Shedding light on the blood-brain barrier in ischemic stroke

Blood-brain barrier (BBB) breakdown is a hallmark of ischemic stroke and has been associated with a poor clinical prognosis for ischemic stroke patients. Tissue- plasminogen activator (tPA), the only approved drug for treatment of ischemic stroke, can cause intracerebral hemorrhage, a severe form of BBB breakdown, and it has been suggested that this is caused by tPA-mediated activation of platelet- derived growth factor (PDGF)-CC/PDGF receptor alpha (PDGFRα) signaling in the neurovascular unit.

In this thesis, an in vivo two-photon imaging pipeline was established which enables studies on ischemic stroke progression in real time. With this technique, BBB breakdown can be assessed and quantified longitudinally in the same animal. We could demonstrate that during the first 7 days after ischemic stroke induction, BBB leakage displays distinct peaks during the acute and the subacute phase, respectively, which could be effectively reduced with the tyrosine kinase inhibitor imatinib, an inhibitor of PDGFRα signaling.

We could further show that phosphorylation of the tight junction protein occludin causes a loss of tight junction integrity in endothelial cells which leads to an increase in BBB permeability. The phosphorylation of occludin was mediated by tPA-induced activation of the PDGF-CC/PDGFRα signaling pathway in perivascular PDGFRα+ cells and inhibiting phosphorylation of occludin reduced BBB permeability.

In addition, our studies established the role of PDGF-CC/PDGFRα signaling in myofibroblast transdifferentiation in the fibrotic scar after experimental ischemic stroke. We showed that inhibiting PDGF-CC/PDGFRα signaling, by pharmacologic or genetic means, preserved cerebrovascular health, reduced the reactive gliosis response and decreased myofibroblast expansion within the fibrotic scar in the early chronic phase after ischemic stroke. Importantly we demonstrate that targeting myofibroblast expansion in the post-acute phase after ischemic stroke onset was associated with improved functional outcome, thus suggesting a novel post-acute treatment opportunity for ischemic stroke patients.

Collectively, this thesis provides further insight into how tPA-mediated PDGF- CC/PDGFRα signaling regulates BBB breakdown and establishes the involvement of PDGF-CC/PDGFRα signaling in myofibroblast scar formation after ischemic stroke.

List of scientific papers

I. Jil Protzmann, Felix Jung, Lars Jakobsson and Linda Fredriksson. Analysis of ischemic stroke-mediated effects on blood-brain barrier properties along the arteriovenous axis assessed by intravital two- photon imaging. Fluids and Barriers of the CNS, 2024, 21, 35 https://doi.org/10.1186/s12987-024-00537-5

II. Jil Protzmann, Manuel Zeitelhofer, Christina Stefanitsch, Daniel Torrente, Milena Z. Adzemovic, Kirils Matjunins, Stella J.I. Randel, Sebastian A. Lewandowski, Lars Muhl, Ulf Eriksson, Ingrid Nilsson, Enming J. Su, Daniel A. Lawrence and Linda Fredriksson. Reduced myofibroblast expansion in the fibrotic scar enhances recovery after ischemic stroke. [Manuscript]

III. Andreia Goncalves, Enming J. Su, Arivalagan Muthusamy, Manuel Zeitelhofer, Daniel Torrente, Ingrid Nilsson, Jil Protzmann, Linda Fredriksson, Ulf Eriksson, David A. Antonetti and Daniel A. Lawrence. Thrombolytic tPA-induced hemorrhagic transformation of ischemic stroke is mediated by PKCß phosphorylation of occludin. Blood, 2022, 140(4):388-400 https://doi.org/10.1182/blood.2021014958