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Ozanimod Preserves Endothelial Health by Attenuating Cerebrovascular MMP‐9 Activity Following Acute Ischemic Injury
Author(s) -
Wendt Trevor,
Arkelius Kajsa,
Ansar Saema,
Gonzales Rayna
Publication year - 2021
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2021.35.s1.03686
Subject(s) - medicine , zymography , stroke (engine) , ischemia , pathology , anesthesia , matrix metalloproteinase , mechanical engineering , engineering
The loss of vascular integrity at the level of the blood brain barrier leads to a vicious cascade of secondary injuries following acute ischemic stroke (AIS). Elevated MMP‐9 activity within the cerebral vasculature has been implicated with severe degradation of the vascular basement membrane leading to abnormal cerebrovascular permeability and detrimental stroke outcome. Sphingosine‐1‐phosphate receptor (S1PR) modulation improves stroke outcome in AIS patients, however the influence of selective S1PR 1 ligands, such as ozanimod, on brain endothelial health and MMP‐9 activity following AIS has not been investigated. Thus, the aim of this study was to determine the impact of acute ischemic injury on MMP‐9 activity in both the rat and the human cerebrovasculature as well as the vascular specific role of ozanimod on human endothelial health and MMP‐9 activity. Using an in vivo thromboembolic stroke model, cerebral vessels were isolated from male Wistar rats that underwent a right middle cerebral artery (MCA) thrombin injection. Sham‐operated animals underwent the same surgical procedure; however, nothing was injected. Vascular MMP‐9 enzymatic activity of was evaluated at 3, 6 and 24h post injury using zymography. Using an in vitro ischemic injury model (HGD; hypoxia plus glucose deprivation), male human brain microvascular endothelial cells (HBMECs; P7) were treated with ozanimod (0.5nM) or vehicle (DMSO) and then exposed to normoxia (21% O 2 ) or HGD (1% O 2 ). In some experiments, W146 (a selective antagonist), verified S1PR 1 dependence. Morphology and vacuole formations were assessed using crystal violet staining. Enzymatic activity of MMP‐9 was evaluated via zymography and the extracellular H 2 O 2 concentration, inducer of MMP‐9 activity, was measured using a colorimetric assay. Following in vivo thromboembolic occlusion, ipsilateral MMP‐9 activity increased at 6h post injury and returned to baseline when compared to sham. In comparison to the contralateral side, thromboembolic occlusion induced a time dependent modulation of ipsilateral MMP‐9 activity compared to the contralateral side with the highest activity peaking at 6h. In our in vitro HBMEC stroke model, MMP‐9 enzymatic activity was increased following 3h HGD exposure, and this response was attenuated by ozanimod. Concomitantly, during HGD, H 2 O 2 production, a partial inducer of MMP‐9 activity, was increased in a time dependent manner when compared to normoxic controls. Furthermore, HGD induced an increase in HBMEC vacuole formations and a decrease in cell viability at 3h. Loss of cell viability was rescued with ozanimod. In conclusion, increased MMP‐9 activity, in part due to H 2 O 2 , is an acute response to ischemic injury in the cerebrovasculature. Specifically, ozanimod's ability to attenuate endothelial MMP‐9 activity may play an important beneficial role in mitigating blood brain barrier integrity loss following an acute ischemic injury.