Healthy Serum-Derived Exosomes Improve Neurological Outcomes and Protect Blood-Brain Barrier by Inhibiting Endothelial Cell Apoptosis and Reversing Autophagy-Mediated Tight Junction Protein Reduction in Rat Stroke Model

Lin-Yan Huang; Jin-Xiu Song; Heng Cai; Pei-Pei Wang; Qi-Long Yin; Yi-De Zhang; Jie Chen; Ming Li; Jia-Jia Song; Yan-Ling Wang; Lan Luo; Wan Wang; Su-Hua Qi

doi:10.3389/fncel.2022.841544

Healthy Serum-Derived Exosomes Improve Neurological Outcomes and Protect Blood-Brain Barrier by Inhibiting Endothelial Cell Apoptosis and Reversing Autophagy-Mediated Tight Junction Protein Reduction in Rat Stroke Model

Front Cell Neurosci. 2022 Mar 3:16:841544. doi: 10.3389/fncel.2022.841544. eCollection 2022.

Authors

Lin-Yan Huang¹, Jin-Xiu Song², Heng Cai², Pei-Pei Wang², Qi-Long Yin², Yi-De Zhang¹, Jie Chen¹, Ming Li¹, Jia-Jia Song¹, Yan-Ling Wang¹, Lan Luo¹, Wan Wang¹, Su-Hua Qi^{1

2}

Affiliations

¹ Xuzhou Key Laboratory of Laboratory Diagnostics, School of Medical Technology, Xuzhou Medical University, Xuzhou, China.
² Pharmacology College, Xuzhou Medical University, Xuzhou, China.

Abstract

Blood-brain barrier (BBB) dysfunction causing edema and hemorrhagic transformation is one of the pathophysiological characteristics of stroke. Protection of BBB integrity has shown great potential in improving stroke outcome. Here, we assessed the efficacy of exosomes extracted from healthy rat serum in protection against ischemic stroke in vivo and in vitro. Exosomes were isolated by gradient centrifugation and ultracentrifugation and exosomes were characterized by transmission electron microscopy (TEM) and nanoparticle tracking video microscope. Exosomes were applied to middle cerebral artery occlusion (MCAO) rats or brain microvascular endothelial cell line (bEnd.3) subjected to oxygen-glucose deprivation (OGD) injury. Serum-derived exosomes were injected intravenously into adult male rats 2 h after transient MCAO. Infarct volume and gross cognitive function were assessed 24 h after reperfusion. Poststroke rats treated with serum-derived exosomes exhibited significantly reduced infarct volumes and enhanced neurological function. Apoptosis was assessed via terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick-end labeling (TUNEL) staining and the expression of B-cell lymphoma-2 (Bcl-2), Bax, and cleaved caspase-3 24 h after injury. Our data showed that serum exosomes treatment strikingly decreased TUNEL⁺ cells in the striatum, enhanced the ratio of Bcl-2 to Bax, and inhibited cleaved caspase-3 production in MCAO rats and OGD/reoxygenation insulted bEnd.3 cells. Under the consistent treatment, the expression of microtubule-associated protein 1 light chain 3B-II (LC3B-II), LC3B-I, and Sequestosome-1 (SQSTM1)/p62 was detected by Western blotting. Autolysosomes were observed via TEM. We found that serum exosomes reversed the ratio of LC3B-II to LC3B-I, prevented SQSTM1/p62 degradation, autolysosome formation, and autophagic flux. Together, these results indicated that exosomes isolated from healthy serum provided neuroprotection against experimental stroke partially via inhibition of endothelial cell apoptosis and autophagy-mediated BBB breakdown. Intravenous serum-derived exosome treatment may, therefore, provide a novel clinical therapeutic strategy for ischemic stroke.

Keywords: BBB; apoptosis; autophagy; serum exosomes; stroke.