Effect and mechanism of sodium ferulate on NLRP3 inflammasomes in spinal cord injury

English Keywords:Spinal cord injury  Sodium ferulate  NLRP3 inflammasomes  Autophagy  Lysosomal

English Abstract:

  【Abstract】 Objectives: To investigate the effect and mechanism of sodium ferulate(SF) on NLRP3 inflammasomes in the rat model of spinal cord injury(SCI). Methods: In vivo studies, the model of SCI was established by clip compression as previously described, and female SD rats were randomly divided into 3 groups (n=12), spinal cords of the Sham group did not suffer injury, the SCI+SF group was treated with SF by intraperitoneal injection(100mg/kg/d) for 7 days in a row, and the SCI group was given the same amount of saline. The expressions of NLRP3, and its effector protein pro-caspase-1, P20, IL-1, autophagy protein P62, LC3Ⅱ were detected by Western blot. Western blot on the 3rd day after surgery, and immunofluorescence staining for NLRP3 was also performed. Nissl staining was used to measure the survival of neurons on the 7th day after surgery. In vitro experiments, rat microglia cells(normal culture was used as control group) were treated with LPS(LPS group) to induce NLRP3 expression. To verify the effect of autophagy and lysosomal function on NLRP3, activation of NLRP3 and lysosomal Cathepsin B(CTSB) were detected after following addition of autophagy blocker Chloroquine(CQ)(LPS+CQ group). Finally, the molecular changes induced by SF(LPS+CQ+SF group) intervention were analyzed to explore the mechanism. Results: In vivo studies, compared with the Sham group, the NLRP3 inflammasomes in the SCI group were increased and activated, leading to the degradation of pro-caspase-1 and the increased production of P20 and IL-1β. Moreover, autophagy markers of p62 and LC3Ⅱ were both at higher levels(P＜0.05). Compared with the SCI group, the expression and activation of NLRP3 inflammasomes in the SCI+SF group were decreased, and so were the markers of p62 and LC3Ⅱ(P＜0.05). Nissl staining showed that the survival of neurons in anterior horn of spinal cord was improved(P＜0.05). In vitro experiments, the result showed that the NLRP3 inflammasomes of microglia in the LPS group were induced to generated compared with the control group(P＜0.05), but their activation were not significant. However, the level of NLRP3 in the LPS+CQ group was much higher, accompanied by the significant degeneration of pro-caspase-1 and the increased production of P20 and IL-1β. Moreover, the autophagy markers of P62 and LC3Ⅱ were abnormally elevated, due to their inhibited degradation. Also the maturation of the lysosomestic Cathepsin B(CTSB) was decreased(P＜0.05). However, compared with the LPS+CQ group, the levels of P62 and LC3Ⅱ in the LPS+CQ+SF group were declined synchronously and the maturation of CTSB was improved(P＜0.05), autophagy flux showed a tendency to recover. Conclusions: Sodium ferulate reduces the generation and activation of NLRP3 inflammasomes by protecting the function of autophagy and lysosome. Thus, the neuronal damage of SCI rats is alleviated at last.