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PubMed2004

Heterophilic interactions of platelet factor 4 and RANTES promote monocyte arrest on endothelium.

The chemokines platelet factor 4 (PF4) and RANTES (regulated on activation normal T cell expressed and secreted) are secreted by activated platelets and influence multiple cell types and biologic processes. For instance, PF4 inhibits progenitor cell proliferation and angiogenesis, while platelet-derived RANTES is involved in vascular recruitment of monocytes. However, little is known about functional interactions of PF4 and RANTES. Here we show that the presence of PF4 enhanced the arrest of RANTES-stimulated monocytes and monocytic cells on activated endothelial cells under flow conditions, while binding of PF4 to the monocyte surface was increased by RANTES. Both RANTES-triggered arrest and PF4 binding involved monocytic chondroitin sulfate. Ligand blots and surface plasmon resonance revealed a robust heterophilic interaction of PF4 with RANTES but not with RANTES variants defective in higher order oligomerization. The tetrameric mutant E26A bound to the monocyte surface without increasing PF4 binding, and monocyte arrest induced by E26A-RANTES was not enhanced by PF4. Stimulation of monocytes with supernatants of activated platelets triggered arrest involving RANTES and PF4, as shown by inhibition studies. Our results suggest that heterophilic interactions with PF4 require structural motifs important in RANTES oligomerization and amplify RANTES-triggered effects on monocyte adhesion. This may have implications for the modulation of inflammatory recruitment by platelet-derived chemokines.

Cell Adhesion↗
PubMed1998

New insights into the negative regulation of hematopoiesis by chemokine platelet factor 4 and related peptides.

Platelet factor 4 (PF4) has been recognized as an inhibitor of myeloid progenitors. However, the mechanism of action of this chemokine remains poorly understood. The present study was designed to determine its structure/function relationship. A series of peptides overlapping the C-terminal and central regions of PF4 were analyzed in vitro for their action on murine hematopoietic progenitor growth to assess the minimal sequence length required for activity. The peptides p17-58 and p34-58 possessed an increased hematopoietic inhibitory activity when compared with PF4, whereas the shorter peptides p47-58 and p47-70 were equivalent to the native molecule and the peptide p58-70 was inactive. The PF4 functional motif DLQ located in 54-56 was required for the activity of these peptides. The peptide p34-58 impaired to a similar extent the growth of colony-forming unit-megakaryocyte (CFU-MK) as well as burst-forming unit-erythroid (BFU-E) and colony-forming unit-granulocyte-macrophage (CFU-GM), whereas PF4 was more active on CFU-MK. In the experiments using purified murine CD34(+) marrow cells, statistically significant inhibition induced by p34-58 was shown at concentrations of 2.2 nmol/L or greater for progenitors of the three lineages, whereas that induced by PF4 was seen at 130 nmol/L for CFU-MK and 650 nmol/L for CFU-GM and BFU-E, indicating that the p34-58 acts directly on hematopoietic progenitors and its activity is approximately 60- to 300-fold higher than PF4. The p34-58, unlike PF4, lacked affinity for heparin and its inhibitory activity could not be abrogated by the addition of heparin. In addition, an antibody recognizing p34-58 neutralized the activity of p34-58 but not whole PF4 molecule. These results demonstrate that PF4 contains a functional domain in its central region, which is independent of the heparin binding properties, and provide evidence for a model of heparin-dependent and independent pathways of PF4 in inhibiting hematopoiesis.

Amino Acid Sequence↗
PubMed1985

Serial studies of platelet factor 4 and beta thromboglobulin during exercise in patients with coronary artery disease.

In vivo activation of platelets can be accurately measured by radioimmunoassays of platelet factor 4 (PF4) and beta thromboglobulin (beta TG). Studies that attempt to correlate increases in PF4 and beta TG levels with exercise-induced myocardial ischemia have yielded conflicting results. To further examine the natural history of release of PF4 and beta TG we used a method of serial samplings of these proteins during and after exercise in nine normal subjects and 24 patients with coronary artery disease (CAD). Mean values for PF4 and beta TG at rest, during each stage, and immediately after treadmill exercise were the same for normal subjects and for patients with positive and negative responses to exercise-tolerance tests (ETTs). However, nonparametric analysis and regression equations disclosed differences in trends of PF4 level during exercise; PF4 levels increased in normal subjects during exercise, while patients with positive ETTs had no change in PF4 levels and patients with negative ETTs actually showed a decrease in PF4. This investigation confirmed that exercise-induced myocardial ischemia is not associated with platelet aggregation as manifested by the release of the platelet-specific proteins PF4 and beta TG. Statistical analysis suggested that prior reports of elevated levels of PF4 during exercise could have been caused by technical and methodologic difficulties that were associated with the collection and handling of the samples.

Adult↗
PubMed1987

A formula for correcting for the in vitro release of platelet beta-thromboglobulin.

The interpretation of platelet beta-thromboglobulin (BTG) and platelet factor 4 (PF4) levels as indicators of in vivo platelet activation is complicated by the artefactual release of these proteins in vitro. A formula was devised to correct for in vitro platelet activation and release of BTG. Blood was collected from normal volunteers by an ideal method and BTG and PF4 levels determined by radioimmunoassay; these were the reference values. Blood from normal volunteers was activated in vitro by standing at room temperature. The BTG and PF4 released was measured at different time intervals. The relationship between BTG and PF4 released was measured at different time intervals. The relationship between BTG and PF4 was measured mathematically best described by a second degree polynomial function. The true plasma BTG value was then calculated by correcting for in vitro release by the general formula: BTG corrected = BTG measured - BtG for PF4 measured + BtG for PF4 reference The validity of the correction formula was tested in 10 normal subjects and in patients with either recent myocardial infarction(n = 10), familial hypercholesterolaemia(n = 10) or arterial prostheses(n = 14). Correction was adequate in normal subjects if the plasma BTG levels did not exceed 260ng/ml. In patients with a thrombotic tendency, the formula overcorrected for in vitro release. This could be ascribed to increased in vivo PF4 levels in these patients, especially those with prostheses. The reference values for PF4 in these patients, and especially those with vascular prostheses, were also higher than normal. The PF4 measured in their plasma thus reflects both in vivo and in vitro released protein. The hypothesis on which the correction formula was based, is therefore not always applicable. It may be possible to improve the correction by establishing formulae for specific disease groups.

Adult↗
PubMed2003

Platelet factor 4 neutralizes heparan sulfate-enhanced antithrombin inactivation of factor Xa by preventing interaction(s) of enzyme with polysaccharide.

Platelet factor 4 (PF4) is a heparin-binding protein which exhibits anti-heparin activities through the inhibition of antithrombin (AT)-dependent reactions with the serine proteases thrombin and factor Xa. PF4 also neutralizes heparan sulfate (HS), a glycosaminoglycan (GAG) present on the surface of endothelial cells, thereby possibly modulating an anticoagulant response. Previous models of PF4 mechanism did not distinguish whether PF4 causes steric hindrance of AT binding to fXa or of AT binding to the surface of the GAG chain. To shed light on the mechanism of PF4, studies of HS/heparin-catalyzed fXa inactivation by AT were undertaken. The results were consistent with PF4 directly interfering with AT binding to fXa rather than AT binding to the GAG chain, since PF4 did not prevent the heparin-dependent increase in AT intrinsic fluorescence. In fact, PF4 mechanism was competitive with respect to AT and non-competitive with respect to fXa, suggesting inhibition of important regulatory/catalytic interactions of fXa with the polysaccharide. Altogether, the results suggested a model by which PF4 bound to proximal (but distinct) sites to AT, resulting in steric interference of fXa binding to both polysaccharide and AT. It is proposed that PF4 inhibited the sequence of events recapitulated in the template mechanism describing heparin-dependent inhibition of fXa.

Anticoagulants↗
PubMed2000

Affinity purification of heparin-dependent antibodies to platelet factor 4 developed in heparin-induced thrombocytopenia: biological characteristics and effects on platelet activation.

Antibodies to heparin platelet factor 4 (H-PF4) complexes were purified from the plasma of three patients with heparin-induced thrombocytopenia (HIT) using affinity chromatography. From each plasma, the largest amount of antibodies was eluted with 2 M NaCl at pH 7.5 (peak 1) and the remainder was obtained using 0.1 M glycine/0. 5 M NaCl at pH 2.5 (peak 2). In an enzyme-linked immunosorbent assay (ELISA), we then showed that each patient had developed antibodies to PF4 displaying different characteristics. In patient 1, peak 1 IgG reacted almost exclusively with H-PF4 complexes, whereas peak 2 IgG had similar reactivity with PF4 whether or not heparin was present. Patient 2 expressed a mixture of IgA, IgM and IgG and both fractions bound to PF4 alone or to H-PF4 complexes. Finally, IgG in patient 3 only bound to H-PF4 and was unreactive with PF4 alone. Using [14C]-serotonin release assays, the antibodies developed in the three patients and exhibiting the strongest ability to activate platelets with heparin were those having the highest affinity to H-PF4. These results strongly support the hypothesis that HIT antibodies to PF4 are heterogeneous regarding their affinity and specificity for target antigens and this may greatly influence their ability to activate platelets and their pathogenicity.

Anticoagulants↗
PubMed1984

Platelet-released proteins as molecular markers for the activation process.

The desire to have a specific, sensitive marker for platelet activation was originally thought to lie in the development of RIAs for BTG and PF4. Although this wish has not been denied, the interpretation of the information obtained from such an analysis has proved far less rewarding. The principal challenge of these procedures is based upon the lack of a cause and effect relationship between a given disease and platelet activation, coupled with the differential clearance rate and mechanism for each of the discussed proteins. Thus, we have seen that the renal clearance rate and mechanism for each of the discussed proteins. Thus, we have seen that the renal clearance of a patient should be noted prior to interpreting the elevation of BTG. Similarly, since PF4 is removed from the circulation so rapidly, its plasma values tend to be lower than BTG by a factor of 5, although the significance of the BTG to PF4 ratio is questioned. Administration of heparin results in a heparin-induced increase in plasma PF4 levels but not for BTG, and this PF4 increase can be as great as 20-fold. PF4 and BTG values are also directly increased by pressure increases. Taken individually, these mediators each compromise the ability to correlate the significance of platelet protein increases with any single pathologic condition. When viewed collectively, an analysis of platelet-released proteins is best interpreted as an indication that the functional integrity of the platelet has been perturbed; the direct relationship of disease processes to platelet release is far from certain and is simply documented by the use of such described procedures. The final note of caution for these assays is to be found in the recent summary analysis of the standardization of both BTG and PF4. In this study, considerably greater variation among laboratories was noted for PF4 than was seen for BTG, and the study directors concluded that comparisons of results between laboratories should be regarded as unreliable due mainly to the use of different standards for each protein in a given laboratory. The final note of caution for these assays is to be found in the recent summary analysis of the standardization of both BTG and PF4. In this study, considerably greater variation among laboratories was noted for PF4 than was seen for BTG, and the study directors concluded that comparisons of results between laboratories should be regarded as unreliable due mainly to the use of different standards for each protein in a given laboratory.

Blood Platelets↗
PubMed1981

Stimulation of human leukocyte elastase by platelet factor 4. Physiologic, morphologic, and biochemical effects on hamster lungs in vitro.

The purpose of this study was to determine if human platelet factor 4 (PF4) stimulates human leukocyte elastase (HLE) against lung elastin. Lung elastin was purified from hamster lungs and tritiated by reduction with NaB3H4. We found that HLE activity against this substrate is increased by concentrations of PF4 as low as 1.6 microgram/ml, and that this stimulation increased linearly with additional PF4. Lungs removed from hamsters and inflated with solutions containing buffer alone, low dose HLE, HLE plus PF4, or PF4 alone were incubated for 2 h at 37 degrees C. Whereas low-dose HLE failed to lower lung elastin when compared to control animals, HLE stimulated by PF4 lowered lung elastin by 20%. PF4 alone had no effect. Furthermore, low-dose HLE failed to alter the mechanical properties of hamster lungs as measured by pressure-volume curves in saline, although there was a significant loss of lung elasticity in the mid- and high-lung volume ranges in lungs treated with HLE and PF4. Morphologic studies revealed that low dose HLE resulted in a minimal emphysemalike lesion whereas HlE plus PF4 caused a significantly more severe lesion. PF4 is capable of stimulating HLE against lung elastin, and this effect may have a role in the pathogenesis of emphysema.

Animals↗
PubMed2003

Platelet factor 4 promotes adhesion of hematopoietic progenitor cells and binds IL-8: novel mechanisms for modulation of hematopoiesis.

Platelet factor 4 (PF4) is an abundant platelet alpha-granule C-X-C chemokine that has weak chemotactic potency but strongly inhibits hematopoiesis through an unknown mechanism. We find that PF4 binds to human CD34+ hematopoietic progenitor cells (HPCs) with a median effective concentration of 1 microg/mL but not after exposure to chondroitinase ABC. PF4 enhances adhesion of HPCs to intact stroma. Committed progenitors also adhere avidly to immobilized PF4. This adhesion is time-dependent, requires metabolic activity, causes cytoskeletal rearrangement, and induces cell-cycle inhibition. Using extracellular acidification rate to indicate transmembrane signaling, we find that interleukin-8 (IL-8), but not PF4, activates CD34+ progenitors, and PF4 blocks IL-8-mediated activation. Surface plasmon resonance analysis shows that PF4 binds IL-8 with high (dissociation constant [Kd] = 42 nM) affinity. Nuclear magnetic resonance analysis of IL-8 and PF4 in solution confirms this interaction. We conclude that PF4 has the capacity to influence hematopoiesis through mechanisms not mediated by a classical high-affinity, 7-transmembrane domain chemokine receptor. Instead, PF4 may modulate the hematopoietic milieu both directly, by promoting progenitor adhesion and quiescence through interaction with an HPC chondroitin sulfate-containing moiety, and indirectly, by binding to or interfering with signaling caused by other, hematopoietically active chemokines, such as IL-8.

Antigens, CD34↗
PubMed2003

Platelet factor 4 enhances generation of activated protein C in vitro and in vivo.

Platelet factor 4 (PF4), an abundant platelet alpha-granule protein, accelerates in vitro generation of activated protein C (APC) by soluble thrombin/thrombomodulin (TM) complexes up to 25-fold. To test the hypothesis that PF4 similarly stimulates endothelium-associated TM, we assessed the influence of human PF4 on thrombin-dependent APC generation by cultured endothelial monolayers. APC generated in the presence of 1 to 100 microg PF4 was up to 5-fold higher than baseline for human umbilical vein endothelial cells, 10-fold higher for microvascular endothelial cells, and unaltered for blood outgrowth endothelial cells. In an in vivo model, cynomolgus monkeys (n = 6, each serving as its own control) were infused with either PF4 (7.5 mg/kg) or vehicle buffer, then with human thrombin (1.0 microg/kg/min) for 10 minutes. Circulating APC levels (baseline 3 ng/mL) peaked at 10 minutes, when PF4-treated and vehicle-treated animals had APC levels of 67 +/- 5 ng/mL and 39 +/- 2 ng/mL, respectively (P <.001). The activated partial thromboplastin time (APTT; baseline, 28 seconds) increased maximally by 27 +/- 6 seconds in PF4-treated animals and by 9 +/- 1 seconds in control animals at 30 minutes (P <.001). PF4-dependent increases in circulating APC and APTT persisted more than 2-fold greater than that of controls from 10 through 120 minutes (P < or =.04). All APTT prolongations were essentially reversed by monoclonal antibody C3, which blocks APC activity. Thus, physiologically relevant concentrations of PF4 stimulate thrombin-dependent APC generation both in vitro by cultured endothelial cells and in vivo in a primate thrombin infusion model. These findings suggest that PF4 may play a previously unsuspected physiologic role in enhancing APC generation.

Animals↗
PubMed1997

Suppression by platelet factor 4 of the myogenic activity of basic fibroblast growth factor.

The effect of platelet factor 4 (PF4) on myoblast cultures with or without basic fibroblast growth factor (bFGF) or other growth factors was investigated in the present in vitro experiments, with reference to bFGF binding to myoblast membrane fraction. When PF4 was added to the culture medium 1 day after myoblast cultivation, the nuclei of both myoblasts and myotubes were markedly reduced in number in a dose-dependent manner, whereas the inhibitory effect of PF4 on myoblast development was not observed when PF4 was added to the culture medium 3, 7, or 14 days after myoblast cultivation. In contrast, bFGF significantly increased the numbers of myoblast and myotube nuclei. When bFGF and PF4 were simultaneously added to the culture medium, PF4 abolished the facilitatory effects of bFGF on myogenesis. The real-time biospecific interaction analysis (BLA) core system showed that the myoblast membrane fraction at 1 day after cultivation contains bFGF-binding elements which are blocked by PF4 in a dose-dependent manner. Moreover, [126I]-bFGF binding experiments indicated the existence of both high and low affinity binding sites on myoblast membranes, although the high affinity binding sites decreased in number and the dissociation constant increased in value as the culture period was prolonged. Among the six other growth factors examined, acidic fibroblast growth factor and platelet-derived growth factor-BB stimulated myogenesis, and their effects were blocked by PF4 treatment. These findings suggest that: 1) PF4 inhibits myoblast proliferation and myotube formation only for a limited initial period of cultivation, possibly because of the time-dependent down-regulation of high affinity bFGF receptors: and 2) PF4 may be used as a tool to investigate the function of endogenous heparin-binding growth factors upregulated transiently at a certain developmental stage or in case of tissue damage and repair, even though it is not monospecific to bFGF.

Animals↗
PubMed1996

Inhibition of mesangial cell proliferation by platelet factor 4.

Platelet factor 4(PF4), an abundant platelet secretory product, is a strong candidate for modulating glomerular pathology. Because PF4 might be released from platelets and influence intrinsic cell growth during glomerular injury, the effect of PF4 on fetal calf serum- and platelet-derived growth factor (PDGF)-induced mesangial cell mitogenesis was examined. Mitogenesis was measured as the amount of 3H-thymidine incorporated into acid-precipitable material as well as by autoradiography. The effect of PF4 on mesangial cell expression of mRNA for PDGF A chain and transforming growth factor-beta (TGF-beta 1) was also examined. Fetal calf serum (10%)- and PDGF (10 ng/mL)-stimulated increases in mesangial cell 3H-thymidine incorporation were inhibited by incremental concentrations of PF4 (1 to 25 micrograms/mL) showing a maximum reduction of approximately 80% at 25 micrograms/mL of PF4. PF4 was effective when added 24 h before and 1, 4, and 8 h, but not 16 h after the addition of PDGF, indicating that inhibition occurred at delayed events in cell-cycle regulation. PF4 inhibited PDGF-induced increments in mRNA encoding PDGF A chain and TGF-beta 1. Also, PF4 did not interfere with PDGF receptor binding. The results of this study show that PF4 is a negative regulator of mesangial cell proliferation and suggest an interference in cell growth by pathways associated with modulation of the autocrine growth factors PDGF and TGF-beta 1.

Animals↗
PubMed1987

Uptake and processing of human platelet factor 4 by hepatocytes.

We previously demonstrated rapid clearance of human platelet factor 4 (PF4) from rabbit and rat blood, its accumulation in the liver, and elimination of PF4 degradation products in urine. The purpose of the present experiments was to characterize interaction of PF4 with cultured rat hepatocytes. 125I-PF4 was taken up by hepatocytes reaching maximum at 180 min. The association of 125I-PF4 with hepatocytes was two times greater at 37 degrees C than at 4 degrees C. At 37 degrees C degradation of 125I-PF4 by hepatocytes was also observed as indicated by the increase of 125I-PF4 radioactivity soluble in 6% trichloroacetic acid. By contrast, no uptake of 125I-beta-thromboglobulin antigen was observed. Autoradiography demonstrated that short incubation (5-20 min) of 125I-PF4 with hepatocytes results in the association of 125I-radioactivity with cell membranes while after longer incubation (60 min) radioactivity was also localized in the endosomes. Heparin inhibited binding and uptake of 125I-PF4 radioactivity by hepatocytes. We propose that part of PF4 released in the circulating blood by activated platelets is bound to the surface of hepatocytes and that it is further processed by these cells.

Animals↗
PubMed2003

[Effect of platelet factor 4 on the adherence of cord blood CD34(+) cells].

OBJECTIVE: To investigate the effects of platelet factor 4 (PF4) on the adherence, and the expressions of adherent molecules CD(49d) and CXCR4 and the receptor of SDF-1 of fresh and expanded cord blood CD(34)(+) cells. METHODS: CD(34)(+) cells were isolated from cord blood using MACS immune magnetic beads. The adherent ability was assayed by using crystal violet staining and the expression of adherent molecule CD(49d) and CXCR4 by FACS. RESULTS: (1) PF4 could increase the adherent ability of the fresh cord blood CD(34)(+) cells, the effect being positively correlated with the dose of PF4. (2) SDF-1 at concentration of 100 ng/ml increased the adherent ability of the fresh cord blood CD(34)(+) cells. (3) The spontaneous and the SDF-1 induced adherent ability of the cord blood CD(34)(+) cells began to decrease after being cultured for 10 days without PF4, while in the presence of PF4 at 100 ng/ml, the ability of the cord blood CD(34)(+) cell adhering to the stroma layer still remained at higher level. At day 14, the adherent ability was (262.04 +/- 64.81)% and (64.35 +/- 8.29)% in PF4 group and control group, respectively, if it was defined as 100% at day 0. SDF-1 at concentration of 100 ng/ml induced adherent ability was (138.31 +/- 32.39)% and (67.66 +/- 12.44)% in PF4 group and control group, respectively. (4) The expression of CD(49d) and CXCR4 increased 13.02% and 17.33%, respectively, when incubated with PF4. CONCLUSIONS: PF4 could increase the adherent ability and promote the expression of CD(49d) and CXCR4 of the cord blood CD(34)(+) cells, suggesting that PF4 promote the circulating stem cells homing to the marrow in the process of stem cells transplantation.

Antigens, CD34↗
PubMed1990

Platelet factor 4 inhibits human megakaryocytopoiesis in vitro.

Platelet factor 4 (PF4) is a multifunctional protein specific to platelets, synthesized in megakaryocytes and stored in alpha granules. This report of our work shows that PF4 potently inhibits human megakaryocyte colony formation in vitro. Colony formation by megakaryocyte progenitor cells from normal bone marrows was studied using the plasma clot culture system and indirect immunoperoxidase staining. Nonadherent mononuclear cells were co-cultured with various concentrations (0 to 20 micrograms/mL) of highly purified human PF4. Statistically significant inhibition of three classes of megakaryocyte progenitor cells, the mixed colony forming unit-megakaryocytes (mCFU-MK), the burst forming unit-megakaryocytes (BFU-MK), and the colony forming unit-megakaryocytes (CFU-MK), was seen at a PF4 concentration of 2.5 micrograms/mL or greater. PF4 had no effect on erythroid (BFU-E) and granulocyte/macrophage (CFU-GM) colony formation except at high concentration (5 micrograms/mL for BFU-E and 10 micrograms/mL for CFU-GM). When a concentration of 5 micrograms/mL PF4 was added at various time points during marrow culture, a reduction of megakaryocyte colony formation also occurred. In the presence of PF4 2.5 micrograms or 5 micrograms/mL, the percentage of mature type of colonies was found to be decreased compared with cultures with no added PF4. These data demonstrate that PF4 inhibits both proliferation and maturation of megakaryocyte progenitor cells in vitro and suggest that PF4 may play a role in autoregulating human megakaryocytopoiesis.

Bone Marrow↗
PubMed1980

In vivo release and turnover of secreted platelet antiheparin proteins in rhesus monkey (Macaca mulatta).

Human and rhesus monkey platelets secrete at least two antiheparin proteins: platelet factor 4 (PF4) and low affinity platelet factor 4 (LA-PF4). Neither of these proteins showed species-related antigenic differences. As determined by radioimmunoassay, the levels of PF4 and LA-PF4 antigen per 10(9) monkey platelets amounted to 10.7 and 20.3 microgram, respectively. One milliliter of monkey plasma prepared from blood collected into an anticoagulant composed of EDTA, prostaglandin E1, and theophylline solution contained 22.4 ng LA-PF4 and 8.0 ng PF4. Concentrations of these two platelet-specific proteins in monkeys closely resembled levels found in human platelets and plasma. Infusion of prostacyclin (PGI2) (100 or 300 ng/kg/min) into monkeys for 15 min resulted in a significant decrease of plasma levels of LA-PF4 antigen and of PF4 by 40%--60% (p < 0.0001). This decrease was related to the inhibitory effect of PGI2 on the secretion of platelets stimulated by a catheter or by venipuncture. Longer infusion of PGI2 did not produce further significant change. The supernate obtained after aggregation of human platelets stimulated by thrombin was injected into monkeys receiving PGI2 infusion. The disappearance of LA-PF4 antigen in monkey plasma followed a biphasic exponential curve with half-lives for the fast and slow components of 8.4 and 63 min. PF4 disappeared faster but followed the same pattern (half-lives for the fast and slow component of 2.1 and 70 min). Analysis of the experimental data suggests that the low levels of secreted platelet proteins in monkey plasma are related to their minimal in vivo release and to their rapid clearance.

Animals↗
PubMed1983

Effect of heparin on the in vivo release and clearance of human platelet factor 4.

Intravenous injection of heparin (100 U/kg) into normal volunteers resulted in an increase of platelet factor 4 (PF4) level in platelet-poor plasma from a mean value of 18.1 +/- 6.6 ng/ml before the injection to 257.9 +/- 68.3 ng/ml at 5 min after injection. PF4 antigen isolated from "postheparin plasma" by adsorption on heparin-agarose and elution with 2.0 M NaCl and "authentic PF4" isolated from human platelets showed identical patterns of migration as determined by sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Material released by washed human platelets was injected intravenously into rats. The clearance of PF4 followed a biphasic exponential pattern. The half-lives (T1/2) for the fast and slow components for control rats were 1.2 and 17.1 min. Heparin significantly extended the half-life of human PF4 in rat circulation. The clearance of PF4 injected together with heparin followed a single component model with a half-life of 27.6 min. Administration of heparin to rats that had been previously injected with human platelet releasate resulted in a 30-fold increase of plasma PF4 level in their circulation. The clearance of PF4 from the circulation of these rats (T1/2 = 45 min) fitted a single component model. We propose that PF4 is originally secreted by platelets into circulation and subsequently bound reversibly to vascular sites from which it can be released back into the circulation by heparin. The fast component of PF4 clearance that is abolished by heparin may reflect binding of this protein to the endothelial cells.

Animals↗
PubMed1986

In vitro effects of platelet factor 4 on normal human neutrophil functions.

Platelet factor (PF4) prepared from human outdated platelets by heparinagarose affinity chromatography was confirmed to be chemotactic for human neutrophils and in a concentration-dependent fashion caused significant release of lysosomal enzymes (myeloperoxidase, lysozyme, beta-glucuronidase) from human neutrophils treated with cytochalasin B. Lysosomal enzyme release from PF4-stimulated neutrophils was rapid and reached a plateau by 1-3 min. PF4 did not cause release of the cytoplasmic enzyme lactate dehydrogenase which indicates that exocytosis of granule-containing lysosomal enzymes did not result from cytolysis. In contrast, superoxide anion generation from human neutrophils stimulated with PF4 was undetectable even at the highest PF4 concentration tested (2 X 10(-5) M). Pretreatment of neutrophils with PF4 caused significant increased adherence of neutrophils to plastic surfaces and cultured pulmonary artery endothelial cells. The concentration of PF4 that elicited neutrophil chemotaxis, lysosomal enzyme release and increased adherence is slightly higher than those concentrations found in normal human sera. However, the results suggest that PF4 may be an important mediator in neutrophil-platelet interactions and the induction of acute inflammation especially at sites of platelet microthrombi where the concentration of PF4 would be elevated.

Animals↗