Discussion On TCM Syndrome And Biological Mechanism Of Central Fatigue Based On Mitochondrial Biosynthesis

ObjectiveThe current research analysised the Chinese medical theory of central fatigue through the aspects of definition,category,symptoms,and syndromes,by ancient literature reviewing and analyzing.Based on the previous research and modern research process,the research discussed the relationship between mitochondrial biogenesis and syndrome of liver stagnation and spleen deficiency of central fatigue.In the experimental study,the research evaluated the efficiency of decoction modified Tifukang(TFK),after the construction and evaluation of rat model of central fatigue.In addition,the research analyzed the biological and pharmacological mechanism of the liver stagnation and spleen deficiency of central fatigue,from the gradient aspects of energy metabolism,oxidative effect,and mitochondrial function.Finally,the research detected the protein and gene changes in the SIRT1-PGC1?-NRF1 signal pathway,which represented the key regulators in mitochondrial biogenesis.Together to discuss the molecular targets of intervention of TKF against central fatigue,and provide research basis for clinical intervention.Methods Theoretic study:This research collated the related terms of central fatigue by searchingthe connotations in reference books.Then searched the key words "pi","lao","juan" in the 5th edition Zhonghuayidian.The corresponding symptoms and syndromes of central fatigue was analyzed.In addition,the research discussed the relationship between central fatigue and syndrome of liver stagnation and spleen deficiency via mitochondrial biogenesis,with the reviewing of literatures.Finally,the research proposed the theoretical basis of the intervention of TFK in central fatigue though the principle of treating liver and spleen.Experimental study:During the construction and evaluation of central fatigue rat model,the rats were randomly devided into 4 groups:the control group,the 5-day group,the 14-day group,the 21-day group.General status observation(such as mental status,hair),behavioral tests(weight-loaded swimming test,open-field test,and elevated plus maze),and ultrastructural changes were used to evaluate the model.In the treating experiment,rats were randomly devided into 6 groups:the control group,the central fatigue group,the low-,middle-,high-dose TFK group,and the coenzyme Q10 group.The general status and behavioral changes were performed for the efficacy evaluation of TFK,and the verification the syndrome model of liver stagnation and spleen deficiency.The creatine kinase(CK),lactate dehydrogenase(LDH),and blood urea nitrogen(BUN)in blood serum,and the malondialdehyde(MDA)and superoxide dismutase(SOD)in the liver were detected for the research of mechanism of energy metabolism and oxidative effect.The mitochondrial ultrastructural changes in hippocampal CA1 region were observed by utilizing a transmission electron microscope.The copy number of the mtDNA was detected by reverse transcription polymerase chain reaction(RT-PCR),together to evaluate the changes of mitochondrial function in the central fatigue rat model of syndrome of liver stagnation and spleen deficiency.In addition,changes of silent information regulator1(SIRT1),proliferator-activated receptor gamma coactivator-1?(PGC-l?),and nuclear transcription factor 1(NRF1)were detected by immunohistochemistry,western blot,and RT-PCR.In order to analyze the biological mechanism of the syndrome of liver stagnation and spleen deficiency of central fatigue,and the molecular targets of intervention of TKF.ResultsTheoretic study:(1)The terms "shenlao" and "zhijuan" have proved to be the most relevant to the connotation of central fatigue in TCM theory.The symptoms mainly represent the decreasing function of mental activity,proprioception,which indicated a decrease in high level cognition,objective control and manipulation of emotion.These results are consist with the manifestation of central fatigue such as decreasing cognition and negative emotions.The pathological mechanism of central fatigue is closely related to yin-yang,blood and qi,and five internal organs.The syndrome generally differs according to the individual differences and pathogenesis.The coordination and harmony of liver and spleen paly important role in central fatigue.The characteristics of functions of these two organs,and the life style of modern population make liver stagnation and spleen deficiency the main syndrome in central fatigue.(2)The mechanism under this process is closely related to mitochondrial biogenesis.During the liver stagnation,body was in the long term stimulation of stress,the protection effect of mitochondrial biogenesis on nerves is weaken.Results in the low efficiency of mitochondrial biogenesis,then triggers the decrease of liquidity in the membrane protential,and the dysfunction of mitochondrial.Then the function of the HPA axis is affected,and the negative emotions appears such as anger and depression.Meanwhile,during the status of spleen deficiency,the process of mitochondrial biogenesis is damaged,which make is difficult to produce new protein and gene.This result leads to a decrease in energy metabolism,ATP production,and the process of nutritions turning into energy.On the other hand,since the mitochondrial biogenesis guarantees the self metabolism of mitochondria,the low efficiency of mitochondrial biogenesis immediately decreases the number of mtDNA,changes the shape of mitochondria to swelling,and this result is consist with the stagnation of fluid-dampness after spleen deficiency.(3)In addition,the research discussed the scientificity and feasibility of the intervention of TFK against syndrome of liver stagnation and spleen deficiency in central fatigue.Experimental study:(1)According to the evaluation results of animal model,in the open field test(OFT),compared with the control group,the 5-(p<0.05)and 14-(p<0.01)day of MMPM increased the central time.In the elevated plus maze(EMP),the 21-day group significantly decreased the ratio of number(p<0.05)and time(p<0.001)in the open arms.In the weight-loaded swimming test(WST),the 14-and 21-day MMPM decreased the swimming time compared with the control group(p>0.05).In addition,the ultrastructural changes of mitochondria in the 21-day group appeared including mitochondrial swelling,broken cristae.(2)After treatment with modified Tifukang(TFK),according to the results of behavioral tests,compared with the central fatigue(CF)group,in the OFT test,the low-,middle-,high-dose TFK and coenzyme Q10(COQ)all decreased the central time(p<0.001),total distance travelled(p<0.01,p<0.05,p>0.05,p>0.05),crossing number(p<0.05,p>0.05,p>0.05,p>0.05),maximum distance travelled(p<0.01,p>0.05,p<0.05,p>0.05),mean velocity(p<0.01,p>0.05,p>0.05,p>0.05);increased the vertical activity(p>0.05,p>0.05,p<0.01,p<0.01),and grooming time(p<0.05,p<0.01,p>0.05,p>0.05)and frequency(p<0.05,p<0.01,p>0.05,p<0.05).During the EMP,compared with the CF group,all treatments increased the ratio of entries(p<0.01,p<0.05,p<0.01,p<0.01)and time(p<0.01,p>0.05,p>0.05,p<0.01)in the open arms,and the central time(p<0.001,p<0.05,p<0.05,p<0.05).The WST test indicated that all treatments increased the swimming time of rats(p<0.001).(3)Compared with the CF group,all treatments significantly decreased the CK(p<0.01,p<0.05,p<0.01,p>0.05)and BUN(p<0.001)in blood serum,increased the activity of liver SOD(p<0.01,p<0.001,p<0.01,p<0.001),and decreased liver MDA(p<0.001,p<0.001,p<0.001,p<0.01)as well.(4)After the observation of transmission electron microscopy(TEM),it is found that all treatment obviously improved the degenerative changes of mitochondria,such as decreasing mitochondrial number,swelling,broken cristae and membranes.In addition,all treatment increased the copy number of mtDNA(p<0.01,p>0.05,p<0.01,p>0.05).(5)The immunohistochemistry test revealed that,the MDA of SIRT1(p<0.05,p<0.05,p<0.05,p<0.01),PGC-la(p>0.05),NRFl(p<0.01,p<0.05,p<0.001,p<0.001)in the hippocampus of the treatment groups were higher than the CF group.The result of western blot indicated that the protein expression of SIRT1(p<0.05,p<0.01,p<0.01,p<0.01),PGC-1?(p<0.05,p<0.001,p<0.001,p<0.001),NRF1(p<0.05,p<0.001,p<0.001,p<0.01)of treatment groups were increased compared with the CF group.The result of the RT-PCR indicated that the mRNA expression of SIRT1(p>0.05),PGC-la(p<0.01,p>0.05,p>0.05,p<0.01),NRF1(p>0.05,p<0.05,p<0.01,p<0.01)increased when compared with the CF group.ConclusionAlthough there is no specific concept and definition of central fatigue in Chinese medical theory,the terms "shenlao" and "zhijuan" in TCM theory are most relevant to the connotation of central fatigue,and main characteristic of syndrome is liver stagnation and spleen deficiency.Central fatigue could be treated by harmonizing the liver and spleen.The biological mechanism of syndrome of liver stagnation and spleen deficiency in central fatigue is closely related to the mitochondrial biogenesis.Behavioral tests,ultrastructural changes,and molecular biological tests verified that TFK can ameliorate central fatigue,employing the decreasing products of energy metabolism and enhancing activity of anti-oxidative enzyme.This process is closely related to the up-regulation of the protein and gene expression of SIRT1,PGC-la,and NRF1 in the hippocampus,which enhance the mitochondrial biogenesis.