Identification of low-density platelet populations with increased reactivity and elevated α-granule content

Milovanovic, Micha; Lysen, J.; Ramström, Sofia; Lindahl, Tomas; Richter, Arina; Järemo, Petter

doi:10.1016/j.thromres.2003.08.019

Cited by 19 publications

(20 citation statements)

References 16 publications

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“…The finding indicates release reactions occurring in vivo. Finally, the study confirms previous work [21] in showing that the lightest platelets were more activated in vivo. They further contained more P-selectin than peak platelets.…”

Section: Discussionsupporting

confidence: 91%

“…We also described that lowdensity populations display more membrane-bound fibrinogen indicating platelet activity. In contrast, the study also demonstrates that peak density platelets were less activated as estimated from in vivo platelet-bound fibrinogen [21] .…”

mentioning

confidence: 54%

“…Earlier work [21] shows that low-density populations have less dense bodies. Thus, low levels of dense granule and glycogen could explain why these cells have low densities despite enhanced ␣ granule content.…”

Section: Discussionmentioning

confidence: 89%

“…Previous work from our laboratory shows that healthy individuals have high-density platelets with more surfacebound fibrinogen in vivo. Consequently, they circulate in a more activated state [21] . We also described that lowdensity populations display more membrane-bound fibrinogen indicating platelet activity.…”

mentioning

confidence: 99%

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Platelet Density Distribution in Essential Thrombocythemia

Milovanovic

Lotfi

Lindahl

et al. 2010

Pathophysiol Haemos Thromb

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Essential thrombocythemia (ET) is characterized by high platelet counts and a slightly increased bleeding risk. Why severe hemorrhage does not occur more frequently is not known. Variations of platelet density (kg/l) depend mainly on cell organelle content in that high-density platelets contain more α and dense granules. This study compares ET patients (n = 2) and healthy volunteers (n = 2) with respect to platelet density subpopulations. A linear Percoll™ gradient containing prostaglandin E₁ was employed to separate platelets according to density. The platelet population was subsequently divided by density into 16 or 17 subpopulations. Determination of platelet counts was carried out. In each density fraction, platelet in vivo activity, i.e. platelet-bound fibrinogen, was measured using a flow cytometer. To further characterize platelet subpopulations, we determined intracellular concentrations of CD40 ligand (CD40L) and P-selectin in all fractions. Patients and controls demonstrated similar density distributions, i.e. 1 density peak. High-density platelets had more surface-bound fibrinogen in conjunction with signs of platelet release reactions, i.e. with few exceptions they contained less CD40L and P-selectin. Peak density platelets showed less surface-bound fibrinogen. These platelets contained less CD40L and P-selectin than nearby denser populations. The light platelets had more surface-bound fibrinogen than peak platelets together with elevated concentrations of CD40L. In ET, the malignant platelet production could exist together with platelets originating from normal megakaryocytes. It is also possible that clonal megakaryocytes produce platelets covering the entire density span. The ‘normal’ density distribution offers a tenable explanation as to why serious bleedings do not occur more frequently.

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Section: Discussionsupporting

confidence: 91%

mentioning

confidence: 54%

Section: Discussionmentioning

confidence: 89%

mentioning

confidence: 99%

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Platelet Density Distribution in Essential Thrombocythemia

Milovanovic

Lotfi

Lindahl

et al. 2010

Pathophysiol Haemos Thromb

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“…Details of the assays for determining platelet reactivity have been explained before [4,5]. In short, platelets were stimulated in whole blood with ADP (1.7 and 8.5 mol/L) and a thrombin receptor activating peptide (TRAP-6) (54 and 74 mol/L).…”

Section: Laboratorymentioning

confidence: 99%

Atrial fibrillation and platelet reactivity

Milovanovic¹,

Fransson²,

Hallert³

et al. 2010

International Journal of Cardiology

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Objectve: Relationships between platelet reactivity and atrial fibrillation (AF) have not been investigated before. Thus, platelet reactions after stimulation were compared with the presence of sinus rhythm (SR) after a surveillance period exceeding two years after elective electrical cardioversion.Merthods: 33 individuals with non-valvular AF subject to elective electrical cardioversion were consented. Immediately before cardioversion determination of markers (see below) reflecting platelet reactivity was carried out. After an average of 26±8(SD) months an ECG was analyzed and platelet laboratory analysis were repeated.A flow cytometry technique was used to determine platelet reactivity i.e. surface bound fibrinogen after stimulation. ADP (1.7 and 8.5 mol/L) and a thrombin receptor activating peptide (TRAP-6) (54 and 74 mol/L) were used as platelet agonists. Results:As the study was brought to an end subjects with AF (n=18) had a trend towards lower platelet reactivity. Compared with day 1 it reached statistical significance (p=0,016) when using 1.7 mol/L ADP to activate platelets. Responses towards the other agonists did not change significantly over time. In contrast, after 26±8(SD) months subjects with SR (n=15) had significant lower platelet reactivity when stimulating with both agonists pairs. The p-values for the two ADP dilutions (1.7 and 8.5 mol/L) were p=0.002 and p=0.031, respectively. The corresponding figures for TRAP-6 (54 and 74 mol/L) proved to be p=0.042 and p=0.006. Conclusion:After 26±8(SD) months patients returning to AF had higher platelet reactivity than those who remained in SR. The finding provides a link between arrhythmia and clot formation.3

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Platelet activation patterns in platelet size sub-populations: differential responses to aspirin in vitro

Mangalpally

Siqueiros-Garcia

Vaduganathan

et al. 2010

J Thromb Thrombolysis

105

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Circulating platelets are heterogeneous in size and structure. Whether this translates into differences in platelet function and efficacy of antiplatelet therapy is unclear. Hence, we decided to investigate the activation patterns among different platelet populations differentiated by size, and to compare the inhibitory effects of aspirin in these populations. Circulating platelets from 9 healthy volunteers were separated by size and stratified into the largest and smallest quintiles. Platelets were stimulated with 75 μM arachidonic acid (AA), 10 μM ADP or 25 μM TRAP. Alpha-granule protein secretion and expression (P-selectin, VWF, fibrinogen), surface-protein activation (activated integrin αIIbβ3) were assessed. Platelet thromboxane B(2) (TxB(2)) synthesis following AA stimulation was measured in vitro before and after incubation with 265 μM aspirin. Reticulated (juvenile) platelets were assessed using thiazole orange staining. A greater number of large platelets in the largest quintile were reticulated compared with the smallest quintile (6.1 ± 2.8% vs. 1.2 ± 1.5% respectively, p < 0.001). Larger platelets also synthesized more TxB(2) than small platelets both before (1348 ± 276 pg/mL vs. 1023 ± 214 pg/mL, respectively, p = 0.01) and after aspirin (1029 ± 190 pg/mL vs. 851 ± 159 pg/mL, respectively, p = 0.03). After stimulation with each agonist, a greater proportion of large platelets bound fibrinogen, VWF, P-selectin and activated integrin αIIbβ3 than small platelets both in the presence and in the absence of in vitro aspirin. In an in vitro setting, large platelets appear to be more active than small platelets and continue to be more active even after in vitro aspirin. Platelets exhibit heterogeneity in size and structure. Whether this translates into platelet function and efficacy of antiplatelet therapy is unclear. We evaluated platelet functional properties and the effects of aspirin on separated platelet subpopulations in an in vitro setting. Platelets were sorted into the largest and smallest size quintiles using flow cytometry forward scatter. Alpha-granule protein release, dense granule content, surface protein activation and thromboxane synthesis were significantly greater in large platelets compared with small platelets, before and after stimulation with arachidonic acid, ADP and TRAP. Even after incubation with aspirin, large platelets continued to be more active than small platelets. In conclusion, large platelets are more active than small platelets and aspirin fails to eliminate these differential activation properties.

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Identification of low-density platelet populations with increased reactivity and elevated α-granule content

Cited by 19 publications

References 16 publications

Platelet Density Distribution in Essential Thrombocythemia

Platelet Density Distribution in Essential Thrombocythemia

Atrial fibrillation and platelet reactivity

Platelet activation patterns in platelet size sub-populations: differential responses to aspirin in vitro

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