Page 24 - 南京医科大学学报自然科学版

南京医科大学学报（自然科学版） 第43卷第10期
·1350 · Journal of Nanjing Medical University（Natural Sciences） 2023年10月

·基础研究·

Notch1在对乙酰氨基酚诱导的肝损伤中的作用及机制

邵宇云，王 涵，王 潇，戴晶晶，李 军，蒋龙凤 *
南京医科大学第一附属医院感染病科，江苏 南京 210029

［摘 要］ 目的：探讨Notch1信号通过转化生长因子β激活激酶1（transforming growth factor⁃β⁃activated kinase 1，TAK1）调控对
乙酰氨基酚（acetyl⁃para⁃aminophenol，APAP）诱导的肝损伤（APAP induced liver injury，AILI）的作用机制。方法：髓系特异性
Notch1敲除（Notch1 M⁃KO ）和对照floxed Notch1（Notch1 FL/FL ）小鼠通过腹腔注射APAP构建AILI模型。留取小鼠血清标本，用全自
动生化分析仪及酶联免疫反应分析法检测肝功能和细胞因子。留取小鼠肝组织标本，HE染色观察肝组织病理损伤情况，使用
Suzuki评分评估肝组织损伤程度，免疫印迹法检测TAK1、磷酸化TAK1（p⁃TAK1）、p65、磷酸化p65（p⁃p65）、Caspase⁃8（Casp⁃8）、
受体相互作用蛋白激酶 1（receptor⁃interacting protein kinase 1，RIPK1）、磷酸化混合谱系激酶域样蛋白（mixed lineage kinase
domain⁃like protein，p⁃MLKL）的表达，免疫荧光染色观察 CD11b、p⁃TAK1 的表达及活性氧（reactive oxygen species ROS）水平。
结果：小鼠腹腔注射APAP后，肝脏病理提示肝细胞体积增大，窦道淤血，出现广泛的坏死。与Notch1 FL/FL 对照组相比，Notch1 M⁃KO
小鼠血清丙氨酸氨基转移酶（alanine aminotransferase，ALT）、天冬氨酸氨基转移酶（aspartate aminotransferase，AST）明显升高，
血清炎性因子水平上升，HE染色显示肝细胞体积增大更明显，伴大面积坏死及炎性细胞浸润，DCF探针检测显示原代肝细胞
内ROS增加。肝组织p⁃TAK1表达增加，Casp⁃8的表达减少，RIPK1、p⁃MLKL表达增加。结论：在AILI中，髓系特异性Notch1敲
除可活化TAK1，降低Casp⁃8水平，激活RIPK1⁃MLKL坏死性凋亡通路，加重肝损伤的发生。
［关键词］ 对乙酰氨基酚；肝损伤；髓系特异性敲除；Notch1；TAK1；坏死性凋亡
［中图分类号］ R994.39 ［文献标志码］ A ［文章编号］ 1007⁃4368（2023）10⁃1350⁃07
doi：10.7655/NYDXBNS20231003

Effects and mechanisms of Notch1 on acetyl⁃para⁃aminophenol induced liver injury
*
SHAO Yuyun，WANG Han，WANG Xiao，DAI Jingjing，LI Jun，JIANG Longfeng
Department of Infectious Diseases，the First Affiliated Hospital of Nanjing Medical University，Nanjing 210029，China

［Abstract］ Objective：To investigate the mechanism by which Notch1 signaling regulates acetyl⁃para⁃aminophenol（APAP）⁃induced
liver injury（AILI）via TAK1. Methods：AILI models were constructed on myeloid⁃specific Notch1 knockout（Notch1 M⁃KO ）and floxed
Notch1（Notch1 FL/FL ）mice by intraperitoneal injection of APAP. Serum samples of mice were collected for detection of liver function
and cytokines by fully automated biochemical analyser and enzyme ⁃ linked immunoassay（ELISA），respectively. The pathological
damage of liver tissue was observed by HE staining，and the degree of liver tissue damage was evaluated by Suzuki score. Western blot
analysis was performed to detect the expression levels of TAK1，phospho⁃TAK1，p65，phospho⁃p65，caspase⁃8，receptor⁃interacting
protein kinase1（RIPK1），and phospho ⁃ mixed lineage kinase domain ⁃ like protein（MLKL）in the liver tissue. The expressions of
CD11b，p ⁃ TAK1 and the level of reactive oxygen species（ROS）were observed by immunofluorescence staining. Results：After
injected with APAP intraperitoneally in mice，liver histopathological examination suggested increased hepatocyte volume，sinus
congestion，and extensive necrosis. Compared with Notch1 FL/FL group，Notch1 M- KO mice showed significantly elevated serum alanine
aminotransferase（ALT） and aspartate aminotransferase（AST），increased inflammatory factor levels. HE staining showed more
pronounced increase in hepatocyte volume，accompanied by extensive necrosis and increased infiltration of inflammatory cells.
Additionally，the primary hepatocytes showed higher levels of ROS when assessed using the DCF probe. The expression of p⁃TAK1
in liver tissue elevated，and the expression of caspase⁃8 was down⁃regulated，while the expressions of RIPK1 and p⁃MLKL were up⁃
regulated. Conclusion：In AILI，myeloid⁃specific Notch1 knockout activates TAK1 expression，which also decreases caspase⁃8 levels

［基金项目］ 江苏省自然科学基金青年基金（BK20161059）；中国肝炎防治基金会——天晴肝病研究基金（CFHPC20132071）
∗
通信作者（Corresponding author），E⁃mail：longfengjiang@njmu.edu.cn