The Intervention Effect And Mechanism Research Of External Therapy Of Warming Channels Activating Blood Circulation Treating Early Knee Osteoarthritis Based On Metabonomics Technique

Objective:To observe the intervention effect of external treatment of warming channels activating blood circulation on KOA,and to search for early serum markers of KOA and the mechanism of external treatment of warming channels activating blood circulation based on metabolomics technology.Methods:1.The intervention effects of warming channels activating blood circulation on cold stimulation pain,mechanical stimulation pain,the IL-1β,TNF-αprotein and mRNA expression in synovial tissue and TRPA1,TRPV4 protein and mRNA expression in dorsal root ganglion.2.The intervention effect of warming channels activating blood circulation on serum metabolome of KOA rats was observed based on GC-MS.3.The intervention effect of warming channels activating blood circulation on serum metabolome of KOA rats was observed based on UHPLC-MS.4.To observe the clinical effect of external treatment of warming channels activating blood circulation on patients with early KOA.5.The intervention effect of warming channels activating blood circulation on serum metabolome of patients with early KOA was observed based on GC-MS.6.The intervention effect of warming channels activating blood circulation on serum metabolome of patients with early KOA was observed based on UHPLC-MS.Results:1.External treatment of warming channels activating blood circulation could significantly increase cold pain threshold and mechanical pain threshold in KOA rats,improve synovial inflammation,and reduce the protein and mRNA expressions of IL-1β,TNF-α in synovial tissue(P<0.05-0.01),and the protein of TRPA1,TRPV4 in drosal root ganglion of KOA rats,with statistical significance(P<0.01).2.Based on GC-MS,the results showed KOA rats compared with the blank control group rats 35 different metabolites,warming channels activating blood circulation of the treatment group and KOA rats compared with 23 differences metabolites,including warming channels activating blood circulation external treatment can significantly callback 2-hydroxy butyric acid,arachidonic acid,asparagine,cholesterol and bile acid,creatine,glucose,glycerol,phenylalanine,serine,threonine,uracil and urea metabolites of 13 kinds of differences,mainly related to metabolism of galactose,glycine,serine and threonine metabolism and biosynthesis of phenylalanine,tyrosine and tryptophan,4 metabolic pathways such as carbamyl-tRNA biosynthesis.3.Based on UHPLC-MS,the results showed that compared with the KOA model group,a total of 57 different lipids were obtained in the blank control group under the positive ion mode,and 11 different lipids were obtained under the negative ion mode,including LPC,PC,PE,PI,etc.In the KOA model group and the easy layer group,36 different lipids were obtained under positive ion mode,mainly including TAG,DAG,LPC,PC,etc.In the negative ion mode,a total of 6 different lipids were obtained:LPC,FAHFA and PE.Among them,23 positive ions and 3 negative ions were recovered significantly by the external treatment of "warming channels activating blood circulation".4.The external treatment of warming channels activating blood circulation could significantly reduce WOMAC score,improve tenderness threshold and temperature pain threshold in early KOA patients,and the difference was statistically significant(P<0.05).5.Based on GC-MS,the results showed that there were 35 different metabolites in the control group compared with the KOA group,and there were 26 different metabolites in the KOA group after treatment compared with before treatment.Among them,the external treatment of warming channels activating blood circulation could significantly callback 16 different metabolites.It mainly involves 9 metabolic pathways,including alanine,aspartate and glutamate,purine metabolism,galactose metabolism,primary bile acid biosynthesis,taurine and taurine metabolism,glycerol metabolism,starch and sucrose metabolism,steroid biosynthesis and steroid hormone biosynthesis.6.Based on UHPLC-MS,the results showed that in the positive ion mode,113 differential lipids were obtained in the blank control group and the KOA group,and 145 differential metabolites were obtained in the KOA group before and after treatment,among which 60 differential metabolites,mainly including LPC,PC,CAR and CE,were significantly recovered by the external treatment of "warming channels activating blood circulation".In the negative ion mode,a total of 32 different lipids were obtained in the blank control group and the KOA group.A total of 46 different metabolites were obtained in the KOA group before and after treatment,among which 13 different metabolites,including FA,LPE,PE and SMGDG,could be significantly recovered by the external treatment of "warm meridian and promote blood circulation".Conclusions:1.The external treatment of warming channels activating blood circulation has a definite effect on KOA,which can significantly increase the pain threshold of cold stimulation and mechanical stimulation of KOA,and improve synovial inflammation.2.KOA serum metabolic changes obviously,metabonomics can screening potential markers and related pathways,External treatment of warming channels activating blood circulation may be affected mainly by LPC/PC metabolism,alanine,glutamic acid and aspartic acid and purine metabolism,galactose metabolism,glycine,serine and threonine metabolism,phenylalanine,tyrosine and tryptophan biosynthesis and related metabolic pathways play a role of treatment.