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环磷酰胺诱导的尼罗罗非鱼体内外肝损伤模型的构建
曹丽萍,杜金梁,贾睿,丁炜东,GalinaJeney,徐跑,殷国俊
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(中国水产科学研究院淡水渔业研究中心,农业农村部淡水渔业和种质资源利用重点实验室,无锡 214081; 中国水产科学研究院淡水渔业研究中心,农业农村部鱼类免疫药理学国际联合实验室,无锡 214081;中国水产科学研究院淡水渔业研究中心,农业农村部鱼类免疫药理学国际联合实验室,无锡 214081; Research Institute for Fisheries,Aquaculture and Irrigation,Ann Light 8,Szarvas H-4440,Hungary)
摘要:
以环磷酰胺(CTX)为诱导物,建立罗非鱼体内外急性肝损伤模型。体外,0、5、10、15、20和25 mmol·L-1 CTX 分别作用于离体培养的罗非鱼精密肝切片(PCLS)6 h 后,测定切片培养上清中谷丙转氨酶(GPT)、谷草转氨酶(GOT)和乳酸脱氢酶(LDH)水平,同时收集肝切片,用噻唑蓝(MTT)法检测肝切片增殖活性,测定切片内总抗氧化能力(T-AOC)、丙二醛(MDA)、过氧化物歧化酶(SOD)和谷胱甘肽(GSH)水平。体内,采用胸腔注射法造模,每千克鱼体重分别注射0、10、25、75和100 mg环磷酰胺(下文以mg·kg-1表示),每隔3 d给药1次,连续3次。末次给药后第4天取血,测定罗非鱼血清中GPT、 GOT 和LDH及肝组织匀浆中T-AOC 、MDA、SOD和GSH水平。结果显示: 20~25 mmol·L-1 CTX作用体外培养精密肝切片6 h及75~100 mg·kg-1 CTX胸腔注射罗非鱼后,均可以导致GPT、GOT和 LDH 活力显著升高;T-AOC、SOD活力及GSH含量显著下降,MDA含量显著提高。同时,20 和25 mmol·L-1 CTX能导致肝切片增殖活性显著下降,分别为空白对照组的67.29%和55.41%,且有剂量依赖性。结果表明:CTX可引起罗非鱼肝损伤;分别采用20 mmol·L-1 CTX 作用体外培养PCLS 6 h或者采用75~100 mg·kg-1 CTX胸腔注射罗非鱼,每隔3 d给药1次,连续3次,均可以构建体内外急性肝损伤模型。
关键词:  环磷酰胺;罗非鱼;肝损伤模型  体内外
DOI:10.13610/j.cnki.1672-352x.20220325.010
基金项目:中央级公益性科研院所基本科研业务费专项资金 (2019JBFM11)和江苏自然科学基金(BK20201143)共同资助。
Establishment of the acute hepatic injury model induced by cyclophosphamide in tilapia (Oreochromis niloticus) in vivo and in vitro
CAO Liping,DU Jinliang,JIA Rui,DING Weidong,Galina Jeney,XU Pao,YIN Guojun
(Key Laboratory of Freshwater Fisheries and Germplasm Resources Utilization, Ministry of Agriculture and Rural Affairs, Freshwater Fisheries Research Center, Chinese Academy of Fishery Sciences, Wuxi 214081, China; International Joint Research Laboratory for Fish Immunopharmacology, Chinese Academy of Fishery Sciences, Wuxi 214081, China;International Joint Research Laboratory for Fish Immunopharmacology, Chinese Academy of Fishery Sciences, Wuxi 214081, China; Research Institute for Fisheries, Aquaculture and Irrigation, Ann Light 8, Szarvas H-4440, Hungary) Abstract; The present study aimed to develop acute liver injury models of tilapia (Oreochromis niloticus) in vivo and in vitro using cyclophosphamide (CTX) as hepatotoxicant. In vitro, the precision cut liver slices (PCLS) were exposed to 6 concentrations of CTX (0, 5, 10, 15, 20 and 25 mmol·L-1) for 6 h, and the levels of glutamate oxalate transaminase (GOT), glutamate pyruvate transaminase (GPT) and lactate dehydrogenase (LDH) in slices culture supernatant were determined. At the same time, the total antioxidant capacity (T-AOC), superoxide dismutase (SOD), glutathione (GSH) and malondialdehyde (MDA) levels in slices and the survival rate of the PCLS were detected. In vivo, tilapias were injected with 0, 10, 25, 50, 75 and 100 mg·kg-1 CTX intrapleurally. It was ad-ministered for intrapleurally once every three days for three times. Samples were taken on the 4th day after the last administration. The GPT, GOT and LDH of serum and T-AOC, SOD, GSH and MDA of liver tissue ho-mogenates were detected. The results showed that both in the PCLS treated with CTX at 20 - 25 mmol·L-1 for 6 h and the tilapias injected with 75- 100 mg·kg-1 CTX, the activities of GPT, GOT and LDH were significantly increased, the MDA production were increased, while the activities of T-AOC, SOD and the GSH content were decreased. Meanwhile, 20 and 25 mmol·L-1)
Abstract:
Key words:  cyclophosphamide  Oreochromis niloticus  hepatic injury model  in vivo and in vitro

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