摘要
目的:利用精密肝切片(PCLS)技术,建立醋氨酚所致肝切片损伤的体外模型,并探讨CYP2E1活性变化对肝损伤的调节作用,为研究和筛选肝保护药物提供实验依据。方法:利用振荡切片机制备PCLS,建立培养系统。将终浓度为500μmol/L的醋氨酚分别与切片共孵育0、2、4、6h,测定培养基和切片中的谷胱甘肽S-转移酶(GST)和乳酸脱氢酶(LDH)活性。在体给予不同剂量(0.25、0.5、1.0g/kg)的CYP2E1诱导剂乙醇,每天一次,连续3d。末次给药8h后处死动物,取其肝脏制备PCLS,再与500μmol/L醋氨酚共同孵育6h,检测ALT和LDH漏出率。正常PCLS与不同浓度的二乙基二硫代氨基甲酸酯盐(DDTC,CYP2E1抑制剂)(5、10、20μmol/L)、500μmol/L醋氨酚共同孵育6h,检测ALT和LDH漏出率。结果:与常规培养组比较,醋氨酚孵育4、6h组的GST和LDH漏出率显著升高(P<0.01)。与醋氨酚模型对照组比较,乙醇中、大剂量组LDH漏出率和小、大剂量组ALT漏出率皆升高(P<0.01,P<0.05);DDTC各浓度组ALT漏出率则降低(P<0.05)。结论:PCLS与500μmol/L醋氨酚共孵育4h即能成功地建立体外肝切片损伤的模型。抑制CYP2E1活性可能对醋氨酚所致的肝损伤有保护作用,而上调CYP2E1活性则可能加重醋氨酚所致的肝损伤。
AIM： To establish an acetaminohpheninduced hepatotoxicity model using precision-cut liver slice （PCLS） technique and observe the regulation of cytochrome P4502E1 （CYP2E1） so as to provide experimental basis on investigating and screening new potential hepatoprotective drugs. METHODS： （ 1 ） The PCLS was prepared by the vibratome and co-incubated with 500 μmol/L acetaminophen for 0, 2, 4 and 6 h. The activities of glutathione S-transferase （GST） and lactic acid dehydrogenase （LDH） were measured in the medium and slices. （2）The rots were intragastrically administrated with ethanol in different dosages （0.25, 0.5, 1.0 g/kg） once a day for three days consecutively, then they were at sacrificed 8 h after the last administration and the slices were prepared. And the slices were co-incubated with 500 μmol/L acetaminophen for 6 h and then the activities of ALT and LDH were detected in the slices and medium. （3）Replace the medium after 1 h pre-incubation, the slices were co-incubated with 500 μmol/L acetaminophen and diethyldithiocarbamate （DDTC） （5, 10, 20 μmol/L） for 6 h and the activities of ALT and LDH in the slices and medium were assayed. RESULTS： Compared with regular culture group, the leakage rates of GST and LDH in acetaminophen co-incubated for 4 and 6 h groups were significantly increased （ P 〈 0.01 ）. Compared with those in acetaminophen model group, the leakage rates of LDH of ethanol at the dosages of 0.5, 1.0 g/kg and the leakage rates of ALT of ethanol at the dosages of 0.25, 1.0 g/kg increased significantly （ P 〈 0.01, P 〈 0.05）. Moreover, the leakage rates of ALT in all concentrations of DDTC decreased remarkably （ P 〈 0.05 ）. CONCLUSION： Co-incubation PCLS with 500 μmol/L acetaminophen for more than 4 h is a successful way to establish hepatotoxicity in vitro model. The reduction of CYP2E1 activity might protect the slices from acetaminophen-induced hepatotoxicity, while the increase of CYP2E1 activity might aggravate it.
出处
《中国临床药理学与治疗学》
CAS
CSCD
2008年第1期46-50,共5页
Chinese Journal of Clinical Pharmacology and Therapeutics
基金
国家自然科学基金资助项目(30371666)
关键词
CYP2E1
乙醇
醋氨酚
精密肝切片技术
肝损伤
CYP2E1
ethanol
acetaminophen
precision-cut liver slice technique
liver injury
