Abstract Number: PB1673
Meeting: ISTH 2020 Congress
Theme: Platelets and Megakaryocytes » Platelet Function and Interactions
Background: Thrombocytopenia is one of the most challenging limitations for lumi-aggregometry assays and even more if platelet size is increase. Interpretation in such situations requires deep knowledge and some cases need complementary assays to get a conclusion. Flow cytometry (FC) assays assess platelet behavior individually and in combination with lumi-aggregometry, provides a better approach about platelet function. Mepacrine uptake/release assay by FC (Mep u/r) is used to characterized storage pool disease (SPD) and evaluates granule content release.
Aims: To evaluate some technical aspects of Mep u/r and the contribution along with lumi-aggregometry in functional characterization of patients with thrombocytopenia.
Methods: We evaluate a normal group (NG, n: 37), a SPD group (SPD, n: 4) as pathologic control, and 14 patients with thrombocytopenia (TP1-14; platelet count < 150.109/L). Mep u/r was carry out as Cai et al. 2016, using platelet rich plasma, Trap-6 60uM as agonist inducing maximum degranulation. Platelet count dependency was tested in 5 individuals from NG at 50-100-300.109platelets/L. ATP release by lumi-aggregometry (ATPr) were performed using ADP, ADR, arachidonic acid and collagen. TP group results was interpreted according to platelet count.
Results: Mep u/r was independent of the platelet count in the evaluated range (table 1). SPD group had normal platelet count, absent ATPr and low Mep u/r. ATPr in PT group was normal in PT1-11 and correlates well with Mep u/r except in 2 cases (PT10-11). Degranulation of PT10-11 was reevaluated with PMA on FC showing normal results. PT12-14 had decrease ATPr or its inconclusive due to big platelet size and were diagnosed as SPD through abnormal Mep u/r (table 2).
Conclusions: Mep u/r can be used on thrombocytopenia and correlates well with ATPr. PT10-11 showed abnormal Mep u/r with TRAP-6 exclusively, suggesting some alteration in its signaling pathway. In such situations it may be necessary use more than one agonist.
| Platelet count | Mepacrine | |
| Uptake | Release ( TRAP-6 60μM) | |
| ( .109/L) | log (mep/aut)/log FSCH | log (mep/mep-r) |
| 0.55±0.09 (mean ±2SD) | 0.55±0.17 (mean± 2SD) | |
| 50 (n=5) | 0.54±0.12 | 0.56±0.11 |
| 100 (n=5) | 0.55±0.04 | 0.55±0.10 |
| 300 (n=5) | 0.53±0.08 | 0.53±0.03 |
| ANOVA | p< 0.05 (ns) | p< 0.05 (ns) |
| aut= platelets MFI without mepacrine ; mep= resting platelets MFI with mepacrine;FSCH= mean of frontal scattering histogram; mep-r= platelet MFI after TRAP-6 stimulation | ||
[Table 1_ Comparation between means]
| Platelet count (range= 150-450) (.109/L) | ATPr* (according to platelet count) | Mepacrine Uptake (range= 0.46-0.64) | Mepacrine Release (range= 0.38-0.72) | |
| NG (n= 37) | 150-490** | Normal | 0.46-0.53** | 0.46-0.70** |
| SPD (n=4) | 150-338** | Absent with all agonist | 0.19-0.33** | 0.04-0.24** |
| PT1-9 (n=9) | 58-146** | Normal | 0.53-0.62** | 0.47-0.79** |
| PT10 | 55 | Normal | 0.59 | 0.08 |
| PT11 | 110 | Normal | 0.60 | 0.15 |
| PT12 | 108 | Absent with all agonits | 0.28 | 0.12 |
| PT13 | 82 | Absent with all agonist | 0.30 | 0.18 |
| PT14 | 74 | Inconclusive | 0.34 | 0.29 |
| *AA: 0,5-1 mM; ADP: 2,5-5 uM; ADR: 10-50 uM: COL1 : 1- 8µg/ml; **:min-max range | ||||
[Table 2_ Lumy-Aggregometry and FC results]
To cite this abstract in AMA style:
Asensio M, Alberto MF, Bermejo E, Kinen A, Agazzoni M, Romero ML, Sanchez-Luceros A. Mepacrine Uptake & Release Assay in Thrombocytopenic Patients [abstract]. Res Pract Thromb Haemost. 2020; 4 (Suppl 1). https://abstracts.isth.org/abstract/mepacrine-uptake-release-assay-in-thrombocytopenic-patients/. Accessed November 28, 2023.« Back to ISTH 2020 Congress
ISTH Congress Abstracts - https://abstracts.isth.org/abstract/mepacrine-uptake-release-assay-in-thrombocytopenic-patients/