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摘要：
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摘要：目的 观察下瘀血汤对酒精性肝病（alcoholic liver disease，ALD）小鼠肝脏炎症
和脂肪变性的改善作用，初步探讨其可能的作用机制。方法 将48只雄性C57BL/6小鼠
随机分为对照组、对照美他多辛组、对照下瘀血汤组、酒精组、酒精美他多辛组和酒
精下瘀血汤组，每组各8只。对照组、对照美他多辛组和对照下瘀血汤组造模过程中全
程给予液体对照饲料。酒精组、酒精美他多辛组和酒精下瘀血汤组采用4周慢性乙醇喂
养加急性乙醇灌胃法造模，造模第3周第1 d起，对照下瘀血汤组和酒精下瘀血汤组小鼠
以0.4678 g/kg下瘀血汤灌胃，对照美他多辛组和酒精美他多辛组小鼠以2.857 mg/kg美
他多辛灌胃。其余组小鼠以等体积蒸馏水灌胃。第32 d酒精组、酒精美他多辛组和酒精
下瘀血汤组小鼠以31.5%乙醇灌胃，对照组、对照美他多辛组、对照下瘀血汤组小鼠
以45%糊精灌胃。9 h后处死小鼠，留取静脉血和肝脏。计算各组小鼠肝体比值，检测
丙氨酸氨基转移酶（alanine aminotransferase，ALT）、血清甘油三酯（triglyceride，
TG）及肝脏TG水平。采用HE染色和油红染色观察肝脏病理形态学变化。采用免疫组
织化学法检测中性粒细胞标志物髓过氧化物酶（myeloperoxidase，MPO）的表达。
采用实时荧光定量聚合酶链式反应（real time polymerase chain reaction，RT-PCR）
检测肝组织白细胞介素-6（interleukin 6，IL-6）、白细胞介素1β（interleukin 1β，IL1β）和单核细胞趋化蛋白1（monocyte chemotactic protein 1，MCP1）mRNA的相对表
达量。采用Western blot和RT-PCR检测脂肪酸合成酶（fatty acid synthase，FAS）、肉
毒碱棕榈酰基转移酶-1α（carnitine palmitoyltransferase 1α，CPT-1α）和过氧化物酶体
增殖激活受体α（peroxisome proliferator-activated receptor alpha，PPARα）的表达。
结果 与对照组相比，酒精组小鼠肝体比值[（4.78 ± 0.48）% vs（3.71 ± 0.36）%]、
ALT [（44.71 ± 25.37）U/L vs（20.41 ± 7.11）U/L]、血清TG [（4.16 ± 1.27）mmol/L
vs（1.44 ± 0.23）mmol/L]和肝脏TG [（27.15 ± 6.43）mmol/g vs（10.74 ± 9.83）mmol/g]
均显著升高（P均＜ 0.05）。与酒精组相比，酒精美他多辛组和酒精下瘀血汤组小鼠ALT
[11.79（10.17，24.48）U/L vs（16.76 ± 1.64）U/L vs（44.71 ± 25.37）U/L]、血清TG
[（1.89 ± 1.54）mmol/L vs 2.40（2.39，2.67）mmol/L vs（4.16 ± 1.27）mmol/L]及肝组织
TG [（14.18 ± 5.88）mmol/g vs 19.77（5.92，20.90）mmol/g vs（27.15 ± 6.43）mmol/g]水平
均显著降低（P均＜ 0.05），肝体比值[（4.49 ± 0.43）% vs（4.82 ± 0.14）% vs（4.78 ±
0.48）%]差异无统计学意义（t值分别为1.099、-0.165，P值分别为0.283、0.871）。HE
和油红染色提示酒精组小鼠肝组织脂肪变性明显。免疫组织化学结果表明酒精组小鼠
肝脏MPO阳染较对照组显著增加。与对照组相比，酒精组小鼠IL-6（1.95 ± 0.74 vs 1.00
± 0.47）、IL-1β（2.06 ± 0.64 vs 1.00 ± 0.26）和MCP1（2.98 ± 1.13 vs 0.99 ± 0.30）及
FAS [2.40 ± 0.53 vs 0.916（0.876，1.221）] mRNA相对表达量均显著升高（z = -2.242，
P = 0.025；z = -3.695，P ＜ 0.001；z = -2.867，P = 0.004；z = -3.838，P ＜ 0.001），
CPT-1α（0.39 ± 0.75 vs 1.00 ± 0.22）和PPARα（0.27 ± 0.08 vs 1.00 ± 0.26）mRNA相对表
达量显著降低（z = -4.392，P ＜ 0.001；z = -4.392，P ＜ 0.001）；与酒精组相比，酒
精美他多辛组和酒精下瘀血汤组IL-6（1.16 ± 0.42 vs 0.93 ± 0.42 vs 1.95 ± 0.74）、IL-1β
（0.75 ± 0.19 vs 0.59 ± 0.07 vs 2.06 ± 0.64）、MCP1（1.27 ± 0.25 vs 1.23 ± 0.50 vs 2.98 ±
1.13）及FAS（1.41 ± 1.05 vs 1.43 ± 0.30 vs 2.40 ± 0.53）mRNA相对表达量均显著降低，
CPT-1α [0.81（0.79，0.81）vs 0.72 ± 0.14 vs 0.39 ± 0.75]和PPARα（0.63 ± 0.30 vs 0.69 ±
0.41 vs 0.27 ± 0.08）mRNA相对表达量显著升高（P均＜ 0.05）。Western blot表明，与
对照组相比，酒精组小鼠FAS蛋白相对表达量（0.56 ± 0.07 vs 0.20 ± 0.02）上调（z =
-2.309，P = 0.021），CPT-1α蛋白相对表达量（0.24 ± 0.02 vs 1.03 ± 0.06）和PPARα蛋
白相对表达量（0.17 ± 0.01 vs 1.02 ± 0.08）下调（P均＜ 0.05）；与酒精组相比，酒精
美他多辛组和酒精下瘀血汤组FAS蛋白相对表达量（0.31 ± 0.02 vs 0.29 ± 0.04 vs 0.56 ±
0.07）均显著降低，CPT-1α蛋白相对表达量（0.43 ± 0.01 vs 0.65 ± 0.10 vs 0.24 ± 0.02）
和PPARα（0.55 ± 0.07 vs 0.39 ± 0.04 vs 0.17 ± 0.01）显著升高（P均＜ 0.05）。结论 下
瘀血汤可通过抑制中性粒细胞浸润及炎症反应减轻酒精引起的脂肪生成并促进脂肪酸
氧化，从而发挥保护肝脏的作用。
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Abstract: Objective To observe the effects of Xia-yu-xue decoction on improving liver
inflammation and steatosis in mice with alcoholic liver disease (ALD) and to investigate its
possible mechanism. Methods A total of 48 male C57/BL6 mice were randomly divided into
blank control group, control metadoxine group, control Xia-yu-xue decoction group, alcohol
group, alcohol metadoxine group and alcohol Xia-yu-xue decoction group, 8 mice in each
group. Mice in control group, control metadoxine group and control Xia-yu-xue decoction
group were given liquid control feed throughout the process of modeling. Mice in alcohol
group, alcohol metadoxine group and alcohol Xia-yu-xue decoction group were fed with
ethanol liquid diet for 4 weeks and gavage with alcohol. From the 1st day of the 3rd week,
mice in control Xia-yu-xue decoction group and the alcohol Xia-yu-xue decoction group were
given 0.4678 g/kg of Xia-yu-xue decoction by gavage, mice in control metadoxine group
and alcohol metadoxine group were given 2.857 mg/kg of metadoxine by gavage. Mice in
other groups were given equal volume of distilled water by gavage. On the 32nd day, mice
in alcohol group, alcohol metadoxine group and alcohol Xia-yu-xue decoction group were
given 31.5% ethanol by gavage, while mice in control group, control metadoxine group and
control Xia-yu-xue decoction group were given 45% dextrin. Then the mice were sacrificed
9 hours later, and venous blood and liver were collected. The ratio of liver to body weight
was calculated and levels of serum alanine aminotransferase (ALT), triglyceride (TG) and
liver TG contents were detected. The pathological changes of liver were observed by HE
staining and oil red staining. Immunohistochemical method was used to detect the expression
of myeloperoxidase (MPO), which was a marker of neutrophils in liver tissue. Realtime polymerase chain reaction (RT-PCR) was used to detect mRNA relative expression levels of interleukin 6 (IL-6), interleukin 1β (IL-1β) and monocytes monocyte chemotactic
protein 1 (MCP1) in liver tissue. The expression of fatty acid synthase (FAS), carnitine
palmitoyltransferase 1α (CPT-1α) and peroxisome proliferator-activated receptor alpha
(PPARα) were detected by RT-PCR and Western blot. Results Compared with those of
control group, the ratio of liver to body weight [(4.78 ± 0.48)% vs (3.71 ± 0.36)%], the
levels of serum ALT [(44.71 ± 25.37) U/L vs (20.41 ± 7.11) U/L], TG [(4.16 ± 1.27) mmol/L
vs (1.44 ± 0.23) mmol/L] and liver TG [(27.15 ± 6.43) mmol/g vs (10.74 ± 9.83) mmol/g]
of mice in alcohol group increased significantly (all P ＜ 0.05). Compared with those of
alcohol group, the serum ALT [11.79 (10.17, 24.48) U/L vs (16.76 ± 1.64) U/L vs (44.71 ±
25.37) U/L], serum TG [(1.89 ± 1.54) mmol/L vs 2.40 (2.39, 2.67) mmol/L vs (4.16 ± 1.27) mmol/L]
and liver TG [(14.18 ± 5.88) mmol/g vs 19.77 (5.92, 20.90) mmol/g vs (27.15 ± 6.43) mmol/g]
levels of mice in alcohol metadoxine group and alcohol Xia-yu-xue decoction group decreased
significantly (all P ＜ 0.05); there were no significant differences in the ratio of liver to body
weight of mice in the three groups [(4.49 ± 0.43)% vs (4.82 ± 0.14)% vs (4.78 ± 0.48)%；
t = 1.099, -0.165, P = 0.283, 0.871]. HE staining and oil red staining showed that liver fatty
degeneration of mice in alcohol group was obvious. The immunohistochemical results showed
that the positive staining of MPO of mice in alcohol group was significantly higher than that
of control group. Compared with those of control group, mRNA relative expression of IL-6
(1.95 ± 0.74 vs 1.00 ± 0.47), IL-1β (2.06 ± 0.64 vs 1.00 ± 0.26), MCP1 (2.98 ± 1.13 vs 0.99 ± 0.30) and
FAS [2.40 ± 0.53 vs 0.916 (0.876，1.221)] of mice in alcohol group increased significantly
(z = -2.242, P = 0.025; z = -3.695, P ＜ 0.001; z = -2.867, P = 0.004; z = -3.838, P ＜ 0.001),
mRNA relative expression of CPT-1α (0.39 ± 0.75 vs 1.00 ± 0.22) and PPARα (0.27 ± 0.08 vs
1.00 ± 0.26) decreased significantly (z = -4.392, P ＜ 0.001; z = -4.392, P ＜ 0.001). Compared
with those of alcohol group, mRNA relative expression of IL-6 (1.16 ± 0.42 vs 0.93 ± 0.42 vs
1.95 ± 0.74), IL-1β (0.75 ± 0.19 vs 0.59 ± 0.07 vs 2.06 ± 0.64), MCP1 (1.27 ± 0.25 vs 1.23 ±
0.50 vs 2.98 ± 1.13) and FAS (1.41 ± 1.05 vs 1.43 ± 0.30 vs 2.40 ± 0.53) of mice in alcohol
metadoxine group and alcohol Xia-yu-xue decoction group decreased significantly, mRNA
relative expression of CPT-1α [0.81 (0.79, 0.81) vs 0.72 ± 0.14 vs 0.39 ± 0.75] and PPARα
(0.63 ± 0.30 vs 0.69 ± 0.41 vs 0.27 ± 0.08) increased significantly (all P ＜ 0.05). Western
blot showed that the FAS protein relative expression up-regulated significantly (0.56 ± 0.07
vs 0.20 ± 0.02; z = -2.309, P = 0.021), whereas CPT-1 protein relative expression (0.24 ± 0.02
vs 1.03 ± 0.06) and PPARα protein relative expression (0.17 ± 0.01 vs 1.02 ± 0.08) decreased
significantly of mice in alcohol group compared with those in control group (all P ＜ 0.05).
Compared with those of alcohol group, FAS protein relative expression of mice in alcohol
metadoxine group and alcohol Xia-yu-xue decoction group decreased significantly (0.31 ± 0.02
vs 0.29 ± 0.04 vs 0.56 ± 0.07), protein relative expression of CPT-1α (0.43 ±0.01 vs 0.65 ± 0.10
vs 0.24 ± 0.02) and PPARα (0.55 ± 0.07 vs 0.39 ± 0.04 vs 0.17 ± 0.01) increased significantly
(all P ＜ 0.05). Conclusions Xia-yu-xue decoction can reduce alcohol-induced lipogenesis
and promote fatty acid oxidation by inhibiting neutrophil infiltration and inflammatory
reaction, thus playing a protective role in liver.
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