前列消汤干预NLRP3炎性小体调控慢性非细菌性前列腺炎Caspase-1/IL-1β/NF-κB p65轴机制研究

Basic Study on the Intervention of Qianliexiao Decoction in the Regulation of the Inflammatory Axis of Caspase-1/IL-1β/NF-κB p65in NLRP3Inflammasome in Chronic nonbacterial prostatitis Model mice

目的探讨前列消汤通过干预NLRP3炎性小体调控慢性非细菌性前列腺炎Caspase-1/IL-1β/NF-κB p65轴机制研究。方法将120只雄性SPF级C57BL/6小鼠按随机数表法分为空白组、模型组、阳性对照组(MCC950)、前列消汤高/中/低剂量组;除空白组外,其它组采用复合因素造模法制备EAP小鼠模型,以湿热证候积分、前列腺湿重指数、HE染色评估模型;免疫荧光检测各组小鼠前列腺组织NLRP3、Caspase-1、IL-1β表达水平;Western Blot检测各组小鼠前列腺组织NLRP3、ASC、Caspase-1、GSDMD、NF-κB p65、NF-κB p65磷酸化蛋白表达水平。结果模型组前列腺湿热证候积分(P<0.01)、前列腺湿重指数较空白组升高(P<0.05);与空白组比较,模型组腺体及间质见大量炎症细胞浸润、腺腔结构紊乱、纤维化增生,各药物组能不同程度改善模型小鼠前列腺的炎性浸润程度,以MCC950组及前列消中、高剂量组最为显著(P<0.01);免疫荧光显示:与空白组比较,模型组小鼠前列腺NLRP3、Caspase-1、IL-1β表达明显增强(P<0.01),与模型组比较,MCC950组、前列消汤高/中/低剂量组均能不同程度减弱模型小鼠前列腺NLRP3、Caspase-1、IL-1β(P<0.05),MCC950组与前列消中/高剂量组比较,差异不具有统计学意义;Western Blot显示:与空白组比较,模型组小鼠前列腺NLRP3、ASC、Caspase-1、GSDMD、NF-κB p65、NF-κB p65磷酸化蛋白表达明显升高(P<0.01),与模型组比较,MCC950组能不同程度下调模型小鼠前列腺NLRP3、ASC、Caspase-1、GSDMD、NF-κB p65、NF-κB p65磷酸化蛋白表达(P<0.01),前列消汤各剂量组下调以中/高剂量组较为显著(P<0.01)。结论前列消汤可通过干预NLRP3炎症小体介导的Caspase-1/IL-1β/NF-κB p65炎症轴发挥抑制慢性非细菌性前列腺炎炎症反应的作用。

Objective To investigate the mechanism of Qianliexiao decoction regulating experimental autoimmune prostatitis through Caspase-1/IL-1β/NF-κB p65 pathway by intervening in NLRP3 inflammasomes.Methods The 120 male SPF-grade C57BL/6 mice were divided into blank group,model group,positive control group(MCC950),and high/medium/low dose group of Qianliexiao decoction according to the random number table method.Except for the blank group,the rest were prepared by composite factor molding method to prepare EAP mouse models,and the model was evaluated by dampness-heat syndrome scores,prostate index,HE staining pathology.Immunofluorescence detected the expression levels of NLRP3,Caspase-1 and IL-1βin prostate tissues of each group.Western Blot detected the expression levels of NLRP3,ASC,Caspase-1,GSDMD,NF-κB p65,and NF-κB p65 phosphorylated proteins in each group of mouse tissue.Results The dampness-heat syndrome scores and prostate index of the prostate of the model was higher than that in the blank group(P<0.05).Compared with the blank group,a large number of inflammatory cell infiltration,glandular cavity structure disorder and fibrosis hyperplasia were seen in the glands and interstitium of the model group,and the degree of inflammatory infiltration of the prostate gland in the model group could be improved to different degrees,with MCC950 group and prostatic elimination and high-dose groups being the most significant.Immunofluorescence displayed that compared with the blank group,the expression of prostate NLRP3,Caspase-1 and IL-1βin the model group was significantly enhanced(P<0.01),and compared with the model group,the high/medium/low dose group of MCC950 group and the high/medium/low dose group of the model mouse could weaken the prostate NLRP3,Caspase-1 and IL-1β(P<0.05)to varying degrees;compared with the prostatic neutralization/high dose group,the difference was not statistically significant.Western Blot showed that compared with the blank group,the expression of prostate NLRP3,ASC,Caspase-1,GSDMD,NF-κB p65 and NF-κB p65 phosphorylated proteins in the model group was significantly increased(P<0.01),and compared with the model group,the MCC950 group could downregulate the prostate NLRP3,ASC,Caspase-1,GSDMD,NF-κB p65,NF-κB to varying degrees in the model group The expression of p65 phosphorylated protein(P<0.01),and the dose group of Qianliexiao decoction was lowered to the medium/high dose group(P<0.01).Conclusion Qianliexiao decoction can intervene in caspase-1/IL-1 mediated by NLRP3 inflammasomeβ/NF-κB p65 inflammatory axis plays an intervention role in inhibiting inflammatory response of chronic nonbacterial prostatitis.