Escin Rescues Blood-brain Barrier and Inhibits NLRP3 Inflammasome-mediated Pyroptosis in Rats with Superior Sagital Sinus Thrombosis.

Objective: The purpose of this study is to investigate the therapeutic value of escin and its potential mechanisms in rats with superior sagittal sinus thrombosis (SSST). Methods: The motor function of rats was assessed using the open field, balance beam, and rotarod tests following one week of escin treatment. Laser Speckle Contrast Imaging (LSCI) was used to evaluate the blood flow in the superior sagittal sinus. Evans blue staining was used to assess the permeability of blood-brain barrier. The protein expression levels of Occludin, ZO-1 and MMP-9 was analyzed by Western blotting to assess blood-brain barrier disruption. Iba1, CD68 and NLRP3 immunofluorescent staining was conducted to evaluate the activation of microglia and NLRP3 inflammasome. The protein expression levels of NLRP3, Caspase-1, IL-1β, IL-18 and GSDMD was analyzed by Western blotting to assess NLRP3 inflammasome activation and pyroptosis. Immunohistochemistry for NeuN was used to evaluate the neuronal pyroptosis. Results: Rats with SSST exhibited decreased movement distance, duration and latency to fall, as well as increased balance latency compared to sham rats, indicating its motor dysfunction. Escin treatment alleviated the motor dysfunction in rats with SSST. LSCI revealed a decrease in the blood flow in the superior sagittal sinus of rats with SSST. There was no discernible change in blood flow between SSST rats treated with escin and those not, suggesting that escin did not promote recanalization of the superior sagittal sinus. By decreasing Evans blue dye content and MMP-9 protein expression and increasing the protein expressions of Occludin and ZO-1, escin treatment improved blood-brain barrier (BBB) disruption and protected tight junction proteins. Escin also reduced the number of microglia-originated macrophage and the number of NLRP3-positive macrophage. Additionally, escin treatment down-regulated the protein expression levels of NLRP3, Caspase-1, IL-1β, IL-18, and GSDMD in the parasagittal cortex of SSST rats, suggesting its ability to inhibit NLRP3 inflammasome activation and therefore reduce pyroptosis. Finally, escin treatment significantly increased the neuronal survival in the parasagittal cortex of rats with SSST. Conclusions: Treatment with escin improved motor function not by recanalizing the SSS. Treatment with escin protected the blood-brain barrier, inhibited the microglia activation and suppressed the NLRP3 inflammasome-mediated pyroptosis in the parasagittal cortex of SSST rats, thereby playing an anti-pyroptosis and neuroprotective effect.