-affinity receptor for immunoglobulin E.
The high-affinity receptor for IgE (Fc epsilon RI) belongs to a class of multimeric receptors associated with nonreceptor tyrosine kinases. It has been assumed that Fc epsilon RI beta and gamma chains, which have extensive cytoplasmic domains, play an important, although undefined role in coupling the receptor to signal transduction mechanisms. The results reviewed here suggest a synergistic effect of these two chains in the initiation of Fc epsilon RI signaling. According to our model, receptor engagement can activate kinase(s), such as lyn, already bound to the receptor under resting conditions. The receptor phosphorylation following this activation can be responsible for recruitment and activation of other signaling molecules, such as syk, which can then activate downstream effector molecules. This model could be extended to include other multimeric receptors, such as the T- and B-cell receptors and the low-affinity receptor for IgG (Fc gamma RIII), that control the activation of cytoplasmic tyrosine kinases.
Amino Acid Sequence↗
Anti-hepatitis C virus serology in patients affected with congenital coagulation defects: a comparative study using three second generation ELISA tests.
We determined the prevalence of anti-hepatitis C virus (HCV) antibodies in 34 patients affected with congenital coagulation disorders attending the Haemophilia Centre of Padua, Italy. Serological tests were carried out by three second generation enzyme linked immunosorbent assays (ELISA), two based on recombinant proteins (Ortho and Abbott) and one based on synthetic peptides (Behring) as antigenic substrate. The repeatedly reactive specimens were further assayed by the supplemental 4-antigen recombinant immunoblot assay (RIBA) (Chiron and Ortho). Moreover, we performed the dot-blot Matrix test (Abbott) on the samples showing discrepant results by the three ELISA tests. Twenty-six patients (76.5%) were anti-HCV positive using all three ELISA tests; 25 were confirmed by the supplemental RIBA test, the other one was indeterminate. Two samples were in a gray-zone only using the anti-HCV ELISA Abbott. These were positive by the RIBA; in contrast, such samples showed no reactivity with the Matrix test. In accordance with the current literature, these data show an equivalence between the 2nd generation screening tests (ELISA), at least when applied to a high risk population as in the present study. Further, these screening tests demonstrated a reliable specificity, since most of the ELISA-reactive specimens were confirmed by the supplemental RIBA test. In contrast, combined use of the anti-HCV tests could be useful when high sensitivity is requested, as in the case of blood donor pretransfusion screening.
Adolescent↗
Measurement of cellular microbicidal activity against Pneumocystis carinii in vitro.
As Pneumocystis carinii cysts cannot be cultivated for enumeration of colony forming cells, two alternative approaches to measuring killing of P. carinii by mouse peritoneal cells were investigated. The cells tested were either normal resident peritoneal cells, or cells which were elicited by intraperitoneal injection of the bacterium Listeria monocytogenes. The latter population showed enhanced antibacterial activity against Listeria organisms and enhanced production of H2O2 in the presence of P. carinii, cysts from immunosuppressed rats. To assess killing of P. carinii, cysts were mixed with peritoneal cells at a ratio of 10:1, and after intervals of incubation the peritoneal cells were lysed by saponin treatment. The viability of the cysts was assessed by staining with vital dyes or by uptake of tritiated uridine over 7 h incubation. Viability of cysts was unaffected by saponin treatment, and there was agreement between the two techniques that the elicited peritoneal cells killed approximately twice the number of cysts over a 20 h incubation period compared to normal resident peritoneal cells.
Animals↗
[Proposal for using mechanical staplers with Polysorb in creation of orthotopic neobladder].
After radical cysto-prostatectomy the bladder substitute is made from an ileal segment, opened along its antimesenteric border and folded; the results are in general good. The success of this procedure demands attention to detail, hemostasis and gentle handling of tissue so this surgery depends an unusual degree of commitment to meticulous technique. The very low rate complications, obtained with mechanical sutures in gastrointestinal surgery, incited the Authors using the staplers GIA and TA Polysorb for detubularized ileal segment. They are very manageable, easy to use and the follow-up shows the absence of urinary stone; so the team approach is less tedious and strenuous. The Authors present their results of 11 neobladder-staplers made with 35 cm of detubularized ileal segment without folding.
Aged↗
Cell surface control of the multiubiquitination and deubiquitination of high-affinity immunoglobulin E receptors.
Multiubiquitination of proteins is a critical step leading to selective degradation for many polypeptides. Therefore, activation-induced multiubiquitination of cell surface receptors, such as the platelet-derived growth factor (PDGF) receptor and the T cell antigen (TCR) receptor, may correspond to a degradation pathway for ligand-receptor complexes. Here we show that the antigen-induced engagement of high-affinity immunoglobulin E receptors (Fc epsilon RI) results in the immediate multiubiquitination of Fc epsilon RI beta and gamma chains. This ubiquitination is independent of receptor phosphorylation and is restricted to activated receptors. Surprisingly, receptor multiubiquitination is immediately reversible when receptors are disengaged. Therefore, multiubiquitination and deubiquitination of Fc epsilon RI receptors is controlled at the cell surface by receptor engagement and disengagement. The rapidity, specificity and, most importantly, the reversibility of the activation-induced receptor multiubiquitination suggest that this process may turn on/off a cell surface receptor signaling function thus far unsuspected.
Animals↗
Mild haemophilia A fails to protect from coronary artery disease: the report of two cases.
The description of two additional cases of coronary artery disease in haemophiliacs is reported. Both patients are affected by mild haemophilia A. The first one was not aware of his coagulation disorder and therefore he had never been treated with FVIII concentrates, despite the occurrence of severe and recurrent articular and gastric bleeding episodes. He underwent cardiac catheterization that gave evidence of extensive coronary atherosclerosis. Percutaneous transluminal coronary angioplasty (PTCA) was then performed. The second patient had an inferior myocardial infarction. Influence of cardiovascular risk factors in haemophiliacs and the possible role of thrombosis in the pathogenesis of atherosclerosis are discussed.
Angioplasty, Balloon, Coronary↗
Effect of recombinant-tissue plasminogen activator, low molecular weight urokinase and unfractionated heparin on platelet aggregation.
We studied the interaction of two thrombolytic agents, recombinant-tissue plasminogen activator (r-TPA) and low molecular weight urokinase (UK), with platelet aggregation in the absence or presence of unfractionated heparin (UH). With platelet rich plasma (PRP), a dose dependent inhibition was seen for both r-TPA and UK. However, this effect was more evident in the presence of r-TPA. UH did not modify this effect. However, it enhanced platelet aggregation induced by ADP and decreased the aggregation induced by collagen (COL) as already shown by us. With washed platelets, only r-TPA decreased platelet aggregation in a dose dependent manner in the presence of COL and only at the highest dose (100 micrograms) in the presence of TH. The presence of ten units of plasminogen (PLG) together with 10 micrograms of r-TPA or 1250 units of UK totally inhibited TH-induced platelet aggregation. UH reversed this effect. In contrast, when COL was the aggregating agent, the inhibition of platelet aggregation in presence of PLG, seems to be further increased by UH. Since UH is an adjunct of thrombolytic therapy to prevent rethrombosis, this double edged sword could partially explain the lack of therapeutic effect in some patients.
Adenosine Diphosphate↗
Phosphorylation/dephosphorylation of high-affinity IgE receptors: a mechanism for coupling/uncoupling a large signaling complex.
Engagement of high-affinity IgE receptors leads to activation of tyrosine and serine/threonine kinases and the immediate phosphorylation of receptor beta (serine and tyrosine) and gamma (threonine and tyrosine) chains. Receptor disengagement leads to dephosphorylation of beta and gamma chains via the action of undefined phosphatases. Here we have identified five distinct polypeptides associated with the high-affinity IgE-receptor tetrameric complex, which apparently become phosphorylated and dephosphorylated in sequence with the beta and gamma chains. Like beta chain, polypeptides pp180, pp48, pp42, and pp28 are phosphorylated on serine and tyrosine, whereas pp125 is only phosphorylated on serine. The phosphorylation of each of these receptor-associated polypeptides is antigen-dose dependent and is restricted to activated receptor complexes. Furthermore the physical association between pp125 and the receptor is quantitatively affected by receptor phosphorylation and dephosphorylation, indicating a coupling-uncoupling mechanism. Finally, in vitro kinase experiments show that activated receptor complexes are also physically associated with tyrosine and serine/threonine kinases as part of a larger complex containing the phosphorylated polypeptides.
Animals↗
Evidence for two distinct phosphorylation pathways activated by high affinity immunoglobulin E receptors.
The high affinity receptor for immunoglobulin (Ig) E on mast cells, along with the antigen receptors on T and B cells and Fc receptors for IgG, belongs to a class of receptors which lack intrinsic kinase activity, but activate non-receptor tyrosine and serine/threonine kinases. Receptor engagement triggers a chain of signaling events leading from protein phosphorylation to activation of phosphatidylinositol-specific phospholipase C, an increase in intracellular calcium levels, and ultimately the activation of more specialized functions. IgE receptor disengagement leads to reversal of phosphorylation by undefined phosphatases and to inhibition of activation pathways. Here we show that phenylarsine oxide, a chemical which reacts with thiol groups and has been reported to inhibit tyrosine phosphatases, uncouples the IgE receptor-mediated phosphorylation signal from activation of phosphatidyl inositol metabolism, the increase in intracellular calcium levels, and serotonin release. Phenylarsine oxide inhibits neither the kinases (tyrosine and serine/threonine) phosphorylating the receptor and various cellular substrates nor, unexpectedly, the phosphatases responsible for the dephosphorylation following receptor disengagement. By contrast, it abolishes the receptor-mediated phosphorylation of phospholipase C-gamma 1, but not phospholipase C activity in vitro. Therefore the phosphorylation and activation of phospholipase C likely requires a phenylarsine oxide-sensitive element. Receptor aggregation thus activates at least two distinct phosphorylation pathways: a phenylarsine oxide-insensitive pathway leading to phosphorylation/dephosphorylation of the receptor and of various substrates and a sensitive pathway leading to phospholipase C-gamma 1 phosphorylation.
Animals↗
Interaction between histidine-rich glycoprotein and platelet factor 4 with dermatan sulfate and low-molecular-weight dermatan sulfate.
There is a controversy about whether or not histidine-rich glycoprotein (HRG), the most abundant plasma protein with glycosaminoglycans-neutralizing capacity, is able to prevent the inhibition of human thrombin by heparin cofactor II (HC II) in the presence of dermatan sulfate (DS). The authors studied the interaction of DS and low molecular weight DS, in a purified system with HRG, platelet factor 4 (PF 4), and with HC II. Their results show that HRG, like PF 4, has an affinity, not only for heparin, but also for DS. However, this affinity seems very weak. In fact, HRG is 10 times less effective than PF 4 in neutralizing the 50% antithrombin activity of HC II in the presence of DS.
Blood Proteins↗
Phosphorylation and dephosphorylation of the high-affinity receptor for immunoglobulin E immediately after receptor engagement and disengagement.
Triggering of mast cells and basophils by immunoglobulin E (IgE) and antigen induces various biochemical signals, including tyrosine kinase activation, which lead to cell degranulation and the release of mediators of the allergic reaction. The high-affinity receptor for IgE (Fc epsilon RI) responsible for initiating these events is a complex structure composed of an IgE-binding alpha-chain, a beta-chain and a homodimer of gamma-chains. It has been assumed that beta and gamma, which have extensive cytoplasmic domains, play an important but undefined role in coupling Fc epsilon RI to signal transduction mechanisms. Here we show that Fc epsilon RI engagement induces immediate in vivo phosphorylation on beta (tyrosine and serine) and gamma (tyrosine and threonine) by at least two different non-receptor kinases. We take advantage of unique features of this receptor system to demonstrate that the phosphorylation signal is restricted to activated receptors and is immediately reversible upon receptor disengagement by undefined phosphatases. Rapid phosphorylation and dephosphorylation may be a general mechanism to couple and uncouple activated receptors to other effector molecules. This could be particularly relevant to other multimeric receptors containing Fc epsilon RI gamma-chains or the related zeta and eta chains such as the T-cell antigen receptor (TCR) and the low-affinity receptor for immunoglobulin G (Fc gamma RIII, CD16).
Animals↗
Heparin released platelet factor 4 in uncomplicated type 1 diabetes mellitus.
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Diabetes Mellitus, Type 1↗
GTP-binding proteins transduce signals generated via human FC gamma receptor IIIA (CD16).
This study demonstrates that GTP-binding proteins regulate Fc gamma RIII-mediated signal transduction and inositol phosphate (IPn) generation in human NK cells. In addition the cross-linking of CD16 by mAb, guanosine 5'-o-3-thiophosphate induced 1,4,5 inositol trisphosphate (IP3) release in permeabilized NK cells and their membranes. By contrast, guanosine 5'-o-2-thiophosphate, almost completely inhibited IP3 generation induced by cross-linking with anti-CD16 mAb. Pretreatment of NK cells with 10 to 100 ng/ml Vibrio cholerae toxin (Ctx) almost completely inhibited the generation of IP3 and of other Ipn as well as Fc gamma RIII-operated cell functions such as antibody-dependent cell-mediated cytotoxicity against antibody-coated P815 mastocytoma cells. Isolated B subunit of Ctx was inactive. Bordetella pertussis toxin (0.1 to 1 microgram/ml) only marginally affected IP3 release and antibody-dependent cell-mediated cytotoxicity. Ctx increased cAMP levels in NK cells. However, inhibition of IP3 release preceded the rise of cAMP. Moreover, cAMP analogues (8-chlor-cAMP, 8-bromo-cAMP, dibutiryl-cAMP), as well as intracellular cAMP-enhancing agents (PGE1, PGE2, and forskolin) did not mimicked the effects of Ctx on IP3 generation, suggesting that the adenylate cyclase pathway is not responsible for the early effects of Ctx on Fc gamma RIII-mediated signalling. Overall these results demonstrate that signal transduction via Fc gamma RIII is mediated by Ctx-sensitive cellular membrane GTP-binding protein.
Adenylate Cyclase Toxin↗
Natural killer activity and antibody-dependent cellular cytotoxicity in progressive systemic sclerosis.
Enhanced natural killer (NK) activity and normal lymphocyte antibody-dependent cellular cytotoxicity (ADCC) were observed in 16 patients with a diagnosis of progressive systemic sclerosis (PSS). Higher NK activity levels were observed against NK-sensitive K562 target cells, while the NK-resistant P815, Daudi and Raji cell lines were not lysed. Cytofluorimetric studies and morphological analysis of peripheral blood lymphocytes (PBL) showed an increased number of CD16 positive cells and large granular lymphocytes (LGL), indicating that the enhancement observed was probably attributable to an increase in the number of circulating NK cells.
Adult↗
Effect of cellulose acetate 0.2 micron filter on platelet specific proteins plasma levels.
We are reporting the effect of a cellulose acetate 0.20 micron filter (Flow Pore D26) on preparation of platelet poor plasma (PPP) for subsequent assay of platelet specific proteins. PPP was obtained using the Edinburgh anticoagulant mixture and a centrifugation at 2,300 g for 20 minutes. The mean levels of platelet factor 4 (PF4) and beta-thromboglobulin (beta TG) before filtration were: PF4 15.7 ng/ml (1.5-42.8 ng/ml); beta TG 62.0 ng/ml (25-140 ng/ml). After filtration we obtained: PF4 4.8 ng/ml (1.0-8.9 ng/ml), beta TG 28.4 ng/ml (8.5-51.1 ng/ml). Thus difference for both the platelet specific proteins was statistically significant (p less than 0.005). The contamination by platelet-like material was also greatly affected by the filter (before filtration 4.3 x 10(6)/ml), (2-15 x 10(6)/ml), after filtration 2.0 x 10(6)/ml (1-5 x 10(6)/ml), p less than 0.005). If a high speed centrifuge is not available, the use of this simple filter method could avoid falsely elevated values of platelet specific proteins.
Adult↗
Steroid sensitivity of thymocyte subpopulations during intrathymic differentiation. Effects of 17 beta-estradiol and dexamethasone on subsets expressing T cell antigen receptor or IL-2 receptor.
We have studied the effects of the steroid hormones, 17 beta-estradiol and dexamethasone, on the relative proportion of thymocyte expression of CD4 (L3T4), CD8 (Ly-2), TCR and IL-2R, used to identify different stages of thymocyte differentiation. After short-term in vivo steroid treatment, a significant decrease in the number and proportion of the CD4+/CD8+, double positive subpopulation was observed in parallel with a proportional increase in the percentage of the CD4+/CD8- single positive, of the CD4-/CD8-, double negative and, to a lesser extent, of the CD8+/CD4- subsets. Either steroid treatment increased the proportion of cells expressing either the epsilon-chain of the CD3 complex and/or the beta-chain of the TCR (beta-TCR) (TCR+/CD3+) and the 55 kDa protein of the IL-2R (IL-2R+), related to the increase of CD4+ SP thymocytes and of DN cells, respectively. Furthermore, the increased proportion of CD3+ cells could also be partially related to the increase of both the CD4+ and DN subsets. A decrease of the effect on either DN/IL-2R+ cells or CD4+ SP cells was selectively observed after long-term treatments with 17 beta-estradiol or DEX, respectively. It is concluded that after short-term administration, different steroid hormones mediate a similar selective depletion of DP, TCR-/CD3-, IL2R- cells presumably in an intermediate stage of differentiation. However, either steroid effects evolve differently in long-term treatment schedules, resulting in different effects on early (DN/IL2R+) and late (SP/IL2R-) steps of thymocyte differentiation.
Animals↗
Effects of cadmium on lymphocyte activation.
The effects of cadmium (Cd) on phytohemoagglutinin or phorbol myristate acetate-induced lymphocyte activation were investigated and a dose-dependent inhibition of cell proliferation was found. Kinetic studies revealed that the Cd-sensitive step is an early event of T cell stimulation. Failure of IL2 secretion and reduction of IL2 receptor expression in the Cd-treated cells are also reported. Regardless of which mechanism is responsible for Cd effects, our studies show that the inhibition of lymphocyte activation is associated with reduced [3H]phorbol dibutyrate binding to Ca2+-phospholipid-dependent protein kinase and altered breakdown of phosphatidylinositols. Thus, Cd interferes with two biochemical events which play a critical role in lymphocyte signal transduction and activation.
Adult↗