摘要
目的观察一贯煎对野百合碱诱导的急性肝窦阻塞综合征(hepatic sinusoidal obstruction syndrome,HSOS)大鼠模型的干预作用并探析其部分作用机制。方法24只雄性SD大鼠随机分为正常对照组、模型组及一贯煎组,每组8只。采用野百合碱90 mg/kg灌胃,制备大鼠HSOS模型。一贯煎组大鼠在造模后6小时及30小时予以浓度为1.139 g/kg一贯煎灌胃,造模48小时后处死取材。检测血清丙氨酸氨基转移酶、天冬氨酸氨基转移酶活性,采用扫描电镜及苏木素—伊红染色观察肝组织病理变化,微板法检测肝组织匀浆谷胱甘肽S转移酶、总谷胱甘肽、丙二醛含量,TUNEL法检测细胞凋亡,实时荧光定量聚合酶链式反应法检测肝组织肿瘤坏死因子α、血红加氧酶1、炎症小体NLRP3及白介素1β的mRNA表达,免疫组化法检测肝组织巨噬细胞标志物F4/80表达,Western Blot法检测肝组织肿瘤坏死因子α、细胞凋亡相关B淋巴细胞瘤-2(B-cell lymphoma-2,Bcl2)和Bcl-2相关X蛋白(Bcl-2 associated X,Bax)的蛋白表达。结果(1)与正常对照组相比,模型组血清谷丙转氨酶和谷草转氨酶活性显著升高(P<0.01),苏木素—伊红染色可见大量肝细胞坏死、炎性细胞浸润及红细胞淤积,电镜观察可见肝血窦扩张,窦内皮窗孔扩大,“筛状”结构消失,甚至脱落,Disse空间暴露。(2)模型组肝组织丙二醛含量显著增高(P<0.05),巨噬细胞标志物F4/80及肿瘤坏死因子α的蛋白表达显著增加(P<0.01),肿瘤坏死因子α、血红加氧酶-1、炎症小体及白介素1β的mRNA表达显著增加(P<0.01),而一贯煎用药可降低肝组织丙二醛含量(P<0.05),下调F4/80及肿瘤坏死因子α蛋白表达(P<0.01)及肿瘤坏死因子α、血红加氧酶1(P<0.001)、炎症小体及白介素1β(P<0.05)的mRNA表达。(3)TUNEL检测模型组肝组织凋亡细胞显著增加,Bcl2/Bax比值显著下降(P<0.05)。结论一贯煎可有效改善野百合碱诱导的急性肝窦阻塞综合征,可显著降低模型大鼠血清谷丙转氨酶及谷草转氨酶的活性,改善肝组织坏死及肝窦内皮损伤,并减少肝组织细胞凋亡。其作用机制可能与抗过氧化、抗炎性损伤、抑制肝细胞凋亡有关。
Objective To observe the intervention effect of Yiguanjian on monocrotaline-induced acute hepatic sinusoidal obstruction syndrome(HSOS)in rats and to explore its mechanism.Methods Twenty-four male SD rats were randomly divided into the control group,the model group and the Yiguanjian treated group(n=8).90 mg/kg body weight of monocrotaline were administered intragastrically to prepare rat HSOS model.Rats in the Yiguanjian treated group were administered intragastrically with 10 ml/kg body weight 6 hours or 30 hours after treated with monocrotaline respectively,and consistency of decoction is 1.139 g/kg.All the rats were sacrificed after 48 hours.The serum and liver tissue samples were collected and serum liver function was measured.Scanning electron microscope and HE staining were used to observe liver pathology.The activity of glutathione S-transferase,total superoxide dismutase and the content of malondialdehyde in liver tissue homogenates were detected by microplate method.Cell apoptosis was detected by TUNEL method,the mRNA expressions of TNF-α,HO-1,NLRP3,IL-1βin liver tissues were detected by qRT-PCR.F4/80 expressions in liver tissues were detected by immunohistochemistry and the expressions of TNF-α,Bcl2 and Bax proteins in liver tissues were detected by Western Blot.Results(1)Compared with the control group,the serum glutamic pyruvic transaminase and glutamic oxaloacetic transaminase activities in the model group were significantly increased(P<0.01).A large amount of hepatocyte necrosis,inflammatory cells infiltration and erythrocyte stasis were observed by HE staining.Electron microscopy showed that hepatic sinus dilatation and endothelial window openings.The“ethmoidal”structure of hepatic sinuses disappeared or even fell off,and the Disse space was exposed.(2)The MDA content,the protein expressions of F4/80 and TNF-α,and the mRNA expressions of TNF-α,HO-1,NLRP3,and IL-1βof liver tissue in the model group was increased significantly(P<0.01),while Yiguanjian could significantly reduce the MDA content(P<0.05),and down-regulate the expressions of F4/80,TNF-α(P<0.01),HO-1(P<0.001),NLRP3(P<0.01)and IL-1β(P<0.05).(3)The TUNEL positive cells were increased significantly in the model group,Bcl2/Bax ratio decreased significantly(P<0.05).Conclusion Yiguanjian can effectively improve monocrotaline-induced acute HSOS,it can significantly reduce the activity of ALT and AST in serum of model rats,reduce liver necrosis and sinusoidal endothelial damage,and reduce liver tissue cell apoptosis.Its mechanism may be related to inhibit oxidative stress and inflammatory injury,and reduce cell apoptosis.
作者
高思琦
肖准
王晓柠
刘伟
陈佳美
刘平
GAO Siqi;XIAO Zhun;WANG Xiaoning;LIU Wei;CHEN Jiamei;LIU Ping(Institute of Interdisciplinary Science,Shanghai University of Traditional Chinese Medicine,Shanghai 201203,China)
出处
《环球中医药》
CAS
2021年第9期1566-1572,共7页
Global Traditional Chinese Medicine
基金
国家自然科学基金(81530101、81973613)
上海市青年科技启明星计划(19QA1408900)。
关键词
一贯煎
肝窦阻塞综合征
野百合碱
抗氧化
抗炎
抗凋亡
效应机制
Yiguanjian
Sinusoidal obstruction syndrome
Monocrotaline
Anti-oxidation
Anti-inflammatory
Anti-apoptosis
Effect mechanism
作者简介
高思琦(1994-),2018级在读博士研究生。研究方向:中医药防治慢性肝病。E-mail:sharryco2@163.com;肖准,博士研究生;通信作者:陈佳美(1987-),博士,副研究员。研究方向:中医药防治慢性肝病的临床与基础研究。E-mail:cjm0102@126.com。
