gms | German Medical Science

Objective: Breakdown of the blood-brain barrier (BBB) including the loss of endothelial tightness and components of the neurovascular unit plays a crucial role in the understanding of subarachnoid hemorrhage (SAH). Under proinflammatory conditions following SAH, interleukin (IL) 6 is considered to have an effect on cerebrovascular leakage and extracellular matrix composition. Our present work focuses on investigating causes and extent of BBB disruption as well as the influence of IL6 on BBB integrity post SAH in mice.

Methods: SAH was induced in wildtype (WT) and IL6 knock out (IL6KO) mice by filament perforation and ruled out by H&E staining and MRI. SAH- and SHAM-operated mice (controls) were decapitated on day 4 after onset. BBB breakdown was assessed by extravasation of fluorescent Evans Blue (EB, 70 kDa) and Fluorescein Isothiocyanate- (FITC)-Dextran (4 kDa). Endothelial expression of tight junction (TJ) molecules and cytokines IL6, tumor-necrosis factor α (TNFα), IL1β were examined in isolated brain capillaries employing qPCR. Apoptotic desmin- and TUNEL-double-positive pericytes were analyzed by immunofluorescence.

Results: A significant IL6 overexpression (2.7±0.5-fold) was detected in WTs on day 4 after SAH. A concomitant BBB breakdown in terms of EB (70 kDa) extravasation was observed in SAH-WTs, which was not present in IL6KOs. FITC-Dextran (4 kDa) extravasation was measured either in WTs and IL6KOs after SAH. A significant reconstitution of occludin (1.3±0.3-fold of SHAM) but not claudin-5 (0.5±0.3-fold) was detected in SAH-IL6KOs compared to WTs. Our analysis revealed TNFα and IL1β to be elevated in WTs post SAH (3.3±1.2-fold and 2.8±1.2-fold of SHAM, respectively), while this overexpression was attenuated in IL6KOs. We found a higher number of apoptotic pericytes in SAH-WTs (3.1±1.0-fold) compared to IL6KOs.

Conclusion: We demonstrated the loss of BBB integrity in WT mice post SAH by means of EB extravasation and lower TJ expression. While extravasation of EB was constricted, FITC-Dextran was not hindered to pass the BBB in IL6KOs seemingly due to lower claudin-5 expression. Pericyte-endothelial interaction is assumed to contribute to BBB breakdown post SAH, too. Showing a lower number of apoptotic pericytes, we conclude that BBB tightness could be obtained better in IL6KOs. We presume that IL6 is critically involved in secondary brain injury post SAH due to its influence on cerebrovascular leakage, its regulation of TJs and its effect on pericytes.