Study The Metabolite And Metabolic Pathways Of Inflammation Induce By LPS And Intervention Of Angelica Sinensis Volatile Oil

As the development of systme biology, we find that the philosophy of system biology andTraditional Chinese Veterinary Medicine(TCVM) is the same. In the theory and practice of TCVM,the body is recognized as a whole system and the status of health or diseases is considered as thebalance status of that system. As the terminal of system biology, metabonomics, defined as―thequantitative measurement of the multiparametric metabolic response of living systems topathophysicological stimuli orgenetic modification‖, is a novel methodology arising from thepost-genomics era for the study of molecular systems biology. The strategy of metabonomicscoincides with TCVM theory and may play an important role in the modern study of TCVM. Inthis study a metabonomic approach is employed for investigation of an animal model induced by ahigh dose of Lipopolysaccharide, and the therapeutic and metabonomic effects of Angelicasinensis Volatile Oil (OVAS) was chosen as the research subject, and gas chromatography-massspectrometry(GC-MS) was used as analytical tools. This research provides the basis forpharmacological activity of OVAS, and gives a certain amount of reference and help forpharmacodynamic study of Chinese medicine containing OVAS. Some studies are as follows,1. OVAS were extracted by steam distillation and organic solvent in the petroleum ether.The main chemical compositions were determined using GC-MS. OVAS yield of extracted bysteam distillation was about0.38%, the petroleum ether extract yield about1%. Chemicalcompositions of Petroleum ether was similar to water vapor distillation extracts of OVAS, Themain chemical compositions including Z-Ligustilide, formic acid,1S--pinene,6-ketone ofundecane, Thujopsene,-cedrene,2-methoxy phenol,(+)-Aromadendrene,-Bisabolene,Eucalyptus oil (-)-leaves enol, Z-Butylidene phthalide, E-Ligustilide and so on. Thesimilarity of15OVAS samples of gansu province were evaluation by principal componentanalysis and the similarity evaluation software, the results show that similarity were greater than0.900, the average was0.951±0.019, angelica sinensis in different areas of the gansu province noobvious difference between volatile oil chemical composition.2. LPS was injected intraperitoneally into rats tobuild inflammation in animal models ofendotoxin. OVAS was acted on the model (OVASL0.088g/kg, OVASM0.176g/kg, OVASH0.352g/kg, control drugs: Dex).8h after LPS injection, we got the blood from the rats and the ratswere killed. We got the mice’s liver, lung organs by routine conservation method. Blood of micewas determined by automated hematology analyzer; separation of serum was determined withaspartate aminotransferase (GOT), alkaline phosphatase (ALP),alanine aminotransferase (GPT)by using the automatic biochemical analyzer. We measured the serum inflammatory cytokinesTNF-a, IL-6, IL-1β、IL-8、IL-4、IL-10content by ELISA;Using classic Griess reagent method,we detected serum NO content. Using GC-MS production biopsy in rats liver and lung, weobserved the changesof Metabolite in rats liver, lung, urine, plasm. we explored the effect andmechanism of OVAS in LPS inflammation in vivo rat model. Model of inflammation could besuccessfully established. LPS infection of the liverand lungs of the rats was subject to moreserious injury. From blood and biochemical parameters, OVAS and dexamethasone couldeffectively inhibit the inflammatory symptoms of the rats, and could effectively protect the liver ofrats, reduce endotoxin to its damage. ELISA results showed that the OVAS and the dexamethasonegroup significantly inhibited levels of TNF-, IL-6, IL-lβ, I L-8protein secretion in serum.OVAS and the dexamethasone group were significantly inhibited the NO、HIS、5-HT. Notably, OVASMexhibited a potential pharmacological effect.3. Gas chromatography-mass spectrometry (GC-MS) combined with pattern recognitionapproaches, namely, principal component analysis (PCA) and partial least squares-discriminantanalysis (PLS-DA), were used to determine differentiating metabolites in plasma,liver tissue, lungtissue, urine of KB, LPS model of inflammation, Dex, OVASL, OVASM, OVASH. PCA andPLS-DA score plots of LPS model of inflammation group clustered separately from that of thecontrol, while groups treated with OVASL、OVASM、OVASH clustered closely with the KB. Thisresult indicates that the metabolic profiles of the OVASL、OVASM、OVASH groups are almostsimilar to those of the control, Notably, OVASM exhibited a potential pharmacological effect byregulating multiple perturbed pathways to the normal state.(1) Difference metabolite (thirty-two in the plasma) were identified. Functional pathwayanalysis revealed that alterations in these metabolites are associated with Fatty acid metabolism,glycolysis, glycine, urea metabolism. VOAS may exert its effectiveness on anti-inflammatoryactivation by restoring the protein synthesis through adjusting the urea content in plasma, restoringthe lipid metabolism disorders and involving the LXRs signaling pathway, increasing glycine inplasma, give play to the role of direct target cells or indirectly modulate inflammatory factors playa cell protection, so as to alleviate inflammation;(2) Difference metabolite (twenty-three in the liver tissue) were identified. Functionalpathway analysis revealed that alterations in these metabolites are associated with Linoleic acidmetabolism, Galactose metabolism, Glycerolipid metabolism, Arachidonic acid metabolism.VOAS may exert its effectiveness on anti-inflammatory activation by Lower the content of freefatty acids and glycerol in the liver, in order to reduce the release of inflammatory cytokines andpromote the anti-inflammatory factor release; Improve the content of galactose, galactosemetabolism in the liver is normal.(3) Difference metabolite (twenty-seven in the lung tissue) were identified. Functionalpathway analysis revealed that alterations in these metabolites are associated with D-Glutamineand D-glutamate metabolism, Linoleic acid metabolism, alpha-Linolenic acid metabolism,Galactose metabolism, Arachidonic acid metabolism, Alanine, aspartate and glutamate metabolism,Pentose phosphate pathway, Glycerolipid metabolism.VOAS may exert its effectiveness onanti-inflammatory activation by recovering of glutamic acid generated to block the accumulationof succinic acid to reduce the release of inflammatory factor, reducing linoleic acid content in lungtissue homogenate, lipid metabolism returned to normal from inhibiting inflammation process, Byincreasing the release of docosahexaenoic acid (DHA), makes the AA to release a quantity toreduce inflammation, restrain the generation of inflammatory mediators.(4) Difference metabolite (twenty-seven in the urine) were identified. Functional pathwayanalysis revealed that alterations in these metabolites are associated with D-Glutamine andD-glutamate metabolism, Glycine, serine and threonine metabolism, Alanine, aspartate andglutamate metabolism, Galactose metabolism, Arachidonic acid metabolism, Pyruvate metabolism,Pentose phosphate pathway, Aminoacyl-tRNA biosynthesis, Citrate cycle (TCA cycle), Glycolysisor Gluconeogenesis.VOAS may exert its effectiveness on anti-inflammatory activation byreducing the content of glutamic acid, succinic acid, pyruvic acid in the urine to restore the Krebscycle disorder, correction make glycolysis, glucose and galactose and so on content carbohydratemetabolism of acid pathway back to normal, increase the synthesis of serine, glycine, reduce the accumulation of glutamic acid, increasing the body’s antioxidant capacity to play a role ofanti-inflammatory.4. Using MetPA database to build six groups of plasma, liver tissue homogenate, lung tissuehomogenate, urine screening all differences metabolites related metabolic pathways, D-Glutamineand D-glutamate metabolism, Galactose metabolism, Glycine, serine and threonine metabolism,Alanine, aspartate and glutamate metabolism, Amino sugar and nucleotide sugar metabolism,Pyruvate metabolism, Pentose and glucuronate interconversions, Linoleic acid metabolismwere as potential pathways of LPS inflammation model, OVASL, OVASM, OVASH. Hotcluster analysis plot of the differential metabolites based on Matlab, various metabolites could beidentified as responsible for the separation betweencontrol and model groups, and these were therefore viewed as potential biomarkers.Inflammation caused by LPS in rats model of a central part of the metabolic disorder may becaused by the Krebs cycle obstacles and promote glycolysis and fatty acid metabolism disorder,make relevant glutamic acid, glutamate metabolism and branched chain amino acid metabolicdisorders, such as angelica naphtha can prompt the Krebs cycle, glycolysis and fatty acidmetabolism disorder, so as to achieve anti-inflammatory effects.5. By using Spss software to detect the differences of metabolites and detection ofinflammatory cytokines, inflammatory mediators index correlation analysis, show that there arebiological significance of succinic acid, pyruvic acid, glucose, lactic acid, acetic acid, arachidonicacid, docosahexaenoic acid biological significance by the screening of PLS-DA.OVAS are multi-component, multi-pathway, and multi-target treatments. It is likely thatOVASL, OVASM, OVASH intervenes the metabolic process of LPS model of inflammationby affecting the D-Glutamine and D-glutamate metabolism, Galactose metabolism, Glycine,serine and threonine metabolism, Alanine, aspartate and glutamate metabolism, Butanoatemetabolism, Amino sugar and nucleotide sugar metabolism, Pyruvate metabolism, Pentose andglucuronate interconversions, Linoleic acid metabolism. Metabonomics is a robust andpromising for the identification of biomarkers and elucidation of the mechanisms of a disease,thereby highlighting its importance in drug discovery.