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PubMed2005

Mechanistic basis of heparin-induced thrombocytopenia.

Heparin-induced thrombocytopenia (HIT) occurs in 1% to 5% of patients treated with unfractionated high-molecular weight heparin and often results in limb- and/or life-threatening thrombotic complications. This disorder involves the formation of antibodies to a complex of Platelet Factor 4 (PF4) with the heparin. We believe that the pathogenesis of this disease begins with having an excess of platelet PF4 release. The freed PF4 is bound to platelet and other vascular membrane surfaces and has three critical roles in the initiation of HIT. (1) Infused heparin neutralizes a portion of excess surface PF4, directly enhancing local thrombosis. (2) The excess PF4 is mobilized into PF4/heparin complexes that stimulates HIT antibody production. (3) The remaining PF4 complexed to heparanoids and heparin on the vascular surfaces now bind to these HIT antibodies and through surface Fc gammaRII receptors leads to more platelet activation and removal, thrombus formation, and inflammation. These events, in turn, further stimulate PF4 release, perpetuating repetitive cycle that results in the clinical manifestations of this disorder.

Dose-Response Relationship, Drug↗

PubMed1985

In vivo studies on the binding of heparin and its fractions with platelet factor 4.

PF4 has a half-life in plasma of less than 3 minutes, and its rapid clearance appears to be a function of binding to the vascular endothelium. Once bound to the endothelium, PF4 can be released by heparin in a time-dependent manner; recovery is greater the sooner heparin is administered following PF4 infusion. This heparin-induced release of PF4 can be abolished if the heparin is first complexed with hexadimethrine bromide. Likewise, this heparin-induced release of PF4 is dependent upon the type of heparin used; low molecular weight heparin fractions and fragments do not cause the PF4 rebound seen with intact heparin. Thus, it would appear that low molecular weight forms of heparin are advantageous in that their in vivo administration would not be mediated by such platelet modulators as PF4.

Animals↗

PubMed2002

Platelet factor 4 enhances the binding of oxidized low-density lipoprotein to vascular wall cells.

Accumulation of low-density lipoprotein (LDL)-derived cholesterol by macrophages in vessel walls is a pathogenomic feature of atherosclerotic lesions. Platelets contribute to lipid uptake by macrophages through mechanisms that are only partially understood. We have previously shown that platelet factor 4 (PF4) inhibits the binding and degradation of LDL through its receptor, a process that could promote the formation of oxidized LDL (ox-LDL). We have now characterized the effect of PF4 on the binding of ox-LDL to vascular cells and macrophages and on the accumulation of cholesterol esters. PF4 bound to ox-LDL directly and also increased ox-LDL binding to vascular cells and macrophages. PF4 did not stimulate ox-LDL binding to cells that do not synthesize glycosaminoglycans or after enzymatic cleavage of cell surface heparan and chondroitin sulfates. The effect of PF4 on binding ox-LDL was dependent on specific lysine residues in its C terminus. Addition of PF4 also caused an approximately 10-fold increase in the amount of ox-LDL esterified by macrophages. Furthermore, PF4 and ox-LDL co-localize in atherosclerotic lesion, especially in macrophage-derived foam cells. These observations offer a potential mechanism by which platelet activation at sites of vascular injury may promote the accumulation of deleterious lipoproteins and offer a new focus for pharmacological intervention in the development of atherosclerosis.

Amino Acid Substitution↗

PubMed2006

Platelet factor 4 (CXCL4) seals blood clots by altering the structure of fibrin.

Platelet factor-4 (PF4/CXCL4) is an orphan chemokine released in large quantities in the vicinity of growing blood clots. Coagulation of plasma supplemented with a matching amount of PF4 results in a translucent jelly-like clot. Saturating amounts of PF4 reduce the porosity of the fibrin network 4.4-fold and decrease the values of the elastic and loss moduli by 31- and 59-fold, respectively. PF4 alters neither the cleavage of fibrinogen by thrombin nor the cross-linking of protofibrils by activated factor XIII but binds to fibrin and dramatically transforms the structure of the ensuing network. Scanning electron microscopy showed that PF4 gives rise to a previously unreported pattern of polymerization where fibrin assembles to form a sealed network. The subunits constituting PF4 form a tetrahedron having at its corners a RPRH motif that mimics (in reverse orientation) the Gly-His-Arg-Pro-amide peptides that co-crystallize with fibrin. Molecular modeling showed that PF4 could be docked to fibrin with remarkable complementarities and absence of steric clashes, allowing the assembly of irregular polymers. Consistent with this hypothesis, as little as 50 microm the QVRPRHIT peptide derived from PF4 affects the polymerization of fibrin.

Blood Coagulation↗

PubMed2004

Bacteriophage and phenotypic variation in Pseudomonas aeruginosa biofilm development.

A current question in biofilm research is whether biofilm-specific genetic processes can lead to differentiation in physiology and function among biofilm cells. In Pseudomonas aeruginosa, phenotypic variants which exhibit a small-colony phenotype on agar media and a markedly accelerated pattern of biofilm development compared to that of the parental strain are often isolated from biofilms. We grew P. aeruginosa biofilms in glass flow cell reactors and observed that the emergence of small-colony variants (SCVs) in the effluent runoff from the biofilms correlated with the emergence of plaque-forming Pf1-like filamentous phage (designated Pf4) from the biofilm. Because several recent studies have shown that bacteriophage genes are among the most highly upregulated groups of genes during biofilm development, we investigated whether Pf4 plays a role in SCV formation during P. aeruginosa biofilm development. We carried out immunoelectron microscopy using anti-Pf4 antibodies and observed that SCV cells, but not parental-type cells, exhibited high densities of Pf4 filaments on the cell surface and that these filaments were often tightly interwoven into complex latticeworks surrounding the cells. Moreover, infection of P. aeruginosa planktonic cultures with Pf4 caused the emergence of SCVs within the culture. These SCVs exhibited enhanced attachment, accelerated biofilm development, and large regions of dead and lysed cells inside microcolonies in a manner identical to that of SCVs obtained from biofilms. We concluded that Pf4 can mediate phenotypic variation in P. aeruginosa biofilms. We also performed partial sequencing and analysis of the Pf4 replicative form and identified a number of open reading frames not previously recognized in the genome of P. aeruginosa, including a putative postsegregational killing operon.

Bacteriophages↗

PubMed2003

Platelet factor 4 localization in carotid atherosclerotic plaques: correlation with clinical parameters.

Emerging evidence supports a role for platelets in the progression of atherosclerosis in addition to an involvement in thrombotic vascular occlusion. Platelet Factor 4 (PF4), a chemokine released by activated platelets, stimulates several pro-atherogenic processes. Therefore, we examined the localization of PF4 and the homologous protein, Neutrophil Activating Protein-2 (NAP-2) in lesions representing the evolution of human atherosclerotic plaques. Carotid plaques from 132 patients with critical carotid stenosis and 6 autopsy specimens were studied. Clinical, histologic and immunohistochemical data were analyzed using a chi(2)-test. PF4 was detected in the cytoplasm of luminal and neovascular endothelium, in macrophages and in regions of plaque calcification. The presence of PF4 in macrophages and neovascular endothelium correlated with lesion grade (p = 0.004; p = 0.044). Staining of macrophages for PF4 correlated with the presence of symptomatic atherosclerotic disease (p = 0.028). In early lesions, PF4 was commonly found in macrophages of early lesions (Grade I/II), whereas NAP-2 was rarely present. In conclusion, correlation between PF4 deposition, lesion severity and symptomatic atherosclerosis suggests that persistent platelet activation may contribute to the evolution of atherosclerotic vascular lesions. These studies support the rationale for the chronic use of anti-platelet therapy in patients at risk for developing symptomatic atherosclerosis.

Adult↗

PubMed2006

Close approximation of two platelet factor 4 tetramers by charge neutralization forms the antigens recognized by HIT antibodies.

OBJECTIVE: Heparin-induced thrombocytopenia (HIT) is a prothrombotic drug reaction caused by antibodies that recognize positively charged platelet factor 4 (PF4), bound to the polyanion, heparin. The resulting immune complexes activate platelets. Unfractionated heparin (UFH) causes HIT more frequently than low-molecular-weight heparin (LMWH), whereas the smallest heparin-like molecule (the pentasaccharide, fondaparinux), induces anti-PF4/heparin antibodies as frequently as LMWH, but without exhibiting cross-reactivity with these antibodies. To better understand these findings, we analyzed the molecular structure of the complexes formed between PF4 and UFH, LMWH, or fondaparinux. METHODS AND RESULTS: By atomic force microscopy and photon correlation spectroscopy, we show that with any of the 3 polyanions, but in the order, UFH>LMWH>>fondaparinux--PF4 forms clusters in which PF4 tetramers become closely apposed, and to which anti-PF4/heparin antibodies bind. By immunoassay, HIT antibodies bind strongly to PF4/H/PF4 complexes, but only weakly to single PF4/heparin molecules. CONCLUSIONS: HIT antigens are formed when charge neutralization by polyanion allows positively charged PF4 tetramers to undergo close approximation. Whereas such a model could explain why all 3 polyanions form antibodies with similar specificities, the striking differences in the relative size and amount of complexes formed likely correspond to the observed differences in immunogenicity (UFH>LMWH approximately fondaparinux) and clinically relevant cross-reactivity (UFH>LMWH>>fondaparinux).

Adsorption↗

PubMed1981

Plasma beta-thromboglobulin and platelet factor 4 in patients with chronic renal failure and effect of hemodialysis.

Significantly increased levels of plasma beta-thromboglobulin (beta-TG) (76.8 +/- 25.5 ng/ml, p less than 0.01) were observed in 24 patients with chronic renal failure (blood urea nitrogen (BUN) greater than 20 mg/100 ml), as compared with normal subjects (13.2 +/- 5.6 ng/ml). The increase in beta-TG was highly correlated with BUN (r = 0.651, p less than 0.01), creatinine (r = 0.778, p less than 0.01) and creatinine clearance ( t = -0.723, p less than 0.01). Plasma platelet factor 4 (PF4) and normal 5.0 +/- 2.0 ng/ml) also increased significantly to 8.5 +/- 3.4 ng/ml (p less than 0.01). However, statistical correlation between beta-TG and PF4 was not found in these patients. The reason is thought to be due to differences in molecular weight (PF4 8,000 MW; beta-TG 36,000 MW) and half-life time (PF4 30 min; beta-TG 100 min), and due to the difficulty in calculating statistically the correlation because of the narrow distribution of PF4 levels. The high levels of beta-TG (89.4 +/- 3.4 ng/ml) showed a further increase (109.4 +/- 5.8 ng/100 ml, p less than 0.01) after dialysis. This is thought to be due to hemoconcentration, because other blood factors such as RBC, WBC, platelets, fibrinogen, etc were elevated by about 20% during hemodialysis and because no adhesion of platelets to the cellulose membrane did occur. The increase in PF4 levels at 15 min (55.2 +/- 19.6 ng/ml, p less than 0.01) and 1 hr (23.7 +/- 8.4 ng/ml, p less than 0.01) of hemodialysis from the level before it (7.7 +/- 1.3 ng/ml) is thought to be caused the effect of heparin infusion. The change in PF4 was not accompanied by the change in beta-TG. During hemodialysis the decrease of other platelet functions such as adhesiveness, aggregation induced by ADP, collagen and PF3 remained unchanged.

Adult↗

PubMed1984

Determination of low affinity platelet factor 4 in frozen and thawed human platelets by the newly developed enzyme immunoassay system.

By the use of a newly developed sandwich enzyme immunoassay method for low affinity platelet factor 4 (LA-PF4), the effects of repeated freeze-thawing on the contents of this protein in platelets were determined and compared with those of Triton-X 100 lysed platelets. The assay system consisted of polystyrene balls covered with immobilized antibody fragments F(ab')2 and the same antibody Fab' labeled with beta-D-galactosidase from E. coli. The assay was specific to LA-PF4 with no significant cross-reactivity with platelet factor 4. Coefficients of variation within-run and between-run for the assay of LA-PF4 were less than 12%. The results obtained with this enzyme immunoassay correlated well with those of a radioimmunoassay of beta-thromboglobulin which is immunologically identical with LA-PF4 (r = 0.961, slope = 1.056, y-intercept = -3.739 ng/ml; n = 22). The contents of LA-PF4 per 10(9) Triton-X 100 lysed platelets in platelet-rich plasma were 23.65 +/- 3.14 micrograms (mean +/- S.D.). The contents of LA-PF4 in platelets were increased from 46% to 95% of Triton-X 100 values by repeated freezing and thawing 1 to 7 times. The present data indicate that the freeze-thawing technique should be done carefully to obtain the reliable determination of LA-PF4 in platelets.

Blood Coagulation Factors↗

PubMed1999

Complexes of heparin and platelet factor 4 specifically stimulate T cells from patients with heparin-induced thrombocytopenia/thrombosis.

Heparin-induced thrombocytopenia with thrombosis (HITT) is associated with antibodies specific for complexes consisting of heparin and platelet factor 4 (PF4). Studies in individual patients with HITT have demonstrated immunoglobulin (Ig) class switching from IgM to the IgG or IgA isotypes. This transition is thought to require helper T cells, but no studies of the cellular or molecular basis of this process have yet been reported. To characterize T-cell involvement in HITT, peripheral blood mononuclear cells (PBMC) from two patients with classical HITT obtained shortly after the acute episode were restimulated with heparin:PF4 complexes, PF4 alone, heparin alone, and medium alone in the presence of autologous antigen-presenting cells (APC). Responding T cells were then examined using the technique of "spectratyping," in which sequences encoding CDR3 domains of individual V beta (BV) families are amplified and separated by gel electrophoresis. After 14 days in culture with antigen (heparin:PF4 complexes), but not after culture with PF4, heparin, or medium alone, patient cells, but not cells from normal subjects, preferentially expressed T-cell receptor (TCR)-containing beta chains of the BV 5.1 family. Nucleotide sequencing of BV 5.1 TCR CDR3 showed that each patient had a personal repertoire, but also shared a tetrapeptide motif (PGTG). These findings provide evidence that the humoral immune response associated with HITT is driven by helper T cells that presumably recognize peptides derived from PF4. Identification of a common beta-chain CDR3 motif in responding T cells from each of two patients suggests that a limited number of helper TCRs may be used to mount an antibody response to heparin:PF4 complexes. TCR spectratyping appears to offer a new way to examine the molecular basis of pathologic immune responses and may be useful in further studies of HITT and other immune-mediated hematologic disorders.

Aged↗

PubMed2002

[Effect of platelet factor 4 on the adhesive property of leukemic CD34+ KG1a cell].

OBJECTIVE: To study the effect of PF4 on the adherence of leukemic CD34+ KG1a cell to human umbilical vein endothelial cell line ECV-304 cell and on the expression of adhesive molecules. METHODS: Adhesion assay and adhesion blocking assay were respectively applied to measure the effect of PF4 and/or adhesion molecule monoclonal antibodies on the adhesion property of KG1a. The expressions of adhesion molecules were determined by RT-PCR and FACS analysis. RESULTS: The adhesion of KG1a cells to ECV-304 was significantly enhanced in the presence of PF4. Such enhancement was also observed when KG1a or ECV-304 cells were separately treated with PF4 before interaction. The adhesion capacity of KG1a cells was reduced when cells were co-incubated with the blocking monoclonal antibodies (MoAbs) against CD49d, CD106, CD54, respectively. In contrast, MoAbs against CD62L, CD62P and CD62E had no such effect. During a period of 3 hours when KG1a or ECV-304 cells were respectively incubated with PF4, the mRNA expressions of CD49 d, CD54 were up-regulated. Furthermore, when KG1a or ECV-304 cells were incubated with PF4 for 2 hours, respectively, the percentages of CD49d+ KG1a cells and CD54+ ECV-304 were increased significantly. CONCLUSION: PF4 can enhance KG1a cell adhesive capacity by increasing the expressions of adhesion molecules.

Antigens, CD34↗

PubMedDate not supplied

Increased releasability of platelet products and reduced heparin-induced platelet factor 4 release from endothelial cells in bronchial asthma.

To determine whether or not platelet activation is involved in the mechanism of exacerbation of bronchial asthma, we evaluated adenosine triphosphate (ATP) release from thrombin-stimulated washed platelets, plasma levels of beta-thromboglobulin (beta-TG) and platelet factor 4 (PF4), and plasma beta-TG/PF4 ratios during symptomatic and asymptomatic periods in 15 patients with bronchial asthma compared with 16 normal control subjects. We also measured these parameters during allergen provocation tests and acetylcholine inhalation tests in 6 allergic asthmatics. ATP release, plasma levels of beta-TG and PF4 were significantly increased during symptomatic periods and after the allergen provocations but not after acetylcholine inhalations. However, these findings were not accompanied by the elevation of plasma beta-TG/PF4 ratios. The heparin-induced PF4 release, which is reported to reflect release of PF4 attached on endothelial cells, was significantly reduced in 12 asymptomatic asthmatic patients compared with 11 normal subjects, and it was much more reduced in 7 symptomatic asthmatic patients, suggesting the possibility of the reduced PF4 binding on endothelial cell surface. This finding may represent the prolonged half life of PF4 in asthmatics. We concluded that 1) increased releasability of platelet products and in vivo release of the platelet granular contents are involved in the mechanism of the exacerbation of bronchial asthma, 2) some functional alteration in platelet-endothelial cell interaction may be involved in bronchial asthma, and 3) plasma beta-TG/PF4 ratios are not elevated possibly because of both increased platelet releasability and prolonged half-life of PF4 in the blood in asthmatic patients.

Acetylcholine↗

PubMed1989

Thromboembolic complications in an asplenic HbE-beta-thalassaemia patient.

A patient, double heterozygous for HbE-beta-thalassaemia, had recurrent thromboembolic complications following splenectomy. In addition to marginally decreased protein C and S plasma levels, laboratory studies revealed a persistent thrombocytosis and markedly elevated plasma concentrations of platelet factor 4 (PF4). PF4 neutralizes the heparin anticoagulant activity. The increased PF4 levels explained the initial heparin resistance observed during anticoagulant treatment in this patient. Subsequent heparin loading tests revealed an abnormal reaction of the PF4 plasma levels, i.e. no increase of PF4. However, upon repeated heparin injections this PF4 response normalized, which may be due to depletion of the endothelial cell associated heparin mobilizable PF4 pool. These observations may be of relevance for the treatment of similar patients.

Adult↗

PubMed1994

Platelet factor 4 stimulates thrombomodulin protein C-activating cofactor activity. A structure-function analysis.

Thrombomodulin (TM) is an anionic (pI approximately 4) protein cofactor that promotes thrombin (THR) cleavage of protein C to generate activated protein C (APC), a potent anticoagulant. We find that the cationic platelet alpha-granule protein platelet factor 4 (PF4) stimulates 4-25-fold the cofactor activity of rabbit TM and two differentially glycanated versions of an extracellular domain human TM polypeptide in which the glycosaminoglycan (GAG) is either present (GAG+ TM) or absent (GAG- TM) with an ED50 of 3.3-10 micrograms/ml. No such stimulation occurs in response to beta-thromboglobulin or thrombospondin, or when protein C lacking its gamma-carboxyglutamic acid (Gla) domain is the substrate. Heparin and chondroitin sulfates A and E reverse PF4 stimulation. PF4 minimally affects the Kd for THR but decreases 30-fold (from 8.3 to 0.3 microM) the Km for protein C of APC generation by GAG+ TM. PF4 also strikingly transforms the [Ca2+] dependence profile of rabbit and GAG+ TM to resemble that of GAG- TM. A potential explanation for this is that PF4, like Ca2+, induces heparin-reversible alterations in native (but not Gla-domainless) protein C conformation as assessed by autofluorescence emission analysis. We conclude that PF4 stimulates TM APC generation by interacting electrostatically with both the TM GAG and the protein C Gla domain to enhance markedly the affinity of the THR.TM complex for protein C. By this mechanism, PF4 may play a previously unsuspected role in the physiologic regulation of clotting.

Blood Platelets↗

PubMed1993

Selection of an HEL-derived cell line expressing high levels of platelet factor 4.

A platelet factor 4 (PF4)-expressing cell line, HELNeo, was derived from the human erythroleukemia cell line, HEL. This was achieved by stable transfection of HEL cells with a construct containing the rat PF4 promoter driving the gene coding for resistance to neomycin, followed by selection of neomycin-resistant clones. HELNeo cells were all nonadhering and about 5% of the cells had polyploid nuclei (> or = 8N), as compared with 1% in HEL cells. Immunohistochemistry showed that about 90% of the HELNeo cells contained PF4, whereas only approximately 5% of the HEL cells contained PF4. No significant parallel enrichment was observed for other megakaryocytic markers, such as the glycoprotein complex IIb/IIIa, von Willebrand factor, and platelet activation-dependent granule to external membrane glycoprotein (PADGEM), which were present to a similar extent in both HEL and HELNeo lines. The increased expression of PF4 in HELNeo cells was confirmed by transient expression assays and was associated with a fivefold increase in trans-acting factors binding to the PF4 promoter. These cells should be a rich source for purifying trans-acting factors binding to the PF4 gene. Moreover, our study shows how a lineage-specific promoter may be used to generate lineage-specific cell lines from a multilineage hematopoietic cell line.

Base Sequence↗

PubMed1981

Human platelet factor 4: preparation from outdated platelet concentrates and application in cerebral vascular disease.

Platelet factor 4 (PF4), the platelet antiheparin protein, was isolated from both the supernatant and the cells of recently outdated platelet concentrates. Following purification by affinity chromatography, a competitive binding radioimmunoassay was developed to detect this protein in human plasma. The normal range was determined to be 9.4 +/- 4.7 ng/ml (mean +/- SD for 52 healthy adults). In order to determine whether individuals with transient ischemic attack (TIA) or stroke had measurable increments of PF4 in their plasma, radioimmunoassay studies were performed on 11 patients with well-documented TIA, 10 patients with well-documented stroke and on 16 age-matched controls hospitalized on a neurology service with disorders unrelated to arterial thrombosis. The 16 hospitalized controls had PF4 levels of 10.3 +/- 9.1 ng/ml, a value not significantly different from the 52 normals (P greater than 0.50). Patients with TIA had PF4 levels of 24.6 +/- 12.1 ng/ml, a value significantly higher than both the 52 normals (P less than 0.001) and the 16 hospitalized control patients (P less than 0.005). Patients with stroke had PF4 levels of 35.4 +/- 29.2 ng/ml, a value significantly higher than both the 42 normals (P less than 0.001) and the 16 hospitalized control patients (P less than 0.005). Outdated platelet concentrates facilitate the development of a reproducible radioimmunoassay for PF4. The elevation of this platelet-derived protein in the plasma of patients with stroke and TIA provides evidence for recent or ongoing platelet activation in the cerebral vascular disease population.

Adult↗

PubMed2004

New laboratory test in flow cytometry for the combined analysis of serologic and cellular parameters in the diagnosis of heparin-induced thrombocytopenia.

BACKGROUND: Heparin-induced thrombocytopenia (HIT) is a major complication of heparin therapy. A quick and reliable laboratory assay for the combined determination of pathogenic anti-heparin and platelet factor 4 (H:PF4) antibodies in the serum and platelet activation is not currently available. METHODS: We developed a new single-tube assay in flow cytometry that combines the detection of antibodies in the serum and their activatory properties on platelets. The assay was tested on 13 serum samples from patients with suspected HIT and six samples from normal donors. The presence of anti-H:PF4 antibody complexes was detected by H:PF4-coated beads, and donor platelet activation induced by HIT sera was determined by Annexin V binding. All data were compared with the patients' clinical setting, laboratory tests, and standard enzyme-linked immunosorbent assay detection of anti-H:PF4 antibodies. RESULTS: This flow cytometry assay allowed unequivocal, simultaneous detection of anti-H:PF4 antibodies in sera and their activatory properties on platelets. All cases for which the diagnosis of HIT was confirmed were detected by the flow assay. CONCLUSIONS: This assay, combining for the first time functional and nonfunctional testing on anti-H:PF4 antibodies, is likely to influence the clinical decision for the management of HIT patients.

Adult↗

PubMed2005

Prevalence of heparin-induced antibodies in patients with chronic renal failure undergoing hemodialysis.

Heparin-induced thrombocytopenia (HIT) type II is a serious complication of heparin therapy. It presents initially as thrombocytopenia, and is associated with thrombosis in 20-50% of the cases. HIT is related to the presence of heparin-induced antibodies (HIA), which show specificity for the PF4-heparin (PF4-H) complex. The FcgammaRIIa receptor has been suggested to participate in the pathogenic mechanism of HIA. Since patients undergoing chronic hemodialysis (HD) are exposed repeatedly to heparin, we studied the prevalence of HIA and their eventual relationship with thrombocytopenia and/or thrombosis, and the possible participation of the FcgammaRIIa polymorphism. We studied 207 patients with chronic renal failure (CRF) undergoing HD. As a control we included 130 blood donors and 28 patients with CRF without HD. The HIA patients were studied with the use of a PF4-H ELISA. Additionally, in some positive cases for the previous test, a 14C- serotonin release assay (14C-SRA) was performed. The polymorphism FcgammaRIIa H/R131 was studied by polymerase chain reaction (PCR) with allele-specific primers. Thirty-seven patients (17.9%) undergoing HD presented with HIA. The majority of these antibodies were IgG, IgM, and IgA. The HIA investigated presented specificity against the PF4-H complex, but not against PF4 alone (P<0.001). Twelve out of 22 (54.5%) PF4-H antibodies were positive when tested with the 14C-SRA. The distribution of the FcgammaRIIa polymorphism in patients and healthy controls was 42.6% and 41.6% for H/H131, 41% and 48.9% for the H/R131 isoform, and 16.4% and 9.5% for the R/R131 isoform, respectively. No statistically significant difference in the FcgammaRIIa isoform distribution was found. Twenty-nine out of 156 patients (18.5%) presented thrombocytopenia, and 21/207 (12.4%) had thrombosis of the native vein arterio-venous fistula (AVF). We did not find any statistically significant between HIA and thrombocytopenia or thrombosis. An important proportion of patients with CRF undergoing HD developed HIA, but these cases were not associated with thrombocytopenia or thrombosis of AVF. The frequency of the FcgammaRIIa polymorphism did not statistically differ between HIT type II and normal controls.

Adult↗