Angiopoietin 1 enhances blood-spinal cord barrier permeability via regulating epidermal growth factor receptor pathway substrate 8 expression
10.3760/cma.j.issn.1671-8925.2015.10.006
- VernacularTitle:血管生成素1通过调节Eps8的表达增强大鼠血-脊髓屏障功能
- Author:
Xinchun LIU
1
;
Yunfei PENG
;
Sen WANG
Author Information
1. 中国医科大学附属第一医院骨科
- Keywords:
Agiopoietin 1;
Epidermal growth factor receptor pathway substrate 8;
Blood-spinal cord barrier;
Actin skeleton regulatory protein;
F-actin
- From:
Chinese Journal of Neuromedicine
2015;14(10):1000-1005
- CountryChina
- Language:Chinese
-
Abstract:
Objective To explore the role of epidermal growth factor receptor pathway substrate 8 (EpsS) in angiopoietin 1 (Ang1)-induced blood-spinal cord barrier (BSCB) function enhancement in rats.Methods Spinal cord microvascular endothelial cells (SCMECs) were primarily isolated and cultured fiom adult rats to set up the in vitro BSCB models.(1) Transendothelial electrical resistance (TEER) values were measured and Eps8 protein level was detected by Western blotting at different time points (0, 4, 8, 12 h) after Ang1 treatment.Cell lysates untreated with Ang1 were set as controls.(2) The Eps8 expression in SCMECs was silenced by siRNA interference, followed by Ang1 treatment again;the SCMECs were grouped as control siRNA, control siRNA+Ang1, Eps8 siRNA and Eps8 siRNA+Ang1;Western blotting was applied to detect the Eps8 level, TEER values were recorded, endothelial permeability was detected by using sodium fluorescein (Na-F) and Evans-blue albumin (EBA), and then, the F-actin distribution was observed by phalloidine staining.Results (1) TEER values in vitro and Eps8 expression in 4, 8 and 12 h Ang1 treatment groups were both significantly elevated as compared with those in the controls (P＜0.05).(2) The protein level of Eps8 was knocked down by 75％ in Eps8 siRNA group as compared with that in control siRNA group, with significant difference (P＜0.05);while no significant changes of Eps8 expression in the Eps8 siRNA+Ang1 group was noted as compared with that in the Eps siRNA group.Moreover, the Eps8 siRNA group had significantly decreased TEER values, and significantly elevated permeability to Na-F or EBA at different time points as compared with control siRNA group (P＜0.05);differently, the TEER values and permeability to Na-F or EBA of the Eps8 siRNA+Ang1 group did not significantly change as compared with those in the Eps8 siRNA group.F-actin staining also revealed no change between Eps8 siRNA+Ang1 group and Eps8 siRNA group at cell-cell interface, while F-actin arrangement was found to be intensified significantly in the control siRNA+Ang1 group as compared with those in the control siRNA group.Conclusion Ang1 regulates F-actin distribution at rat BSCB by altering the Eps8 expression, which further modulates the barrier function of BSCB.
