凉血解毒化瘀方预防慢性肝损伤小鼠向急性肝衰竭转化及其对TLR4的调控作用  

Preventive Effect of Liangxue Jiedu Huayu Decoction on the Transformation from Chronic Liver Injury to Acute Liver Failure in Mice and Its Regulation on TLR4

作　　者：刘政芳 余绍雄 廖乃顺 LIU Zhengfang;YU Shaoxiong;LIAO Naishun(Mengchao Hepatobiliary Hospital of Fujian Medical University,Fuzhou,Fujian 350028,China;The United Innovation of Mengchao Hepatobiliary Technology Key Laboratory of Fujian Province,Fuzhou,Fujian 350028,China)

机构地区：[1]福建医科大学孟超肝胆医院,福建福州350028 [2]福建省孟超肝胆技术联合创新重点实验室,福建福州350028

出　　处：《福建中医药》2024年第7期7-10,16,共5页Fujian Journal of Traditional Chinese Medicine

基　　金：福建省自然科学基金项目(2023J011465)。

摘　　要：目的探讨凉血解毒化瘀方对慢性肝损伤小鼠向急性肝衰竭转化的预防效果及其对Toll样受体4(TLR4)的调控作用。方法取6周龄SPF级雄性BALB/c小鼠60只,采用随机数字表法分为正常组、模型组、低剂量组、中剂量组、高剂量组,每组12只。除正常组外,其余4组按体质量5 mL/kg腹腔注射20%四氯化碳(CCl4)以构建慢性肝损伤小鼠模型,正常组腹腔注射等剂量生理盐水,2次/周,连续8周。经组织病理学与血清生化分析证实慢性肝损伤造模成功后,低、中、高剂量组按体质量10 mL/(kg·d)分别灌胃0.75、1.50、3.00 g/mL凉血解毒化瘀方药液,正常组、模型组灌胃等剂量生理盐水,连续灌胃28 d后,模型组与各剂量组分别按体质量5 mL/kg一次性腹腔注射2μg/mL脂多糖(LPS)和0.2 g/mL D-氨基半乳糖(D-GalN)以诱导慢性肝损伤小鼠向急性肝衰竭转化,正常组腹腔注射等剂量生理盐水。3 d后采用生化分析法检测5组小鼠血清丙氨酸转氨酶(ALT)、天冬氨酸转氨酶(AST)、总胆红素(TBIL)含量,HE染色观察5组小鼠肝组织病理形态变化,qPCR检测5组小鼠肝组织肿瘤坏死因子-α(TNF-α)、白细胞介素-6(IL-6)、白细胞介素-1β(IL-1β)、Toll样受体4(TLR4)mRNA转录水平,采用免疫荧光染色检测5组小鼠肝组织TLR4蛋白表达水平。结果与正常组比较,模型组血清ALT、AST、TBIL含量显著提高(P<0.05),HE染色结果显示肝组织出现肝细胞坏死,伴随大量纤维化改变及炎症细胞浸润,肝组织TNF-α、IL-6、IL-1β、TLR4 mRNA转录水平升高(P<0.05),肝组织TLR4蛋白表达水平增高。与模型组比较,各剂量组血清ALT、AST、TBIL含量降低(P<0.05),HE染色结果显示肝细胞坏死、纤维化,炎症细胞浸润程度减少,肝组织TNF-α、IL-6、IL-1β、TLR4 mRNA转录水平降低(P<0.05),肝组织TLR4蛋白表达量减少。与低剂量组比较,中、高剂量组血清ALT、AST水平降低更明显(P<0.05),HE染色结果显示肝细胞坏死�Objective:The effect and the TLR4 regulation of Liangxue Jiedu Huayu Decoction(LXJD)on preventing mice form chronic liver injury to acute liver failure.Methods:A total of sixty SPF male BALB/c mice aged 6 weeks were divided into normal group,model group,LXJD groups of low-,medium-and high-dose by random number table method,with 12 mice in each group.Apart from the normal group,all mice were intraperitoneally injected with 5 mL/kg 20%carbon tetrachloride(CCl4)twice a week for eight weeks,and mice in the normal group were administrated with the same dose of normal saline.After the successful models of chronic liver injury were confirmed by histopathology and serum biochemical analysis,the low-,medium-,and high-dose groups were given intragastric administration of 0.75,1.50 and 3.00 g/mL LXJD according to body mass 10 mL/(kg·d)respectively.The normal and model groups were given the same dose of normal saline.After continuous intragastric administration for 28 days,the model group and each dose group were intraperitoneally injected with 2μg/mL lipopolysaccharide(LPS)and 0.2 g/mL D-amino galactose(D-GalN)at a dose of 5 mL/kg to induce the transformation from chronic liver injury to acute liver failure.While the normal mice were given an equal volume of normal saline.Three days later,serum and liver tissue samples were collected for the following evaluation.Biochemical analysis was performed to measure serum levels of alanine aminotransferase(ALT),aspartate aminotransferase(AST),and total bilirubin(TBIL)in all groups.Hematoxylin-eosin(HE)staining was used to evaluate the histopathological changes of liver tissue in all groups.Quantitative PCR(qPCR)was used to detect mRNA levels of TNF-α,IL-6,IL-1β,and TLR4 in liver tissues from all groups.Immunofluorescence staining was used to analyze the protein expression of TLR4 in liver tissues from all groups.Results:Compared with the normal group,the model group showed a significant increase in serum ALT,AST,and TBIL content(P<0.05),as well as the typical hepatocyte necrosis

关 键 词：慢性肝损伤 急性肝衰竭 凉血解毒化瘀方 炎症反应 TLR4 

分 类 号：R285.5[医药卫生—中药学]