Indirubin regulates M1/M2 polarization and inhibits ferroptosis in dextran sulfate sodium induced colitis and in cultured THP-1 cells

Context: Macrophages play a critical role in the pathogenesis of ulcerative colitis (UC). Indirubin (IDR), a natural ligand of the Aryl Hydrocarbon Receptor (AhR), has been shown to ameliorate DSS-induced colitis in our previous study (Liu Z et al.).

Objective: To investigate whether IDR exerts its protective effects by regulating M1/M2 polarization and inhibiting Ferroptosis in macrophages.

Materials and methods: Immunohistochemistry staining targeting CD206 and F4/80 was performed to evaluate the effect of IDR on the polarization of M1/M2 macrophages in colitis. Subsequently, the effects of IDR on the M1- or M2-polarization of THP-1-derived macrophages were investigated. Furthermore, the effects of IDR on Ferroptosis in the colon tissue of mice and on RSL3-induced Ferroptosis in THP-1-derived macrophages were assessed. The results were verified in mouse peritoneal macrophages.

Results: In addition to reducing the infiltrated macrophages, IDR treatment preserved CD206+ macrophages in DSS-induced colitis. Using cultured THP-1 cells, we demonstrated that IDR inhibited M1 polarization and prompted M2 polarization. Furthermore, we showed that IDR treatment decreased levels of 4-HNE while increasing GPX4 and NRF2 in DSS-induced colitis and THP-1 cells. IDR treatments also reduced cellular Reactive Oxygen Species (ROS) and iron content, and mitigated RSL3-induced Ferroptosis in THP-1-derived macrophages. Similarly, IDR augmented M2-polarization and alleviated Ferroptosis in peritoneal macrophages.

Discussion and conclusion: IDR skews the polarization of macrophages from M1 to M2. Furthermore, it inhibits Ferroptosis in both mice and THP-1-derived macrophages. These mechanisms may contribute to the therapeutic effects of IDR in the treatment of UC.

Ferroptosis; indirubin (IDR); macrophage polarization; reactive oxygen species (ROS).