Attenuation of urokinase activity during experimental ischaemia protects the cerebral barrier from damage through regulation of matrix metalloproteinase-2 and NAD(P)H oxidase

Rakkar, Kamini; Srivastava, Kirtiman; Bayraktutan, Ulvi

Abstract

Ischaemic injury impairs the integrity of the blood–brain barrier (BBB). In this study, we investigated the molecular causes of this defect with regard to the putative correlations among NAD(P)H oxidase, plasminogen–plasmin system components, and matrix metalloproteinases. Hence, the activities of NAD(P)H oxidase, matrix metalloproteinase-2, urokinase-type plasminogen activator (uPA), and tissue-type plasminogen activator (tPA), and superoxide anion levels, were assessed in human brain microvascular endothelial cells (HBMECs) exposed to oxygen–glucose deprivation (OGD) alone or OGD followed by reperfusion (OGD + R). The integrity of an in vitro model of BBB comprising HBMECs and astrocytes was studied by measuring transendothelial electrical resistance and the paracellular flux of albumin. OGD with or without reperfusion (OGD ± R) radically perturbed barrier function while concurrently enhancing uPA, tPA and NAD(P)H oxidase activities and superoxide anion release in HBMECs. Pharmacological inactivation of NAD(P)H oxidase attenuated OGD ± R-mediated BBB damage through modulation of matrix metalloproteinase-2 and tPA, but not uPA activity. Overactivation of NAD(P)H oxidase in HBMECs via cDNA electroporation of its p22-phox subunit confirmed the involvement of tPA in oxidase-mediated BBB disruption. Interestingly, blockade of uPA or uPA receptor preserved normal BBB function by neutralizing both NAD(P)H oxidase and matrix metalloproteinase-2 activities. Hence, selective targeting of uPA after ischaemic strokes may protect cerebral barrier integrity and function by concomitantly attenuating basement membrane degradation and oxidative stress.

Citation

Rakkar, K., Srivastava, K., & Bayraktutan, U. (2014). Attenuation of urokinase activity during experimental ischaemia protects the cerebral barrier from damage through regulation of matrix metalloproteinase-2 and NAD(P)H oxidase. European Journal of Neuroscience, 39(12), https://doi.org/10.1111/ejn.12552

• Journal Article Type: Article
• Acceptance Date: Feb 8, 2014
• Online Publication Date: Mar 20, 2014
• Publication Date: Jun 15, 2014
• Deposit Date: Oct 27, 2016
• Publicly Available Date: Oct 27, 2016
• Journal: European Journal of Neuroscience
• Print ISSN: 0953-816X
• Electronic ISSN: 1460-9568
• Publisher: Wiley
• Peer Reviewed: Peer Reviewed
• Volume: 39
• Issue: 12
• DOI: https://doi.org/10.1111/ejn.12552
• Keywords: Reperfusion injury; Superoxide anion; Blood-brain barrier; p22-phox ; Matrix metalloproteinase
• Public URL: https://nottingham-repository.worktribe.com/output/730691
• Publisher URL: http://dx.doi.org/10.1111/ejn.12552
• Additional Information: This is the peer reviewed version of the following article: Rakkar, K., Srivastava, K. and Bayraktutan, U. (2014), Attenuation of urokinase activity during experimental ischaemia protects the cerebral barrier from damage through regulation of matrix metalloproteinase-2 and NAD(P)H oxidase. Eur J Neurosci, 39: 2119–2128. doi:10.1111/ejn.12552, which has been published in final form at http://dx.doi.org/10.1111/ejn.12552. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Self-Archiving.
• Contract Date: Oct 27, 2016