Liver, Spleen, Liver And Spleen Side Of Stagnation And Spleen Deficiency Syndrome Model Animals Hpa Axis Regulation

"The correlation between formula and syndrome" is not only the core principle of treatment based on different syndromes but also the logic foundation of formulary knowledge in traditional Chinese medicine (TCM). Study of formulary is not only to consider the composition, should also take into account to the adaptation indication of formula. Based on finding of significant changes in HPA axis of former animal model, this study is in advance to observe the variation of the HPA axis in the rat model of liver stagnation and spleen deficiency induced by the chronic astricting + excess fatigue + diet and the influences of three relating formulae on the animal model to compare similarities and differences of action of Chaihu Shugan San, Sijunzi Tang and Chaishu Sijun Tang which is a typical formula respectively for function of smoothing liver, invigorating spleen and soothing liver and invigorating spleen, and probe into the effective mechanism of them on HPA axis of the medel.The dissertation contains both of documental summarization and experimental study. Recent research developmentes of clinic and experiment about the syndrome of stagnation of liver and deficiency of spleen, Chaihu Shugan San, Sijunzi Tang and Chaishu Sijun Tang are summed up in this review. The apparent symptom, HPA axis-related hormones and central some receptor expressions in syndrome of liver stagnation and spleen deficiency in rat model are investigated in the experimental study.Experimental research: The effective mechanism of the three formulae on the HPAA of model rat with syndrome of liver-qi stagnation and spleen-qi deficiency.Methods: Rats were randomly divided into normal control group, stagnation of liver deficiency model group, Chaihu Shugan San group, Sijunzi Tang group and Chaishu Sijun Tang group , 10 rats in each group. The four groups of animals used chronic restraint stress +excess fatigue + diet method to form the model, the model rats were astricted for 3 hours at every morning, swim for 10 minutes in water temperated 22±1°C and fed every other day, for 28d; normal control group without any stimulation, natural feeding. On the 15^th day, medicine group rats were given Chaishu Sijunzi Tang 4. 27g/kg, Chaihu Shugan San 4. 2g/kg, Sijunzi Tang 4.53g/kg, 1 time/d; model group and normal group were given distilled water for 14 days. Observe and record all of the emotional behavior in rats, hair color, diet, appearance of urine, and so on. Measured the body weight and body temperature every week(temperature at every Sunday AM 8:00-8:30 determination;weight in the determination of PM 2:00-2:30 every Sunday). On the 28^th day, the 10% of chloral hydrate anesthesia animals, abdominal aortic blood, separation test plasma CRH, ACTH, COR. Seperate the hypothalamus and pituitary gland rapidly on the ice plate, situated at a liquid nitrogen filling in the preservation, use of RT-PCR method to determine the pituitary CRHR, GR mRNA, hypothalamus ACTHR, GR mRNA. Some of the hypothalamus under the boil, after weighing the saline (1ml) in 3min, coupled with glacial acetic acid 0.5ml of 1N browser absorbed in the homogenate Pulp extraction, reuse and 1NNaOH 0.5ml, low temperature conditions, 3000rpm 30min, supernatant taken at -20℃preservation, test the brain CRH.Statistical methods: the data were discriped by (x|-)±SD and the number of samples compared between groups used one-way ANOVA (one-way ANOVA) test; non-normal data using non-parametric statistical analysis. All data used SPSS 13.0 statistics software.Experimental results:1. Effects of different prescriptions on the changes in appearance: Compared with normal group, two weeks later, the model rats shown that the level of activity reduced, emotional instability, reduced diet and easy to wake up and so on. After four weeks, the model animals became depression status, the struggle reduced, the expression was indifference, etc. After two weeks' treatment, Chaihu Shugan San group' s rats increased the level of activity, emotional state of stability; Sijunzi Tang group' s rats improved on the skin, diet and significantly improved the state of stool; Chaishusijun Tang rats regardless of the skin, the level of activity, emotional reaction, sleep or diet all see the improvement clearly.2. Effects of different prescriptions on the rats' body weight:The normal rats' weight increased gradually, while the model group and the three treatment groups increased significantly slower, in which the model group rats showed a downward trend in the 4^th week. Compared with normal group, model group, Chaihu Shugan San group , Sijunzi Tang group, ChaiShu Sijun Tang rats' weight decreased from the first week to the fourth, the difference was significant (P<0.05). Compared with model group, after two weeks' treatment (the fourth week of the experiment), Chaishu Sijun Tang group, Chaihu Shugan San group and Sijunzi Tang group' s weight were increased to varying degrees, of which Chaishu Sijuntang group' s difference was significant(P<0. 05). Compared with the Chaihu Shugan San group, Chaishu Sijun Tang group and Sijunzi Tang group had no significant difference in body weight (P>0. 05);Chaishu Sijun Tang and Sijunzi Tang rats had no significant differences in body weight (P>0. 05). 3.Effects of different prescriptions on the rats' body temperature:The normal group rats had no significant changes in body temperature, while the model group rats' temperature decreased in trend. The three treatment groups' temperature was also decrease before the treatment, but for two weeks after the treatment (the 3-4th week), its temperature was increasing. Compared with normal group, model group' s temperature was decreased in the first 2-4 weeks, the difference was significant(P<0.01); Chaihu Shugan San group temperature decreased constantly, the difference was significant(P<0. 05);Sijunzi Tang group rats had no significant differences in body temperature(P>0. 05), of which Chaishu Sijun Tang group' s temperature had reached the level of normal group. Compared with model group, three treatment groups showed a body temperature of rats increased to varying degrees, the differences were significant(P<0.01). Compared with the Chaihu Shugan San group, Chaishu Sijun Tang group rats significantly increased in body temperature, the difference was significant (P<0. 05) ;Sijunzi Tang group had no significant differences in body temperature(P>0. 05). Compared with the Sijunzi Tang group, there were no significant differences in body temperature(P>0.05).4. Effects of different prescriptions on the hypothalamus CRH content: Compared with normal group, model group plasma CRH content in the hypothalamus of rats significantly increased, the difference was significant(P<0. 05); Chaihu Shugan San group and Chaishu Sijun Tang group rats decreased in hypothalamic CRH, the Sijunzi Tang group increased, but the difference was not significant(P>0.05). Compared with model group, three treatment groups' CRH content of rat hypothalamus was reduced to different degrees, of which Chaishu Sijun Tang group and Chaihushugan San group reduced significantly, the differences were significant (P<0. 01).And compared with Sijunzi Tang group, Chaihushugan San and Chaishu Sijun Tang rats' hypothalamic CRH content decreased, the difference was significant (P<0.05). Chaihushugan San and Chaishu Sijun Tang had no significant differences(P>0.05).5. Effects of different prescriptions on the plasma CRH, ACTH, COR content:Compared with normal group, model group rats were significantly elevated in plasma CRH, the difference was significant (P<0.01) ; Chaihu Shugan San group and Sijunzi Tang group CRH content in rats plasma were reduced to varying degrees, but the difference was not significant (P>0.05); between the three treatment groups differences were not significant (P>0. 05). Compared with normal group, plasma ACTH in model group was lower, the difference was significant(P<0. 01). Compared with model group, two weeks after delivery, Chaihu Shugan San group, Sijunzi Tang group and Chaishu Sijun Tang group in rats plasma ACTH concent increased, the differences were significant (P<0.05);among the three treatment groups, the differences was not significant (P>0.05). Compared with normal group, plasma COR in model group rats was significantly increased, the difference was significant(P<0.01); Compared with model group, two weeks after delivery, Chaihu Shugan San group, Sijunzi Tang group and Chaishu Sijun Tang group rats plasma concentration of COR were reduced to different degrees, of which Chaishu Sijun Tang group difference was significant(P<0. 05);there were no significant differences among the three treatment groups(P>0.05).6. Effects of different prescriptions on the expression of the rat hypothalamus ACTHR and GR mRNA:Compared with normal group, the hypothalamus ACTHR mRNA were significantly decreased (P<0. 01)in model group; the three treatment groups were seen ACTHR mRNA decline to varying degrees, of which Chaihu Shugan San group and Sijunzi Tang group decreased, the difference was significant (P<0. 05). Compared with model group, Chaihu Shugan San group and Chaishu Sijun Tang group rats' hypothalamus ACTHR mRNA increased, in which Chaishu Sijun Tang group increased significantly, the difference was significant(P<0.01); Sijunzi Tang group had no significant change. Compared with the Chaishu Sijun Tang group, Chaihu Shugan San group and the Sijunzi Tang group ACTHR mRNA were all lower, the difference was significant (P<0.05); there were no significant difference between Chaihu Shugan San group and Sijunzi Tang group(P>0.05). Compared with normal group, model group GR mRNA were significantly lower, the difference was significant(P<0.01); Sijunzi Tang GR mRNA were significantly lower than the normal group, the difference was significant (P<0.05); Chaihu Shugan San and Sijunzi Tang group rats GR mRNA were closer to the normal group, the difference was not significant(P>0.05). Compared with model group, Chaihu Shugan San group, Sijunzi Tang group, Chaishu Sijun Tang group rats' GR mRNA increased in varying degrees, of which Chaihushugansan group and Chaishusijuntang group increased significantly,the difference was significant(P<0.01), Sijunzi Tang had no significant difference (P>0.05). Compared with the Chaishu Sijun Tang group, Sijunzi Tang GR mRNA were significantly lower, the difference was significant (P<0.05), Chaihu Shugan San had no significant difference (P>0. 05). And compagred with Sijunzi Tang group, Chaihu Shugan San GR mRNA were significantly increased, the difference was significant(P<0.05).7. Effects of different prescriptions on the expression of the rat pituitary CRHR and GR mRNA: Compared with normal group, the pituitary CRHR mRNA expression were significantly decreased (P<0. 05) in model group; the pituitary CRHR mRNA expression increased in Chaihu Shugan San group and Sijunzi Tang group but the differences were not significant(P>0.05). There was no obvious difference between Chaishu Sijun Tang group and normal group. Compared with model group, Chaihu Shugan San group, Sijunzi Tang group, Chaishu Sijun Tang group rats' pituitary CRHR mRNA expression were increasesd in varying degrees, the differences were significant or very significant(P<0.01 or P<0.05). Among the three Chinese medicine groups, there were no significant difference(P>0.05). Compared with normal group, model group rats pituitary GR mRNA expression were significantly decreased(P<0.01); Compared with model group, the pituitary GR mRNA expression were significantly increased (P<0.01) in three Chinese medicine groups. There was no significant difference among three Chinese medicine groups (P>0.05).Conclusion:There were body temperature, weight, the apparent effects of group, HPA axis abnormalities by Chronic restraint stress + excess fatigue + diet treatment of the rats. Chaihu Shugan San, Sijunzi Tang, Chaishu Sijun Tang could improve the symdromes and laboratory findings of the model in some degrees, while Chaishu Sijun Tang had the best effect. The results showes that this model matches with Chinese medicine liver stagnation and spleen deficiency and the rat model exists the abnormalities of HPAA with sympathetic nerve dysfunction. The best effect to the model is of Chaishu Sijun Tang. This result is consistent with the traditional Chinese medicine experience.Inference: The rat model of HPA axis dysfunction in the hypothalamus may be characterized of abnormally reduces of central ACTHR, GR and pituitary CRHR, GR mRNA expression. Chaishu Sijun Tang had the function on hypothalamic and the pituitary while Sijunzi Tang on pituitary. Similar to Chaishu Sijun Tang, Chaihu Shugan San acted only on hypothalamus selectively.This study provided some objective evidence on" the correlation between prescription and syndrome ". It revealed the meaning of liver stagnation and spleen deficiency. And it provided an important experimental basis for the study of mechanism of prescription and Chinese medicine.