Background  The sodium-potassium-chloride cotransporter is closely related to the damage of the blood-brain barrier (BBB), but the specific regulatory mechanism remains to be studied.

  Objective  To investigate the regulatory mechanism of sodium potassium chloride cotransporter on BBB permeability by in vitro model.

  Methods  The BBB model was established in Transwell chambers using bEnd3 mouse brain microvascular endothelial cells and BV2 mouse microglia. The BBB function was detected by trans-epithelium electrical resistant (TEER) and fluorescein sodium permeation assay, the content of inflammasome NLRP3 protein was detected by Western blot, the levels of IL-1β and MMP-9 were detected by ELISA, and the integrity of the tight junction protein ZO-1 between vascular endothelial cells was assessed by immunofluorescence.

  Results  The activation of microglia in inflammatory environment could destroy the integrity of the blood-brain barrier model, reduce the TEER value, and increase the permeability of the blood-brain barrier model (P＜0.05). In the co-culture model, the sodium-potassium-chloride symporter-specific inhibitor significantly ameliorated the LPS-induced BBB damage (P＜0.05), while this phenomenon was not observed in the BBB model constructed by endothelial cell monoculture. The specific inhibitor of sodium potassium chloride symporter also reduced the expression level of NLRP3 inflammasome in microglial and inhibited the secretion of IL-1β and MMP-9 in microglial cells (P＜0.05), in the co-culture model, NLRP3 inflammasome-specific inhibitor ameliorated LPS-induced BBB damage, increased TEER value, and decreased barrier permeability (P＜0.05).

  Conclusion  The regulation of the sodium-potassium-chloride cotransporter on BBB permeability may be mediated by NLRP3 inflammasome derived from microglia.