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Journal of Chinese Integrative Medicine ›› 2009, Vol. 7 ›› Issue (7): 642-650.doi: 10.3736/jcim20090708

• Original Experimental Research • Previous Articles     Next Articles

Effects of Qinggan Huoxue Recipe and its separated recipes on urokinase-type plasminogen activator and plasminogen activator inhibitor-1 fibrinolytic system in rats with alcoholic liver fibrosis

 Jun-ming Chen, Lei Wang, Tao Liu, Lian-jun Xing, Pei-yong Zheng, Guang Ji   

  1. Institute of Digestive Diseases and Department of Gastroenterology, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China
  • Received:2009-04-13 Accepted:2009-05-26 Online:2009-07-20 Published:2009-07-15
  • Contact: Guang Ji E-mail:jiliver@vip.sina.com

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Objective

To study the action mechanisms of Qinggan Huoxue Recipe (QGHXR), a compound traditional Chinese herbal medicine, and its separated recipes by observing their effects on expressions of urokinase-type plasminogen activator (uPA) and plasminogen activator inhibitor-1 (PAI-1) in rats with alcoholic liver fibrosis (ALF).
Methods

A total of 150 SD rats were divided into three groups: blank group, carbon tetrachloride (CCl4) group and ALF-inducing group. Rats in the ALF-inducing group were administered with a mixture diet (56% alcohol 10 mL/kg, corn oil 2 mL/kg, pyrazole 25 mg/kg) once daily and intraperitoneally injected with 0.3 mL/kg 25% solution of CCl4 in olive oil twice weekly. The CCl4 group was intraperitoneally injected with equal volume of CCl4 and olive oil as the ALF-inducing group and ingested normal saline (12 mL/kg per day). The blank group was intraperitoneally injected with and ingested saline in equal volumes of the above. At the end of the eighth week, the survived rats in the ALF-inducing group were divided into four subgroups: untreated group, QGHXR group, Qinggan Recipe (QGR) group and Huoxue Recipe (HXR) group. The three treated groups were given corresponding drugs respectively (4.75, 1.5, 3.25 g/kg). The blank group, CCl4 group and untreated group were given normal saline in equal volume (5 mL/kg per day). All rats were anaesthetized and killed at the end of the twelfth week. The activities of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in the serum were analyzed. Pathological changes in liver tissues were observed by hematoxylin and eosin staining and Masson staining. The expressions of uPA and PAI-1 were evaluated with Western blotting, immunofluorescence, and real-time polymerase chain reaction.
Results

There existed obvious liver fibrosis in liver tissues in the untreated group as compared with the blank group (P<0.01), and the activities of ALT and AST and the expressions of uPA and PAI-1 also increased in the untreated group. QGHXR and its separated recipes could improve the degree of liver fibrosis (P<0.01). QGHXR and its separated recipes could degrade the activity of ALT as compared with the untreated group; QGHXR and its separated recipes advanced the expression of uPA, and decreased the expression of PAI-1 significantly as compared with the untreated group. The effect of QGHXR was the best among the three recipes.
Conclusion

QGHXR and its separated recipes may improve ALF in rats by decreasing the expression of PAI-1 and advance the expression of uPA. The effect of QGHXR is the best among them.

Key words: Liver cirrhosis, Alcoholic, Qinggan Huoxue Recipe, Separated recipes, Urinary plasminogen activator, Plasminogen activator inhibitor-1, Rats

Table 1

Primer subsequences of PAI-1, uPA and GAPDH"

Target gene Primer subsequence Size of product (bp)
PAI-1 Forward: 5'-GTGGTTCGGCACAATCCAACAGAG-3' 441
Reverse: 5'-GCAATGGAGGACGATACAAGAGGT-3'
uPA Forward: 5'-AAGGTCTGGGATACTAGAGCC-3' 144
Reverse: 5'-CAGGATATCCTCAGTCTCCACC-3'
GAPDH Forward: 5'-GGTCGGAGTCAACGGATTTG-3' 320
Reverse: 5'-ATGAGCCCCCAGCCTTCTCCA-3'

Table 2

Effects of QGHXR and its separated recipes on activities of ALT and AST in rats with ALF ($\bar{x}$±s, U/L)"

Group n ALT AST
Blank 10 33.90±9.81 119.57±28.63
CCl4 10 27.58±3.03 116.90±20.47
Untreated 12 138.34±43.35** 257.41±162.31**
QGR 12 41.95±26.78△△ 102.74±23.55△
HXR 12 46.63±21.00△△ 111.50±21.26△
QGHXR 12 37.57±27.85△△ 83.72±33.81△△

Figure 1

Pathological changes of liver tissues in different groups observed by HE staining (Light microscopy, ×400) A: Blank group; B: CCl4 group; C: Untreated group; D: QGR group; E: HXR group; F: QGHXR group."

Figure 2

Liver fibrosis in different groups observed by Masson staining (Light microscopy, ×200) A: Blank group; B: CCl4 group; C: Untreated group; D: QGR group; E: HXR group; F: QGHXR group."

Table 3

Effects of QGHXR and its separated recipes on degree of liver fibrosis of rats with ALF"

Group n Number of rats with different
degrees of liver fibrosis
– + ++ +++
Blank 10 10 0 0 0
CCl4 10 7 3 0 0
Untreated 12 0 3 5 4
QGR 12 0 8 3 1
HXR 12 0 7 3 2
QGHXR 12 1 8 2 1

Figure 3

Effects of QGHXR and its separated recipes on protein expressions of uPA and PAI-1 in liver tissues of rats with ALF A: Blank group; B: CCl4 group; C: Untreated group; D: QGR group; E: HXR group; F: QGHXR group."

Table 4

Effects of QGHXR and its separated recipes on expressions of uPA and PAI-1 proteins in liver tissues of rats with ALF ($\bar{x}$±s)"

Group n uPA PAI-1
Blank 10 0.15±0.01 0.30±0.05
CCl4 10 0.17±0.01 0.36±0.08
Untreated 12 0.31±0.16** 1.36±0.15**
QGR 12 0.98±0.09△▲▲ 0.76±0.12△▲
HXR 12 0.75±0.06△▲▲ 0.87±0.18△▲▲
QGHXR 12 1.98±0.24△△ 0.41±0.06△△

Figure 4

Effects of QGHXR and its separated recipes on expressions of uPA and PAI-1 proteins in liver tissues of rats with ALF (Immumofluorescence method,×400) a: The green fluorescence denotes uPA protein positive expression; b: The red fluorescence denotes PAI-1 protein positive expression; c: The yellow fluorescence denotes uPA and PAI-1 protein positive expression."

Figure 5

Effects of QGHXR and its separated recipes on the expressions of uPA, PAI-1 mRNAs in the liver of rats with ALF *P<0.05, **P<0.01, vs blank group; △P<0.05, △△P<0.01, vs untreated group; ▲P<0.05, ▲▲P<0.01, vs QGHXR group. Data were represented as $\bar{x}$±s. The numbers of the rats were 10 in blank and CCl4 groups and 12 in the other groups."

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