PB0677 - Itaconate derivative 4-OI inhibits M1 macrophage polarization and restores its immunomodulatory function in immune thrombocytopenia through metabolic reprogramming

Background: Aberrant macrophage polarization is involved in the pathogenesis of immune thrombocytopenia (ITP). Itaconate, a Krebs cycle-derived immunometabolite, is synthesized by myeloid cells and could modulate cellular metabolism and inflammatory response.

Aims: This study aimed to evaluate the immune-regulatory effects of 4-octyl itaconate (OI), a cell-permeable derivative of itaconate, on macrophages in patients with ITP.

Methods: Human monocytes derived macrophages were induced with M-CSF and treated with or without 4-OI. The cytokines TNF-α and IL-6 were assayed with ELISA. The phenotypes of macrophages were measured by flow cytometry (FCM). CFSE labelled CD4+ T cells were cocultured with macrophages, and the proliferation of T cells was analyzed by FCM. Besides, platelets were separated and stained with CMFDA, and macrophage phagocytosis was evaluated by FCM. The aerobic glycolysis of macrophage in ITP patients were detected by Seahorse XFe96 Analyzer and qPCR.

Results: We demonstrated that 4-OI decreased the expression of co-stimulatory molecules CD80 and CD86 in M1 macrophages. Besides, 4-OI inhibited the pro-inflammatory cytokines TNF-α and IL-6 secreted by M1 macrophages, indicating 4-OI might inhibit the M1 polarization of macrophages. We identified that 4-OI pre-modulated macrophages suppressed the proliferation of CD4+ T cells and promoted the differentiation of Treg cells compared with unmodulated macrophages. In addition, 4-OI treatment deceased phagocytic capacity of M1 macrophage towards antibody-coated platelets in ITP patients. Furthermore, M1 from ITP patients showed lower extracellular acidification rate and higher glycolysis rates, glycolytic capacity, and glycolytic reserve compared to those from healthy controls, which could be corrected by 4-OI treatment. Glycolysis inhibitor 2-DG could also inhibit the polarization of M1 macrophage and restore its immunomodulatory functions.

Conclusion(s): Our results showed itaconate derivative 4-OI inhibits M1 polarization and restores its immunomodulatory function through metabolic reprogramming. The study provides a novel therapeutic option for the treatment of ITP.