gms | German Medical Science

Objective: Interleukin 6 (IL6) is implicated in the compromised blood-brain barrier (BBB) by dysregulation of cellular junctions and in inflammatory processes, i.e. due to secondary brain injury after subarachnoid hemorrhage (SAH). Although IL6 has been shown to exert pro-inflammatory action on brain microvascular endothelial cells (ECs), the expression of one of the IL6 receptors (IL6R) in these cells is controversially discussed.

Methods: SAH was induced in wildtype mice by filament perforation. Gene expression of IL6 and its receptors, IL6R and glycoprotein 130 (gp130) was tested in mRNA isolated from cerebral capillaries employing qPCR. Vascular expression of IL6R was evaluated in coronal brain slices by immunohistochemistry. Moreover, an in vitro model of the BBB, the cEND cell line administered with recombinant IL6 protein (50 ng/mL) and subjected with or without a blocking antibody directed to the IL6R (0.2 µg/mL) was used to analyze the involvement of the receptor on cerebrovascular barrier properties by ELISA, measurements of the transendothelial electrical resistance (TEER), qPCR, Western Blot and functional cell culture assays.

Results: We are the first to demonstrate a basal expression of the IL6R in BBB capillaries being markedly increased 2 days after experimental SAH (2.9±0.8-fold of SHAM). SAH caused a significant overexpression of IL6 transcript (2.2±0.1-fold of SHAM) without influencing the gp130. A robust upregulation of the IL6R was confirmed in coronal brain slices post SAH and in cEND cells following recombinant IL6 administration for 48 hours (2.1±0.1-fold of the untreated control). IL6 treatment over 48 hours resulted in overexpressed IL6 protein, an autocrine IL6 secretion into the extracellular compartment and a concomitantly lowered TEER (5.8±2.7-fold, 3.5±0.7-fold and 0.6±0.05-fold of the control, respectively) due to reduced cellular junction proteins: claudin-5, occludin and vascular-endothelial (VE-) cadherin. Functional assays revealed that the IL6-induced loss of cEND maintenance could be restored almost to the control level by neutralizing the IL6R. Briefly, treatment with the IL6R blocking antibody improved claudin-5, occludin and VE-cadherin expression up to 1.05±0.1, 0.9±0.1 and 0.9±0.2-fold of the control.

Conclusion: Our findings add depth to the current understanding of the involvement of IL6R in the IL6-induced loss of EC integrity implicating novel therapy options for BBB inflammation occurring due to secondary brain injury after SAH.