谷甾醇通过STAT3信号通路治疗尿路感染的机制研究

Basic study on sitosterol treatment of urinary tract infection via STAT3 pathway

目的分析谷甾醇治疗尿路感染(UTI)的作用机制,为临床治疗提供指导意义。方法通过TCMSP数据库收集木通活性成分并筛选出谷甾醇作为研究对象,脂多糖(LPS)联合三磷酸腺苷(ATP)诱导人尿路上皮细胞(SV-HUC-1)建立体外尿路感染模型组,分别设对照组、模型+谷甾醇组和对照+谷甾醇组;通过GeneCards数据库收集UTI相关的疾病靶点,并与药物活性成分靶点取交集得到共同靶点,采用细胞增殖检测实验(CCK8)检测四组细胞的活力。采用免疫印迹实验(Western blot)检测核苷酸结合寡聚化结构域样受体蛋白3(NLRP3)、gasdermin D-N端(GSDMD-N)、半胱天冬酶-1(Caspase-1)、磷酸化信号转导因子和转录激活因子3(P-STAT3)和STAT3的蛋白表达水平。采用酶联免疫吸附试验(ELISA)检测细胞上清液白介素-1β(IL-1β)、白介素-18(IL-18)的含量。结果通过TCMSP数据库分析,选择谷甾醇作为研究药物,Western blot结果显示,与对照组比较,模型组的NLRP3、DSDMD-N、Caspase-1和P-STAT3蛋白表达增加,细胞上清液中IL-1β、IL-18的含量增加(均P<0.05);与模型组比较,模型+谷甾醇组的NLRP3、DSDMD-N、Caspase-1和P-STAT3蛋白表达降低,细胞上清液中IL-1β、IL-18的含量降低(均P<0.05)。结论谷甾醇可通过抑制STAT3信号转导减弱脂多糖诱导的炎症和细胞焦亡,对治疗UTI具有一定的临床应用价值。

Objective To explore the mechanism of action of sitosterol in the treatment of urinary tract infection(UTI)and to provide guidance for clinical treatment.Methods The active constituents of xylosterol were obtained from the Traditional Chinese Medicine Systems Pharmacology(TCMSP)database,and sitosterol was selected as the study compound.A UTI model was established in vitro using SV-HUC-1 cells induced by lipopolysaccharide(LPS)and adenosine triphosphate(ATP).SV-HUC-1 was set as control group,model+sitosterol group and control+sitosterol group.UTI-related disease targets were collected from the GeneCards database,and the targets of the drug active ingredients were obtained by intersecting with them.Cell viability was measured using the cell counting kit-8(CCK-8)assay.Western blot assay was used to detect the protein expressions of nucleotide-binding oligomerization domain-like receptor protein 3(NLRP3),gasdermin D-N terminal(GSDMD-N),Caspase-1,phosphorylated signal transducer and activator of transcription 3(P-STAT3),and STAT3.The levels of interleukin-1β(IL-1β)and interleukin-18(IL-18)in cell supernatants were measured using enzyme-linked immunosorbent assay(ELISA).Results Sitosterol was found to be the main active component of xylosterol.The Western blot results showed that compared with the control group,the protein expressions of NLRP3,GSDMD-N,Caspase-1,and P-STAT3 were significantly increased in the model group(all P<0.05),and the levels of IL-1βand IL-18 in cell supernatants were also elevated(all P<0.05).In contrast,compared with the model group,the protein expressions of NLRP3,GSDMD-N,Caspase-1,and P-STAT3 were significantly reduced(all P<0.05),and the levels of IL-1βand IL-18 in cell supernatants were also decreased in the model+sitosterol group(all P<0.05).Conclusions Sitosterol may reduce inflammation and pyroptosis induced by lipopolysaccharide by inhibiting the STAT3 signaling pathway.Therefore,sitosterol has potential clinical value in the treatment of UTI.