Study On Syndrome Evidence Of TCM Syndrome Of Type 2 Diabetic Rats And Correlative Study Of Traditional Chinese Medicine Intervention

In Chinese medicine system, "Correlation between Formula and Syndrome" is not only an important academic proposition in formula of TCM but also the experiential foundation of clinical treatment based on syndrome differentiation. Currently, the studies on correlation between formula and syndrome are from the perspective of static state of TCM syndromes. There are few researches carrying out it from dynamic evolutionary of syndromes or the development of disease along with time.Our discipline has adopted high glucose and fat diet together with peritoneal injection of STZ to copy Type 2 Diabetes model before. The research has observed that the model presented different TCM syndromes during model development which contained that the Pathology background was insulin resistance, the basic pathogenesis was simultaneous deficiency of both qi and yin, and the evolution tendency were simultaneous deficiency of both qi and yin, simultaneous deficiency of both qi and yin concurrent phlegm turbidness, simultaneous deficiency of both qi and yin concurrent phlegm turbidness and blood stasis. The results hinted that this model could be a useful model which was combined disease and syndrome model that presented dynamic evolutionary of TCM syndromes in different phases during the pathological development on Type 2 Diabetes. The research takes modern research of etiology and pathology of Type 2 Diabetes in western medicine, modern connotation of TCM syndromes and effective formulas of TCM on Type 2 Diabetes as logic basis. During replicated the rats model of Type 2 Diabetes, by observing appearance and behavior of the rats model and analyzing multiple indicators of experiment in biological system concomitantly corresponding TCM syndromes, we explored the dynamic evolutionary progression of syndromes and pathology; On this basis, we selected some formulas which correlated with TCM syndromes on some extent and influenced on the rats model in different phases to discuss the modern connotation of TCM syndromes and the biological foundation of "Correlation between Formula and Syndrome" and the mechanism of effective formulas.This paper is divided into two parts:literature review and experimental study. The literature review mainly includes two aspects:the research progress on mechanism of Type 2 Diabetes; TCM syndromes of Type 2 Diabetes and the relationship between syndromes and physiological modern indexes. Experimental study mainly include three aspects:the changes in different phases of TCM syndrome on rat model of Type 2 Diabetes; study on characterization of TCM syndrome in different phases in the evolution of rat model of Type 2 Diabetes; comparison of formulas effect on rat model of Type 2 Diabetes in different phases after dynamic intervention.Research 1:Dynamic evolutionary of TCM syndromes in different phases on rat model of Type 2 DiabetesMethods:The male SD rats were randomly divided into normal group and model group with 75 each group. The control group was treated with basic diet and the model group was fed with high fat and high glucose diet, continuous 5w. STZ was injected in model rats in the 6 week. After 3 days, rats with FBG≥11.1mmol/L were selected into model group.12 rats were randomly selected from the normal group and model group, and serum, plasma or tissues were prepared by abdominal aortic blood. Fins, IDE, GC, SS, iNOS, Leptin, CRP, NF-κB, TC, TG, HDL-C, LDL-C, Hcy, Renin, RPB4, cAMP/cGMP, SOD, FFA, Apo-A, FIB, TXB2,6-keto-PGF1a, Na+-K+-ATPase Muscle-Ins R, Muscle- GLUT4, Liver-Ins R, Liver-GLUT2, Fat-Ins R, Fat-GLUT4 were measured. After the model group were fed with high fat and high glucose diet after 8w, 10w and 12w, Fins, HbA1c, IDE, GC, SS, iNOS, Leptin, CRP, NF-RB, TC, TG, HDL-C, LDL-C, Hey, Renin, RPB4, Resistin, Musclin, Amylin, CORT, ACTH, cAMP/cGMP, SOD, FFA, Apo-A, FIB, TXB2,6-keto-PGF1α, Na+-K+-ATPase, Muscle-Ins R, Muscle-GLUT4, Liver-Ins R, Liver-GLUT2, Fat-Ins R, Fat-GLUT4 were measured. The appearance and behavior such as fur, behavior, tongue, tail, weight, grip strength, BP, diet intake, water intake, defecation output and urine output were observed.Results:(1) Appearance and behavior:Compared with the normal group, the body weight and grip strength in model group were decreased significantly during 10-12w (P< 0.01), and the blood pressure was increased significantly during the 5w+3d-12w. Compared with the normal group, the amounts of diet intake, water intake, Defecation and urine in model group were significantly increased during 5w+3d-12w (P< 0.01). Compared with the normal group, the changes of integral in fur, behavior, tongue, tail were increased significantly during 5w+3d-12w (P<0.01).(2)Based indicators:Compared with the normal group, FPG, IRI were significantly increased (P<0.01) during 5w+3d-12w, and Fins, HbAlc were significantly increased (P<0.01) during 8-12w, IAI was decreased significantly (P<0.01) during 5w+ 3d-12w.(3)Regulation factors of insulin:Compared with the normal group, the IDE and GC in model group were increased significantly, and SS was decreased significantly (P<0.01) during 8-12w.(4)Lipid metabolism:Compared with the normal group, TG, TC, LDL-C during 10-12w were significantly increased (P<0.01), HDL-C was significantly during 10-12w (P< 0.01); Compared with the normal group, FFA was decreased significantly in model group during 5w+3d-12w (P<0.01), and Aop-A was decreased significantly in model group during 10-12w (P<0.01).(5) Blood enzyme:Compared with the normal group, Na+-K+-ATPase, SOD in model group were significantly decreased during 5w+3d-12w (P<0.01), iNOS was increased significantly during 5w+3d-12w (P<0.01).(6)Cell factors:Compared with the normal group, model group rats CRP, Leptin were significantly increased in model group during 5w+3d-12w (P<0.01); NF-κB was significantly increased in model group during 8-12w (P<0.01); RPB4 was significantly increased in model group during 10-12w (P <0.01).(7)Vascular regulation factors:Compared with the normal group, Hcy, FIB were significantly increased in model group during 10-12w (P<0.01); TXB2, Renin were increased significantly in model group at 12w (P<0.01); 6-keto-PGF1 was decreased significantly in model group at 12w (P<0.05).(8) The HPA axis:Compared with the normal group, ACTH, CORT were significantly increased in model group during 8-12w (P<0.01). (9)Others:Compared with the normal group, cAMP was decreased significantly in model group during 8-12w (P<0.01); cGMP, cAMP/cGMP, Resistin, Musclin were significantly increased in model group during 8-12w (P<0.01).(10)Indexes related IR:Compared with the normal group, Glut-2 and Ins R of liver were decreased significantly in model group during 5w+3d-12w (P<0.01); GLUT4 and Ins R of fat were decreased significantly in model group during 10-12w (P<0.01); GLUT4 and Ins R of muscle were decreased significantly in model group at12w (P<0.01).Conclusion:The rats model of T2DM which feeding high glucose and fat diet together with peritoneal injection of STZ appeared typical characteristics of T2DM in clinic:"A little more than three", hyperglycemia, hyperinsulinemia, high glucagon, hyperlipidemia, hypertension, and with grip reduce. The organ has appeared IR from liver, adipose tissue to muscle tissue. The dynamic evolutionary progression of syndromes and pathology in different phaseses are simultaneous deficiency of both qi and yin→simultaneous deficiency of both qi and yin concurrent phlegm turbidness→simultaneous deficiency of both qi and yin concurrent phlegm turbidness and blood stasis.Research 2:Study on characterization of TCM syndrome in different phases in the evolution of rat model of Type 2 Diabetes based on data miningMethods:Intuitive analysis, decision tree and information entropy analysis were adopted to analyze the data of Type 2 Diabetes in different latitudes.Results:(1) Intuitive analysis:1)There is a significant difference between the normal group and model group in BP, Food, Water, Defecation, Urine, FPG, IAI, IRI, FFA, Leptin, iNOS, Na+-K+- ATP, SOD,CRP,Liver-GLUT2,Liver-InsR,TC,Fat-Ins R and IDE at 5w+3d; 2) There is a significant difference between the normal group and model group in BP, Food, Water, Defecation, Urine, FPG, IAI, IRI, FFA, Leptin, iNOS, Na+-K+-ATPase, SOD, CRP, Liver-GLUT2, Liver-Ins R, Fins, GC, cAMP/cGMP, NF-κB, HbA1c, CORT, Resistin, Musclin, ACTH, IDE, SS at 8w; 3) There is a significant difference between the normal group and model group in BP, Food, Water, Defecation, Urine, FPG, IAI, IRI, FFA, Leptin, iNOS, Na+-K+-ATPase, SOD, CRP, Liver-GLUT2, Liver-Ins R, Fins, GC, cAMP/cGMP, NF-κB, HbAlc, CORT, Resistin, Musclin, ACTH, IDE, SS, Fat-Ins R, weight, Grip, TG, TC, HDL-C, LDL-C, Apo-A, FIB, Hcy, RPB4, Fat-GLUT4 at 10w;4) There is a significant difference between the normal group and model group in BP, Food, Water, Defecation, Urine, FPG, IAI, IRI, FFA, Leptin, iNOS, Na+-K+-ATP, SOD, CRP, Liver- GLUT2, Liver-Ins R, Fins, GC, cAMP/cGMP, NF-κB, HbAlc, CORT, Resistin, Musclin, ACTH, IDE, SS, Fat-Ins R, weight, Grip, TG, TC, HDL-C, LDL-C, Apo-A, FIB, Hcy, RPB4, Fat-GLUT4, TXB2,6-keto-PGF1a, Renin, Muscle-Ins R, Muscle-GLUT4, Amylin at 12w.Compared with 5w+3d, the differences of BP, Weight, Water, Urine, IAI, IRI, SOD, Liver-Ins R, Defecation, cAMP/cGMP, iNOS, FPG, Fins, Renin, Food in 8w were significant; Compared with 8w, the differences of Weight, BP, Grip, Water, Food, Urine, IAI, IRI, TG,TC,HDL-C, LDL-C, Apo-A,FIB, SOD, Liver-Ins R in 10w were significant; Compared with 10w, the differences of BP, Grip, TXB2, LDL-C, Fat-GLUT4, Muscle-Ins R, Muscle-GLUT4, Amylin in 12w were significant. (2) Decision tree analysis:There were four nodes in normal group and model group, and IAI, BP, Weight, cAMP/cGMP were involved. The normal group and four model groups could be separated by IAI; M1 (the first phase of model group) and M2(the second phase of model group) could be separated by IAI and cAMP/cGMP; M3(the third phase of model group) and M2(the second phase of model group) could be separated by IAI, BP, Weight; M3(the third phase of model group) and M4(the fourth phase of model group) could be separated by IAI, BP. The correct classification rate was 98.75%. There were three nodes in models group, and BP, Weight were involved. M3 and M2 could be separated by IBP, Weight; M4 and M1 could be separated by BP.(3) Information entropy analysis:indexes with a strong discrimination between normal groups and model groups:1) M1 and N1 could be distinguished by IRI, IAI, FGP, SOD, CRP, Leptin, Water, Gripe, iNOS, Defecation; 2) M2and N2 could be distinguished by IRI, HbAlc, IAI, BP, FPG, cAMP/cGMP, Resistin, CORT, FFA, iNOS, CRP, SOD, Musclin, Leptin, Fins, NF-κB, Gripe, Defecation, Na+-K+-ATPase, Urine, Food, Water; 3) M3and N3 could be distinguished by IRI, BP, Weight, IAI, CORT, CRP, HbAlc, FPG, cAMP/cGMP, iNOS, Resistin, Leptin, TG, FIB, FFA, LDL-C, NF-κB, HDL, TC, Gripe, Apo-A, Food, Na+-K+-ATPase, SOD, Urine, Defecation, Water; 4) M4 and N4 could be distinguished by IRI, BP, IAI, HbAlc, Weight, FPG, Resistin, GC, Fins, TC, CORT, cAMP/ cGMP, Fat-GLUT4, TXB2, TG, Fat-InsR, iNOS, SOD, FFA, Liver-GLUT-2, Muscle-InsR, IDE, Muscle-GLUT4, Defecation, Urine, Water, Gripe, Food, Na+-K+-ATPase, FIB, Apo-A. Indexes with a strong discrimination between model groups:M1 and M2 could be distinguished by cAMP/cGMP, BP, Weight, FPG, IRI, Urine, Food, Gripe, IAI, Water; M2 and M3 could be distinguished by Weight, FPG, Apo-A, TG, LDL, FIB, Gripe, Urine, Water; M3 and M4 could be distinguished by BP, Muscle-GLUT4, Muscle-InsR.Conclusion:Based on the evolvement characteristics of TCM syndrome in model, the indexes groups through different phases were correlated with TCM syndromes. And the pathophysiological significances of those indexes could provide the connotation for Type 2 Diabetes. Corresponding to the modern TCM syndromes, characterizations could be indexes groups which contain one or multiple cross-system indexes. It is worth to further study that still is there a correlation between pathophysiological characteristics.Research 3:Effect of formulas on the rat model with Type 2 Diabetes in different phases after dynamic intervention.Methods:The male SD rats were randomly divided into normal group and model group. The control group was treated with basic diet and the model group was fed with high fat and high glucose diet, continuous 5w. STZ was injected in model rats in the 6 week. After 3 days, FBG≥11.1mmol/Lo rats were selected into model group.12 rats were randomly selected from the normal group and model group, and serum or plasma was prepared by abdominal aortic blood. From the 7th week, the rats of first phase intervention of Chinese medicine were divided into formula A group, formula B group, formula C group, the western medicine group was administrated Pioglitazone for two weeks. From the 9th week, the rats of first phase intervention of Chinese medicine were divided into formula A group, formula B group, formula C group, the western medicine group was administrated Pioglitazone for two weeks. From the 11th week, the rats of first phase intervention of Chinese medicine were divided into formula A group, formula B group, formula C group, the western medicine group was administrated Pioglitazone for two weeks. Serum, plasma indexes were measured at different phases.Results:At the first phase (7-8w), compared with the normal group, BP,the amounts of diet intake, water intake, Defecation, urine, FGB、Fins、IRI,IDE, GC, iNOS,CRP, Leptin, FFA, cGMP were increased significantly; IAI, SS, Na+-K+-ATPase, SOD, cAMP, cAMP/ cGMP were decreased significantly. Formula A could improve the BP, FGB, Fins, IRI; IDE, GC; Na+-K+-ATPase, SOD, iNOS, CRP, Leptin, FFA, cAMP, cGMP, cAMP/cGMP, which had changed significantly in model group. Formula B could improve the BP; Fins, IRI, IDE; Na+-K+-ATPase, SOD, iNOS; CRP; cGMP, cAMP/cGMP, which had changed significantly in model group. Formula C could improve the BP, FGB, IRI, Na+-K+-ATPase, SOD, iNOS, CRP, cGMP, cAMP/cGMP, which had changed significantly in model group.At the second phase (9-10w), compared with the normal group, BP, the amounts of diet intake, water intake, Defecation, urine, FGB, Fins, IRI, IDE, GC, iNOS, CRP, Leptin, RPB4, TG, TC, LDL-C, FFA, FIB, Hcy, cGMP were increased significantly; weight, grip, IAI, SS, Na+-K+- ATPase, SOD, HDL-C, Apo-A, cAMP, cAMP/cGMP were decreased significantly. Formula A could improve the weight, BP, Fins, IRI, Na+-K+-ATPase, SOD, iNOS, CRP, which had changed significantly in model group. Formula B could improve the weight, diet intake, BP, FGB, Fins, IRI, IDE, SS, Na+-K+-ATPase, SOD, iNOS, CRP, Leptin, TG, TC, LDL-C, HDL-C, RPB4, FFA, Apo-A, Hcy, cGMP, cAMP/cGMP, which had changed significantly in model group. Formula C could improve the weight, BP, Fins, IRI, IAI, IDE, SS, Na+-K+-ATPase, SOD, iNOS, CRP, Leptin, TG, TC, LDL-C, FFA, cGMP which had changed significantly in model group.At the third phase (11-12w), compared with the normal group, BP, the amounts of diet intake, water intake, Defecation, urine, FGB, Fins, IRI, IDE, GC, iNOS, CRP, Leptin, RPB4, TG, TC, LDL-C, FFA, FIB, Hcy, TXB2, cGMP were increased significantly; weight, grip, IAI, SS, Na+-K+-ATPase, SOD, HDL-C, Apo-A,6-Keto-PGF1a, cAMP, cAMP/cGMP were decreased significantly. Formula A could improve the BP, grip, diet intake, water intake, FGB, IRI, Na+-K+-ATPase, SOD, iNOS, which had changed significantly in model group. Formula B could improve the weight, diet intake, BP, grip, diet intake, water intake, urine, defecation, FGB, Fins, IRI, IAI, Na+-K+-ATPase, SOD, iNOS, TG,TC, FIB, cGMP, cAMP/ cGMP, which had changed significantly in model group. Formula C could improve the weight, grip, diet intake, water intake, Defecation, urine, BP, FGB, Fins, IRI, IAI, IDE, GC, SS, Na+-K+- ATPase, SOD, iNOS, Leptin, TG,TC,LDL-C, HDL-C,FFA, Apo-A, FIB, Hcy, TXB2, 6-K.eto-PGFla, cAMP, cGMP cAMP/cGMP which had changed significantly in model group.Conclusion:There is a certain effect of Formula A, Formula B and Formula C on syndrome in three stages. Compared the whole affection of Formula A, Formula B and Formula C, we found that the intervention of formula A are better than Formula B and Formula C in the first phase, the intervention of formula B are better than Formula A and Formula C in the second phase, the intervention of formula C are better than Formula A and Formula A in the third phase. The results show that, Formula A, Formula B and Formula C have certain relevance with syndromes in three stages, while the correlation degree is different. Formula A, Formula B and Formula C has higher correlation degree with syndromes in the first stages, the second stages, the third stages respectively. The result confirms the experience of prevention and treatment of tonifying qi and yin for deficiency of both qi and yin syndrome, simultaneous tonifying qi and yin concurrent reduce phlegm for simultaneous deficiency of both qi and yin concurrent phlegm turbidness, simultaneous tonifying qi and yin concurrent reduce phlegm invigorate the circulation of blood for simultaneous deficiency of both qi and yin concurrent phlegm turbidness and blood stasis.