实用医学杂志2023，Vol.39Issue(23)：3034-3042,9.DOI:10.3969/j.issn.1006-5725.2023.23.003
茵陈蒿汤协同脐带间充质干细胞所释放的外泌体对急性肝衰竭及肝细胞焦亡的影响
Effect of Yinchenhao Decoction combined with exosomes derived from umbilical cord mesenchymal stem cells on acute liver failure and hepatocyte pyroptosis
摘要
Abstract
Objective To investigate the hepatoprotective effect and anti-hepatocyte pyroptosis effect of Yinchenhao decoction synergized with umbilical cord mesenchymal stem cells(UcMSCs)in vitro and vivo models of acute liver failure(ALF),as compared to single-drug treatment.Methods UcMSCs were extracted from umbilical cords and identified using electron microscopy and flow cytometry.Then the exosomes released from ucMSCs(UcMSCs-exo)were isolated through ultracentrifugation,and ucMSCs-exo were identified by nanoparticle tracking analysis(NTA),transmission electron microscopy,and Western blot for CD63 and CD9.Human hepatocyte cell line LO2 cells were stimulated by 100 ng/mL lipopolysaccharide(LPS)and 5 mmol/L adenosine triphosphate(ATP)to construct a vitro model of ALF.The cells were randomly divided into the following groups:a normal control group,a model group,a Yinchenhao decoction treatment group,a ucMSCs-exo treatment group,and a combination treatment group.The morphological changes of LO2 were observed under the electron microscope,the mortality of LO2 cells was detected by flow analysis,the expression of pyroptosis-related proteins Caspase-1 and Cleaved-Caspase-1 was detected by Western blot,and the expression of pyroptosis-related cytokines IL-1β and IL-18 was detected by ELISA.Then,fifty 6-8-week-old male C57BL/6 mice were randomly divided into the following groups:normal control group,model group,Yinchenhao decoction treatment group,ucMSCs-exo treatment group,and combined treat-ment group.Each group consisted of 10 mice.The ALF mouse model was constructed by intraperitoneal injection of 10 μg/kg LPS and 800 mg/kg D-galactosamine(D-GalN).After successful modeling,each group was given the appropriate drug interventions,and the ucMSCs-exo and combined treatment groups were injected with ucMSCs-exo interventions in the tail vein,and the drugs were administered continuously for 3 days.After 12 hours of intervention,serum specimens and liver tissues were collected from each group.The expression levels of ALT and AST were detected,and the liver tissues were stained with HE to observe the pathological changes.Results In the ALF cell model,hepatic cell death was inhibited in the Yinchenhao decoction treatment group,the ucMSCs-exo treatment group,and the combination group(P<0.05).The rate of hepatic cell death in the combination treatment group was significantly reduced(P<0.05).Initially,the study aimed to explore the effect on hepatic cell pyroptosis,and it was found that the combination treatment group could effectively suppress hepatocellular cell death more effectively compared to the single treatment groups.The combined treatment group significantly reduced the expression of the pyroptosis-related protein Cleaved-Caspase-1,as well as the cytokines IL-1β and IL-18(P<0.05).However,there was no statistically significant difference in the expression of Caspase-1(P>0.05).In the vivo model,the single Yinchenhao decoction treatment group,the ucMSCs-exo treatment group,and the combined treatment group were able to effectively reduce the serum ALT and AST levels compared to the model group(P<0.05).Additionally,these treatments were found to attenuate hepatocellular necrosis and the infiltration of inflammatory cells.Compared to the group treated with only Yinchenhao decoction or only ucMSCs-Exo,the group that received combined treat-ment showed a greater decrease in serum ALT and AST levels.Additionally,there was a reduction in the infiltration of inflammatory cells and the extent of cell death in the liver tissues(P<0.05).Conclusions The combination of Yinchenhao decoction and ucMSCs-exo demonstrated a significantly better hepatoprotective effect in the ALF vivo model compared to the individual treatments of Yinchenhao decoction or ucMSCs-exo alone.Furthermore,this combination treatment was able to effectively inhibit hepatocyte pyroptosis.关键词
急性肝衰竭/茵陈蒿汤/脐带间充质干细胞/肝细胞焦亡/外泌体Key words
acute liver failure/Yinchenhao decoction/umbilical cord mesenchymal stem cells/hepatocyte pyroptosis/exosomes分类
医药卫生引用本文复制引用
谢丹,欧阳石..茵陈蒿汤协同脐带间充质干细胞所释放的外泌体对急性肝衰竭及肝细胞焦亡的影响[J].实用医学杂志,2023,39(23):3034-3042,9.基金项目
国家自然科学基金资助项目(编号:81803884) （编号:81803884）
广州医科大学科研能力提升项目(编号:02-410-2302092XM) （编号:02-410-2302092XM）
广州医科大学2022年学生创新能力提升计划项目(编号:02-408-2203-2064) （编号:02-408-2203-2064）
2022年广州医科大学学科建设项目(编号:02-410-2206013) （编号:02-410-2206013）
广州医科大学本科生能力提升创新项目(编号:2022JXA003) （编号:2022JXA003）
2023年市校(院)企联合资助项目(编号:2023A03J0421) （院）
2023年北京肝胆相照公益基金会资助项目(编号:iGandanF-1082023-RGG023) （编号:iGandanF-1082023-RGG023）
广州市医学重点学科[2021-2023]项目 （）
广东高校生物靶向诊治与康复重点实验室(编号:.2021KSYS009) （编号:.2021KSYS009）
