Transcriptome Analysis And Mechanism Of Baoyuan Jiedu Decoction In Improving Cancer-induced Muscle Atrophy

Objective:In this study,Apcmin/+ cancer cachexia model was used as the research object.By using the methods of transcriptome and molecular biology,the main target,action pathway and mechanism of Baoyuan Jiedu Decoction to improve the muscle atrophy of cancer cachexia were discussed,which provided experimental basis for the development of effective Chinese medicine to intervene the muscle atrophy of cancer cachexia.Method:1.The components of Baoyuan Jiedu Decoction were detected by UHPLC-Q Exactive orbital hydrazine high resolution mass spectrometry,and the detected compounds were analyzed qualitatively.2.After the intervention of Baoyuan Jiedu Decoction on Apcmin/+mice for 12 weeks,the general situation of the animals,the changes of food intake,water intake and body weight,the weight of gastrocnemius muscle and HE staining,the number,volume and shape of intestinal adenoma were observed by methylene blue staining,the morphological structure of gastrocnemius muscle mitochondria was observed by electron microscopy,the mRNA and protein expression of muscle atrophic protein(Atrogin-1,MuRP-1)were detected by RT-PCR and Western blot.To evaluate the effect of Baoyuan Jiedu Decoction on cancer-related muscular atrophy3.After the intervention of Baoyuan Jiedu Decoction,the whole gene expression profile of gastrocnemius muscle of Apcmin/+ mice was analyzed by transcriptome sequencing,and the differential genes were screened.Combined with GO function annotation and KEGG enrichment pathway analysis,the main target and action pathway of Baoyuan Jiedu Decoction to improve cancer-related muscle atrophy were screened.4.Based on p38MAPK/PGC-1 signal pathway and mitochondrial function,the mechanism of Baoyuan Jiedu Decoction to improve cancer-related muscular atrophy was studied.The levels of TNF-? and IL-6 in serum,p38MAPK,PGC-1? mRNA and protein,ATP in gastrocnemius muscle,mtDNA copy by RT-PCR,Nrf1,Nrf2,TFAM mRNA and protein expression by ELISA,COX-?,Cyclochrome C mRNA and protein expression levels;UCP-2 and UCP-3 mRNA and protein expression levels related to mitochondrial uncoupling,Mfn1,Mfn2 and Fis1 mRNA and protein expression levels related to mitochondrial polymerization and cleavage;to explore the mechanism of Baoyuan Jiedu Decoction regulating mitochondrial function through p38MAPK/PGC-1 pathway and improving cancer-related muscular tube atrophyResults:1.UHPLC-Q Exactive orbital hydrazine high resolution mass spectrometry analysis showed that there were 12 compounds in the decoction of Baoyuan Jiedu Decoction,including ginsenoside Rg1,aconitine,astragaloside a,chlorogenic acid,ferulic acid and liquiritigenin,which indicated that Baoyuan Jiedu Decoction ensured the good retention of the effective components in the decoction process.2.After the intervention of Baoyuan Jiedu Decoction,the weight of gastrocnemius muscle increased(P<0.05),the diameter of muscle fiber increased,the amount of food intake and water intake increased(P<0.05),and the number and volume of intestinal adenoma in Apcmin/+ mice were effectively reduced,the morphology of gastrocnemius mitochondria was improved,and the expression of Atrogin-1 and Murf-1 were inhibited(P<0.05)3.Through the analysis of transcriptome sequencing technology,we found that compared with the blank control group,model group mice screened 1423 significantly different genes,of which 494 were up-regulated and 929 were down regulated;compared with model group,Baoyuan Jiedu Decoction group screened 355 significantly different genes,of which 319 were up regulated and 36 were down regulated.Among the top 100 differential genes,pdhx,rxrg,ugp2,PGP,tuba8,tnks2,mylk2,lrrc39,UCP-3,mfnl and FIS 1 were found to be related to mitochondrial energy metabolism(including oxidative phosphorylation,TCA cycle,fatty acid metabolism and glycolysis/gluconeogenesis),skeletal muscle structure,mitochondrial uncoupling,mitochondrion fusion and division.The key genes such as IL-6,TNF-? and PGC1 ?were further screened.The analysis of differential gene GO function showed that ATP binding,cytokine receptor activity and other entries;KEGG pathway analysis showed that compared with the normal control group,the main enrichment pathways of the model group were:protein digestion and absorption,PI3K Akt signal pathway,relaxin signal pathway,Apelin signal pathway,focal adhesion,Rapl signal pathway,proteoglycan and cancer,cancer Pathway,GABAergic synapse,MAPK signaling pathway.After treatment with Baoyuan Jiedu Decoction,compared with the model group,the main enrichment pathways of significant difference genes were:cytokine cytokine receptor interaction,hematopoietic cell line,p53 signal pathway,cell cycle,cell aging,primary immunodeficiency,leukocyte migration through endothelial cells,platinum drug resistance,T cell receptor signal pathway,allograft rejection,F Oxo signaling pathway,apoptosis,JAK STAT signaling pathway,MAPK signaling pathway,cancer pathway,DNA replication pathway,etc.MAPK signal pathway is a common pathway of both,which suggests that the pathogenesis of cancerous cachexia muscular atrophy and the therapeutic mechanism of Baoyuan Jiedu Decoction on cancerous cachexia muscular atrophy may be related to this pathway.4,Baoyuan Jiedu Decoction can significantly reduce the content of TNF-? and IL-6 in the serum of Apcmin/+(P<0.05),effectively regulate the expression of p38MAPK/PGC-1? pathway protein(P<0.05),significantly increase the content of ATP and the copy number of mtDNA in the gastrocnemius muscle of mice(P<0.05),promote the expression of Nrf1,Nrf2 and TFAM,which are the key proteins of mitochondrial biosynthesis(P<0.05),and significantly enhance the expression of COXIV and Cyclochrome C expression(P<0.05)improved mitochondrial oxidative phosphorylation.The expression of UCP-2 and UCP-3 mRNA and protein was significantly decreased(P<0.05),and the expression of Mfn1 and Mfn2 was up-regulated,and the expression of Fis1 was inhibited.Conclusion:In conclusion,Baoyuan Jiedu Decoction can effectively regulate p38MAPK/PGC-1 a pathway,inhibit the expression of E3 ubiquitin ligase Atrogin-1 and MuRF-1,promote mitochondrial production,restore the normal metabolism of mitochondrial energy,improve mitochondrial production,mitochondrial oxidative phosphorylation and other functions,inhibit mitochondrial uncoupling and coordinate the dynamic level of mitotic fusion by reducing the expression of cytokines Balance is the possible mechanism of improving cancer-related cachexia muscular atrophy.