Neuroprotection From Intracerebral Hemorrhage Following Pharmacological Inhibition of GSK3β Depends on HFE Gene Status.

Iron release from hemoglobin breakdown following an intracerebral hemorrhage (ICH) is a key mediator in stroke-induced cytotoxicity. We have previously demonstrated that mice carrying the H67D mutation in the homeostatic iron regulatory gene (HFE) experience marked neuroprotection following ICH. This improvement is likely due to an endogenous upregulation in the Nrf2 antioxidant system. Prior studies in H67D mice discovered decreased activity in GSK3β, a kinase that functions to break down Nrf2. Interestingly, pharmacological inhibition of GSK3β has been shown to vastly improve outcomes in ICH animal models. However, it remains unclear whether this pathway is responsible for the enhanced antioxidant response in H67D animals. In this study, H67D and WT mice received daily injections of intraperitoneal SB216763, a selective inhibitor of GSK3β, 14 days prior to ICH. The functional motor recovery of each animal was assessed by rotarod and neurodegeneration was measured using Fluorojade-B. Immunoblotting assessed the antioxidant response and GSK3β activity through Nrf2, GPX4, FTH1, and β-Catenin. At 3 days post-ICH, SB216763-treated WT mice display enhanced functional recovery, decreased degenerated neurons, and increased brain levels of Nrf2 and GPX4 compared to WT-Vehicle-Controls. Further, SB216763 treatment in H67D mice did not result in any significant changes in measured outcomes compared to H67D-Vehicle-Controls. In conclusion, WT mice benefit from GSK3β inhibition following ICH whereas H67D animals do not. This suggests that the regulation of the antioxidant response may have reached its biological limit in H67D animals. Importantly, these data suggest that clinical trials aimed towards improving ICH outcomes, especially through GSK3β inhibition, must take into account HFE genotype as this mutation, present in nearly 20% of individuals worldwide, may alter ICH recovery regardless of therapy.