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Increases in intracellular calcium perturb blood–brain barrier via protein kinase C-alpha and apoptosis

Rakkar, Kamini; Bayraktutan, Ulvi

Authors



Abstract

An increase in intracellular calcium represents one of the early events during an ischaemic stroke. It triggers many downstream processes which promote the formation of brain oedema, the leading cause of death after an ischaemic stroke. As impairment of blood–brain barrier (BBB) accounts for much of oedema formation, the current study explored the impact of intracellular calcium on barrier integrity in relation to protein kinase C, caspase-3/7, plasminogen activators and the pro-oxidant enzyme NADPH oxidase. Human brain microvascular endothelial cells alone or in co-culture with human astrocytes were subjected to 4 h of oxygen–glucose deprivation alone or followed by 20 h of reperfusion (OGD ± R) in the absence or presence of inhibitors for urokinase plasminogen activator (amiloride), NADPH oxidase (apocynin), intracellular calcium (BAPTA-AM) and protein kinase C-α (RO-32-0432). Endothelial cells with protein kinase C-α knockdown, achieved by siRNA, were also exposed to the above conditions. BBB permeability was measured by transendothelial electrical resistance and Evan's blue-albumin and sodium fluorescein flux. Intracellular calcium and total superoxide anion levels, caspase-3/7, NADPH oxidase, plasminogen activator and protein kinase C activities, stress fibre formation, the rate of apoptosis and BBB permeability were increased by OGD ± R. Treatment with the specific inhibitors or knockdown of protein kinase C-α attenuated them. This study reveals successive increases in intracellular calcium levels and protein kinase C-α activity are key mechanisms in OGD ± R-mediated impairment of BBB. Furthermore inhibition of protein kinase C-α may be therapeutic in restoring BBB function by reducing the rate of cytoskeletal reorganisation, oxidative stress and apoptosis.

Citation

Rakkar, K., & Bayraktutan, U. (2016). Increases in intracellular calcium perturb blood–brain barrier via protein kinase C-alpha and apoptosis. BBA - Molecular Basis of Disease, 1862(1), https://doi.org/10.1016/j.bbadis.2015.10.016

Journal Article Type Article
Acceptance Date Oct 20, 2015
Online Publication Date Oct 23, 2015
Publication Date Jan 31, 2016
Deposit Date Oct 27, 2016
Publicly Available Date Oct 27, 2016
Journal Biochimica et Biophysica Acta - Molecular Basis of Disease
Electronic ISSN 0925-4439
Publisher Elsevier
Peer Reviewed Peer Reviewed
Volume 1862
Issue 1
DOI https://doi.org/10.1016/j.bbadis.2015.10.016
Keywords Apoptosis; Blood-brain barrier; Intracellular calcium; NADPH oxidase; Protein kinase C; Urokinase
Public URL http://eprints.nottingham.ac.uk/id/eprint/37969
Publisher URL http://dx.doi.org/10.1016/j.bbadis.2015.10.016
Copyright Statement Copyright information regarding this work can be found at the following address: http://creativecommons.org/licenses/by-nc-nd/4.0

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Copyright Statement
Copyright information regarding this work can be found at the following address: http://creativecommons.org/licenses/by-nc-nd/4.0





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