| Objective:Qi-blood are the foundation and core of the human body.Human body is nothing but Qi-blood.It is said in The Eight of Classic on Medical problems that "Qi is the essence of human beings." As the origin of qi "attached to the Qi-blood,kill on Qi-blood first".Primordial Qi is the source of all kinds of qi.The sufficient qi contributes to the prosperity and decline of blood.The qi due to blood deficiency is also deficient.The prosperity and decline of Qi-blood is also the reflection of the prosperity and decline of internal primordial Qi.Stem cell research is the medical field with the most rapid development,the most in-depth research and the most abundant application results in the past 50 years,and it is still developing at a rapid rate and changing with each passing day.Primordial qi is highly similar to stem cells in terms of name,source,distribution,and function.Primordial qi generates blood by generating qi.Hematopoietic stem cells are the founder and source of blood.Primitive qi is sealed and stored in kidney,which is the "kidney-to-kidney qi movement".Hematopoietic stem cells settle in bone marrow,which controls marrow.Bone marrow is also a component of kidney in traditional Chinese medicine.In this study,we constructed the mouse model of primordial qi deficiency through the deficiency of qi caused by exercise fatigue and the blood deficiency caused by the combination of acetylphenylhydrazine and cyclophosphamide,and analyzed the changes before and after treatment with Renshen Yangrong Decoction,to explore the intervention and effect mechanism of Yiqi Yangxue Method on primordial qi deficiency.Method:1.Theoretical discussion: collate the ancient literature and development context of "The Theory of Vitality in Traditional Chinese Medicine",analyze the relationship between vitality and Qi-blood,the relationship between vitality and stem cells(especially hematopoietic stem cells),and the relationship between vitality deficiency and modern diseases(especially blood-related diseases),and learn from animal models of qi deficiency and blood deficiency to provide theoretical basis and guidance for the construction of animal models of vitality deficiency.2.Experimental study: Six-week-old female Kunming mice were used to establish the animal model of qi deficiency by running exercise fatigue and injection of acetylphenylhydrazine and cyclophosphamide.The mice were intervened by Renshen Yangrong Decoction,a classic famous prescription of benefiting qi and nourishing blood.They were randomly divided into the blank group,model group,Renshen Yangrong Decoction low dose group(Ronglow group),Renshen Yangrong Decoction medium dose group(Rongzhong group)and Renshen Yangrong Decoction high dose group(Ronggao group).The general state changes of the mice before and after modeling and administration were observed.Through spontaneous activity and Morris water maze test,we explored the changes and recovery of behavior and cognitive ability in animal models with vigour deficiency,detected the changes of peripheral red blood cell count(RBC),white blood cell count(WBC),hemoglobin concentration(HGB)and platelet count(PLT),detected the expression changes of erythropoietin EPO,interleukin IL-3 and IL-6 by ELISA,and detected the bone marrow stem cells by flow cytometry,as well as the action targets of Renyangrong Decoction on anemia by network pharmacology.Results:1?General symptoms and body weight.After modeling,the mice showed such phenomena as decreased diet,slow action,malaise,loose and less lustrous fur,decreased stool excretion,and mania in some mice.After treatment with Renshen Yangrong Decoction,the mice of Ronglow group,Rongzhong group and Ronggao group showed different degrees of state recovery.As for the body weight,there was no difference among the groups on the first day before modeling(P < 0.05).After modeling,the mouse body weight was decreased(P < 0.05);After treatment with Renshen Yangrong Decoction,the body weights of mice in Ronglow group,Rongzhong group and Ronggao group were still low compared with the blank group(P < 0.05),but compared with the model group,the body weight recovery was more significant(P < 0.05).There was no significant difference in the body weights of mice among the three groups(P < 0.05).2?Experiment of autonomous activity.Compared with the mice in the blank control group at the same time,the total distance and the average speed of the mice in the model group were significantly reduced(P < 0.01),and the rest time and central time were significantly prolonged(P < 0.01).Compared with the model group,the total distance and the average speed of Rongzhong group were significantly increased(P < 0.01),but they were still lower than those of the blank group(P < 0.05).The rest time and central time were shortened(P < 0.01),but they were still longer than those in the blank control group(P < 0.05).Compared with the mice in the model group at the same time,the total distance traveled and the average speed in the Ronggao group were significantly increased(P < 0.01),which were close to those in the blank control group(P < 0.01).The rest time and central time were shortened(P <0.01),which was also close to those in the blank control group(P < 0.01).3?Morris water maze experiment.Compared with the mice in the blank control group at the same time,the total mileage,average speed and times of crossing the platform of the mice in the model group were significantly reduced,and the time to reach the platform for the first time was significantly prolonged(P < 0.01).Compared with the mice in the blank control group at the same time,the total mileage,average speed and target-platform crossing times of mice in the Ronglow group,Rongzhong group and Ronggao group were still lower,and the time to reach the target-platform for the first time was still long(P < 0.05).However,compared with the mice in the model group at the same time,the total mileage,average speed and target-platform crossing times of mice in the Ronglow group,Rongzhong group and Ronggao group were significantly increased,and the time to reach the target-platform for the first time was significantly shortened(P < 0.01).4?Blood test.Compared with the counterparts in the blank control group,the red blood cell count(RBC),white blood cell count(WBC),hemoglobin concentration(HGB),and platelet count(PLT)of the mice in the model group were significantly reduced(P < 0.05).Compared with the mice in the model group at the same time,the RBC,WBC,HGB and PLT of mice in Ronglow group,Rongzhong group and Ronggao group were significantly increased(P <0.05).Compared with the mice in the blank control group at the same time,the expression levels of EPO,IL-3 and IL-6 in the model group were significantly decreased(P < 0.01);Compared with the mice in the model group at the same time,the expression levels of EPO,IL-3 and IL-6 in the Ronglow group,Rongzhong group and Ronggao group were increased,all P < 0.05.The expression levels of EPO,IL-3 and IL-6 in the Ronggao group were higher than those in the blank group(P < 0.05).5?Hematopoietic stem cell testing.Compared with the blank group,the proportion of bone marrow hematopoietic stem cells in the empty medicine group(blank dosing group)of normal mice treated with Renshen Yangrong Decoction(the same dose as Rongzhong Decoction)was increased,P < 0.05.Compared with the blank group,the proportion of bone marrow hematopoietic stem cells in the model group was significantly decreased(P < 0.05).After treatment with Renshen Yangrong Decoction,the proportion of bone marrow hematopoietic stem cells in Ronglow group,Rongzhong group and Ronggao group was increased(P < 0.05).The proportion in Ronggao group was comparable to that in the air-drug group(P < 0.05).6?Network pharmacology.A total of 217 active components of Renshen Yangrong Decoction were screened out,including 10 core chemical active components such as quercetin,kaempferol,feruloyl tyramine and kaempferol.167 potential action targets for the treatment of anemia were obtained,of which,STAT3,IL6,VEGFA,AKT1 and EGFR were the five core targets,and20 significantly enriched signaling pathways were identified,where STAT3 and IL6 were highly correlated with the regulation of hematopoietic stem cells.Conclusion:1.Human beings are based on qi and blood.Vital qi is the source and root of qi and blood.The mouse model caused by qi deficiency due to running fatigue and blood deficiency due to injection of acetylphenylhydrazine and cyclophosphamide conformed to the syndrome of primordial qi deficiency and could be selected as one of the animal models with primordial qi deficiency.2.The method of replenishing qi and nourishing blood can treat the syndrome of primordial qi deficiency,which may be related to increasing the proportion and number of hematopoietic stem cells.Ginseng Yangrong Decoction can increase the proportion of bone marrow hematopoietic stem cells in normal mice and has a significant intervention effect on some symptoms of mice with qi deficiency,which may be related to the increase of the proportion of bone marrow hematopoietic stem cells.3.Renshen Yangrong Decoction had many active components and extensive effects.The action targets related to hematological diseases by network pharmacology analysis might be STAT3,IL-6,VEGFA,AKT1,and EGFR,which were consistent with the results of total IL-6 content in blood detected by ELISA.4.It is possible to characterize primordial qi with the quantity and activity of hematopoietic stem cells.With the in-depth study on the proportion relationship between hematopoietic stem cells and each cell in peripheral blood,it is possible for the status of relevant indicators in peripheral blood to reflect the status of primordial qi,thus greatly improving the development and application of TCM theory of primordial qi. |
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