Hemoglobin-related molecules injure blood-brain barrier via iron accumulation

[Speaker] Takahiko Imai:1
[Co-author] Sena Iwata:1, Tasuku Hirayama:2, Hideko Nagasawa:2, Shinsuke Nakamura:1, Masamitsu Shimazawa:1, Hideaki Hara:1
1:Molecular Pharmacology, Gifu Pharmaceutical University, Japan, 2:Pharmaceutical and Medicinal Chemistry, Gifu Pharmaceutical University, Japan

After hemorrhagic stroke, hemoglobin (Hb)-related molecules (HRMs) exist in hematoma and damage brain tissues. The toxic mechanism is related to oxidative stress and inflammatory reactions. Previous reports suggest that HRMs have detrimental effect on neurons, however, the effects on blood-brain barrier (BBB) components such as endothelial cells and pericytes are still unknown. This study demonstrated the harmful effects of HRMs on BBB components in vitro and in vivo.

In vitro study, human brain microvascular endothelial cells (HBMVECs) and pericytes (HBMVPs) were incubated with HRMs (Hb or hemin), and then evaluated cellular functions. In vivo study, we evaluated BBB permeability and neurological function with hemin intracerebral injected mouse model. Moreover, to investigate the contribution of iron accumulation in HRMs damage, we used the bivalence iron (Fe2+)-detectable fluorescence probe, Si-RhoNox-1.

HRMs induced cell death approximately 10 % via increasing reactive oxygen species production by 3 to 10 folds in both cells, and increased the intracellular Fe2+ accumulation by 2 to 8 folds. These changes were canceled by co-treatment with iron chelator (Bipyridil). In vivo study, the hemin injection caused neuronal deficit, and aggravated BBB permeability more than 10 times compared with Sham group at 3 days after injection. Expression of BBB components proteins, tight junction proteins (vascular endothelial-cadherin, occludin) and platelet-derived growth factor-β(PDGFR-β) were decreased. Moreover, Fe2+ was detected in peri-hematoma region.

HRMs released from hematoma damage BBB via oxidative stress, and these mechanisms may involve in the intracellular Fe2+ accumulation. Investigation the intracellular movement of iron may lead to new therapeutic target in BBB damage after hemorrhagic stroke. This study might help that iron chelators and compounds affected to the iron are applied in clinical stage.

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