NS1619 Alleviate Brain-Derived Extracellular Vesicle-Induced Brain Injury by Regulating BKca Channel and Nrf2/HO-1/NF-ĸB Pathway

Brain induced extracellular vesicle (BDEV) elevates after traumatic brain injury (TBI) and contributes to secondary brain injury. However, the role of BDEV in TBI remains unclear. In this study, we determined the mechanisms of BDEV in brain injury and explored whether neuroprotective drug BKca channel opener NS1619 may attenuate BDEV-induced brain injury. We injected BDEV and lactadherin, respectively, to mimic the up and downregulation of BDEV after TBI and illustrated the role of BDEV in vivo. In vitro, the membrane potential and calcium concentration of HT-22, bEnd3, and BV-2 were measured by fluorescent staining. The effects of BDEV and NS1619 on HT-22 were evaluated by CCK-8, LDH release assay, Na(+)/k(+)-ATPase activity, JC-1 staining, DHE staining, and 4-HNE staining, respectively. The role of BDEV and NS1619 on the Nrf2/HO-1/p65 pathway was also evaluated in HT-22. Finally, we administrated TBI mice with NS1619 to clarify the role of NS1619 against BDEV in vivo. Our results suggested that BDEV aggravated and lactadherin mitigated TBI-induced EB leakage, brain edema, neuronal degeneration, apoptosis, ROS level, microgliosis, MMP-9 activity, and NF-κB activation. In vitro, BDEV-caused depolarized membrane potential and calcium overload were significantly attenuated by NS1619 in HT-22, bEnd3, and BV-2. BDEV markedly decreased cell viability, Na(+)/k(+)-ATPase activity, and caused mitochondrial dysregulation, oxidative stress, and NF-ĸB activation. NS1619 pretreatment alleviated above process and enhanced antioxidant system Nrf2/HO-1 in HT-22. Finally, NS1619 administration significantly inhibited neuroinflammation response and improved TBI outcome after TBI. NS1619 treatment also reduced 4-HNE content and NF-ĸB activation and enhanced Nrf2/HO-1 pathway. Our data showed that BDEV aggravated brain injury by perturbing cell membrane potential, calcium homeostasis, oxidative stress, and neuroinflammation. The BKca channel opener NS1619 attenuated BDEV-induced pathological process in vitro and in vivo by modulating the BKca channel and Nrf2/HO-1/NF-ĸB pathway.