基于一水草酸钙诱导HK-2细胞损伤模型筛选广金钱草药效成分及其作用机制  

Screening for bioactive components of Desmodium styracifolium and its mechanisms study based on calcium oxalate monohydrate-induced HK-2 damage model

作　　者：李明慧 王术玲[1] 曹骋[1] 陈丰连[1] LI Ming-hui;WANG Shu-ling;CAO Cheng;CHEN Feng-lian(School of Pharmaceutical Sciences,Guangzhou University of Chinese Medicine,Guangzhou 510006,China)

机构地区：[1]广州中医药大学中药学院,广东广州510006

出　　处：《药学学报》2020年第10期2375-2380,共6页Acta Pharmaceutica Sinica

基　　金：国家中医药管理局项目(201507002);广东省科技计划项目(2015B090901033)。

摘　　要：研究广金钱草保护一水草酸钙(calcium oxalate monohydrate,COM)诱导人近端肾小管上皮细胞(human proximaltubular epithelial cell,HK-2)损伤模型的药效成分,并进一步探讨其作用机制。首先建立COM诱导HK-2细胞损伤模型,通过MTT法对广金钱草中所含8种黄酮成分(夏佛塔苷、异夏佛塔苷、维采宁-2、异牡荆苷、异荭草苷、芹菜素、木犀草素和染料木素)进行药效筛选,检测不同处理组细胞上清液中乳酸脱氢酶(lactate dehydrogenase,LDH)释放量、细胞内超氧化物歧化酶(superoxide dismutase,SOD)活力、细胞内活性氧(reactive oxygen species,ROS)产生量,Western blot检测HK-2细胞中NLRP3炎症小体相关蛋白(NLRP3、caspase-1、HMGB1)的表达水平,进一步探讨药效成分的作用机制。研究结果表明,与模型组相比,广金钱草总黄酮(total flavonoids of Desmodium styracifolium,TFDS)及4个黄酮单体(异荭草苷、芹菜素、染料木素和木犀草素)干预后的细胞活力显著上升,均可减少细胞上清液中LDH释放量,提高细胞内SOD活性,减少ROS产生量;COM造模后细胞内NLRP3炎症小体相关蛋白(NLRP3、caspase-1、HMGB1)表达均升高,给药组细胞中表达均有不同程度的回调。综上,TFDS及芹菜素、异荭草苷、木犀草素、染料木素可保护COM诱导HK-2损伤细胞,其作用机制包括提高细胞活力、保护细胞膜完整性和提高细胞抗氧化应激能力,并且可调控NLRP3炎症小体相关蛋白的表达。To screen active components of Desmodium styracifolium in protecting calcium oxalate monohydrate(COM)-induced human proximaltubular epithelial cell(HK-2)damage model,and furtherly explore its mechanism of action,total flavonoids of Desmodium styracifolium(TFDS)and eight flavonoids(schaftoside,isoschaftoside,vicenin-2,isovitexin,isoorientin,apigenin,luteolin and genistein)were tested by COM-induced HK-2 damage model.MTT assay was used to detect the effects of different components on the cell viability of COM-induced HK-2 damage model.The lactate dehydrogenase(LDH)release in the cell supernatant and the activity level of superoxide dismutase(SOD)and reactive oxygen species(ROS)of cell were detected by the kit.Western blot was used to detect the expression levels of NLRP3,caspase-1,HMGB1 in HK-2 of different groups.Compared with the model group,the cell activity was significantly increased after 24 h co-culture with TFDS and four flavonoids(isoorientin,apigenin,genistein and luteolin).These active components can reduce the LDH leakage and ROS in cell supernatant and increase the activity of SOD,with regulating the expression of NLRP3,caspase-1,HMGB1.TFDS,apigenin,isoorientin,luteolin and genistein can protect COM-induced HK-2 cell damage,including enhancing cell viability,protecting cell membrane integrity and enhancing oxidative stress,and regulate the expression of proteins related to NLRP3 inflammasome.

关 键 词：广金钱草 草酸钙结石 活性成分 黄酮 炎症因子 

分 类 号：R285[医药卫生—中药学]