Low-Dose Decitabine Could Improve the Imbalance of M1/M2 Ratio in Immune Thrombocytopenia

Background

Primary immune thrombocytopenia (ITP) is an acquired autoimmune hemorrhagic disease characterized by increased immune-mediated platelet destruction and insufficient platelet production. As the impaired balance of M1 macrophage and M2 macrophage subsets increases platelet destruction, the M1/M2 ratio has become the focus of attention. The demethylating agent, decitabine (DAC) has been successfully used in the treatment of myelodysplastic syndrome with a moderate response in thrombocytopenia. Interestingly, at low doses, DAC induces megakaryocyte differentiation and maturation in vitro via inducing hypomethylation, thus having the potential to promote platelet release in ITP patients. In addition, several reports suggest that DAC can promote the polarization of macrophages to M2 macrophages. Therefore, we hypothesized that low dose of DAC treatment could lead to shrinkage of platelet destruction in murine model of ITP via promoting the M2 macrophage polarization.

Methods

Platelets isolated from Wistar rats were transferred into CBA/Ht mice via intraperitoneal injection weekly to induce a marine model of primary ITP. Starting on day 8, ITP mice were treated with low-dose DAC (0.25 mg/kg/d, n = 8) or PBS (n = 8) for two consecutive weeks intraperitoneally. Platelet counts were recorded every three days. The spleens were removed at day 10, from which M1 macrophages (F4/80⁺ CD11c⁺) and M2 macrophages (F4/80⁺ CD206⁺) were identified by flow cytometry analysis and immunofluorescence.

Results

The platelet levels in ITP group began to descend sharply at the second week after transferring platelets when compared with the control group (735.0 ± 25.2 × 10⁹vs. 390.3 ± 70.0 × 10⁹, p < 0.05). Mice in DAC-treated group recovered from thrombocytopenia by day 10 (713.8 ± 29.1 × 10⁹vs. 311.0 ± 4.5 × 10⁹, p < 0.001). In the following days, it still showed significantly higher level of platelet in DAC-treated group than ITP group despite the trend to decline. Comparing with the ITP group, immunofluorescence revealed that spleen samples showed stronger M2 expression pattern in DAC group, and the ratio of M1/M2 declined regardless of the fact that both the percentage of M1 macrophage and M2 macrophage was increased when DAC was applied.

Conclusions

Low dose DAC can promote the polarization of both M1 and M2 macrophage, especially M2 macrophage, thus reversing the unbalanced M1/M2 ratio in ITP group. Further studies are required to investigate the mechanism underlies the specialized polarization of macrophages.