4-Octyl itaconate ameliorates immune thrombocytopenia by modulating megakaryocyte autophagy and apoptosis

Objective: This study aimed to investigate the effects of 4-Octyl itaconate (4-OI) on immune thrombocytopenia (ITP) mice model and elucidate the underlying mechanism.Methods: An ITP mouse model was established by intraperitoneal injection of the monoclonal antibody MWReg30 and treated by 4-OI with/without chloroquine (CQ). The mice were divided into four groups: Control, ITP, ITP+4-OI and ITP+4-OI + CQ. Platelet (PLT) content was detected and bone marrow megakaryocytes were quantified using Giemsa staining. Apoptosis was evaluated by TUNEL staining, protein expression of Bax and Bcl-2 in bone marrow tissues was detected by western blotting, and megakaryocyte apoptosis ratio was assessed by flow cytometry detection of CD61+ cells. Peripheral blood mononuclear cells (PBMCs) were isolated from peripheral blood, and the mRNA expression of LC3II, Becin-1, and SQSTM1 were detected by qRT-PCR; Immunofluorescence evaluated LC3II, Beclin-1, and SQSTM1 expression in bone marrow, as well as the ratios of CD41+Beclin-1+ and CD41+LC3II+ megakaryocytes.Results: Compared to the ITP group, 4-OI treatment significantly increased PLT counts, while reduced the spleen index and bone marrow megakaryocyte numbers. 4-OI also increased the apoptosis rate of megakaryocytes by increasing Bax protein expression and reducing Bcl-2 expression in the bone marrow. LC3II and Beclin-1 expression increased in PBMCs and bone marrow tissues, whereas SQSTM1 expression decreased. Megakaryocytes exhibited reduced LC3II and Beclin-1. The autophagy inhibitor chloroquine (CQ) suppressed all the 4-OI-induced effects.Conclusion: 4-OI ameliorated ITP-induced thrombocytopenia by modulating autophagy-related proteins in megakaryocytes, inducing autophagy, and promoting apoptosis.