The Role Of Tight Junction In Emp-induced Blood Brain Barrier Opening And Its Mechanism

Electromagnetic Pulse (EMP) is a short high-voltage pulse with an extremely fast rising time and a broad bandwidth. This kind of signal can be generated by nuclear bomb explosion. EMP signals also exist in certain occupational conditions, for example, Pulse Power Technology Lab, in which the strong electrical field apparatus such as high pressure gas switch and Tesla transformer generator can generate EMP. The unusual properties of EMP have raised concerns about their biological effects and possible health hazard to humans, especially to some workers or researchers who work with or can be exposed to this kind of electromagnetic field in their working environment. In our previous study we found that brain is one of the sensitive target of EMP, under certain conditions, EMP exposure could increase the permeability of rat Blood-Brain Barrier (BBB). However, the mechanism is unclear.Since Tight Junction (TJ) between brain endothelial cells plays an important role in maintaining the integrity of the BBB. In this study, we investigated the role of TJ in EMP-induced BBB opening and its mechanismAdult male SD rats were sham or whole-body exposed to EMP at 200 kV/m for 200 or 400 pulses. The permeability of BBB in rat cerebral cortex was examined by using Evans Blue (EB) and lanthanum nitrate as vascular tracers. The localization and expression of TJ proteins (ZO-1, occludin) and cytoskeleton protein (actin) were assessed by Western Blotting and immunofluorescence analysis, respectively. It was found that comparing to sham group the BBB permeability began to increase at 0.5 h after EMP exposure, and reached the peak at 3h after EMP exposure, then began to recover at 6 h, and finally recovered to sham levels at 12 h after EMP exposure. Transmission electronic microscopic (TEM) examination showed that the predominant route of lanthanum extravasation was by the interendothelial pathway. In addition, we found that the expression level of ZO-1 in both cerebral cortex homogenate and cerebral cortex microvessel homogenate was significantly decreased at 1 h and 3 h after EMP exposure. The immunofluorescence assay showed that the staining intensity for ZO-1 appeared to be reduced after EMP exposure, and this is consistent with Western Blotting data. In addition, the alterations in ZO-1 protein localization occurred in cerebral cortex microvessel after EMP exposure. In sham animals, ZO-1 showed a predominant pattern of continuous staining along the margins of cell–cell contact. However, at 3 h after EMP exposure, ZO-1 lost its distribution in continuity and well defined filaments. Immunofluorescence and Western-Blotting analysis documented no significant alterations in the immunoreactivity or the expression of occludin and actin in cerebral cortex of rats exposed to EMP. Therefore, the data obtained from our investigation indicated changes in both ZO-1 expression and the cellular localization were associated with increased BBB permeability after EMP exposure.It was reported that PKC was associated with BBB permeability as well as BBB TJ proteins translocation under pathological conditions. To determine the role of protein kinase C signaling in EMP-induced BBB permeability change in rats. The protein level of total PKC and two PKC isoforms (PKC- , and PKC-βⅡ) were determined in brain cerebral cortex microvessels by Western Blotting after exposing rats to EMP at 200 kV/m for 200 pulses with 1 Hz repetition rate. It was found that the protein level of PKC and PKC-βⅡ(but not PKC- ) in cerebral cortex microvessels increased significantly at 0.5 h and 1 h after EMP exposure compared with sham-exposed animals and then recovered at 3 h.A specific PKC antagonist (H7) almost blocked EMP-induced BBB permeability change. EMP-induced BBB tight junction protein ZO-1 translocation was also inhibited. Our data indicated that PKC signaling was involved in EMP-induced BBB permeability change and ZO-1 translocation in rat. In general, these results indicated that: TJ injury played an important role in EMP-induced BBB opening; changes in both ZO-1 expression and the cellular localization were associated with increased BBB permeability after EMP exposure; PKC signaling was involved in EMP-induced BBB permeability change and ZO-1 translocation in rat.