| BackgroundWilson’s disease is also called Hepatolenticular degeneration,which is an autosomal recessively inherited disorder that leads to copper deposition in the liv er, spleen, brain, kidney, cornea and other tissues.The clinical features are liver cirr hosis, hypersplenism, proteinuria, nervous system symptoms and Kayser–Fleischer ring in Wilson’s disease patients.Not only the original defect organ but a direct t arget organ involvement with liver,so liver injury symptoms are the most comm on symptoms of WD.WD is one of the few neural genetic diseases can be treated.Treatment wit h metal chelating agent mainly lead to low blood counts, drug-induced liver damag e and other toxic or side effect which seriously affect the patient’s condition and prognosis.Since 70’s last century, Neurological research of Anhui University of t raditional Chinese take Qing Re Jie Du, Tong Fu Li Shi treatment with rhubarb, Coptis chinensis, Curcumin, rhizoma alismatis, Lysimachia christinae Hance,and radix n otoginseng for the group of Gandou Decoction in the treatment of WD patients.Th e clinical symptoms of the patients were improved obviously.The researchers altho ugh confirmed Gandou Decoction has a significant copper excretion function, but b ecause the traditional Chinese medicine compound composition is complicated,th e side effect and the optimal dose combination is not yet clear, and between the effe ct of Chinese medicine whether synergistic or constraining role has not yet been determined.This study used ICP-AES and Western-blot technology, the application of mathematical model of "uniform design" and "uniform dispersion" characteristi cs, screening and researching on mechanism of effect of Chinese medicine on Gan dou Decoction in treating Wilson’s disease of liver damage in TX mice.Objective1.To screen the effective drugs of the compound traditional Chinese medicine G andou Decoction on Wilson Disease-TX mice of liver damage on different path ways based on uniform design experimentation,and investigating its theoretical si gnificance.2.To observe the change of trace elements in serum and liver tissue of TX mice,and screening the effective drugs of the compound traditional Chinese medicine Gandou Decoction and investigating its theoretical significance.3.To observe the expression of P38, ERK, BCL-2, BAX protein in liver tissue of TX mice,and to investigate its function in TX mice of liver damage.To screen t he effective drugs of the compound traditional Chinese medicine of Gandou De coctionand and investigating its theoretical significance.Methods1.Selecting 2-month-old TX mice(120 mice) and dividing into 11 medicine gro ups,and a model group(10 mice) randomly. Selecting 2-month-old DL mice as c ontrol group,randomly.The control group and model group was fed the same fod der with the medicine groups with normal 0.2ml/10 g saline twice a day for 30 days by gastric perfusion.The medicine groups was gavaged with normal 0.2ml/10 g medicine a day for 30 days by gastric perfusion.2.To detect the content of Cu, Zn, Fe in TX mice liver tissue and serum by ICP-AES,screening the effect of Chinese medicine Gandou Decoction, and to expl ore its possible mechanisms.3.To detect the expression levels of P38, ERK, BCL-2 and BAX in liver tissue of TX mice and screening the effect of Chinese medicine Gandou Decoction by Western-Blot method, and to explore its possible mechanisms.Results1.Result of trace elements1.1 The trace elements of serum in TX mice:The serum copper level of me dicine group 4,5 were significantly higher than model group(P<0.05).The seru m copper level of model group TX mice was lower than control group DL mic e(P<0.05).The serum copper level of other medicine groups were higher than model group,but it has no statistical significance(P>0.05).The serum zinc level of medicine group 1,2,3,4,5,6,7,8,9,10 were significantly higher than model gro up(P<0.05).The serum zinc level of model group was lower than control gro up,but it has no statistical significance(P>0.05).The serum iron level of of me dicine group 1、8、10 were lower than model group(P<0.05).The serum iron level of model group TX mice was lower than control group DL mice(P<0.05).1.2 By multiple regression analysis:the effect Chinese medicine of Gandou decoction that increased the serum copper and zinc level of TX mice was turm eric.The maximum dose of turmeric was 0.04 g. The components of Gandou deco ction on Fe elements were not responded to effect Chinese medicine in serum m etabolic.1.3 The trace elements in TX mice liver: The liver copper level of medicin e group 1,3,5,8,9,10,11 were significantly lower than model group(P<0.05).The liver copper level of model group TX mice was higher than control group DL mice(P<0.05).The liver copper level of other medicine groups were lower than model group,but it has no statistical significance(P>0.05).The liver zine lev el of medicine group 2,3,8,10 were significantly lower than model group(P<0.05).The liver zine level of model group was significantly higher than control g roup(P<0.05).The liver iron level of of medicine group 3,7,10 were lower th an model group(P<0.05).The liver iron level of model group TX mice was h igher than control group DL mice(P<0.05).1.4 By multiple regression analysis:the effect Chinese medicine of Gandou decoction that reduced the liver copper and zinc level of TX mice was radix n otoginseng.The maximum dose of radix notoginseng was 0.006 g. The component s of Gandou decoction on Fe elements were not responded to effect Chinese med icine in TX mice liver.2 Expression of P38, ERK, BAX, BCL-2 protein in liver tissue of TX mice2.1 P38:The expressions of P38 in TX mice liver of medicine group 3,4,6,8,10,11 were lower than model group and control group(P<0.05). Compared wi th control group DL mice,the expressions of P38 in TX mice liver of model gr oup was not statistical significance(P>0.05).2.2 ERK:The expressions of ERK in TX mice liver of medicine group 1,4,5,6,7,10 were lower than model group(P<0.05). The expressions of ERK in T X mice liver of model group was higher than model group and control group(P<0.05).The expressions of ERK in TX mice liver of medicine group 1~11 wer e higher than model group,but it has no statistical significance(P>0.05).2.3 BAX: The expressions of BAX in TX mice liver of medicine group 1、4,5,6,9,10 were higher than model group(P<0.05). The expressions of BAX in TX mice liver of model group was slightly lower than control group DL mice,but it has no statistical significance(P>0.05).The expressions of BAX in TX mice liver of medicine group 1,4 were was lower than control group(P<0.05).2.4 BCL-2:The expressions of BCL-2 in TX mice liver of medicine group1,3,4,6,9,11 were higher than model group(P<0.05). The expressions of BCL-2 in TX mice liver of model group was higher than control group DL mice,but it has no statistical significance(P>0.05).The expressions of BCL-2 in TX mi ce liver of medicine group 1,3,4,6,9,11 were was higher than control group(P<0.05).2.5 By multiple regression analysis, the effect Chinese medicine of Gandou decoction that suppressed expression of P38 in TX mice liver were rhubarb andradix notoginseng. The maximum dose of Dahuang was 0.04 g, and the maxim um dose of radix notoginseng was 0.006 g. The effect Chinese medicine of Gan dou decoction that suppressed expression of ERK in TX mice liver was radix no toginseng.The maximum dose of radix notoginseng was 0.006 g.The effect Chine se medicine of Gandou decoction that promoted expression of BCL-2 in TX mic e liver was rhizoma alismatis.The maximum dose of rhizoma alismatis was 0.048 g.The effect Chinese medicine of Gandou decoction that suppressed expression of BAX in TX mice liver was radix notoginseng.The maximum dose of radix n otoginseng was 0.006 g.Conclusion1.The turmeric can increase the content of serum copper and zinc in TX mice, and the effect of elevated serum zinc was more serious.The radix notoginseng can r educe the copper, zinc content of liver tissue in TX mice, and reduce the effect of copper was more significant.Probably,we can increase the effect of copper extracti on by adding the dose of turmeric and radix notoginseng,to reduce the degree of liver damage.2.The rhubarb and radix notoginseng can inhibit the expression of P38 and radix n otoginseng can inhibit the expression of ERK.Probably,we can increase the effect of treating liver fibrosis by adding the dose of rhubarb and radix notoginseng,to reduce the degree of liver damage.3.The rhizoma alismatis can promote the expression of BCL-2, and the radix notog inseng can inhibit the expression of BAX.Probably,we can increase the effect of i nhibition of hepatic apoptosis by adding the dose of rhizoma alismatis and radix notoginseng,to reduce the degree of liver damage.4.In conclusion,this four drugs rhubarb,rhizoma alismatis and radix notoginseng in Gandou Decoction play a major role in the repair of liver damage in the proces s,which especially the radix notoginseng,all play a major role in the three approach.Rhizoma coptidis and herba lysimachiae in each pathway has no significant eff ect.Later verification if it is conforming with scientific facts,we can removal of r hizoma coptidis and herba lysimachiae from Gandou Decoction to optimize com ponent. |
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