| Objective:Reference of description about primordial qi in TCM ancient classics,we consider resection of right kidney as model of deficiency of vital energy,and consider resection of left kidney as the control group.We used Guben Jiannao prescription as a intervention,to observe the klotho(kl)protein and related protein expression in kidney and hippocampus.To explore the effects of deficiency of vital energy on renal function and memory function,and the intervention effect of consolidating vital base,supplementing and brain tonic method on kidney and hippocampus and mechanism.Method:1.Theoretic discussion: By sorting out the ancient literature of TCM and research progress,we analysis the development and function of the vitality of TCM,the relationship between modern diseases and vital energy,and the reference significance of other deficiency animal models of TCM for the establishment of the animal model of deficiency of vital energy,and providing a theoretical basis for the establishment of the animal model.2.Experimental study: The blank group was used as the control group,and the unilateral nephrectomy rats were used as the animal model of deficiency of vital energy.The rats were randomly divided into blank group,model group,left kidney group,Guben Jiannao prescription high-dose group(Gugao group),Guben Jiannao prescription medium-dose group(Guzhong group)and Guben Jiannao prescription low-dose group(Gudi group).To explore the feasibility of unilateral nephrectomy as an animal model of deficiency of vital energy with kl protein as the entry point;to explore the relationship between vital gate and kidney with left and right nephrectomy respectively;to observe the changes of renal function,blood routine,kl protein,TNF-a,IL-6 and 8-OHDG in unilateral nephrectomy and the intervention effect of Guben Jiannao prescription;to explore the animal model of deficiency of vital energy through spontaneous activity and Morris water maze.The expression of kl,NF-kappa B and MAPK1 in kidney,hippocampus and the pathological changes of kidney and hippocampus were observed by PCR and Western blot.Results:1.Generally,the hair of unilateral nephrectomy rats is withered and yellow,the activity is reduced,some of them are restless,loose stool and wheezing.There is no obvious difference between left nephrectomy and right nephrectomy.There was no significant change in the blank group.No mice died in the blank group,three in the model group,one in the left kidney group,one in the Gugao group and one in the Gudi group,and two in the Guzhong group.(2)The comparison of body weight and kidney weight at different time points in each group: there was no significant difference in body weight before operation(p>0.05);one week after operation,the body weight of Gugao group was lower than that of Gudi group(p<0.05);and there was no significant change in body weight between groups at 2,4,6 and 8 weeks after operation(p>0.05).After 8 weeks of execution,the kidney weight of model group,left kidney group,Gugao group,Guzhong group and Gudi group were higher than that of blank group(p<0.05).(3)There was no significant difference in renal function,blood routine test and model group between each group(p>0.05).Compared with the model group,left kidney group and Gudi group were higher than blank group(p<0.05);left kidney group and Gudi group were lower than model group(p<0.05);Guzhong group and Gugao group were lower than model group,left kidney group and Gudi group(p<0.05).There was no difference in WBC and RBC among the groups(p>0.05);Hb in model group and left kidney group was lower than that in blank group(p<0.05);Gugao group and Guzhong group were higher than that in model group and left kidney group(p<0.05).2.Behavioral study and intervention effect of consolidating vital base,supplementing and brain tonic method in rats with deficiency of vital energy:(1)The total distance of autonomous activity in model group and left kidney group was lower than that in blank group(p<0.01),while that in Gugao group,Guzhong group and Gudi group was lower than that in blank group(p<0.05),and that in Gugao group and Guzhong group was higher than that in model group and left kidney group(p <0.05).The rest time of model group and left kidney group was higher than that of blank group(p<0.01),while that of Gugao group,Guzhong group and Gudi group was higher than that of model group and left kidney group(p< 0.05),while that of Gugao group,Guzhong group and Gudi group was higher than that of blank group(p<0.01),and that of Gugao group,Guzhong group and Gudi group was lower than that of model group and left kidney group(p< 0.05).Fewer than the blank group(p<0.01),the Gugao group,the Guzhong group and the Gudigroup were fewer than the blank group,and more than the model group and the left kidney group(p<0.05).(2)The results of Morris water maze positioning navigation and space exploration experiment in rats: the total distance of finding platform in 90 s swimming was higher in model group and left kidney group than in blank group(p<0.01),higher in Gugao group,Guzhong group and Guzhong group than in blank group,lower than in model group and left kidney group(p<0.05);the average speed was slower in model group and left kidney group than in blank group(p<0.01),higher in Guzhong group and Guzhong group than in blank group(p<0.01).The first time arrival time of model group and left kidney group was more than that of blank group(p<0.01),while that of Gugao group,Guzhong group and Gudi group was slower than that of blank group(p<0.05),faster than that of model group and left kidney group(p<0.05);the times of crossing platform were less in model group and left kidney group than that in blank group(p<0.01),Gugao group,Guzhong group and Gudi group were less than blank group(p< 0.05),Gugao group and Guzhong group were more than model group and left kidney group(p<0.05).3.Inflammatory factors,oxidative stress response and intervention of consolidating vital base,supplementing and brain tonic method in rats with deficiency of vital energy: Compared with the blank group,the levels of serum kl protein in model group and left kidney group were lower than those in blank group(p<0.01),while those in Gugao group,Guzhong group and Gudi group were lower than those in blank group,higher than those in model group and left kidney group(p<0.05).TNF-a in model group and left kidney group was higher than that in blank group(p< 0.01),while that in Gugao group,Guzhong group and Gudi group was lower than that in model group and left kidney group(p<0.05).IL-6 in model group and left kidney group was higher than that in blank group(p< 0.01,0.05),in Gugao group,Guzhong group and Gudi group was higher than that in blank group and model group(p<0.05),and in Gugao group was lower than that in left kidney group(p<0.05).Compared with the blank group,8-OHDG in model group and left kidney group was higher(p<0.01);Gugao group was lower than model group and left kidney group and Gugao group(p<0.05);Guzhong group and Gudi group were higher than blank group,lower than model group and left kidney group(p<0.05).4.The expression of kl,MAPK1 and NF-kappa B in kidney and hippocampus of rats with deficiency of vital energy and the intervention effect of consolidating vital base,supplementing and brain tonic methodReal-time PCR comparison of kidneys:(1)Compared with model group,left kidney group,Gudi group were lower than blank group(p<0.01),Guzhong group and Gugao group were lower than blank group(p<0.05),Guzhong group and Gugao group were higher than model group and left kidney group(p<0.05).(2)Compared with the blank group,the expression of NF-kappa B in model group and left kidney group was higher(p<0.01),while that in Gugao group,Guzhong group and Gudi group was higher(p<0.05),lower than that in model group and left kidney group(p<0.05).(3)Compared with the blank group,the expression of MAPK1 in model group and left kidney group was higher than that in blank group(p<0.01),while that in Guzhong group,Guzhong group and Gudi group was higher than that in blank group(p<0.05),while that in Guzhong group and Gugao group was lower than that in model group and left kidney group(p<0.05).Real-time PCR comparison of hippocampus:(1)Compared with model group,left kidney group,Guzhong group,Guzhong group were lower than blank group(p<0.01),Gugao group was lower than blank group(p<0.05),Guzhong group,Gugao group were higher than model group(p<0.05),Guzhong group and Gugao group were higher than left kidney group(p<0.05).(2)Compared with the blank group,the expression of NF-kappa B in model group and left kidney group was higher than that in blank group(p<0.01),while that in Gugao group,Guzhong group and Gudi group was higher than that in blank group,lower than that in left kidney group(p<0.05).(3)Compared with the blank group,the expression of MAPK1 in the model group and left kidney group was higher(p<0.05),while that in the low,middle and high groups was lower(p<0.05).Western blot comparison of kidneys:(1)Compared with the blank group,the model group and left kidney group were lower than the blank group(p< 0.01),while Gudi group,Guzhong group and Gugao group were lower than the blank group(p<0.05);Guzhong group and Gugao group were higher than the model group and the left kidney group(p<0.05).(2)Compared with the blank group,the levels of NF-?B P65 in the model group and left kidney group were higher than those in the blank group(p<0.01),while those in Gudi group,Guzhong group and Gugao group were higher than those in the blank group(p< 0.05),and lower than those in the model group and the left kidney group(p< 0.05).(3)Compared with the blank group,the model group and blank group were higher than the blank group(p<0.05),while Gudi group,Guzhong group and Gugao group were lower than the model group and the left kidney group(p<0.05).Western blot comparison of hippocampus:(1)Kl in model group and left kidney group was lower than that in blank group(p<0.01),Gudi group and Guzhong group were lower than that in blank group(p<0.05);Gudi group,Guzhong group and Gugao group were higher than that in model group and left kidney group(p<0.05).(2)Compared with the blank group,the level of NF-kappa B in the model group and left kidney group was higher than that in the blank group(p<0.01),while that in Gudi group was higher than that in the blank group(p<0.05),and that in Gudi group,Guzhong group and Gugao group was lower than that in the model group and the left kidney group(p<0.05).(3)Compared with the blank group,the levels of MAPK1 in the model group,left kidney group,Gudi group,Guzhong group and Gugao group were higher than those in the blank group(p<0.05).5.Pathological changes of kidney and hippocampus in rats with unilateral nephrectomy and the intervention effect of consolidating vital base,supplementing and brain tonic methodKidney light microscopy: The glomeruli and tubules of rats in the blank group were normal.In model group,glomerular proliferation,mesangial matrix proliferation,cell proliferation,erythrocyte granules in cytoplasm,necrosis and swelling of proximal convoluted tubular epithelial cells were observed.In left kidney group,glomerular hypertrophy,cell proliferation,Baumann's cavity stenosis,renal tubule morphology was basically normal,brush-like margin was clear.The glomerular hypertrophy,erythrocyte proliferation,endothelial cell proliferation,telangiectasia and renal interstitial no significant changes were observed in the Gudi group.In Guzhong group,glomerular hypertrophy,mesangial matrix hyperplasia and renal interstitium did not change significantly.In Gugao group,glomerular hypertrophy,mesangial hyperplasia,capillary endothelial cell proliferation and interstitial hyperemia were observed.Hippocampal light microscopy: The pyramidal cells in the blank group were arranged regularly without obvious pathological changes;the pyramidal cells in the model group were not arranged regularly,and the pyramidal cells degenerated(cell swelling,cytoplasmic light staining),necrosis(nuclear pyknosis);the pyramidal cells in the left kidney group were arranged at a level that was still acceptable,and the degree of degeneration and necrosis was lighter than that in the model group;the pyramidal cells in the Gudi group were still capable of degeneration and necrosis.The degree of necrosis was lighter than that of model group and left kidney group;the pyramidal cell layers in Guzhong group were still acceptable,and the degeneration and necrosis were lighter than those in Guzhong group;the pyramidal cell layers in Gugao group were acceptable,and the degree of degeneration and necrosis was lighter.Conclusion:1.Unilateral nephrectomy rats showed similar symptoms to deficiency of vital energy,which could be used as one of animal models of deficiency of vital energy.2.Consolidating vital base,supplementing and brain tonic method can improve the function of kidney and brain in rats with deficiency of vital energy.Its mechanism may be related to the promotion of the expression of Kl protein,thus inhibiting inflammation and oxidative stress.3.Kl protein may be used as one of the reference indicators of deficiency of vital energy,and inflammation and oxidation may be the main causes of damage to vital energy.4.There are not differences between the left kidney and the right kidney,so the theory that the left kidney is the kidney and the right kidney is vital gate still needs further research to confirm. |
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