復(fù)方黃芪多苷在抵抗大鼠急性肝損傷發(fā)揮保護(hù)作用外文翻譯原文

1、 BRIEF REPORTS Protective effect of fufanghuangqiduogan against acute liver injury in miceShuang-Ying Gui, Wei Wei, Hua Wang, Li Wu, Wu-Yi Sun, Cheng-Yi WuELSEVIERPO Box 2345, Beijing 100023, China World J Gastroenterol 2005;11(19:2984-2989 World Journal of Gastroenterology ISSN 1007-9327wjg 2005 Th

2、e WJG Press and Elsevier Inc. All rights reserved.Shuang-Ying Gui, Wei Wei, Hua Wang, Li Wu, Wu-Yi Sun Cheng-Yi Wu, Institute of Clinical Pharmacology, Anhui Medical University, Hefei 230032, Anhui Province, ChinaShuang-Ying Gui, Department of Pharmacy, Anhui College of TCM, Hefei 230031, Anhui Prov

3、ince, ChinaSupported by the State High Technology Research and Development Program of China (863 Program, No. 2002AA2Z3235Correspondence to: Professor Wei Wei, Institute of Clinical Pharmacology, Anhui Medical University, Hefei 230032, Anhui Province, China. wweiTelephone: +86-551-5161208 Fax: +86-5

4、51-5161208Received: 2004-07-19 Accepted: 2004-09-04AbstractAIM: To study the effects and possible mechanisms of fufanghuangqiduogan (FFHQ in mice with acute liver injury (ALI.METHODS: ALI was successfully induced by injecting carbon tetrachloride (CCl 4 intraperitoneally and by tail vein injection o

5、f Bacillus Calmette Guerin (BCG and lipopolysaccharide (LPS in mice, respectively. Each of the two model groups was divided into normal group, model group, FFHQ (60, 120 and 240 mg/kg treatment groups,and bifendate treatment group. At the end of the experiment,levels of alanine aminotransferase (ALT

6、 and aspartate aminotransferase (AST, content of malondialdehyde (MDA,activities of superoxide dismutase (SOD and glutathione peroxidase (GSH-px in liver homogenate were measured by biochemical methods. The activities of tumor necrosis factor- (TNF- and interleukin-1 (IL-1 were determined by radio-i

7、mmunoassay. Hepatic tissue sections were stained with hematoxylin and eosin and examined under a light microscope.RESULTS: In the two models of ALI, FFHQ (60, 120,240 mg/kg was found to significantly decrease the serum transaminase (ALT, AST activities. Meanwhile, FFHQ decreased MDA contents and upr

8、egulated the lower SOD and GSH-px levels in liver homogenate. Furthermore, in immunologic liver injury model, FFHQ decreased levels of TNF- and IL-1 in serum. Histologic examination showed that FFHQ could attenuate the area and extent of necrosis, reduce the immigration of inflammatory cells.CONCLUS

9、ION: FFHQ had protective effect on liver injury induced by either CCl 4 or BCG+LPS in mice, and its mecha-nisms were related to free radical scavenging, increasing SOD and GSH-px activities and inhibiting the production of proinflammatory mediators. 2005 The WJG Press and Elsevier Inc. All rights re

10、served.Key words: Fufanghuangqiduogan; Radix Paeonia Pall ;Radix Astragali ; Acute liver injuryGui SY, Wei W, Wang H, Wu L, Sun WY,Wu CY . Protective effect of fufanghuangqiduogan against acute liver injury in mice. World J Gastroenterol 2005; 11(19: 2984-2989INTRODUCTIONAcute liver injury (ALI is a

11、 co-operative consequence of endotoxemia, microcirculation dysfunction as well as inflammatory cells (such as macrophage, lymphocyte that release inflammatory mediators and cytokines (such as tumor necrosis factor- (TNF-, interleukin-1 (IL-1 when stimulated. ALI is mostly induced by viral hepatitis,

12、 alcoholism,iron overload, or drug toxicity. It has a very high morbidity and mortality. The treatment might be anti-inflammatory or antioxidant action. Many modern Western medicines have been used to remedy ALI, but strategies are difficult to achieve satisfied outcomes due to their side effects.Ho

13、wever, some traditional Chinese herbs (such as Radix Paeonia Pall , Radix Astragali, Radix Salviae Miltiorrhizae ,Cordycep sinensis , Ginkgo biloba , Picrorhiza scrophulariflora have been found to have particular advantages in therapeutic research of ALI and other liver disease for their definite ef

14、fectiveness, cheap prices and negligible side effects 1-6.Traditional Chinese medicine (TCM treatment is based on overall analysis of symptoms and signs, and the physical condition of the patient 7. Fufanghuangqiduogan (FFHQis an extract of prescription TCM consisting of Radix Astragali ,Radix Paeon

15、ia lactiflora , etc . The present study aims at exploring the effects of FFHQ on the prevention of immunologic ALI induced by Bacillus Calmette Guerin (BCG+ lipopolysaccharide (LPS in mice and chemical ALI induced by CCl 4 in mice,and the content of malondialdehyde (MDA and the activities of superox

16、ide dismutase (SOD and glutathione peroxidase (GSH-px in mice liver homogenate were determined in order to investigate its possible mechanisms.MA MATERIALS AND METHODS TERIALS AND METHODS Drugs and materialsCCl 4, purchased from Beijing Chemical Factory, was diluted to 0.1% in vegetable oil. LPS fro

17、m Escherichia coli was obtained from Sigma Chemical Co. (St. Louis, MO, USA.BCG was purchased from Institute of Shanghai Biological Products. FFHQ is an extract of traditional Chinese herbsconsisting of Radix Astragali , Radix Paeonia lactiflora and Radix Glycyrrhizae purchased from Anhui Heyitang P

18、harmacy,China. These herbs mixed up in specified ratio (1:4:0.8were boiled with water and extracted by alcohol: 95% alcohol in liquid of these herbs by the volume proportion of 2:1 was mixed and stored at 0-4 for 24 h, then the sediments were filtered and the suspension including the protein and amy

19、lum was finally heated at 90-95 to evaporate the remaining alcohol and to obtain yellow brown powders. FFHQ was mainly composed of the total glucosides of Paeony (TGP,the total astragalosides (TAS, the total flavonoids of astragalus (TFA, astragalus polysaccharides and so on. TGP and TAS, accounting

20、 for 59.3%, were main effective components of FFHQ. FFHQ was dissolved into 0.5% sodium carboxymethylcellulose (CMC-Na solutions before use.Commercial kits used for determining lipid peroxidation and SOD activity were obtained from the Jiancheng Institute of Biotechnology (Nanjing, China. Other chem

21、icals used in these experiments were of analytical grade from comm-ercial sources.AnimalsMale Kunming mice (202 g were obtained from the Animal Department of Anhui Medical University. Mice were maint-ained on 12-h light/dark cycles. Animals were allowed free access to food and water. All mice were f

22、asted for 16 h prior to blood/tissue sampling. All experiments were performed in accordance with the institutional ethical guideline.Establishment of chemical liver injury model 8-10A CCl 4 0.1% vegetable oil solution was injected intraperitoneally into each animal in a dose of 10 mL/kg body weight.

23、 All the mice were anesthetized with ether, then killed by cervical dislocation 16 h after CCl 4 injection and trunk blood was collected into heparinized tubes (50 U/mL and centrifuged (1 500 r/min, 10 min, 4 . Serum was aspirated and stored at -70 until assayed as described below. The liver was als

24、o removed and stored at -70 until use.Establishment of immunological liver injury model 11A 2.5 mg dose of BCG (viable bacilli suspended in 0.2 mL saline was injected via the tail vein into each animal, and 10 d later, injected with 7.5 g LPS dissolved in 0.2 mL saline.The mice were anesthetized wit

25、h ether, and then killed by cervical dislocation 16 h after LPS injection. The other method adopted in the case of pretreatment studies was the same as in CCl 4-induced liver injury mentioned above.Drug treatmentIn the two model experiments, the animals were equally divided into six groups randomly

26、which included normal,model control, FFHQ groups (three different doses and bifendate. The mice in FFHQ groups received daily doses of 60, 120 or 240 mg/kg b.w. of FFHQ using an 18-gauge stainless steel animal feeding needle for 10 d prior to LPS injection and for 7 d prior to CCl 4 injection, respe

27、ctively.Mice in normal and model control group in the two model experiments were fed only with the same volume of vehicle.Measurement of serum ALT , AST, TNF- and IL-1Serum alanine aminotransferase (ALT and aspartate aminot-ransferase (AST were determined using commercial kits produced by Jiancheng

28、Institute of Biotechnology (Nanjing,China. The activities of ALT and AST were expressed as an international unit (U/L. Serum TNF- and IL-1 were measured using commercial kits produced by Beijing Biotechnology Co., Ltd, and their levels were expressed as nanogram per milliliter.Measurement of MDA, SO

29、D and GSH-px in liver homogenate Liver was thawed, weighed and homogenized in Tris-HCl (5 mmol/L containing 2 mmol/L EDTA, pH 7.4. Homog- enates were centrifuged (1 000 r/min, 10 min, 4 and the supernatant was used immediately for the assays of MDA and SOD . MDA, SOD and GSH-px were determined follo

30、wing the instructions on the kit. In brief, MDA in liver tissue was determined by the thiobarbituric acid method.All samples were assayed in triplicates. The content of MDA was expressed as nanomole per gram liver tissue. The assay for total SOD was based on its ability to inhibit the oxidation of o

31、xyamine by the xanthine-xanthine oxidase system. The red product (nitrite produced by the oxidation of oxyamine had an absorbance at 550 nm. One unit (U of SOD activity was defined as the amount that reduced the absorbance at 550 nm by 50%. All samples were assayed in triplicates.Results were expres

32、sed as unit per gram liver tissue. GSH-px was measured by the DTNB method, and its content was expressed as unit per milligram protein.Histologic analysisFormalin-fixed specimens were embedded in paraffin and stained with hematoxylin and eosin for conventional morphologic evaluation. After decapitat

33、ion of rats, small liver specimens were placed in 100 mL/L formalin solution and processed routinely by embedding in paraffin. Tissue sections (4-5 m were stained with hematoxylin and eosin and examined under light microscope (Olympus, Japan.An experienced histologist who was unaware of the treatmen

34、t conditions made histologic assessments.Statistical analysisAll values were presented as meanSD. Statistical analysis of the data for multiple comparisons was performed by one-way analysis of variance followed by Duncans test. For a single comparison, the significance of differences between means w

35、as determined by Students t -test. A level of P 0.05was taken as statistically significant.RESUL RESULTSTS Effect of FFHQ on serum ALT, AST, TNF- and IL-1Activities of both serum AST and ALT, indices of hepatic cell damage, were significantly higher in BCG+LPS-induced group and CCl 4 group than in t

36、he control group in the two models. FFHQ (60, 120, 240 mg/kg b.w. significantly reduced the activities of serum AST and ALT. Levels of TNF- and IL-1 in serum were significantly higher in BCG+LPS-induced group than in the control group in the immunologic mice model. FFHQ (60, 120, 240 mg/kg b.w. sign

37、ificantly reduced the levels of serum TNF- and IL-1 in the immunologic mice model (Tables 1 and 2.Gui SY et al . Fufanghuangqiduogan on acute liver injury 2985Table 2 Effects of FFHQ on serum ALT and AST in chemical ALI in mice (n = 10, meanSDGroup Dose ALT AST (mg/kg b.w. (U/L (U/LNormal- 26.48.2 3

38、4.19.3 Model-222.435.9d231.840.0d FFHQ 60181.630.7a182.035.8a 120153.134.8b161.239.8b240144.746.5b153.530.9b Bifendate 100103.529.8b139.331.6b d P0.01 vs normal group; a P0.05, b P0.01 vs model group.Effect of FFHQ on liver homogenate MDA, total SOD and GSH-px Liver homogenate MDA content in BCG+LPS

39、-induced group and CCl4 group was significantly higher than that in the control group in the two models, while liver homogenate total SOD activity and the GSH-px level were sharply decreased. FFHQ (60, 120, 240 mg/kg b.w. could not only significantly attenuate MDA generation, but evidently increased

40、 the liver total SOD activity and the GSH-px level in the two mice models (Tables 3 and 4.Table 3 Effects of FFHQ on MDA, SOD and GSH-px of immunologi-cal ALI mices liver homogenate (n = 10, meanSDGroup Dose MDA SOD GSH-px (mg/kg b.w. (nmol/g tissue (U/g tissue (U/mg proteinNormal - 6.541.82 396.660

41、.6 141.427.5 Model - 15.183.57d 181.340.7 d 97.924.7d FFHQ 60 12.573.15a 212.339.8a 116.529.1a 120 10.742.34b 252.142.0b 129.530.5b240 10.082.28b 269.052.7b 133.134.1b Bifendate 100 12.033.21a 193.840.5 107.630.5 a P0.05, b P0.01 vs model group; d P0.01 vs normal group.Table 4 Effects of FFHQ on MDA

42、, SOD and GSH-px of chemical ALI mices liver homogenate (n = 10, meanSDGroup Dose MDA SOD GSH-px (mg/kg b.w. (nmol/g tissue (U/g tissue (U/mg proteinNormal - 6.762.3 440.551.8136.424.5 Model - 18.684.72d 204.334.1d 91.921.2d FFHQ 60 13.212.88b 310.135.5 b105.523.6a 120 12.363.19b 321.339.2b121.124.6

43、b240 12.083.28b 337.035.1b124.327.9b Bifendate 100 12.292.67b 221.638.5 98.322.5a P0.05,b P0.01 vs model group; d P0.01 vs normal group.Histologic resultsIn the two models, normal mice had no pathologic abnormality. Liver parenchyma was in good morphology and hepatocytes were arranged around the cen

44、tral vein. No congestion and inflammation were noticed in the sinusoids (Figures 1A and 2A. In the two models, model group mice had severe pathologic abnormality. Hepatocytes were prominent with marked vacuolization; moreover, hepatocytes necrosis, striped necrosis, bridging necrosis appeared and in

45、flammatory cells were arranged around the necrotic tissue. Congestions in liver sinusoids were significant with scattering immersion of inflammatory cells (Figures 1B and 2B. In the two models, the area and extent of necrosis in FFHQ-treated groups attenuated and the immigration of inflammatory cell

46、s reduced. Liver parenchyma was well preserved with radially arranged hepatocytes around the central vein. Regular sinusoidal structures were noticed without congestion (Figures 1C and 2C. DISCUSSIONFFHQ was an extract of Chinese herbs prescription that has various kinds of pharmacologic actions. In

47、 the prescription, the main Chinese herbs such as Radix Astragali and Radix Paeonia lactiflora have been used to relieve the pain and be an effective prescription for treatment of liver disease and other diseases1,2,7,12,13. FFHQ has some active compounds, such as TGP (consist of paeoniflorin, albif

48、lorin, benzoylpaeoniflorin, oxypaeoniflorin, paeonin, etc., TAS (consist of astragaloside I-VI, soyasaponin, etc., TFA, astragalus polysaccharides and so on. The previous results from our laboratory showed that TGP was effective against ALI induced by CCl4, D-galactosamine (D-GalN and BCG+LPS in mic

49、e and chronic liver1. In vivo and in vitro, TGP showed obvious anti-inflammatory and antioxidative activities in other diseases besides in liver disease. For example, it was found that treatment of AA rats with TGP (50 mg/kg, ig (14-28 d could inhibit the elevated level of MDA and NO, and upregulate

50、d the lowered activities of SOD and GSH-px14,15. In vitro, TGP could scavenge OH and O216,17. It was reported13 that TAS could protect liver from chemical injury induced by CCl4, D-GalN and acetaminophen in mice. TAS could impede the elevation of ALT level, decrease the MDA content and increase the

51、GSH concentration in mice liver homogenate. Obvious improvements of histologic changes were also observed. In vitro, TAS (0.75 mol/L-0.18 mmol/L could decrease elevated ALT level in hepatocytes separated from rats. Previous studies of our institute showed that TAS had an antinociceptive effect on fo

52、rmalin test in mice that related to its inhibitory effect on the production of NO. Besides, astragalus polysaccharide was found to have immunoregulatoryTable 1 Effects of FFHQ on serum ALT, AST, TNF- and IL-1 in immunologic liver injury in mice (n = 10, meanSDGroup Dose (mg/kg b.w.ALT (U/LAST (U/L T

53、NF- (ng/mL IL-1 ng/mLNormal - 8.5 1.290.410.1570.054Model -204.349.6d197.642.3d 3.951.24d0.3410.101d FFHQ 60171.633.7a172.035.8a 3.000.98a0.2890.068a 120131.124.8b151.239.8b 2.760.63b0.2440.049b240129.726.5b143.530.9b 2.610.55b0.2730.051b Bifendate100 89.621.3b95.428.3b 2.890.92a0.2790.04

54、6aa P0.05,b P0.01 vs model group; d P0.01 vs normal group.2986 ISSN 1007-9327 CN 14-1219/ R World J Gastroenterol May 21, 2005 Volume 11 Number 19activity and was used in various kinds of immunologic diseases 17. In our previous study, the optimum proportion of herbs in FFHQ prescription was obtaine

55、d by uniform design in ALI mice. On the basis of the optimum proportion,FFHQ extracts were produced with the method stated in the part of “Drugs and materials”. In the present study, the two kinds of ALI models in mice were successfully established,namely immunologic ALI model induced by BCG+LPS and

56、 chemical ALI model induced by CCl 4 to observe the protective effects and its probable mechanisms of FFHQ.CCl 4 is a well-known hepatotoxic chemical 8,10,18,19. The main cause of ALI by CCl 4 is free radicals of its metabolites.By the activation of liver cytochrome P-450, CCl 4 generates methyltric

57、hloride radicals (CCl 3, which are highly unstable and immediately react with membrane components. They form covalent bonds with unsaturated fatty acids, or take a hydrogen atom from the unsaturated fatty acids of membrane lipids, resulting in the production of chloroform and lipid radicals. The lip

58、id radicals react with molecular oxygen, which initiates peroxidative decomposition of phospholipids in the endoplasmic reticulum. The peroxidation process results in the release of soluble products that may affect cell membrane. Cell membrane integrity is broken and the enzymes (such as ALT, AST, etc . in cell plasma leak out. The free radicals and its triggered lipid peroxidation were involved in the main mechanisms by which CCl 4 induced ALI 20-25. MDA was one of the main lipid peroxidation products, its elevated levels could reflec