Heparin-induced thrombocytopenia: towards consensus.
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Anticoagulants↗
Biomedical subjects
B H Chong
Publications and source records attributed to B H Chong.
Localization and regulation of thrombopoietin mRNa expression in human kidney, liver, bone marrow, and spleen using in situ hybridization.
Thrombopoietin (TPO) is the primary hematopoietic growth factor involved in the regulation of platelet production. Although the kidney, liver, bone marrow (BM), and spleen have been identified as the major sources of TPO production, the precise cellular location of TPO mRNA expression in these tissues remains unknown. We have identified the cells expressing TPO mRNA in the human kidney, liver, and BM using an in situ hybridization assay. In the BM of individuals with normal platelet counts, the hybridization signal was too weak to allow identification of the TPO mRNA expressing cells. However, in thrombocytopenic subjects with aplastic anemia, postchemotherapy marrow aplasia, and immune thrombocytopenia, the stromal cells showed strong TPO mRNA expression. In the human subjects with normal platelet counts, the cells of the proximal convoluted tubules of the kidney showed consistent positive staining whereas the signal in the cells of the distal convoluted tubules was less consistent. Strong hybridization signal was also evident in the hepatocytes. The hybridization signal in the spleen, even in thrombocytopenic subjects, was too weak to allow confident identification of the cells expressing TPO mRNA. In all subjects, the interstitial cells and endothelial cells of the liver and spleen, the renal peritubular cells, and the hematopoietic precursor cells of the BM showed no TPO mRNA expression. Our data suggest that TPO mRNA expression in the human BM may be modulated by platelet mass.
Bone Marrow↗
The platelet proaggregating and potentiating effects of unfractionated heparin, low molecular weight heparin and heparinoid in intensive care patients and healthy controls.
Heparin binds to platelets and can cause platelet proaggregating and potentiating effects, possibly causing thrombocytopenia, particularly in patients in intensive care with hyperaggregable platelets. In this study we compared the platelet proaggregating and potentiating effects of unfractionated heparin (UH), 2 low molecular weight (LMW) heparins, enoxaparin and dalteparin, and a heparinoid, danaparoid sodium (orgaran), to platelets of an ICU patient population and a normal control group. In both populations UH caused platelet aggregation in a dose-dependent manner. This occurred in the therapeutic range of the drug, with as little as 0.5 U/ml UH. The LMW heparins caused less and the heparinoid least platelet aggregation. Generally, the aggregation observed in ICU patients was greater than in the normal population. The potentiating effects of the 4 drugs in association with physiological agonists was examined. Similar patterns of potentiation were observed in both populations, with UH causing significant enhancement of platelet aggregation, the LMW heparins intermediate and heparinoid least enhancement. There was substantial variability in the individuals' platelets' reactions to the drugs, in particular to UH. Our findings suggest that UH has the greatest effect, the low molecular weight heparins an intermediate effect and the heparinoid the least propensity to cause platelet activation.
Adenosine Diphosphate↗
Plasma macrophage colony-stimulating factor and P-selectin levels in malaria-associated thrombocytopenia.
Thrombocytopenia is a common finding in malaria. In clinical trials, recombinant macrophage colony-stimulating factor (M-CSF) causes a reversible, dose-dependent thrombocytopenia, and high M-CSF has been reported in autoimmune thrombocytopenias. P-selectin, which is secreted into the plasma following platelet/endothelial activation or damage, is elevated in certain consumptive thrombocytopenic disorders. The relationships between thrombocytopenia, M-CSF and P-selectin were analysed in 63 patients with severe (n = 13) or uncomplicated (n = 26) P. falciparum (PF) or P. vivax (PV) malaria (n = 24). On admission, 69% of PF patients and 75% of PV patients were thrombocytopenic (platelets < 150 x 10(9)/l). M-CSF was elevated in PF (3021 +/- 1844 pg/ml) and PV (2602 +/- 1668 pg/ml) patients, compared to controls (589 +/- 200 pg/ml). The platelet count was inversely correlated with M-CSF in PF (r = -0.681), and in PV malaria (r = -0.548). Elevated P-selectin was found in severe PF malaria, but not in PV malaria. Severe PF malaria was associated with marked thrombocytopenia, very high M-CSF, elevated P-selectin and compelling evidence of disseminated intravascular coagulopathy (DIC). Platelet counts, M-CSF and P-selectin returned to control values in 7-14 days. These data suggest that elevated M-CSF in malaria, by enhancing macrophage activity, may result in increased macrophage-mediated platelet destruction. Further, platelet/endothelial activation or damage, as measured by P-selectin, or DIC could intensify thrombocytopenia in severe PF malaria, but does not appear to contribute to thrombocytopenia in uncomplicated PF or PV malaria.
Acute Disease↗
Identification of PDGF receptors on human megakaryocytes and megakaryocytic cell lines.
Platelet-derived growth factor (PDGF) is a potent chemotactic and mitogenic factor implicated to play important roles in a variety of normal and pathophysiologic settings. We investigated PDGF receptor expression on human megakaryocytes and several megakaryocytic cell lines (CHRF, DAMI, Meg-01, M-07e) using enzyme-linked immunosorbent assay (ELISA), flow cytometry and immunocytochemical staining. Both PDGF receptor subtypes were identified on CHRF, DAMI, and Meg-01 cells by ELISA; PDGF beta-receptor levels exceeded alpha-receptor levels. Flow cytometry revealed that beta-receptor levels on CHRF and DAMI cells exceeded those on Meg-01 cells, and that M-07e expressed neither receptor. Immunocytochemical staining confirmed these findings and determined that bone marrow megakaryocytes also expressed PDGF receptors. Exposure of megakaryocytes to PDGF-BB dramatically induced the expression of the immediate-early gene, c-fos, within 30 min. Moreover, PDGF-BB significantly stimulated CHRF proliferation and colony formation. The present study demonstrates the presence of functional PDGF receptors on human megakaryocytes and their ability to mediate a mitogenic response.
Cell Line↗
Prospective evaluation of the clinical usefulness of an antigen-specific assay (MAIPA) in idiopathic thrombocytopenic purpura and other immune thrombocytopenias.
The diagnosis of idiopathic immune thrombocytopenia remains a clinical diagnosis based on the exclusion of other causes of immune and nonimmune thrombocytopenia. Measurement of platelet-associated Ig (PAIg), while sensitive, is nonspecific for the diagnosis of immune thrombocytopenia. Published experience of antigen capture assays (including monoclonal antibody immobilization of platelet antigens or MAIPA) suggest a high sensitivity and specificity (70% to 80%) in selected groups of patients. In a prospective evaluation of 158 patients with thrombocytopenia from all causes, we report a sensitivity of 51% and specificity of 80% for direct MAIPA assays. MAIPA was considerably better in discriminating immune from nonimmune thrombocytopenia than two assays of PAIgG. Antiplatelet antibodies detected by MAIPA were more frequently directed against the glycoprotein (GP) IIb/IIIa than the GP Ib/IX complex. Our experience suggests that MAIPA assays are useful in the laboratory assessment of thrombocytopenia, should be performed before therapy, and that some patients with 'nonimmune' thrombocytopenia may have genuine antiplatelet antibodies.
Antibodies, Monoclonal↗
Prevention of postoperative venous thromboembolism.
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Humans↗
Cross-reactivity study of low molecular weight heparins and heparinoid in heparin-induced thrombocytopenia.
Previous studies suggested that the cross-reactivity rates of low molecular weight (LMW) heparins with the antibody in heparin-induced thrombocytopenia (HIT) are more than 80%, whilst that of the LMW heparinoid (Orgaran) is relatively low at about 10%. These earlier studies were limited either in the number of patients studied, or in investigating only a single drug. They were also inadequate due to non-standardisation of testing conditions. This study compares three LMW heparins (Fragmin, Clexane, Fraxiparin) and a heparinoid (Orgaran) in their cross-reactivity rates with the HIT antibody. The sera of 45 HIT patients were tested using platelet aggregometry under standardised conditions. The cross-reactivity rates are: 7% (Orgaran), 89% (Fragmin), 83% (Clexane) and 86% (Fraxiparin). Although there are no controlled trials to determine the in vivo cross-reactivity rates of LMW heparins and heparinoid, the low in vitro cross-reactivity rate of the latter favours its use in HIT. Nevertheless, the therapeutic use of these drugs in HIT should be preceded by exclusion of in vitro cross-reactivity.
Adult↗
Characterization of IgG Fc receptors on CD34 antigen-expressing cell lines (KG-1 and KG-1a).
Although Fc gamma receptors (Fc gamma R) on mature blood cells have been extensively studied, there are only limited data on Fc gamma R expression in the early haematopoietic progenitor cells. In this study, we used the stem cell antigen (CD34)-expressing cell line (KG-1) and its less differentiated subline (KG-1a) as a model for the study of Fc gamma R in the early haematopoietic progenitors. Flow cytometry and immunoprecipitation studies on KG-1 and KG-1a cells with anti-Fc gamma R mAb showed that Fc gamma RII is the only Fc gamma R expressed on the cell surface. Analysis of the steady-state levels of Fc gamma R mRNA in KG-1 and KG-1a cells using a quantitative in situ hybridization assay revealed the presence of only Fc gamma RII mRNA. On further analysis Fc gamma RIIA mRNA but no Fc gamma RIIB or Fc gamma RIIC transcripts were found in these cells; Fc gamma RIIA transcripts with and without the transmembrane exon were present in approximately equal amounts. These findings are surprisingly similar to those observed previously with Fc gamma R in platelets and megakaryocytic cells but different from those found with Fc gamma R in cells of other lineages. These data suggest that the Fc gamma R transcript distribution pattern observed in the early haematopoietic progenitors (KG-1 cells) is retained in later stages of haematopoietic differentiation only in cells of megakaryocytic lineage.
Antigens, CD34↗
Beta 2-glycoprotein I in thrombosis: evidence for a role as a natural anticoagulant.
Although the physiological role of beta2-glycoprotein (B2GPI) is unknown, in vitro evidence indicates that B2GPI may be a natural anticoagulant. In this study we have examined whether fluctuations of plasma B2GPI occur in in vivo coagulation. Serial measurements of B2GPI and other anticoagulant proteins were performed in 51 patients with thrombotic (group 1: six patients with disseminated intravascular coagulation (DIC), group 2: venous (n = 4) or arterial (n = 170 thrombosis) and non-thrombotic disease (group 3: 24 patients undergoing elective surgery). Reductions in plasma B2GPI levels were seen in most patients which were roughly proportional to the severity of their illness. Particularly striking reductions of B2GPI, protein C (PC) and antithrombin III (AT-III) (mean +/- 95% CI: 42.7 +/- 8.6%, 42.1 +/- 14.8%, 39.1 +/- 28.4% respectively) were seen in group 1. The reductions in plasma B2GPI were significantly greater in group 1 than in the other groups. Dilutional factors explain most of the reductions in B2GPI, PC and AT-III in groups 2 and 3, but contribute little to group 1. In conclusion, although B2GPI behaves as a 'negative acute phase reactant', the magnitude of reduction of plasma B2GPI levels, accompanied by reductions in other anticoagulant proteins in patients with DIC, suggests specific consumption of B2GPI in in vivo coagulation. This study provides further evidence that B2GPI is an anticoagulant of physiological importance.
Acute Disease↗
Serotonin stimulates megakaryocytopoiesis via the 5-HT2 receptor.
It is known that platelet alpha-granule constituents including platelet-derived growth factor (PDGF), platelet factor 4 (PF4) and transforming growth factor-beta (TGF-beta) can affect megakaryocytopoiesis. Serotonin, a platelet dense granule constituent has been shown to have a mitogenic effect on fibroblasts and smooth muscle cells but whether it has the same effect on megakaryocytes remains unclear. In this study, we investigated the effect of serotonin on megakaryocytopoiesis and the possible mechanism of its effect using the mouse plasma clot culture method. The results show that: (a) serotonin significantly stimulates megakaryocyte colony formation with maximum stimulation at 100 nM; (b) enhanced action is found between serotonin and interleukin-3 (IL-3), interleukin-6 (IL-6), granulocyte macrophage-colony stimulating factor (GM-CSF), erythropoietin (EPO) and PDGF; (c) ketanserin, a 5-HT2 receptor antagonist, blocks the mitogenic effect of serotonin on megakaryocytopoiesis; and (d) Meg-01 cells (a megakaryocyte cell line) express 5-HT2 receptors. This study demonstrates that serotonin has a mitogenic effect on megakaryocytopoiesis and this effect may be mediated via the 5-HT2 receptor which is known to be coupled to G protein. It is suggested that serotonin may also be involved in the feedback control of megakaryocytopoiesis.
Animals↗
Activated protein C resistance--low incidence in glaucomatous optic disc haemorrhage and central retinal vein occlusion.
PURPOSE: Activated protein C (APC) resistance has recently been reported as conferring a sevenfold increase in the risk of venous thrombosis. It is linked to a genetic mutation in the factor V gene which occurs commonly (about 2% to 4% of the community have the mutation). Glaucoma patients with nerve fibre layer (NFL) haemorrhages on the optic disc and patients with central retinal vein occlusion (CRVO) were tested for APC resistance to determine if there was an association. METHODS: Twenty-three patients with glaucomatous NFL haemorrhages and 23 patients with CRVO were tested. The CRVO cases included 11 with relatively young age of onset (mean 45.1 +/- 6.9 years) without conventional vascular risk factors. Eighty randomly selected Red Cross blood donor samples and 33 staff members were tested as controls. Clotting times with and without exogenous APC were recorded and an APC ratio determined. Cases with APC resistance were tested to confirm that they had the factor V Leiden gene. RESULTS: No cases of APC resistance were identified in the glaucoma patients and only one of the younger CRVO patients tested positive, but four of 113 controls tested positive. The difference in prevalence between groups is not significant. The mean APC ratios for the three groups were very similar: NFL haemorrhages 5.46(+/- 1.62), CRVO 5.70(+/- 1.56), controls 5.34 ( +/- 1.19) p > 0.5. CONCLUSION: There was not clear association detected between glaucomatous NFL haemorrhages or CRVO and APC resistance in this sample of patients. This negative finding is important due its known association with venous thrombosis elsewhere in the body.
Adult↗
Anticoagulation.
This review of anticoagulation discusses not only drugs that inhibit the formation of blood clots but also drugs that induce clot lysis. It includes warfarin, heparin and streptokinase as well as newer agents such as low molecular weight (LMW) heparins and recombinant tissue plasminogen activator (r-tPA).
Aged↗
Characterization of Fc gamma receptors on human megakaryocytes.
Megakaryocyte and platelet Fc gamma receptors (FcR) are of importance in the pathophysiology of immune complex-mediated thrombocytopenias such as heparin-induced thrombocytopenia. In this study, Fc gamma R proteins and mRNAs in normal human megakaryocytes were examined. Fc gamma R proteins were studied with immunocytochemical staining, dual colour flow cytometry and immunoprecipitation using monoclonal antibodies against Fc gamma R I, Fc gamma R II and Fc gamma R III. Fc gamma R mRNAs were measured with biotinylated cDNA of oligonucleotide probes using a novel quantitative in situ hybridization technique. Using these techniques, Fc gamma R II protein and mRNA, but not Fc gamma R I and Fc gamma R III proteins and transcripts were detected in megakaryocytes. Further, transcript analysis showed that megakaryocytes contain only the transcript of Fc gamma R IIA gene but no transcripts of Fc gamma R IIB nor Fc gamma R IIC genes; Fc gamma R IIA transcripts with and without the transmembrane (TM) exon are present in approximately equal proportions. In contrast, neutrophils and macrophages also contain Fc gamma R IIA transcript but Fc gamma R IIA transcript with the TM exon predominates suggesting cell lineage-specific Fc gamma R IIA expression. Fc gamma R IIA transcript lacking the TM exon predicts the presence of a potential soluble form of Fc gamma R in platelets and megakaryocytes which may have a physiological role as it can compete with the membrane-bound Fc gamma R IIA for binding of IgG-containing immune complexes and thus protect these cells from excessive binding and injurious effects of immune complexes.
Base Sequence↗
Serotonin uptake, storage and metabolism in megakaryoblasts.
Serotonin (5-hydroxytryptamine, 5-HT) uptake, storage and metabolism in human megakaryoblastic cell line (Meg-01) which acts as a model for megakaryocyte precursors, megakaryoblasts were investigated by using biochemical (HPLC) and morphological (electron microscope, EM) techniques. Results showed that Meg-01 cells were able to take up 5-HT. The intracellular 5-HT level was 2.8 +/- 0.4 and 51.8 +/- 4.9 (1 h) or 59.0 +/- 4.4 (2 h) ng/10(6) cells, before and after incubation with 5-HT, respectively, but no dense bodies were visualized after incubation with excess 5-HT by electron microscope observation. This uptake was inhibited by 28% on pre-incubation with fluoxetine and 60% of 5-HT in the cells was released on incubation with reserpine. 5-Hydroxyindoleacetic acid (5-HIAA) concentration of Meg-01 cells was increased after incubation of the cells with 5-HT (0 and 17.6 +/- 2.1 or 19.9 +/- 1.9 ng/10(6) cells, before and after incubation 1 or 2 h, respectively). The study suggests that: (1) 5-HT uptake ability is well established in megakaryocytes precursors, megakaryoblasts and the uptake ability is affected by reserpine and fluoxetine; (2) however, the capacity to store the amine is not well developed in megakaryoblasts and (3) megakaryoblasts may contain monoamine oxidase (MAO) which converts 5-HT to 5-HIAA.
Cells, Cultured↗
Diagnosis, treatment and pathophysiology of autoimmune thrombocytopenias.
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Child↗
Antiphospholipid antibodies and platelets.
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Antibodies, Antiphospholipid↗
Quantitation of soluble and membrane-bound Fc gamma RIIA (CD32A) mRNA in platelets and megakaryoblastic cell line (Meg-01).
Fc gamma receptors (Fc gamma Rs) are glycoproteins on platelet surface that bind IgG-containing immune complexes. However, excessive binding of immune complexes leads to platelet activation and thrombosis or increased platelet clearance and thrombocytopenia. In this study, Fc gamma R transcripts in platelets and megakaryoblastic cell line (Meg-01) were investigated using specifically designed oligonucleotides and a new quantitative in situ hybridization assay. Platelets and Meg-01 cells were found to express only Fc gamma RII transcripts. Of Fc gamma RIIA mRNA isoforms (Fc gamma RIIA, B and C), Fc gamma RIIA mRNA predominates in these cells. Platelets and Meg-01 cells contain both alternative spliced forms of Fc gamma RIIA mRNA, those with and without the transmembrane (TM) exon and both forms were present in near equal amounts. In contrast, Fc gamma RIIA transcript with the TM exon predominates in neutrophils and monocytes, suggesting that the splicing of the TM exon is under lineage-specific control.
Antigens, CD↗