In vitro study of the direct effect of extracellular hemoglobin on myelin components.

There is a relationship between cerebral vasculature and multiple sclerosis (MS) lesions: abnormal accumulations of iron have been found in the walls of dilated veins in MS plaques. The sources of this iron can be varied, but capillary and venous hemorrhages leading to blood extravasation have been recorded, and could result in the release of hemoglobin extracellularly. Extracellular hemoglobin oxidizes quickly and is known to become a reactive molecule that triggers low-density lipoprotein oxidation and plays a pivotal role in atherogenesis. In MS, it could lead to local oxidative stress, inflammation, and tissue damage. Here, we investigated whether extracellular hemoglobin and its breakdown products can cause direct oxidative damage to myelin components in a peroxidative environment such as occurs in inflamed tissue. Oxidation of lipids was assessed by the formation of fluorescent peroxidized lipid-protein covalent adducts, by the increase in conjugated diene and malondialdehyde. Oxidation of proteins was analyzed by the change in protein mass. The results suggest that the globin radical could be a trigger of myelin basic protein oxidative cross-linking, and that heme transferred to the lipids is involved in lipid peroxidation. This study provides new insight into the mechanism by which hemoglobin exerts its pathological oxidative activity towards myelin components. This work supports further research into the vascular pathology in MS, to gain insight into the origin and role of iron deposits in disease pathogenesis, or in stimulation of different comorbidities such as cardiovascular disease.

Potential role of ferric hemoglobin in MS pathogenesis: Effects of oxidative stress and extracellular methemoglobin or its degradation products on myelin components.

There is a well-documented relationship between cerebral vasculature and multiple sclerosis (MS) lesions: abnormal accumulations of iron have been found in the walls of the dilated veins in cerebral MS plaques. The source of this iron is unknown, but could be related to the recognized phenomenon of capillary and venous hemorrhages leading to blood extravasation. In turn, hemorrhaging leading to hemolysis results in extracellular release of hemoglobin, a reactive molecule that could induce local oxidative stress, inflammation, and tissue damage. Our previous studies with a reduced form of hemoglobin (oxyHb) have demonstrated its ability to cause extensive lipid and protein oxidation in vitro, which would result in membrane destabilization. Here, we investigated in further detail the mechanism by which the more abundant oxidized form of extracellular hemoglobin (metHb), and dissociated hemin, cause direct oxidative damage to myelin components, specifically membrane-mimetic lipid vesicles and myelin basic protein (MBP), a highly-abundant protein in the CNS. Oxidation of lipids was assessed by the formation of conjugated diene/triene and malondialdehyde, and oxidation of MBP was demonstrated by the bityrosine formation and by the change in protein mass. Our results show that metHb causes oxidative damage to MBP and myelin lipids, partly by transferring its hemin moiety to protein and lipid, but mostly as an intact protein possibly via formation of a ferryl radical. These results elucidating the mechanism of extracellular hemoglobin-induced oxidative damage to myelin components support the need for further research into vascular pathology in MS pathogenesis, to gain insight into the role of iron deposits and/or in stimulation of different comorbidities associated with the disease.