The placenta is crucial for successful human pregnancy and fetal health. During pregnancy, the placenta plays pivotal roles in endorsing the mother’s physiological adaptation to immunological acceptance, gas and nutrient exchange, and support of the developing embryo. Iron is an essential micronutrient through its participation in the structure of hemoproteins and its role in redox reactions. Free iron is a harmful pro-oxidant via the Fenton reaction that forms free hydroxyl radicals and increases oxidative stress and lipid peroxidation. Some pregnancy diseases have been associated with dysregulation in iron metabolism: pre-eclampsia is associated with higher iron maternal and ferritin concentrations. Ferroptosis is an iron-dependent, non-apoptotic programmed cell death. Emerging evidence supports the idea that ferroptosis may play a key role in placental dysfunction.

In this work, we aimed to explore the mechanism of heme and ferroptosis toxicity in human placental cells and to consider their potential reversibility using either antioxidant (Ferrostatin-1; Fer-1) or iron chelators (Deferoxamine, DFO). Therefore, we evaluated on BeWo cells (human trophoblastic cells model) treated either with heme (heme arginate 0.075 mg/mL) or ferroptosis inducer (erastin 5µM) the cell viability, the expression of markers specific to ferroptosis, and the effects on heme anabolism/catabolism and iron acquisition and storage.

Treatment with heme arginate and erastin showed a 40% decrease in BeWo cell viability assessed by MTT assay associated with an LDH release three times greater than for the untreated control group. DFO and Fer-1 partially reversed the cytotoxic effect of heme and erastin, respectively. Glutathione peroxidase 4 expression decreased by nearly 50% following heme arginate and erastin treatment. Lipid peroxidation was increased by erastin (45%), and co-treatment with 10 μM Fer-1 reduced the lipid peroxidation to the initial level, as evidenced using the BODIPY 581/591 C11 probe. Analysis revealed a significant change in the heme and iron metabolism pathways at transcriptional and post-transcriptional levels, with distinct patterns for heme and ferroptosis inducer treatments. Interestingly, compared to the untreated group, the expression of transferrin receptor 1 (TFR1) was upregulated by 1.5-fold in erastin-treated BeWo despite iron availability in this model. As expected, TFR1 protein and transcript markedly decreased in heme-treated cells; DFO significantly reversed this effect.

This study will help decipher heme dysmetabolism’s impact on placental pathologies and develop personalized treatments.