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摘要：
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摘要：目的 探讨急性肝衰竭大鼠血清激活蛋白C（activated protein C，APC）的变化及其意义。方法
本研究实验动物为健康普通级雄性6周龄Sprague-Dawley（SD）大鼠，体质量150～180 g。实验分为两
部分，先将30只SD大鼠采用随机数字表法分为对照组、模型组及干预组，每组10只，观察各组大鼠一
般状况，计算各组生存率。然后再将另外55只SD大鼠采用随机数据表法分为对照组（5只）、模型组
（25只）和干预组（25只）。所有大鼠适应性饲养3 d，禁食12 h后，模型组和干预组大鼠腹腔注射D-
氨基半乳糖（剂量为1000 mg/kg）和脂多糖（剂量为30 μg/kg），建立急性肝衰竭大鼠模型，正常对照
组腹腔注射等量0.9%氯化钠溶液（5 ml/kg）。干预组于造模后10 min尾静脉注射10 μg/ml的APC溶液
（剂量为50 μg/kg），模型组和对照组尾静脉注射等量0.9%氯化钠溶液（5 ml/kg）。模型组和干预组
分别于造模后1 h、2 h、4 h、9 h、12 h腹腔注射10%水合氯醛（0.3 ml/100 g）麻醉处死，每个时间点
处死5只。预实验提示对照组大鼠在等量生理盐水处理前后无明显变化，故于处理后统一时间点处死
5只。检测各组大鼠不同时间点血清丙氨酸氨基转移酶（alanine aminotransferase，ALT）、天门冬氨
酸氨基转移酶（aspertate aminotranferase，AST）、APC和肿瘤坏死因子-α（tumor necrosis factor-α，
TNF-α）水平。光学显微镜下观察各时间点大鼠肝组织病理学变化。采用Pearson相关性分析血清
TNF-α与APC水平的相关性。结果 大鼠肝组织HE染色示模型组和干预组大鼠肝细胞均出现坏死、出
血和炎细胞浸润，并随时间加重，但同一时间点干预组大鼠肝细胞损伤较模型组轻。因前期预实验提
示对照组大鼠各指标在处理前后无明显变化，根据动物实验“3R”原则中的减少原则，将对照组大鼠
各指标的数据视为模型组和干预组大鼠的基线数据。模型组大鼠的生存率为20%（2/10），干预组大鼠
的生存率为40%（4/10）。模型组和干预组基线[ALT：（45.6 ± 7.1）U/L vs（45.6 ± 7.1）U/L，AST：
（107.8 ± 27.2）U/L vs（107.8 ± 27.2）U/L]、1 h [ALT：（48.2 ± 5.9）U/L vs（47.4 ± 6.2）U/L，AST：
（141.0 ± 44.8）U/L vs（134.0 ± 34.9）U/L]和2 h [ALT：（59.8 ± 10.5）U/L vs（53.6 ± 9.6）U/L，AST：
（144.0 ± 39.7）U/L vs（163.2 ± 33.4）U/L]血清ALT和AST水平差异无统计学意义（P均＞ 0.05），
4 h [ALT：（546.6 ± 287.9）U/L vs（310.0 ± 153.5）U/L，AST：（1075.0 ± 840.2）U/L vs（437.4 ±
171.7）U/L]、9 h [ALT：（929.6 ± 630.6）U/L vs（565.4 ± 289.1）U/L，AST：（3078.0 ± 2044.1）U/L
vs（1003.2 ± 452.5）U/L]和12 h [ALT：（528.6 ± 221.6）U/L vs（306.0 ± 146.2）U/L，AST：（1105.0 ±
464.1）U/L vs（518.2 ± 262.1）U/L] ALT和AST水平差异有统计学意义（P均＜ 0.05），4 h、9 h和12 h
干预组ALT和AST水平显著低于模型组（P均＜ 0.05）。建模后1 h，模型组大鼠APC水平迅速下降至
（32.242 ± 2.649）ng/L，与其他时间点相比，差异有统计学意义（P均＜ 0.05），建模后2 h进一步下
降至（23.482 ± 3.033）ng/L，此后APC维持在相对稳定水平[4 h：（24.340 ± 3.367）ng/L，9 h：（19.992 ±
3.238）ng/L，12 h：（22.100 ± 3.950）ng/L]。建模后模型组所有时间点APC水平均低于基线[（99.015 ±
11.543）ng/L]，差异有统计学意义（P均＜ 0.05）。干预组APC较模型组有升高趋势，干预组1 h
[（61.137 ± 6.088）ng/L]和12 h [（27.743 ± 2.623）ng/L] APC水平与模型组差异有统计学意义（P
均＜ 0.05）。建模后模型组和干预组大鼠血清TNF-α水平逐渐升高，均于9 h时达峰值[（177.190 ±
78.473）ng/L、（170.475 ± 75.353）ng/L]。干预组1 h [（24.177 ± 5.037）ng/L vs（57.012 ± 6.231）ng/L]、4 h
[（27.455 ± 6.698）ng/L vs（79.533 ± 5.651）ng/L]和12 h [（53.785 ± 11.501）ng/L vs（89.295 ±
4.188）ng/L] TNF-α水平显著低于模型组（P均＜ 0.001）。模型组大鼠血清APC水平与TNF-α水平呈
负相关（r = -0.5364，P = 0.0013）。结论 APC可减轻急性肝衰竭大鼠的肝细胞损伤，对肝细胞有保护
作用，其机制可能与APC能够抑制TNF-α水平有关。
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Abstract: Objective To investigate the changes and effects of serum activated protein C (APC) in rats with
acute liver failure. Methods The experimental animals were 6-week-old healthy male Sprague-Dawley
(SD) rats with body mass of 150~180 g. A total of 30 SD rats were divided into control group, model group
and intervention group by random digital table method, 10 rats in each group. The survival rates of each
group were calculated. Another 55 rats were also divided into control group (5 rats), model group (25 rats)
and intervention group (25 rats) by random digital table method. All rats were adapted feeding for 3 days.
D-galactose (1000 mg/kg) and lipopolysaccharide (30 μg/kg) were injected intraperitoneally into rats in
model group and intervention group after 12 h of fasting to establish the rat model of acute liver failure. Rats
in control group were intraperitoneally injected with equal amount of 0.9% sodium chloride solution (5 ml/kg). Rats in
intervention group were given 10 μg/ml of APC solution (50 μg/kg) 10 min after modeling by tail intravenous
injection, and rats in model group and control group were given 0.9% sodium chloride solution (5 ml/kg). Rats
in model group and intervention group were injected with 10% chloral hydrate (0.3 ml/100 g) intraperitoneally
and sacrificed at 1 h, 2h, 4h, 9h, 12h after modeling, 5 rats at each time point. The pre-experiment showed
that there were no obvious changes before and after the same amount of normal saline treatment in the control
group, so 5 rats were sacrificed uniformly after treatment. Serum levels of alanine aminotransferase (ALT),
aspartate aminotransferase (AST), APC and tumor necrosis factor-α (TNF-α) were measured at different
time points in each group. The histopathological changes of liver were observed by optical microscope.
Pearson correlation analysis was used to analyze the correlation of serim TNF-α and APC. Results HE
staining showed that necrosis, hemorrhage and inflammatory cell infiltration occurred in both model group
and intervention group, the hepatic cell injury in intervention group was lighter than that in model group
at the same time. As the pre-experiment showed that there were no obvious changes in the indexes of rats
in the control group before and after treatment, according to the reduction in the “3R” principle of animal
experiment, the data of each index in the control group were regarded as the baseline data of the model group
and the intervention group. The survival rate of rats in model group and intervention group were 20% (2/10)
and 40% (4/10), respectively. There were no significant differences in serum ALT and AST levels of rats in
model group and the intervention group at baseline [ALT: (45.6 ± 7.1) U/L vs (45.6 ± 7.1) U/L, AST: (107.8 ±
27.2) U/L vs (107.8 ± 27.2) U/L], 1 h [ALT: (48.2 ± 5.9) U/L vs (47.4 ± 6.2) U/L, AST: (141.0 ± 44.8) U/L vs
(134.0 ± 34.9) U/L] and 2 h [ALT:(59.8 ± 10.5) U/L vs (53.6 ± 9.6) U/L, AST: (144.0 ± 39.7) U/L vs (163.2 ±
33.4) U/L] (P ＞ 0.05). There were significant differences in ALT and AST levels of rats in model group and
the intervention group at 4 h [ALT: (546.6 ± 287.9) U/L vs (310.0 ± 153.5) U/L, AST: (1075.0 ± 840.2) U/L
vs (437.4 ± 171.7) U/L], 9 h [ALT:(929.6 ± 630.6) U/L vs (565.4 ± 289.1) U/L, AST: (3078.0 ± 2044.1) U/L
vs (1003.2 ± 452.5) U/L] and 12 h [ALT: (528.6 ± 221.6) U/L vs (306.0 ± 146.2) U/L, AST: (1105.0 ± 464.1) U/L vs
(518.2 ± 262.1) U/L] (P ＜ 0.05). The serum ALT and AST levels of rats in intervention group were
significantly lower than those in model group at 4 h, 9 h and 12 h (P ＜ 0.05). The APC level of rats in
model group rapidly decreased to (32.242 ± 2.649) ng/L 1 h after modeled, which was significantly lower
than those at other time points (all P ＜ 0.001). The serum APC level of rats in model group furtherly decreased to
(23.482 ± 3.033) ng/L 2 h after modeled, and then the levels of APC remained relatively stable [4 h: (24.340 ±
3.367) ng/L, 9 h: (19.992 ± 3.238) ng/L, 12 h: (22.100 ± 3.950) ng/L]. After modeling, the APC levels
of rats in model group at all time points were lower than those of the baseline [(99.015 ± 11.543) ng/L], the
differences were statistically significant (all P ＜ 0.05). Compared with those in model group, the APC level
of rats in intervention group had an increased trend. The APC levels of rats in intervention group at 1 h
[(61.137 ± 6.088) ng/L] and 12 h [(27.743 ± 2.623) ng/L] were statistically significant compared with those
in model group. The serum TNF-α levels of rats in model group and intervention group increased gradually
after modeling, and reached the peak value at 9 h [(177.190 ± 78.473) ng/L, (170.475 ± 75.353) ng/L].
The serum TNF-α levels of rats in intervention group at 1 h [(24.177 ± 5.037) ng/L vs (57.012 ± 6.231) ng/L],
4 h [(27.455 ± 6.698) ng/L vs (79.533 ± 5.651) ng/L] and 12 h [(53.785 ± 11.501) ng/L vs (89.295 ± 4.188) ng/L] were
significantly lower than those in model group (P ＜ 0.001). There was a negative correlation between
serum APC and TNF-α of ratsin model group (r = -0.5364, P = 0.0013). Conclusions APC can reduce the
hepatocyte damage in rats with acute liver failure and has a protective effect on hepatocytes. The mechanism
may be related to the inhibition of TNF-α.
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