GLP-1通过miR-22/NLRP3途径减轻ox-LDL诱导的内皮细胞焦亡

GLP-1 alleviates ox-LDL-induced endothelial cell pyroptosis via miR-22/NLRP3 pathway

目的探讨胰高血糖素样肽1(GLP-1)对氧化型低密度脂蛋白(ox-LDL)诱导人脐静脉内皮细胞(HUVEC)焦亡的调节作用及分子机制。方法培养HUVEC并分为对照组、ox-LDL组、不同剂量GLP-1组(10 nmol/L、100 nmol/L、1 000 nmol/L)、GLP-1+miR-22抑制物组、miR-22抑制物组、miR-22模拟物组、阴性对照(NC)抑制物组、阴性对照模拟物组。流式细胞术检测细胞凋亡率,荧光定量PCR检测miR-22表达量,Western blot检测NOD样受体蛋白3(NLRP3)、凋亡相关点样蛋白(ASC)、含半胱氨酸的天冬氨酸蛋白水解酶1(Caspase-1)的表达量,酶联免疫吸附法检测白细胞介素1β(IL-1β)、IL-18的含量。结果与对照组比较,ox-LDL组细胞凋亡率和细胞中NLRP3、ASC、Caspase-1的表达以及培养基中IL-1β、IL-18含量均明显增加,细胞中miR-22的表达明显减少(P<0.05);与ox-LDL组比较,GLP-1组细胞凋亡率和细胞中NLRP3、ASC、Caspase-1的表达以及培养基中IL-1β、IL-18含量均明显减少,细胞中miR-22的表达明显增加(P<0.05);与GLP-1组比较,GLP-1+miR-22抑制物组细胞凋亡率和细胞中NLRP3、ASC、Caspase-1的表达以及培养基中IL-1β、IL-18的含量均明显增加,细胞中miR-22的表达明显减少(P<0.05);miR-22抑制物组细胞中NLRP3、ASC、Caspase-1的表达及培养基中IL-1β、IL-18的含量明显高于阴性对照抑制物组,miR-22模拟物组细胞中NLRP3、ASC、Caspase-1的表达及培养基中IL-1β、IL-18的含量明显低于阴性对照模拟物组(P<0.05)。结论 GLP-1通过miR-22/NLRP3途径能够减轻ox-LDL诱导的内皮细胞焦亡。

Aim To study the regulatory effect and molecular mechanism of glucagon-like peptide-1 (GLP-1) on pyroptosis of umbilical vein endothelial cells (HUVEC) induced by oxidized low density lipoprotein (ox-LDL). Methods HUVEC was cultured and divided into control group, ox-LDL group, GLP-1 group (10 nmol/L, 100 nmol/L, 1 000 nmol/L), GLP-1+miR-22 inhibitor group, miR-22 inhibitor group, miR-22 mimic group, negative control (NC) inhibitor group and NC mimic group. Flow cytometry was used to detect apoptotic rate, fluorescence quantitative PCR was used to detect the expression of microRNA-22(miR-22), Western blot was used to detect the expression of NOD-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein containing CARD (ASC) and caspase-1, enzyme-linked immunosorbent assay was used to detect the content of interleukin 1beta (IL-1β) and interleukin-18(IL-18). Results Compared with the control group, the apoptotic rate, the expression of NLRP3, ASC, caspase-1 in cells and the levels of IL-1β and IL-18 in the culture medium of ox-LDL group significantly increased, while the expression of miR-22 significantly decreased (P<0.05). Compared with ox-LDL group, the apoptotic rate, the expression of NLRP3, ASC, caspase-1 in cells and the levels of IL-1β and IL-18 in the culture medium of GLP-1 group significantly decreased, while the expression of miR-22 significantly increased (P<0.05). Compared with GLP-1 group, the apoptotic rate, the expression of NLRP3, ASC, caspase-1 in cells and the levels of IL-1β and IL-18 in the culture medium of GLP-1+miR-22 inhibitor group significantly increased, while the expression of miR-22 significantly decreased (P<0.05). The expression of NLRP3, ASC, caspase-1 in cells and the levels of IL-1β and IL-18 in the culture medium of miR-22 inhibitor group were significantly higher than those of NC inhibitor group, while the expression of NLRP3, ASC, caspase-1 in cells and the levels of IL-1β and IL-18 in the culture medium of miR-22 mimic group were significantly higher than those of NC mimic group. Conclusion GLP-1 can alleviate ox-LDL-induced endothelial cell pyroptosis through the miR-22/NLRP3 pathway.