| The first part:Establishment of animal models of hyperuricemia with uric acid excretion disorder.Purpose: There are many methods to establish the animal model of hyperuricemia,but there are three principles: 1.Increase the intake of uric acid and promote the formation of uric acid;2.Inhibits or eliminates uric acid enzyme activity,thus reducing uric acid decomposition;3.Inhibit uric acid excretion.According to the serum uric acid level and urine uric acid excretion,the hyperuricemia can be divided into three types :(1)the excretion of uric acid;(2)excessive uric acid formation;(3)mixed type.Clinical results show that 90% of primary hyperuricemia is caused by uric acid excretion disorder.Moreover,most people with hyperuricemia had no significant abnormalities in liver function and kidney function in a long period of time.Therefore,it is of great significance to establish an animal model of uric acid excretion with hyperuricemia which is compatible with clinical hyperuricemia.In the course of evolution,humans eliminated uric acid oxidase,which caused uric acid to become the final product of purine metabolism.Uric acid oxidase is an important enzyme in the decomposition of uric acid.Therefore,the application of uric acid enzyme inhibitors is one of the methods to eliminate the human and rodent hyperuricemia model animals.Pyrazinamide and ethambutol are both anti-tuberculosis drugs,which can inhibit the excretion of uric acid in the kidneys.This experiment uses uricase inhibitor(oxygen oxazine acid potassium)combined inhibition of renal excretion of uric acid drugs(pyrazinamide and ethambutol),in order to establish a more ideal high uric acid hematic disease animal model.This model is also a rat model which is more closely related to human clinical uric acid excretion disorder hyperuricemia.And this experiment compare the advantages and disadvantages of the two methods,and lay the foundation for optimizing and regulating the establishment of such models.Material and method: 30 male SD rats were randomly divided into blank control group(hereinafter referred to as blank group),oxygen oxazine acid potassium + ethambutol group(hereinafter referred to as model ? group),oxygen oxazine acid potassium + pyrazinamide group(hereinafter referred to as model ? group),respectively with 0.9% sodium chloride solution,oxygen oxazine acid potassium 300 mg/kg + ethambutol oxazine 250 mg/kg,oxygen acid potassium 300 mg/kg + pyrazinamide 300 mg/kg.After 0 weeks,1 week,3 weeks,5 weeks,the venous blood was collected,and 24 h urine and feces were collected.After the animals were killed,we took part of the liver tissue and the left part of the kidney.We use manual method(colorimetry)to detect serum uric acid,urine uric acid,feces uric,liver function and kidney function.We biopsy the liver and kidney to observe the pathological changes of the liver and kidney.Results: Compared with blank control group,after the experiment 1 w,model ? group and model ? group rats serum uric acid levels were significantly elevated,the difference was statistically significant(P < 0.05).In experiment 3 w,5 w,model ? groups of serum uric acid levels continued to decline,compared with the experiment 1 w blood uric acid difference was statistically significant(P < 0.05).Model ? group during the whole experiment serum uric acid levels in the same high level,there was no statistically significant difference compared with the experiment 1 w(P > 0.05).There was no significant difference in liver function and renal function between the two groups(P > 0.05).The results of pathological examination of liver and kidney showed that compared with the blank control group,no obvious abnormality was observed,which was consistent with the blood test results.Conclusion: The model of uric acid excretion disorder hyperuricemia can be effectively established by the two methods: potassium oxazide + ethylamine butanol and potassium oxazine + pyrazinamide.The model established by potassium oxazine + pyrazinamide has better stability of serum uric acid and is more consistent with the pathogenesis of human uric acid excretion disorder hyperuricemia.Therefore,it is more suitable for long-term application of hyperuricemia.The second part: An experimental study on the treatment of uric acid excretion disorder hyperuricemia by Jiangniaosuan Decoction.Purpose: Hyperuricemia is a great threat to human health.More attention has been paid to the study of hyperuricemia.Currently,there are three kinds of drugs to treat hyperuricemia in clinical application:1.Drugs that can inhibit the production of uric acid,mainly XO inhibitors,including Allopurinol and Febuxostat;2.Drugs to promote the excretion of uric acid.It is mainly URAT1 inhibitors,including benzbromarone and propane.3.Uric acid oxidase,which mainly includes labriase and pegoloxidase,has not been approved for listing in China.There are numerous cases of hyperuricemia treated with traditional Chinese medicine in clinical practice,and there are many related reports in the literature,including the effective ingredients of traditional Chinese medicine,single herb medicine and compound prescription of traditional Chinese medicine.Jiangniaosuan Decoction is the essence of clinical application to treat hyperuricemia and the clinical curative effect is clear.On the basis of the successful establishment of the rat model of uric acid excretion of hyperuricemia,Jiangniaosuan Decoction was used to intervene.To observe whether Jiangniaosuan Decoction has a quantitative effect on the efficacy of the model to determine the optimal dosage of Jiangniaosuan Decoction.At the same time,by comparison between Jiangniaosuan Decoction and benzene bromine Malone to observe both on uric acid reduction,on the regulation of blood lipids,and what are the similarities and differences on the influence of the liver and kidney function,to determine the right dose drugs,and reveal whether they have the effect of adjusting blood fat.Material and method: Before experiment,randomly selected 10 healthy male SD rats as blank control group,and then application modeling method: multiple subcutaneous injection of oxygen by intraperitoneal injection of oxazine acid potassium 300 mg/kg + pyrazinamide 300 mg/kg for the model.A week later,the rats with high uric acid hematic disease were randomly divided into Chinese medicine low dose group(hereinafter referred to as low group),Chinese medicine middle dose group(hereinafter referred to as the middle group),Chinese medicine high dose group(hereinafter referred to as the high group),benzene bromide Malone(hereinafter referred to as the western medicine group)and model group?It was administered with 25g/kg of Chinese medicine,50g/kg of Chinese medicine,100g/kg of traditional Chinese medicine,4.5mg/kg benzbromalon and 0.9% sodium chloride solution for 4 weeks.Before and after the experiment 1w,3w,5w,blood was collected after the eyeball,24 h urine and feces were collected too.Some liver,left part of the kidney and part of the small intestine were extracted after anaesthesia.The serum uric acid,urinary uric acid,fecal uric acid,blood lipid,liver function and kidney function were detected by hand method(colorimetry),and the pathological section of liver and kidney was performed.Results: 1.In the 1w experiment,the serum uric acid level in each group was increased(P <0.01)compared with the blank group,but there was no statistically significant difference between the groups(P>0.05).In experiment 3w and 5w experiment,serum uric acid content in each group was reduced(P<0.05)compared with the model group.Dose in low dose group and the middle group,high dose group,western medicine group,the difference was statistically significant(P<0.05),while in the middle group,the high group and western medicine group,the serum uric acid levels are very close,there was no statistically significant difference(P > 0.05).2.Compared with the blank control group,the level of TG,TC and ldl-c in the model group and the western medicine group increased at 5w,and the difference was statistically significant(P<0.05).There was no statistically significant difference between the model group and the western medicine group(P>0.05).The levels of TC,TG and ldl-c in the middle and high groups decreased,and the difference was statistically significant(P<0.05).There was no statistically significant difference between middle and middle group(P>0.05).The changes of TG,TC and ldl-c in the model group and the western medicine group were statistically significant(P<0.05).In the whole experiment,the changes of hdl-c were not significant,and the difference was not statistically significant(P>0.05).3.Compared with the blank control group,there was no significant difference in liver function and renal function in each group(P BBB 0 0.05).No obvious abnormalities were observed in the pathological sections of liver and kidney tissues.Conclusion: The effect of the drug on the animal model of hyperuricemia of uric acid excretion was significant,but it was not obvious.The medium dose group,the high dose group and the benzbromolone group reduced the serum uric acid capacity almost the same.The Chinese medicine group has the ability to regulate blood lipid,the capacity is related to dose,and the influence of western medicine group on blood lipid is not obvious.No significant liver and kidney damage was observed in the experimental groups.The third part: The study on the curative effect of Jiangniaosuan Decoction for the treatment of uric acid excretion with hyperuricemia.Purpose: Different anti-hyperuricemia drugs may reduce serum uric acid level,but the target of uric acid is different,and the effect size of uric acid is strong and weak.Allopurinol and he mainly by inhibiting the activity of xanthine oxidase in the liver,thus reducing the generation of uric acid and lowering serum uric acid level.However,benzbromarone and prosulfonate reduce the reabsorption of uric acid by inhibiting the expression of URAT1,which can promote the excretion of uric acid and reduce serum uric acid level.Uric acid oxidase is a kind of can directly to uric acid oxidation and decomposition of allantoin enzyme,and allantoin is an inert and water-soluble purine metabolites allantoin easily after renal excretion,also can reduce the body's blood uric acid levels.The effect of single herb on uric acid can inhibit the formation of uric acid,and also promote the excretion of uric acid.Such as Bi Xie total saponins [4] can be expressed through cut URAT1,raised the expression of OAT1 and OAT3 restrain heavy absorption process of uric acid in the kidney,promote in renal excretion of uric acid,lowering serum uric acid levels.The compound of traditional Chinese medicine is also multiple because of its variety of ingredients,so as long as the compatibility is reasonable,the effect of reducing uric acid of Chinese medicine compound should be better.Chinese medicine is more likely to be a powerful weapon in the treatment of hyperuricemia.In this paper,the expression of the liver XO activity of rat liver,renal glut-9,ABCG2,URAT1 and intestinal ABCG2 m RNA were detected to reveal the possible mechanism of Jiangniaosuan Decoction effect.This will provide theoretical support for the development of new therapeutic drugs for multiple targets.Material and method: The experimental animal group and intervention method were the same as the second part.The activity of xanthine oxidase was detected by Western-Blot technique.The expression of glut-9,ABCG2 and URAT1 m RNA in left kidney tissues was detected by RT-PCR.The expression of ABCG2 m RNA in intestinal tissues was detected by RT-PCR.To explore the possible mechanism of the effect of Jiangniaosuan Decoction on hyperuricemia.Results: 1.Western Blot method to determine the liver xanthine oxidase activity results: compared with blank control group,model group slightly reduce the XO expression,Western medicine group have no obvious changes,there were no statistically significant difference(P > 0.05);The XO expression in each dose group of traditional Chinese medicine decreased(P < 0.05),and the reduction degree increased with the increase of dose.However,there was no significant difference between the middle and middle groups(P>0.05).2.RT-PCR method results:(1)the kidney urate transporter URAT1 protein and m RNA expression,compared with the blank control group,traditional Chinese medicine(TCM)for metrology group and western medicine are cut changes(P < 0.05),the biggest declines in western medicine group,there was significant difference(P < 0.01).The differences between the two groups were similar,and the differences were statistically significant(P < 0.05)compared with those in the western medicine group and the middle and lower groups.The expression of the model group was significantly increased,and the difference was statistically significant(P<0.05).2 renal glucose transporter GLUT9 protein and m RNA expression,compared with the blank control group,model group(P > 0.05),there is no change for metrology group and western medicine group in Chinese traditional medicine by changes(P < 0.05),larger declines in western medicine group,the difference was statistically significant(P< 0.05),the differences between groups and high cut,compared with western medicine group,the low and middle group,the difference was statistically significant(P < 0.05);(3)in the kidney and intestine adenosine triphosphate combined transport protein and m RNA expression of ABCG2 protein,compared with blank control group,western medicine group and model group were no significant change(P > 0.05),compared with western medicine group,Chinese medicine for metrology group have all been raised to different degrees,and in the group,the high group in the ABCG2 raised the difference was statistically significant(P < 0.05),although there are low and middle group rise trend,but there was no statistically significant difference(P > 0.05).Conclusion: Jiangniaosuan Decoction by inhibiting liver XO activity in order to reduce the generation of uric acid,by cutting the kidney URAT1,GLUT-9 m RNA expression,reduce renal reabsorption of uric acid,by raising the kidneys and intestines of ABCG2 m RNA expression,promote the uric acid in the kidney and intestinal excretion,thereby reducing the blood uric acid levels.Jiangniaosuan Decoction is a multi-action target,multi-channel diuretic drug. |
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