四氯化碳诱导C57BL/6J小鼠肝纤维化模型建立方法及优化
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1.辽宁中医药大学附属医院;2.中国中医科学院广安门医院;3.中国人民武装警察部队海警总队医院

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沈阳市科技局公共卫生研发专项(21-173-9-37)


Establishment and optimization of C57BL/6J mice liver fibrosis model induced by carbon tetrachloride
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1.Affiliated Hospital of Liaoning University of Traditional Chinese Medicine;2.Guang '3.'4.anmen Hospital, China Academy of Chinese Medical Sciences;5.Marine Police Corps Hospital of Chinese People'6.s Armed Police Force

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Public Health Research and Development Project, Shenyang Science and Technology Bureau(21-173-9-37)

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    摘要:

    目的 通过不同给药方式不同剂量四氯化碳诱导的C57BL/6J小鼠肝纤维化模型,通过影像学、分子生物学及组织病理学等方法进行比较,以此优化四氯化碳诱导的小鼠肝纤维化模型。方法 36只健康C57BL/6J雄性小鼠,适应性饲养1周后,随机分为空白组、2周组、3周组、4周组、6周组及8周组共6组(n=6)。空白组以橄榄油0.2 mL腹腔注射,每周3次,各阳性对照组以以20% CCl4-橄榄油溶液0.2 mL腹腔注射,每周3次。记录各组小鼠体重变化情况。分别于第15天、22天、29天、43天及57天测量小鼠肝脏弹性值后取血,分别测量小鼠谷丙转氨酶(ALT)、天门冬氨酸氨基转移酶(AST)、透明质酸(HA)、层黏蛋白(LN)、前Ⅲ型前胶原(PC-Ⅲ)及Ⅳ型胶原(Ⅳ-C)含量,对肝组织进行HE、Masson及天狼星红染色,Metavir评分系统评估肝纤维化程度。结果 与空白组相比,各阳性对照组小鼠精神萎靡,扎堆喜卧。在小鼠体质量方面,4周组、6周组、8周组与空白组相比均明显下降,而2周组与空白组相比体明显升高。肝脏弹性超声显示,随给药时间增加,弹性值呈进行性升高。生化结果显示,与空白组相比较,各阳性对照组ALT、AST水平均显著升高,随给药时间增加,各阳性对照组HA、LN、PC-Ⅲ及Ⅳ-C水平呈升高趋势。病理结果显示,随着给药时间增加,小鼠肝脏纤维化程度呈进行性加重,4周时符合肝纤维化病理诊断,6周时有假小叶形成趋势,而8周时已形成假小叶,提示早期肝硬化。结论 20%CCl4-橄榄油溶液以每周三次、连续4周腹腔注射的方法能成功建立C57BL/6J小鼠肝纤维化模型,稳定性好、成模较快,可以作为肝纤维化模型制造的优化方案。

    Abstract:

    Objective To optimize C57BL/6J mouse liver fibrosis model induced by different doses of carbon tetrachloride through imaging, molecular biology and pathology methods.Methods Thirty-six healthy C57BL/6J male mice were randomly divided into control group, 2 weeks group, 3 weeks group, 4 weeks group, 6 weeks group and 8 weeks group (n=6) after adaptive feeding for 1 week. The control group was intraperitoneally injected with 0.2 mL olive oil injection three times a week, and the positive control groups were intraperitoneally injected with 0.2 mL 20% CCl4-olive oil solution three times a week. The changes in body weight of mice in each group were recorded. Liver stiffness was measured on days 15, 22, 29, 43 and 57, and blood samples were collected, the mice were measured cereal third transaminase (ALT), aspartate aminotransferase (AST), hyaluronic acid (HA), layer mucins (LN), type Ⅲ before collagen (PC-Ⅲ) and collagen type Ⅳ (Ⅳ-C) content, The liver tissues were stained with HE, Masson and Sirius red.The Metavir scoring system was used to evaluate the degree of liver fibrosis.Results Compared with the control group, the mice in the positive control groups were listless and clustered together. In terms of body weight, the 4-week group, 6-week group and 8-week group were significantly lower than the control group, while the 2-week group was significantly higher than the control group. Liver elastography showed a progressive increase in stiffness with the increase of administration time. Biochemical results show that, compared with the control group, the other groups ALT, AST levels were significantly higher, with the increase of drug delivery time, the positive control group HA, LN, PC-Ⅲ and Ⅳ-C levels showed a trend of rise. The pathological results showed that the degree of liver fibrosis was progressively aggravated with the increase of administration time. At 4 weeks, the pathological diagnosis of liver fibrosis was consistent with that of liver fibrosis, there was a trend of pseudolobules formation at 6 weeks, and pseudolobules were formed at 8 weeks, suggesting early cirrhosis.Conclusions The liver fibrosis model of C57BL/6J mice can be successfully established by intraperitoneal injection of 20% CCl4-olive oil solution three times a week for 4 consecutive weeks, which has good stability and rapid modeling. It can be used as an optimized scheme for the establishment of liver fibrosis model.

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  • 收稿日期:2023-12-05
  • 最后修改日期:2024-04-23
  • 录用日期:2024-05-08
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