Background: Immune thrombocytopenia (ITP) is an autoimmune bleeding disorder characterized by low platelets counts and a mostly mild- and in rare occasion life threatening bleeding symptoms. Previous studies, also from our group, have demonstrated a role of platelet apoptosis in the pathogenesis of childhood ITP. A mechanistic understanding of the ITP pathogenesis is still lacking, and most treatments increase the platelet counts and prevent the severe clinical manifestations a limited number of the patients only.

Aims: In our present study, we aimed to investigate apoptosis molecular mechanisms in ITP platelets and their precursors, the megakaryocytes (MKs).

Methods: We isolated platelets in vitro as described before also by our group (1,2). To differentiate MKs from PBMCs peripheral blood from acute and chronic ITP patients and controls, we used an adapted protocol from Salunke et al (3). As control cells we used the human megakaryoblastic cell line MEG-01, treated for 4h with plasma from acute and chronic ITP patients and healthy controls. We performed silencing experiments in control and ITP platelets and MKs, MEG-01 by using siRNA caspase-3 or suppressing apoptosis by treating with pan-caspase inhibitors (Z-VAD-FMK), and, in comparison, with the apoptosis inducer ABT-737. We determined the expression at mRNA levels of apoptosis pathway regulatory genes (Bax, Bcl-2, Bid, Bak, Apaf-1, cytochrome-c, caspase-3, -8, -9 and clusterin (CLU)) by qRT-PCR.

Results: We could demonstrate increased expression levels of some apoptotic genes such as CLU, cytochrome-c, Bax and Bak in ITP platelets and MKs compared to healthy controls from 5 ITP patients as well as in MEG-01 treated cells that could be silenced by using siRNA caspase-3.

Conclusion(s): Our results indicate a possible impairment of apoptosis pathway via upregulation of Clusterin and Bax in platelets and in their producers MKs that can lead to platelet destruction in ITP disease.

« Back to ISTH 2022 Congress

ISTH Congress Abstracts - https://abstracts.isth.org/abstract/molecular-mechanisms-of-apoptosis-in-platelets-and-megakaryocytes-in-pediatric-immune-thrombocytopenia/