摘要
目的观察柴胡皂苷d(SSd)对人肝细胞损伤的保护作用及抗肝纤维化机制,探讨L02细胞氧化应激损伤与细胞焦亡、上皮-间质转化(EMT)的关系,为SSd在治疗肝纤维化疾病的应用提供临床依据。方法体外培养人正常肝细胞L02,采用H_(2)O_(2)诱导L02细胞损伤模型,干预组加入中药单体SSd(低、中、高剂量组)孵育24 h后再经过H_(2)O_(2)处理细胞,CCK8法观察SSd干预的损伤L02细胞后的活力、丙二醛(MDA)水平、上清液谷丙转氨酶(ALT)、谷草转氨酶(AST)检测,Western Blot法、RT-PCR法检测细胞焦亡相关指标半胱氨酸蛋白酶1(Caspase-1)、白细胞介素1β(IL-1β)、白细胞介素18(IL-18)及上皮间质转化(EMT)相关指标转化生长因子(TGF-β1)、钙黏附蛋白(Ecadherin)、波形蛋白(Vimentin)、肌劲蛋白(α-SMA)的表达。结果SSd能抑制L02细胞损伤引起的细胞活力下降,在有效剂量内无明显肝毒性(ALT、AST未见明显升高);与对照组比较,氧化应激损伤的L02细胞焦亡相关指标Caspase-1、IL-1β、IL-18表达明显升高(P<0.05,P<0.01),EMT相关指标TGF-β1、Vimentin、α-SMA的表达升高,Ecadherin表达下调(P<0.05,P<0.01);SSd干预后,Caspase-1、IL-1β、IL-18、TGF-β1、Vimentin、α-SMA的表达明显下调,Ecadherin表达上调(P<0.05,P<0.01)。结论氧化应激损伤的L02细胞活力下降可能与细胞焦亡相关,SSd有抑制损伤的L02细胞焦亡、控制EMT过程,发挥抗肝纤维化的作用。
Objective To observe the protective effect of Bupleurin D(SSd)on human hepatocyte injury and the mechanism of anti-liver fibrosis and explore the relationship between oxidative stress injury of L02 cells and pyroptosis and epithelial-interstitial transformation(EMT)so as to provide clinical basis for the application of SSd in the treatment of liver fibrosis.Methods Normal human liver cells L02 were cultured in vitro and the damage model of L02 cells was induced by H_(2)O_(2).The intervention group was incubated with SSd(low,medium and high dose groups)for 24 h and then the cells were treated with H_(2)O_(2).CCK8 method was used to observe the activity,MDA concentrations,ALT and AST of the damaged L02 cells after SSd intervention.The expressions of caspase-1,IL-1β,IL-18 and EMT-related indicators,TGF-1β,Ecadherents,Vimentin andα-SMA were detected by Western Blot and RT-PCR respectively.Results SSd could inhibit the decrease of cell activity caused by L02 cell injury,and there was no significant hepatotoxicity in the effective dose(ALT and AST were not significantly increased).The expressions of caspase-1,IL-1β,and IL-18 in L02 cells with oxidative stress injury were significantly increased(P<0.05,P<0.01).The expression levels of EMT-related indicators TGF-1β,Vimentin andα-SMA were increased and the Ecadherents expression was up-regulated(P<0.05,P<0.01).After SSd intervention,the expressions of caspase-1,IL-1β,IL-18,TGF-1β,Vimentin andα-SMA were significantly down-regulated and Ecadherin expression was up-regulated(P<0.05,P<0.01).Conclusion The decreased activity of L02 cells injured by oxidative stress may be related to pyroptosis,and SSd can inhibit the pyroptosis of L02 cells injured,control EMT process and play an anti-liver fibrosis role.
作者
张娜
李勇
ZHANG Na;LI Yong(Shanghai Hospital of Traditional Chinese Medicine,Shanghai University of Traditional Chinese Medicine,Shanghai 200071,China)
出处
《中华中医药学刊》
CAS
北大核心
2021年第12期21-27,I0029,共8页
Chinese Archives of Traditional Chinese Medicine
基金
国家自然科学基金(81573775)。
关键词
柴胡皂苷D
人肝细胞细胞焦亡上皮-间质转化
肝纤维化
bupleuronin D
human hepatocytes
pyroptosis
epithelial-interstitial transformation
liver fibrosis
