Targeting gliovascular connexins prevents inflammatory blood-brain barrier leakage and astrogliosis

Now we just need our stroke leadership to get human testing going on this. But since we have NO LEADERSHIP, nothing will occur.

Targeting gliovascular connexins prevents inflammatory blood-brain barrier leakage and astrogliosis

Marijke De Bock1§, Maarten De Smet1§, Stijn Verwaerde1, Hanane Tahiri1, Steffi Schumacher1,5
Valérie Van Haver1, Katja Witschas1, Christian Steinhäuser2, Nathalie Rouach3, Roosmarijn E.6
Vandenbroucke4 and Luc Leybaert17
§ are co-first authors8
9
1Physiology group, Department of Basic and Applied Medical Sciences, Ghent University, Ghent,10
Belgium11
2Institute of Cellular Neurosciences, University of Bonn, Medical School, Bonn, Germany12
3Center for Interdisciplinary Research in Biology, College de France, CNRS, Inserm, Université13
PSL, Paris, France14
4Department of Biomedical Molecular Biology, Ghent University and Inflammation Research15
Center, VIB, Ghent, Belgium16
17
The authors have declared that no conflict of interest exists.18
19
Address correspondence to: Luc Leybaert, Department of Basic and Applied Medical Sciences -20
Physiology group, Faculty of Medicine and Health Sciences, Ghent University, C. Heymanslaan21
10, Block B (entrance 36) - 3rd floor, 9000 Ghent, Belgium; Tel: +32.9.332.33.66, E-mail:22
luc.Leybaert@UGent.be. ORCID ID: 0000-0001-6452-6982

 Abstract

The blood-brain barrier is formed by capillary endothelial cells expressing Cx37, Cx40 and Cx43,
and is joined by closely apposed astrocytes expressing Cx43 and Cx30. We investigated whether
connexin-targeting peptides could limit barrier leakage triggered by LPS-induced systemic
inflammation in mice. Intraperitoneal LPS increased endothelial and astrocytic Cx43 expression,
elevated TNFα, IL1β, IFNγ and IL6 in plasma and IL6 in the brain, and induced barrier leakage
recorded over 24h. Barrier leakage was largely prevented by global Cx43 knockdown and
Cx43/Cx30 double-knockout in astrocytes, slightly diminished by endothelial Cx43 knockout and
not protected by global Cx30 knockout. Intravenous administration of Gap27 or Tat-Gap19 just
before LPS also prevented barrier leakage, and intravenous BAPTA-AM to chelate intracellular
calcium was equally effective. Patch-clamp experiments demonstrated LPS-induced Cx42
hemichannel opening in endothelial cells, which was suppressed by Gap27, Gap19 and BAPTA.
LPS additionally triggered astrogliosis that was prevented by intravenous Tat-Gap19 or BAPTA-
AM. Cortically applied Tat-Gap19 or BAPTA-AM to primarily target astrocytes, also strongly
diminished barrier leakage. In vivo dye uptake and in vitro patch-clamp showed Cx43
hemichannel opening in astrocytes that was induced by IL6 in a calcium-dependent manner. We
conclude that targeting endothelial and astrocytic connexins is a powerful approach to limit
barrier failure and astrogliosis.