| [Background]Yin-yang theory is the foundation and source of traditional Chinese medicine theory, however, its scientific connotation has not been fully expounded so far. Plain question recorded that Yin and Yang is the way of heaven and earth. Social order and law of all things, parents of change, the beginning of life and death, the house of spirit. So it should be searched for the primary cause of disease in treatment. It points out that the movement of the opposites and unity of Yin and Yang are the root cause of the occurrence of all things int the world, including the development, changes and die. Wang bing said that the spirit root of Yang-qi was in Yin, Yin-qi root was in Yang, Yang cannot born without Yin, Yin cannot born without Yang. It means Yin and Yang not only have opposites relations, also have the relationship of unity, It’s the famous theory of interdependence of Yin and Yang. In Ming dynasty Zhang Jing-yue put forward the famous idea of generating Yin by activating Yang and generating Yang by activating Yin. At the same time he produced Zuogui pill and Yougui pill to treat the Kidney-Yin deficiency syndrome and Kidney-Yang deficiency syndrome respectively. Currently the mechanism of mutual root interoperability between Yin and Yang of kidney has not yet been elucidated.The most concerned problem is cAMP and cGMP hypothesis in mordern research of Yin-yang theory which put forward for the first time in 1975 by American biologist Glodberg. Followed by famous scientific journals of Cell,Science, Nat Cell Biol, Nat Rev Rheumatol, Proc Natl Acad Sci USA and Circ Res published to explore the immune and inflammation, oxidation and anti-oxidation, protein phosphorylation signaling pathways, nuclear transcription factor, etc. The results shows the universality of the theory of Yin and Yang.There were a lot of reports about the function of antagonistic substances and seldom relationship of mutual rooting of Yin-Yang theory.The basic theory of traditional Chinese medicine is classic, mainly depends on its effects could be conducted repectedly and tested in clinic. As modern scholars, we should observe the effects,genes,immunity to explore the mechanism of basic theory of traditional Chinese medicine.The syndrome of Traditional Chinese Medicine is the summarization of etiology and pathogenesis in different stage of diseases. Since the same symptome have common clinical manifestation and pathological mechanism, then they may have common material foundation. According to the central dogma of life, the genetype(DNA or gene) is the most primary and most essential matter of life. The traditional Chinese medicine considers that the kidney is the congenital foundation, stores the spirit, charges the development, growth, reproduction and fluid metabolism. Therefore,gene expression offered unprecedented opportunity for research the essence of TCM Syndrome.In our prior study works we successfully set up cDNA Lib of Kidney-yin deficiency (including diabetes, chronic nephritis, lupus nephropathia and sub-health), developed study on gene expression profiles of Kidney-yin deficiency and Kidney-yang deficiency by some techniques and methods, such as RNA microamount-amplification,differential hybridization and gene cloning. The results showed that Kidney-yin deficiency of the different diseases in the subtractive library card there is an obvious difference, and Kidney-yin deficiency and evidence of the disease must be interference. There is characteristic profiles of differential expressed genes between Kidney-yin deficiency group and normal control group, a initial evidence of Kidney-yin deficiency-related genes, a total of 79.There is characteristic profiles of differential expressed genes between Kidney-yang deficiency group and normal control group, a initial evidence of Kidney-yang deficiency-related genes, a total of 75, there are 9 common differential expressed genes of Kidney-yin and Kidney-yang deficiency syndrome.The results showed that the Kidney-Yin deficiency and Kidney-Yang deficiency syndrome were of different gene expression regulation network, show the opposite restriction relationship; However, they have a common gene expression regulation network, showed that kidney Yin and Yang have a common material base, which means the relationship of mutual root interoperability.To sum up, this study proposed to using modern genetic techniques,combined with clinic observation of Zuogui pill and Yougui pill and decomposition study to treat the Kidney-Yin deficiency syndrome and Kidney-Yang deficiency syndrome, further verify the theory of yin-yang mutual root of interoperability, explore the essence of kidney deficiency, provided the scientific basis for the research of TCM syndrome.[Methods]1.To make systematic review and summary of the Chinese traditional medicine theory of Mutual Rooting of Yin-Yang Theory in Kidney and the present situation of the Kidney-Yang deficiency and Kidney-Yin deficiency syndrome;2.Using the method of "disease combined with syndrome",the patients with chronic glomerulonephritis and diabetes mellitus of Kidney-Yin deficiency, Kidney-Yang deficiency syndrome, healthy normal people with similar demographic data such as gender and age, were chosen as object of study. According to the method of generating Yin by activating Yang and generating Yang by activating Yin, we developed the clinic observation of Zuogui pill and Yougui pill and decomposition study to treat the Kidney-Yin deficiency syndrome and Kidney-Yang deficiency syndrome; the patients of Kidney-Yin deficiency syndrome were randomly divided into treatment group of generating Yin by activating Yang(namely Zuogui pill) and treatment group of nourishing Yin; the patients of Kidney-Yang deficiency syndrome were divided into treatment group of generating Yang by activating Yin(namely Yougui pill) and treatment group of nourishing Yang.3.By lymphocyte transformation test, determination of NK cell activity, to observation the Zuogui pill group,nourishing Yin group, Yougui pill group, nourishing Yang group and blank control group, carry out the immune analysis of the experimental groups.4.According to the theory of Chinese medicine measuring Chinese syndrome, the Yougui pill and nourishing Yin compound, Zuogui pill and nourishing Yang compound were divided into treatment group, those healthy people were divided into normal control group.Collecting samples and using the RT-PCR to observe gene expression, we study the gene expression spectrum of white blood cell of Kidney-Yin deficiency syndrome and kidney-Yang deficiency syndrome, we obtained the therapeutical effect related genes of Yougui pill and Yougui pill by statistical analysis.[Results]1. Summarized the renal mutual rooting of Yin-Yang theory in traditional Chinesemedicine;2.Clinical observations: (i) the patients of Kidney-Yin deficiency syndrome were randomly divided into treatment group of generating Yin by activating Yang(namely Zuogui pill) and control group of nourishing Yin for clinical observation:(a) Observation of the main clinical symptoms and tongue and pulse picture: The main clinical symptoms of 60 patients of kidney-yin deficiency syndrome were as follows:52 cases of feverish feeling in palms and soles(86.7%),33 cases (55%) of waist ache, soft legs and knees 46 cases (76.7%),43 cases (71.7%) of dysphoria with feverish sensation in chest, palms and soles; 35 cases (58.3%)of hot flash,28 cases (46.7%)of night sweat,23 cases (38.3%) of dry throat,15 cases (25%) of insomnia, 18 cases (60%) of tinnitus. In addition, there was a little bit of loose teeth, memory recession,etc.Tongue picture:Within the 60 cases of kidney-yin deficiency syndrome,46 cases(76.7%) with reddish tongue,9 cases(15%) with wrinkled tongue,5 cases of (8.3%) with dark red tongue and tongue petechiae. Tongue coating appears most frequently thin white with 43 cases(71.7%), followed by absence of tongue coating with 11 cases(18.3%), thin yellow tongue coating with 5 cases(8.3%) and yellow greassy moss with 1 cases(1.7%).Pulse condition:Within the 60 cases of kidney-yin deficiency syndrome,28 cases appeared rapid pulse(46.7%),21 cases with thready pulse(35%),8 cases(13.3%) of string pulse and 3 cases(5%) with slow pulse. According to the nature of the pulse condition, kidney-yin deficiency syndrome pulse characterized by thin and rapid pulse.(b) Comparsion of Clinic effects of the two groups:By analysis of the rank inspection, Z=-1.975, P=0.048<0.05, it had a statistically significant difference between the two group, showed that the effects of Zuogui pill group was significantly superior than nourishing Yin group (P<0.05)Comparsion of symptoms integral in the two groups pretherapy and post-treatment:The treatment of symptom integral compared with paired t-test in the group and ANOVA between the groups. Comparion of post-treatment symptom integral using analysis of covariance. The results show that the symptom integral had statistical significance pretherapy and post-treatment of the two groups (tt=18.139, tc=7.732, P<0.01). There is statistically significant difference between the treatment and control groups pretherapy and post-treatment (F=62.413, P<0.01).It showed that the treatment group was superior to control groups in changes of symptoms,(ii)the patients of Kidney-Yang deficiency syndrome were divided into treatment group of generating Yang by activating Yin(namely Yougui pill) and control group of nourishing Yang for clinical observation:(a) Observation of the main clinical symptoms and tongue and pulse picture: The main clinical symptoms of 60 patients of kidney-yang deficiency syndrome were as follows:55 cases intolerance of cold(91.7%), waist ache,51 cases (85%), soft legs and knees 48 cases (80%), fatigue and weakness,42 cases (70%), loss of hair,36 cases (60%), lower limbs edema,22 cases (36.7%), loss of libido,21 cases (35%), limbs cold 17 cases (28.3%), nocturia more than 16 cases (26.7%). In addition, there was a little bit of loose teeth, memory recession,etc.Tongue picture:Within the 60 cases of kidney-yang deficiency syndrome,38 cases(63.3%) with reddish tongue,12 cases(20%) with light tongue,7 cases(11.7%) with pale tongue and teeth marks,3 cases(5%) with dark red tongue and tongue petechiae. Tongue coating appears most frequently thin white with 47 cases(78.3%), followed by thin yellow with 6 cases(10%), lesee moss with 6 cases(10%) and yellow greassy moss with 1 cases(1.7%).Pulse condition:Within the 60 cases of kidney-yang deficiency syndrome,32 cases appeared deep pulse(53.3%),22 cases with thready pulse(36.7%),4 cases(6.7%) with thin and weak pulse and 2 cases(3.3%) with slow pulse. According to the nature of the pulse condition, kidney-yang deficiency syndrome pulse characterized by deficiency pulse,heavy pulse,thin and weak pulse and slow pulse. (b)Comparsion of Clinic effects of the two groups:By analysis of the rank inspection, Z=-2.062, P=0.039<0.05, it had a statistically significant difference between the two group, showed that the effects of Yougui pill group was significantly superior than nourishing Yang group.Comparsion of symptoms integral in the two groups pretherapy and post-treatment:The treatment of symptom integral compared with paired t-test in the group and ANOVA between the groups. Comparion of post-treatment symptom integral using analysis of covariance. The results show that the symptom integral had statistical significance pretherapy and post-treatment of the two groups (P<0.01) There is statistically significant difference between the treatment and control groups pretherapy and post-treatment (F=84.257, P<0.01).It showed that the treatment group was superior to control groups in changes of symptoms kidney-yang.3.The result of immune function test:(i)The lymphocyte transformation test shows:The result of lymphocyte transformation test about Zuogui pill group, nourishing Yin group and blank control group showed:Both of the SI of Zuogui pill group and nourishing Yin group decreased and there was significant difference compared with blank control group pretherapy (P<0.05). post-treatment there was no statistical significance between the group of Zuogui pill and blank control group, while there was statistical significance between the group of nourishing Yin group and blank control group.It showed that the effect of Zuogui pill group was significantly superior than nourishing Yin group. The stimulation index(SI) of lymphocyte transformation test about Yougui pill group, nourishing Yang group and blank control group showed:Both of the SI of Yougui pill group and nourishing Yang group decreased and there was significant difference compared with blank control group pretherapy (P<0.05). post-treatment there was no statistical significance between the group of Yougui pill and blank control group, while there was statistical difference between the group of nourishing Yang group and blank control group.It showed that the effect of Yougui pill group was significantly superior than nourishing Yin group.(ii) Determination of NK cell activity:The determination of NK cell activity about the three groups of Zuogui pill experiment showed that both the NK cell activity(%) of Zuogui pill and nourishing Yin group decreased pretherapy, and there was statistical difference compared with the blank control group.There was no statistical difference between the Zuogui pill and the blank control group post-treatment while there was statistical difference between the nourishing Yin group and blank control group still. It showed that the effect of Zuogui pill group was significantly superior than nourishing Yin group. The determination of NK cell activity about the three groups of Yougui pill experiment showed that both the NK cell activity(%) of Yougui pill and nourishing Yang group decreased pretherapy, and there was statistical difference compared with the blank control group.There was no statistical difference between the Yougui pill and the blank control group post-treatment while there was statistical difference between the nourishing Yang group and blank control group still. It showed that the effect of Yougui pill group was significantly superior than nourishing Yang group.(iii) Peripheral blood leukocytes, serum antibodies, complement and lymphocyte subgroup observations:T-test result of comparsion of peripheral white blood cells within the two groups showed statistically difference pretherapy and post-treatment(tt=-7.227, tc=-4.401, P<0.01); there was no statistically difference pretherapy between the two groups(F=0.229,P>0.05), and the statistically difference appeared post-treatment(F=7.708, P<0.05).It proved that Zuogui pill and nourishing Yin group can promote peripheral white blood cells, and the treatment group was better than the control group. T-test result of comparsion of peripheral white blood cells within the Yougui two groups showed statistically difference pretherapy and post-treatment(tt=-11.794, tc=-8.094, P<0.01); there was no statistically difference pretherapy between the two groups(F=2.150, P>0.05), and the statistically difference appeared post-treatment(F=48.724, P<0.05).It proved that Yougui pill and nourishing Yang group can promote peripheral white blood cells, and the treatment group was better than the control group.Comparsion of serum antibody and complement with the Zuogui two groups pretherapy and post-treatment:T-test result showes that there was no statistical significance within the two groups of serum antibody IgG, IgA, IgM and C3 and C4 comparative(P>0.05). It showed that there was not significance imfluence of serum antibody and comparative with the two groups.Comparsion of lymphocyte subsets pretherapy and post-treatment result:According to the t-test result, there was no statistical significance of lymphocyte subsets CD3, CD4, CD8 within the two groups(P>0.05). It showed that there was little effects on lymphocyte subsets for those two groups. Comparsion of serum antibody and complement with the Yougui two groups pretherapy and post-treatment:T-test result showes that there was no statistical significance within the two groups of serum antibody IgG, IgA, IgM and C3 and C4 comparative(P>0.05). It showed that there was not significance imfluence of serum antibody and comparative with the two groups.4.Fluorescence quantitative PCR test of gene expression:There were 6 common genes expression decreased between the Kidney-Yang and Kidney-Yin deficiency syndrome group pretherapy. post-treatment, both of the 6 genes expression increased of Zuogui pill and nourishing Yin group, yet still failed to reach the normal level, the statistically difference appeared post-treatment.The genes expression of Zuigui pill group increased more obvious than Nourishing Yin group, post-treatment, both of the 6 genes expression increased of Yougui pill and nourishing Yang group, yet still failed to reach the normal level.The genes expression of Yougui pill group increased more obvious than Nourishing Yang group, the statistically difference appeared post-treatment.[Conclusions]1. The clinical effect of Zuogui pill group was superior to nourishing Yin group in the treatment of Kidney-Yin, which showed that the method of generating Yin by activating Yang was superior to pure nourishing Yin. The clinical effect of Yougui pill group was superior to nourishing Yang group in the treatment of Kidney-Yang, which showed that the method of generating Yang by activating Yin was superior to pure nourishing Yang. The theory of Zhang Jingyue was verified scientifically.2. As proved in the perspective of immunology and gene expression, the Zuogui pill group was superior to nourishing Yin group and the Yougui pill group was superior to nourishing Yang group. The theory of Mutual Rooting of Yin-Yang in Kidney Deficiency was verified in immune and gene level. |
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