Subarachnoid hemorrhage (SAH) by a rupture of cerebral aneurysms remains the most devastating cerebrovascular disease. Early brain injury (EBI) is increasingly recognized to be the primary determinant for poor outcomes, and also considered to cause delayed cerebral ischemia (DCI) after SAH. Both clinical and experimental literatures emphasize the impact of global cerebral edema in EBI as negative prognostic and direct pathological factors. The nature of the global cerebral edema is a mixture of cytotoxic and vasogenic edema, both of which may be caused by post‐SAH induction of tenascin‐C (TNC) that is an inducible, non‐structural, secreted and multifunctional matricellular protein. Experimental SAH induces TNC in brain parenchyma in rats and mice. TNC knockout suppressed EBI in terms of brain edema, blood‐brain barrier disruption, neuronal apoptosis and neuroinflammation, associated with the inhibition of post‐SAH activation of mitogen‐activated protein kinases and nuclear factor‐kappa B in mice. In a clinical setting, more severe SAH increases more TNC in cerebrospinal fluid and peripheral blood, which could be a surrogate marker of EBI and predict DCI development and outcomes. In addition, cilostazol, a selective inhibitor of phosphodiesterase type III that is a clinically available anti‐platelet agent and is known to suppress TNC induction, dose‐dependently inhibited delayed cerebral infarction and improved outcomes in a pilot clinical study. Thus, further studies may facilitate application of TNC as biomarkers for non‐invasive diagnosis or assessment of EBI and DCI, and lead to development of a molecular target drug against TNC, contributing to the improvement of post‐SAH outcomes.

There are no direct evidences showing the linkage between Toll-like receptor 4 (TLR4) and blood-brain barrier (BBB) disruption after subarachnoid hemorrhage (SAH). The purpose of this study was to examine if selective blockage of TLR4 prevents BBB disruption after SAH in mice and if the TLR4 signaling involves mitogen-activated protein kinases (MAPKs). One hundred and fifty-one C57BL/6 male mice underwent sham or endovascular perforation SAH operation, randomly followed by an intracerebroventricular infusion of vehicle or two dosages (117 or 585 ng) of a selective TLR4 antagonist IAXO-102 at 30 min post-operation. The effects were evaluated by survival rates, neurological scores, and brain water content at 24-72 h and immunoglobulin G immunostaining and Western blotting at 24 h post-SAH. IAXO-102 significantly prevented post-SAH neurological impairments, brain edema, and BBB disruption, resulting in improved survival rates. IAXO-102 also significantly suppressed post-SAH activation of a major isoform of MAPK p46 c-Jun N-terminal kinase (JNK) and matrix metalloproteinase-9 as well as periostin induction and preserved tight junction protein zona occludens-1. Another selective TLR4 antagonist TAK-242, which has a different binding site from IAXO-102, also showed similar effects to IAXO-102. This study first provided the evidence that TLR4 signaling is involved in post-SAH acute BBB disruption and that the signaling is mediated at least partly by JNK activation. TLR4-targeted therapy may be promising to reduce post-SAH morbidities and mortalities.

No abstract

Background and Purpose— Plasma levels of galectin-3—a matricellular protein—are increased after aneurysmal subarachnoid hemorrhage (SAH), but the functional significance remains undetermined. This study was conducted to evaluate whether modified citrus pectin (MCP; galectin-3 inhibitor) prevents post-SAH early brain injury, focusing on blood-brain barrier disruption. Methods— C57BL/6 male adult mice (n=251) underwent sham or filament perforation SAH modeling, followed by a random intracerebroventricular injection of vehicle or drug at 30 minutes post-modeling. First, vehicle-treated and 0.8, 4, 16, or 32 µg MCP-treated mice were assessed by neuroscore and brain water content at 24 and 48 hours post-modeling. Second, Evans blue extravasation, Western blotting, coimmunoprecipitation and immunostaining were performed in vehicle-treated or 4 µg MCP-treated mice at 24 hours post-modeling. Third, vehicle or R-galectin-3 (recombinant galectin-3) was administered to SAH mice simultaneously with vehicle or MCP, and neuroscore and Evans blue extravasation were evaluated at 24 hours post-modeling. Fourth, vehicle or R-galectin-3 was administered to MCP-treated SAH mice at 24 hours, and neuroscore and IgG immunostaining were evaluated at 48 hours post-SAH. Results— Among tested dosages, 4 µg MCP showed the best neuroprotective effects as to preventing neurological impairments and brain edema at 24 to 48 hours post-SAH. Four micrograms MCP attenuated post-SAH blood-brain barrier disruption and galectin-3 upregulation in brain capillary endothelial cells, associated with inactivation of ERK (extracellular signal-related kinase) 1/2, STAT (signal transducer and activator of transcription)-3, and MMP (matrix metalloproteinase)-9, and the consequent preservation of a tight junction protein ZO-1 (zonula occludens-1). Coimmunoprecipitation assay demonstrated physical interactions between galectin-3 and TLR (Toll-like receptor) 4. R-galectin-3 blocked the neuroprotective effects of MCP. Conclusions— MCP prevents post-SAH blood-brain barrier disruption possibly by inhibiting galectin-3, of which the mechanisms may include binding to TLR4 and activating ERK1/2, STAT-3, and MMP-9. This study suggests galectin-3 to be a novel therapeutic target against post-SAH early brain injury.

Tenascin-C (TNC), a matricellular protein, is upregulated in brain parenchyma after experimental subarachnoid hemorrhage (SAH). Recent studies emphasize that early brain injury (EBI) should be overcome to improve post-SAH outcomes. The aim of this study was to investigate effects of TNC knockout (TNKO) on neuronal apoptosis and neuroinflammation, both of which are important constituents of EBI after SAH. C57BL/6 wild-type (WT) mice or TNKO mice underwent sham or filament perforation SAH modeling. Twenty-five WT mice and 25 TNKO mice were randomly divided into sham+WT (n = 10), sham+TNKO (n = 8), SAH+WT (n = 15), and SAH+TNKO (n = 17) groups. Beam balance test, neurological score, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling staining, immunostaining of Toll-like receptor 4 (TLR4), and Western blotting were performed to evaluate neurobehavioral impairments, neuronal apoptosis, and neuroinflammation at 24 h post-SAH. Deficiency of TNC significantly alleviated post-SAH neurobehavioral impairments and neuronal apoptosis. The protective effects of TNKO on neurons were associated with the inhibition of a caspase-dependent apoptotic pathway, which was at least partly mediated by TLR4/nuclear factor-κB/interleukin-1β and interleukin-6 signaling cascades. This study first provided the direct evidence that TNC causes post-SAH neuronal apoptosis and neuroinflammation, potentially leading to the development of a new molecular targeted therapy against EBI.