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A Research about the Influence of Yi Jia Decoction on Serum NO, GSH-PX Levels of Immunological Liver Injury in Mice
12 3
Sui Fangyu Qu Miao Zhang Miao
Heilongjiang University of Chinese Medicine
Chinese Herbal Teaching and Research Section Harbin China
Abstracts: Objective: To explore the functional mechanism of Yi JiaDecoction in treating immunological liver damage in mice through the test on serum NO, GSH-PX levels.
Methods: 120 mice were randomly allocated into six groups which areblank, model, western medicine, high dose of Chinese medicine, medium dose of Chinese medicine and low dose of Chinese medicine groups. Combining BCG and LPS induces liver injury in mice. After two weeks'treatment, mice were testedon serum NO, GSH-PX.
Results: Compared with the model group, serum NO levelof the treatment groups tended to decrease, especially for the medium dose of Chinese medicine group (P<0.01), and GSH-PX level of treatment groups tended to increase, especially for the high dose of Chinese medicine group (P <0.01). Conclusion: Yi Jia Decoction can treat the liver injury in mice and the functional mechanism may be related to the adjustment the serum NO, GSH-PX levels of mice.
Key words: Yi Jia Decoction; Nitric Oxide; Immunological Liver Injury
1 Introduction
Immunological Liver Injury(ILI) is a common and worldwide disease, regardless of race, region and other restrictions. At present, most academics believe that pathological mechanisms of inducing liver injury in mice by BCG and LPS are quite similar to the pathological mechanisms of human hepatitis immune dysfunction. So combination of BCG and LPS is one of the ideal methods to choose and do research on hepatoprotective medicine.[1]Yi Jia Decoction which includes raw oyster shell, raw rhizoma atractylodis macrocephalae, stir-fried barley sprout, schisandra, is a popular formula for treating immunological liver injury. The author chosemice as subjects via BCG and LPS to induce liver injury and exploredthe therapeutic mechanism of ILI.
2 Materials and methods
2.1 Animals and groups
Kunming mice, weighing 18-22g, 60 males and 60 females (All from Heilongjiang University of Chinese Medicine Experimental Animal Center). Experimental environment: Kept the room ventilated, dry, temperature 22-25 °C and humidity50-70%. Made sure that the mice can drink and eat freely. The mice were randomly divided into six groups: blank group, model group, DDB treatment group (western medicine group), high, medium and low dose of Yi Jia Decoction groups and each group included 20 mice.
2.2 Experimental medicine
(1) Yi Jia Decoction which includes oysters, Rhizoma Atractylodis Macrocephalae,fried malt,Schisandra composition, purchased in Heilongjiang University of Chinese Medicine, Traditional Chinese Medicine BureauofFirst Affiliated Hospital ; (2) DDB pills: dissolved in distilled water to 0.5 g / ml.Purchased from Zhejiang Medicine Co. Ltd.;(3)BCG was produced by the Shanghai Biological Institute;(4) lipopolysaccharide (LPS) was produced by Sigma.
2.3 Preparation of decoction
Dissolved medicine in distilled water for 2h, then boiled for 30min, filtered and got the first time decoction. After that, dissolved dregs in distilled water for 1h, then boiled for 30min, filtered and got the second time decoction. Put the twice decoction together and then filtered one more time and concentrated it under the conditionof 80 °C to the required concentration. The concentrationsof low, medium and high dose Yi Jia Decoction were equivalent to 0.34g, 0.68g, 1.36g per 1ml.
2.4 modeling and administration
Combined BCG and LPS to induce liver injury modelin mice. Injected BCG5 x 107 bacteria / mouse to the subjections via the tail vein except the blank group and started to treat mice by gavage from the next day.Blank group and modelgroup were given 0.2ml distilled water; western medicine group were given DDB solution 20ml/kg; Chinese compound medicine groupswere given water extract 10ml/kg.12 days later, injected LPS7.5ug / mouse, and got the serum for NO, GSH-PX levels' test 12h after intravenous injection.
2.5 Observation of indicators and test methods
NO, GSH-PX reagent kit was provided by Nanjing Jianchengreagent company. Test methods followed bythe introduction.
2.6 Analytical method ofexperimental results
All results data are showed as mean ± standard deviation (x ± S), and compared the data between two different groups by t or t'test.
3 Results
The serumNO, GSH-PX levels of different groups (see chart1)
Chart 1 The serumNO, GSH-PX levels of different groups
Groups Dose numbers of mice NO GSH-PX
g/moused n nmol /mgprot U/ mgprot
Blank 20 31.73±6.47 352.62±65.55
Model 20 61.06±9.01AA 139.18±61.68A
Western medicine 0.003 20 48.08±11.09** 239.01±80.30**
High dose 1.36 20 54.33±9.01**A 269.58±31.87*A
Medium dose 0.68 20 42.79±12.71*A 249.34±69.42** A
Low dose 0.34 20 52.88±13.87** A 221.31±64.82**A
Note: Compared with the blank group ▲ P <0.01, ▲ ▲ P <0.05; compared with the model group:
* P <0.01, ** P <0.05; Each Chinese Medicine group compared with the positive group, △ P> 0.05.
4 Discussion
Main clinical manifestationsof ILI patients are malaise,anorexia, thin sloppy stool, etc, which correspond to spleen deficiency. Yi Jia Decoction is composed ofraw oyster shell, raw rhizoma atractylodis macrocephalae, stir-fried barley sprout, schisandra, in order to cure the spleen deficiency. The raw oyster shell with mild resistance performs as a monarch drug in the prescription, and plays effect in spleen. It can eliminate the stagnant pathogenic factors of spleen, meanwhile soften hardness and dissipate binds. The raw rhizoma atractylodis macrocephalaestrengthens the function of spleen-in-TCM, and the schisandra astringes spleen essence, both of which are deemed to ministerial drug in the prescription. The stir-fried barley sprout as an assistant drug, not only tonifies spleen together with raw rhizoma atractylodis
macrocephalae, but also prevents oyster shell from impairing stomach. The best indication of this prescription is spleen deficiency, whose performance is mild, but effectiveness in regulating spleen and dissipating binds is well.
The change of NOn GSH-PX level may effect the progress of ILI. NO synthetizes from NO synase (NOS) withL-arginine as itssubstrate. Both constitutive NO synthase (CNOS) and inducible NO synthase( iNOS ) are NOS. The former and NO synthetized from it can injure hepatocyte by the toxicitymediated from kupffer's cell. Such toxicity may inhibit mitochondria respiration, activate NF-kB, and result in necrocytosis at last.[2]GSH-PX is an important enzyme that decomposed from peroxide and wildly found in the body. It eliminates intracellularhazardous substance that metabolized by peroxide, and blocks the chain reactions of lipid peroxide, so as to keep structure and function of cytomembrane integrated. Once its level decreases, hepatocyte will be injured.
Conclusions
Experimental results show that NO level of the mice in model group increased significantly, and compared with the blank group, there were significant differences (P <0.05); Compared with the model group, NO level of the mice in medium dose of Yi Jia Decoctiongroup decreased significantly (P <0.01), the difference had statistical significance; compared with the model group, NO level of the mice in positive control group (western medicine group), high dose of Yi Jia Decoctiongroup and low dose of Yi Jia Decoctiongroup decreased (P <0.05), the difference had statistical significance; compare the 3 Chinese medicine groups with the positive control group, (△P> 0.05), the difference didn't have statistical significance; Comparison between treatment groups had no difference. These statistical results indicate that the treatment groups could decrease the NO level of ILI mice, but the curative effect of the medium dose of Yi Jia Decoction group is better than others.
Experimental results show thatGSH-PX level of the mice in model group decreased significantly, and compared with the blank group, there were significant differences(P <0.01); compared with model group, GSH-PX level of the mice in high dose of Yi Jia Decoctiongroup increased significantly (P <0.01), the difference had statistical significance; compared with model group, GSH-PX level of the mice in positive control group, medium dose and low dose of Yi Jia Decoctiongroups increased (P <0.05), the difference had statistical significance; compared the different doses of Yi Jia Decoction groups with the positive control group, (△ P> 0.05), the difference didn't have statistical significance;
(there is no significant difference between each treatment group.) comparison between treatment groups, no difference. These statistical results indicate that the treatment groups could increase the GSH-PX level of ILI mice, but the curative effect of the high dose of Yi Jia Decoction group is better than others.
In conclusion, Yi Jia Decoction may decrease serum NO level and increase serum GSH-PX level to prevent and treat Immunological Liver Injury.
References:
[1] Wu Liping, Liu Rong, Bi Yushun, etc. The protection on immunological liver injury mice from Fu Ganling pills [J]. Shandong University Journal: Medical Sciences, 2008,3 (3) :237-240.
[2] Xiong Jingbo, Kang Gefei. The function of two kinds of nitric oxide synthase inhibitors on endotoxin liver injury. Chinese Hepatology magazine, 1996; 7 (2): 118.
