Background: Human platelets vary in size, function, and age. These platelet features change when platelet turnover increases (e.g. by major surgery, sepsis). We established a platelet apheresis model to deplete platelets from healthy volunteers with subsequent production of young platelets and minimal interfering factors or comorbidities.

Aims: We aim to further characterize large and small human platelets during normal and increased platelet generation regarding age and function.

Methods: Blood was obtained from healthy plateletpheresis donors before and on days 3, 7, 10 after apheresis. Platelets were isolated and separated into large and small platelet fractions by differential centrifugation. We analyzed reticulated platelets labelled with Cy5-conjugated oligo-dT and oligo-dA, HLA-I expression by binding of anti-HLA-A, B, C-APC antibodies and desialylation of platelet glycoproteins by binding of Fluorescein-labelled lectins (RCA, ECL), using flow cytometry.

Results: The overall proportion of RNA-rich young platelets increased after plateletpheresis (p=0.0059, Figure 1A) and was more pronounced in small platelets on day 7 and 10 compared to large ones (Figure 2A).

Likewise, HLA-I expression, as a marker for young platelets, was significantly increased after plateletpheresis in unseparated (day 3: p=0.0062, Figure1B), large (day 3: p=0.0039) and small platelets (day 7: p=0.0156, Figure 2B).

Interestingly, platelet desialylation, which is increased during aging of platelets, was not significantly changed after plateletpheresis. Large platelets were more desialylated compared to small platelets at all time points (Figure 2C).

Conclusion(s): Platelet loss and consecutive platelet production induced by apheresis, increased the proportion of reticulated platelets with higher HLA-I expression in small and large platelets. This indicates simultaneous synthesis of young large and small platelets. The assumption that large platelets always represent younger platelets has to be revised when platelet turnover increases.

Figure 1: The proportion of reticulated platelets and HLA-I expression is increased after plateletpheresis in the whole platelet population

The proportion of reticulated platelets is given in percentage was significantly increased on day 3 after apheresis compared to before [A]. HLA-I expression is shown by relative Mean fluorescence intensity -MFI- and was significantly increased on day 3 and 7 after plateletpheresis [B]. Results are shown as mean with SEM. **P < .01, *P < .05; n>9

Figure 2: Distribution of age markers in large and small platelets

The proportion of reticulated platelets after apheresis was in both platelet fraction in and increase was more persistent in the smaller platelet fraction [A]. Expression of HLA-I on platelet surface is shown by MFI. Due to the larger surface area of the large platelet fraction, higher HLA-I levels were measured for this population through the whole observation period. However, apheresis significantly increased HLA-I levels in large and small platelets [B]. Binding of the lectin RCA displays the status of desialylation of glycoproteins on platelet surface and is shown by MFI in flow cytometry. Results are shown as mean with SEM. Statistical significance was tested with Wilcoxon rank sum test **P < .01, *P < .05; n>9

« Back to ISTH 2022 Congress

ISTH Congress Abstracts - https://abstracts.isth.org/abstract/under-increased-platelet-turnover-large-and-small-platelets-are-produced-simultaneously/