基于NLRP3/Caspase-1/GSDMD信号通路探讨芍药苷对雄性大鼠急性辐射诱导的心脏损伤的作用和机制

Exploration of the effects and mechanism of paeoniflorin on the acute radiation-induced heart disease in male rats based on NLRP3/Caspase-1/GSDMD signaling pathway

目的基于NLRP3/Caspase-1/GSDMD信号通路探讨芍药苷(PF)对雄性大鼠急性辐射诱导的心脏损伤的作用和机制。方法将雄性Wistar大鼠随机分为空白对照组(Con组)、单纯照射20Gy组(Mod组)和芍药苷低剂量+20Gy照射组(L组)、芍药苷中剂量+20Gy照射组(M组)、PF高剂量+20Gy照射组(H组)。采用6 MV X线经心前区照射(单次20 Gy)制作心肌损伤模型。PF组自照射前7 d开始至照射后7 d,隔天灌胃(芍药苷低剂量组为25.0 mg/kg、中剂量组为50.0 mg/kg、高剂量组为75.0 mg/kg),Con组和Mod组以同样方式予生理盐水。用ELISA法检测血清中的IL-1β、IL-18表达水平;用HE染色法观察心肌组织的病理损伤情况;用WB法和IHC法检测心肌组织中的NLRP3、caspase-1、GSDMD、IL-1β蛋白表达水平。结果与Con组相比,Mod组血清中的IL-1β、IL-18表达水平明显升高(P<0.05)。与Mod组比,PF组血清中的IL-1β、IL-18表达水平降低;与Con组比,Mod组的心肌细胞广泛大片状坏死、断裂,排列紊乱,心肌间质有大量炎性细胞浸润,横纹肌消失;与Mod组比,随着PF剂量升高,细胞逐渐恢复正常,排列整齐,心肌炎性细胞浸润减轻;与Con组相比,PF组心肌组织中的NLRP3、Caspase-1、GSDMD、IL-1β表达水平明显升高(P<0.05);与Mod组比,随着PF剂量升高,Mod组心肌组织中的NLRP3、Caspase-1、GSDMD、IL-1β表达水平降低。结论芍药苷可减少辐射诱导的心肌损伤引起的心肌炎症及心肌细胞焦亡,起到心肌保护作用。NLRP3炎症小体可作为辐射诱导的心肌损伤潜在治疗靶点。

Objective To explore the effects and mechanism of paeoniflorin(PF)on the acute radiation-induced heart disease(RIHD)in male rats based on the NLRP3/Caspase-1/GSDMD signaling pathway.Methods Male Wistar rats were randomly divided into 5 groups:blank control group(Con group),pure irradiation of 20 Gy group(Mod group),PF low dose+20 Gy irradiation group(L group),PF medium dose+20 Gy irradiation group(M group),PF high dose+20 Gy irradiation group(H group).A myocardial injury model was made by single 20 Gy precordial irradiation with 6 MVX ray.The PF groups were gavaged with PF low-dose(25.0 mg/kg),mediumdose(50.0 mg/kg)and high-dose(75.0 mg/kg)on every other day starting from 7 d before irradiation to 7 d after irradiation,while saline was gavaged in the Con group and the Mod group.ELISA was performed to detect the levels of IL-1βand IL-18 in serum.HE staining was adopted to observe the pathological damage of myocardial tissue.WB and IHC were used to detect the expressions of NLRP3,GAPDH,caspase-1,GSDMD and IL-1βprotein in myocardial tissue.Results Compared with the Con group,the expression levels of IL-1βand IL-18 in serum were significantly higher in the Mod group(P<0.05).Compared with the Mod group,the expression levels of IL-1βand IL-18 in serum were reduced in the PF groups.Compared with the Con group,the cardiomyocytes in the Mod group were extensively necrotic in large sheets,broken and disordered in arrangement,with a large number of inflammatory cells infiltrating the myocardial interstitium and the disappearance of transverse myocardium.Compared with the Mod group,as the PF dose increased,the cells gradually returned to normal and arranged neatly.Besides,the infiltration of myocarditis cells decreased.Compared with the Con group,the expression levels of NLRP3,Caspase-1,GSDMD and IL-1βin myocardial tissue were significantly higher in the Mod group with significant difference(P<0.05).Compared with the Mod group,the expressions of NLRP3,Caspase-1,GSDMD and IL-1βin myocardial tissue of the PF group decreased with the increase of PF dose.Conclusion Paeoniflorin can reduce myocardial inflammation and myocardial cell death caused by radiation-induced myocardial injury,thereby protecting the myocardium.NLRP3 inflammasome can serve as a potential therapeutic target for radiation-induced myocardial injury.