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Autophagy-mediated occludin degradation contributes to blood-brain barrier disruption during ischemia in bEnd.3 brain endothelial cells and rat ischemic stroke models.

ABSTRACT: BACKGROUND:The blood-brain barrier (BBB) maintains homeostasis of the brain environment by tightly regulating the entry of substances from systemic circulation. A breach in the BBB results in increased permeability to potentially toxic substances and is an important contributor to amplification of ischemic brain damage. The precise molecular pathways that result in impairment of BBB integrity remain to be elucidated. Autophagy is a degradation pathway that clears damaged or unnecessary proteins from cells. However, excessive autophagy can lead to cellular dysfunction and death under pathological conditions. METHODS:In this study, we investigated whether autophagy is involved in BBB disruption in ischemia, using in vitro cells and in vivo rat models. We used brain endothelial bEnd.3 cells and oxygen glucose deprivation (OGD) to simulate ischemia in culture, along with a rat ischemic stroke model to evaluate the role of autophagy in BBB disruption during cerebral ischemia. RESULTS:OGD 18 h induced cellular dysfunction, and increased permeability with degradation of occludin and activation of autophagy pathways in brain endothelial cells. Immunostaining revealed that occludin degradation is co-localized with ischemic autophagosomes. OGD-induced occludin degradation and permeability changes were significantly decreased by inhibition of autophagy using 3-methyladenine (3-MA). Enhanced autophagic activity and loss of occludin were also observed in brain capillaries isolated from rats with middle cerebral artery occlusion (MCAO). Intravenous administration of 3-MA inhibited these molecular changes in brain capillaries, and recovered the increased permeability as determined using Evans blue. CONCLUSIONS:Our findings provide evidence that autophagy plays an important role in ischemia-induced occludin degradation and loss of BBB integrity.

SUBMITTER: Kim KA

PROVIDER: S-EPMC7071658 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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Autophagy-mediated occludin degradation contributes to blood-brain barrier disruption during ischemia in bEnd.3 brain endothelial cells and rat ischemic stroke models.

Kim Kyeong-A KA Kim Donghyun D Kim Jeong-Hyeon JH Shin Young-Jun YJ Kim Eun-Sun ES Akram Muhammad M Kim Eun-Hye EH Majid Arshad A Baek Seung-Hoon SH Bae Ok-Nam ON

Fluids and barriers of the CNS 20200314 1

<h4>Background</h4>The blood-brain barrier (BBB) maintains homeostasis of the brain environment by tightly regulating the entry of substances from systemic circulation. A breach in the BBB results in increased permeability to potentially toxic substances and is an important contributor to amplification of ischemic brain damage. The precise molecular pathways that result in impairment of BBB integrity remain to be elucidated. Autophagy is a degradation pathway that clears damaged or unnecessary p ...[more]

PMID: 32169114

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Project description:Perioperative neurocognitive disorders (PND) occur frequently after surgery, especially in aged patients. Surgery-induced neuroinflammation and blood-brain barrier (BBB) dysfunction play a crucial role in the pathogenesis of PND. Interleukin-17A (IL-17A) increases after surgical stress and will be involved in BBB dysfunction. However, the effect of IL-17A on BBB function during PND remains poorly understood. Male wild-type C57BL/6J mice (15?months old) received tibial fracture surgery and fixation to establish the PND model. All the mice were injected intraperitoneally with an IL-17A-neutralizing antibody (Abs) or isotype-control Abs 30?min before tibial fracture surgery. Animal behaviour tests conducted 24?h after surgery included the contextual fear conditioning and Y maze tests. Serum and hippocampus IL-17A levels and hippocampus IL-6 and IL-1? levels were detected by ELISA. BBB function was detected by Evans blue (EB) test. Hippocampus matrix metalloproteinase-2 (MMP-2)- and MMP-9-positive cells were detected by immunohistochemistry. Hippocampus albumin, occludin, claudin-5 and IL-17A receptors were detected by Western blot. For the in vitro experiment, bEnd.3 cells were incubated with IL-17A. Cell IL-17A receptors were detected by immunofluorescence. Cellular MMP-2, MMP-9, occludin, and claudin-5 were detected by Western blot. Tibial fracture surgery promoted memory impairment, increased levels of IL-17A and IL-17A receptors, inflammatory factor production and BBB dysfunction. IL-17A Abs inhibited this effect, including improving memory function, decreasing inflammatory factor production and alleviating BBB disruption, indicated by decreased tight junctions (TJs) and increased MMPs after surgery. The in vitro study suggested that recombinant IL-17A could upregulate the expression of IL-17A receptors, decrease TJs and increase the level of MMPs in bEnd.3 cells. Our results suggested that IL-17A-promoted BBB disruption might play an important role in the pathogenesis of PND.

Dataset Information

Autophagy-mediated occludin degradation contributes to blood-brain barrier disruption during ischemia in bEnd.3 brain endothelial cells and rat ischemic stroke models.

Publications

Autophagy-mediated occludin degradation contributes to blood-brain barrier disruption during ischemia in bEnd.3 brain endothelial cells and rat ischemic stroke models.

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