Mechanisms of osteopontin-induced stabilization of blood-brain barrier disruption after subarachnoid hemorrhage in rats.
OBJECTIVE
Osteopontin (OPN) is an inducible, multifunctional, extracellular matrix protein that may be protective against blood-brain barrier (BBB) disruption after subarachnoid hemorrhage (SAH). However, the protective mechanisms remain unclear.


METHODS
We produced the endovascular perforation model of SAH in rats and studied the time course of OPN induction in brains by Western blotting and immunofluorescence (n=50). Then, 34 rats were randomly assigned to sham (n=3), sham+OPN small interfering RNA (siRNA, n=3), SAH+negative control siRNA (n=14), and SAH+OPN siRNA (n=14) groups, and 109 rats were allocated to sham+vehicle (n=17), sham+recombinant OPN (n=17), SAH+vehicle (n=33), SAH+recombinant OPN (n=31), and SAH+recombinant OPN+L-arginyl-glycyl-L-aspartate motif-containing hexapeptide (n=11) groups. The effects of OPN siRNA or recombinant OPN on BBB disruption and related proteins were studied.


RESULTS
OPN was significantly induced in reactive astrocytes and capillary endothelial cells, peaking at 72 hours after SAH, during the recovery phase of BBB disruption. Blockage of endogenous OPN induction exacerbated BBB disruption and was associated with a reduction of angiopoietin-1 and mitogen-activated protein kinase (MAPK) phosphatase-1 (an endogenous MAPK inhibitor), activation of MAPKs, and induction of vascular endothelial growth factor-A at 72 hours after SAH, whereas recombinant OPN treatment improved it and was associated with MAPK phosphatase-1 induction, MAPK inactivation, and vascular endothelial growth factor-A reduction, which was blocked by L-arginyl-glycyl-L-aspartate motif-containing hexapeptide at 24 hours after SAH. Vascular endothelial growth factor-B and angiopoietin-2 levels were unchanged.


CONCLUSIONS
OPN may increase MAPK phosphatase-1 that inactivates MAPKs, upstream and downstream of vascular endothelial growth factor-A, by binding to L-arginyl-glycyl-L-aspartate-dependent integrin receptors, suggesting a novel mechanism of OPN-induced post-SAH BBB protection.
Suzuki
Hidenori
H
Department of Physiology, Loma Linda University School of Medicine, Loma Linda, Calif 92354, USA.
Hasegawa
Yu
Y
Kanamaru
Kenji
K
Zhang
John H
JH
D000704:Analysis of Variance; D042683:Angiopoietin-1 / Q000378:metabolism; D000818:Animals; D001253:Astrocytes / Q000378:metabolism* / Q000473:pathology; D001812:Blood-Brain Barrier / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology; D015153:Blotting, Western; D001921:Brain / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology; D054638:Dual Specificity Phosphatase 1 / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D008297:Male; D053495:Osteopontin / Q000378:metabolism*; D034741:RNA, Small Interfering; D011897:Random Allocation; D051381:Rats; D017207:Rats, Sprague-Dawley; D012720:Severity of Illness Index; D018709:Statistics, Nonparametric; D013345:Subarachnoid Hemorrhage / Q000378:metabolism* / Q000473:pathology / Q000503:physiopathology; D013997:Time Factors; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism
D042683:Angiopoietin-1; D034741:RNA, Small Interfering; D042461:Vascular Endothelial Growth Factor A; D053495:Osteopontin; D054638:Dual Specificity Phosphatase 1