Myocardial Iron Overload in an Experimental Model of Sudden Unexpected Death in Epilepsy

Abstract

Uncontrolled repetitive generalized tonic-clonic seizures (GTCS) are the main risk factor for sudden unexpected death in epilepsy (SUDEP). GTCS can be observed in models such as Pentylenetetrazole kindling (PTZ-K) or pilocarpine-induced Status Epilepticus (SE-P), which share similar alterations in cardiac function, with a high risk of SUDEP. Terminal cardiac arrhythmia in SUDEP can develop as a result of a high rate of hypoxic stress-induced by convulsions with excessive sympathetic overstimulation that triggers a neurocardiogenic injury, recently defined as "Epileptic Heart" and characterized by heart rhythm disturbances, such as bradycardia and lengthening of the QT interval. Recently, an iron overload-dependent form of non-apoptotic cell death called ferroptosis was described at the brain level in both the PTZ-K and SE-P experimental models. However, seizure-related cardiac ferroptosis has not yet been reported. Iron overload cardiomyopathy (IOC) results from the accumulation of iron in the myocardium, with high production of reactive oxygen species (ROS), lipid peroxidation, and accumulation of hemosiderin as the final biomarker related to cardiomyocyte ferroptosis. Iron overload cardiomyopathy is the leading cause of death in patients with iron overload secondary to chronic blood transfusion therapy; it is also described in hereditary hemochromatosis. GTCS, through repeated hypoxic stress, can increase ROS production in the heart and cause cardiomyocyte ferroptosis. We hypothesized that iron accumulation in the "Epileptic Heart" could be associated with a terminal cardiac arrhythmia described in the IOC and the development of state-potentially in the development of SUDEP. Using the aforementioned PTZ-K and SE-P experimental models, after SUDEP-related repetitive GTCS, we observed an increase in the cardiac expression of hypoxic inducible factor 1α, indicating hypoxic-ischemic damage, and both necrotic cells and hemorrhagic areas were related to the possible hemosiderin production in the PTZ-K model. Furthermore, we demonstrated for the first time an accumulation of hemosiderin in the heart in the SE-P model. These results suggest that uncontrolled recurrent seizures, as described in refractory epilepsy, can give rise to high hypoxic stress in the heart, thus inducing hemosiderin accumulation as in IOC, and can act as an underlying hidden mechanism contributing to the development of a terminal cardiac arrhythmia in SUDEP. Because iron accumulation in tissues can be detected by non-invasive imaging methods, cardiac iron overload in refractory epilepsy patients could be treated with chelation therapy to reduce the risk of SUDEP.

Keywords: SUDEP; epilepsy; ferroptosis; heart failure; iron overload cardiomyopathy.

Figure 1

Hif 1-α immunoreactivity staining images and graphics. Immunoreactive areas are shown with a black arrow. Compared with the control, ^ap < 0.05 was considered significant. x400.

Figure 2

Plots of Hematoxylin-eosin staining image and cardiomyocyte length. Necrotic cells are shown with a yellow arrow and hemorrhagic areas with a black arrow. Compared with the control ^ap < 0.05 was considered significant.

Figure 3

Iron deposition in cardiomyocytes. (A) Micrograph at x400 showing cardiomyocites (red) and iron deposition (blue). (B) Quantitation of positive iron deposition in myocardial coronal sections of four rats after the third SE *p < 0.05.

Figure 4

Proposed mechanism for cardiac iron overload related to postconvulsive hypoxia-ischemia. Iron transported by holotransferrin is recognized by the plasma membrane receptor (Trf-R). After endocytosis, Fe³⁺ is reduced to Fe²⁺ in endosomes by the metalloreductase six-transmembrane epithelial antigen of prostate 3 (Steap3) and released from endosomes by divalent metal transporter 1 (DMT1). After the Fenton reaction, iron promotes lipid peroxidation via the activation of lipoxygenases, resulting in hemosiderin accumulation. On the other hand, the axis composed of the cystine/glutamate antiporter system X_c, Glutation (GSH) and Glutation peroxidase 4 (GPX4) is as a key regulator for tissue iron overload and ferroptosis. Other possible mechanisms of cardiomyocyte death are listed on the left side of the picture.