Selective inhibition of brain endothelial Rho-kinase-2 provides optimal protection of an in vitro blood-brain barrier from tissue-type plasminogen activator and plasmin

• Published in: PloS one 2017-05, Vol.12 (5), p.e0177332-e0177332
• Main Authors: Niego, Be'eri, Lee, Natasha, Larsson, Pia, De Silva, T Michael, Au, Amanda E-Ling, McCutcheon, Fiona, Medcalf, Robert L
• Format: Article
• Language: English
• Subjects: Acetylation; Activation; Affinity; Amino acids; Anatomy; Animal models; Animals; Astrocytes; Astrocytes - drug effects; Astrocytes - metabolism; Atorvastatin; Biology and Life Sciences; Biotechnology; Blood coagulation; Blood diseases; Blood flow; Blood-brain barrier; Blood-Brain Barrier - metabolism; Bradykinin; Brain - metabolism; Brain research; Cancer; Cardiology; Cardiovascular diseases; Care and treatment; Cell Line; Cell lines; Cerebral edema; Clinical medicine; Coagulation; Comparative analysis; Diseases; Dosage and administration; Driving ability; Edema; Endothelial Cells - metabolism; Endothelium; Fetuses; Fibrinolysin - metabolism; Fibrinolytic agents; Flow; Gene Expression; Glucose; Health aspects; Hemoglobin; Hemorrhage; Heterocyclic Compounds, 4 or More Rings - pharmacology; Homology; Humans; Hypertension; Immune system; Infiltration; Inhibition; Inhibitors; Intracerebral hemorrhage; Ischemia; Isoforms; Kinases; Light emitting diodes; Medicine; Medicine and Health Sciences; Metabolism; Mice; Microbiology; Monocyte chemoattractant protein 1; Multiplication; Neuronal-glial interactions; Neurons; Noma; Pathology; Patient outcomes; Permeability; Pharmacology; Physical Sciences; Protein Isoforms; Protein kinase; Protein-serine/threonine kinase; Research and Analysis Methods; rho-Associated Kinases - antagonists & inhibitors; rho-Associated Kinases - genetics; rho-Associated Kinases - metabolism; Ribonucleic acid; Risk factors; RNA; Rodents; Signal Transduction; Smooth muscle; Stroke; Thromboembolism; Tissue Plasminogen Activator - metabolism; Tissue Plasminogen Activator - pharmacology; Viruses
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0177332

Rho-kinase (ROCK) inhibition, broadly utilised in cardiovascular disease, may protect the blood-brain barrier (BBB) during thrombolysis from rt-PA-induced damage. While the use of nonselective ROCK inhibitors like fasudil together with rt-PA may be hindered by possible hypotensive side-effects and inadequate capacity to block detrimental rt-PA activity in brain endothelial cells (BECs), selective ROCK-2 inhibition may overcome these limitations. Here, we examined ROCK-2 expression in major brain cells and compared the ability of fasudil and KD025, a selective ROCK-2 inhibitor, to attenuate rt-PA-induced BBB impairment in an in vitro human model. ROCK-2 was highly expressed relative to ROCK-1 in all human and mouse brain cell types and particularly enriched in rodent brain endothelial cells and astrocytes compared to neurons. KD025 was more potent than fasudil in attenuation of rt-PA- and plasminogen-induced BBB permeation under normoxia, but especially under stroke-like conditions. Importantly, only KD025, but not fasudil, was able to block rt-PA-dependent permeability increases, morphology changes and tight junction degradation in isolated BECs. Selective ROCK-2 inhibition further diminished rt-PA-triggered myosin phosphorylation, shape alterations and matrix metalloprotease activation in astrocytes. These findings highlight ROCK-2 as the key isoform driving BBB impairment and brain endothelial damage by rt-PA and the potential of KD025 to optimally protect the BBB during thrombolysis.