| Purpose:Observing the effect of Guilu Erxian Gum on postmenopausal osteoporosis(PMOP)rat models,based on non targeted metabolomics,liquid chromatography-mass spectrometry(LC-MS/MS)technology was used to detect the changes in metabolites in PMOP rat models before and after intervention with Guilu Erxian Gum,Furthermore,the pharmacological mechanism of Guilu Erxian Gum in preventing PMOP was further explored,in order to fully utilize the role of traditional Chinese medicine and provide new ideas and approaches for clinical prevention of PMOP.Material and method:In this experiment,50 8-week-old unmatched SD female rats were randomly divided into 5 groups:sham surgery group,castrated model group,castrated Guilu Erxian glue(high,medium,and low dose)medication group.Guilu Erxian glue was selected as the experimental drug.The rats were fed for 1 week and then castrated for modeling.After successful modeling for 1 week,gavage administration was started.After 8 weeks of continuous medication,samples were taken and bone tissue biomechanical testing was performed on the femoral tissue;The Micro-CT scanner analyzes the microstructure and structure of bone trabeculae,and detects parameters such as the number,thickness,separation,and bone density of bone trabeculae;HE staining was used to detect the micro structure,morphology,and quantity of bone;Using ELISA method to detect serum alkaline phosphatase(ALP),bone formation index(P1NP),and bone resorption index(β-CTX);Immunohistochemical(IHC)analysis;Using liquid chromatography-mass spectrometry(LC-MS/MS)technology to construct a rat blood metabolite map and observe the distribution differences of metabolites and related pathways.Results:1.biomechanical tests:the maximum loading force was the highest in the sham-operated group,the maximum loading force decreased with decreasing dose in the tortoise and deer Erxian Gum group,and the lowest in the model group;2.Analysis of the 3D imaging results of the Micro-CT machine scan showed that:the bone volume of femoral tissue in each group was ranked from high to low:sham-operated group>Turtle Deer Erxian Gum high-dose group>model group;bone density was ranked from high to low:sham-operated group>Turtle Deer Erxian Gum high-dose group>model group;cortical bone volume test results showed that there was no significant difference between the groups;cortical bone density:lowest in the model group,no significant difference between the Turtle Deer Erxian Gum group and the model group,and highest in the sham-operated group.The cortical bone density was the lowest in the model group,the highest in the sham-operated group,and the highest in the sham-operated group;the bone trabecular volume was ranked from high to low as sham-operated group>tortoise and deer Erxian gum high-dose group>model group.Bone trabecular density was ranked from high to low:sham-operated group>Turtle and Deer Erxian Gum high-dose group>model group,and there was no significant difference between Turtle and Deer Erxian Gum high-dose group and model group;3.HE staining showed that the bone trabeculae in the sham-operated group were tightly connected and reticulated;in the model group,the bone trabeculae were thin,broken and most of them were non-reticulated;in the low-dose group,the bone trabeculae were thickened,broken and reticulated;in the medium-dose group,the bone trabeculae were thicker and more densely distributed;in the high-dose group,the bone trabeculae were thickened,reticulated and densely arranged.The bone trabeculae in the high dose group were thickened,reticulated and densely arranged;4.serum bone metabolic indexes:P-CTX expression was the highest in the model group,the lowest in the sham-operated group,and decreased with increasing concentrations in the Turtle and Deer Erxian Gum group;ALP and P1NP expression was the highest in the sham-operated group,followed by the Turtle and Deer Erxian Gum group,which decreased with decreasing concentrations,and the lowest in the model group;5.Immunohistochemical(IHC)analysis showed that the expression of Runx 2 in rat femur was highest in the sham-operated group and lowest in the model group,and decreased with decreasing dose in the Kamehameha Erxian Gum group,and Kamehameha Erxian Gum could increase the expression of Runx 2 to promote bone formation;6.According to the screening results of differential metabolites,a total of 108 differential metabolites were identified between the Turtle Deer Erxian Gum group and the model group,among which the metabolite contents of glutamic acid,threonine,arginine,aspartic acid,fructose,lysophosphatidic acid,indole-3-acetic acid and ornithine were significantly up-regulated in the Turtle Deer Erxian Gum medium-dose group compared with the model group;while the metabolite contents of 3-hydroxybutyric acid,methylhistidine,biotin,cystine and prostaglandin G were significantly up-regulated compared with the model group.The levels of metabolites such as 3-hydroxybutyric acid,methylhistidine,biotin,cystine,prostaglandin G2,styrene,glycine and taurine were down-regulated;four metabolites,3-(3-methoxyphenyl)propionic acid,acetamide,Equol and 1-oleoyl-sn-glycero-3-phosphocholine,were co-regulated in the sham-operated group,the high-dose group and the model group,with high expression in the sham-operated group and low expression in the model group.lowered to low expression and returned to high expression in the tortoise and deer ercian gum group;7.The main pathways involved between the sham-operated and model groups were steroid hormone biosynthesis,cortisol synthesis and secretion,and Cushing’s syndrome;the pathways involved between the Kamehameha Erxian Gum group and the model group mainly included tryptophan metabolism,phenylalanine metabolism,galactose metabolism,alanine,aspartate and glutamate metabolism,and histidine metabolism.Conclusion:1.Through animal experiments,it has been found that Guilu Erxian Gum can significantly enhance the continuity and stability of bone trabeculae in the femur of PMOP model rats,promote bone formation,improve bone density,and achieve the goal of preventing PMOP.Its mechanism of action may be through regulating various metabolites and pathways to prevent PMOP;2.PMOP model rats showed high expression of glutamic acid,threonine,arginine,aspartic acid,fructose,lysophosphatidic acid,indole-3-acetic acid,ornithine,3-(3-methoxyphenyl)propionic acid,acetamide,Equol,1-oleoyl-Sn-glycerol-3-phosphocholine,etc;The levels of metabolites such as 3-hydroxybutyric acid,methylhistidine,biotin,cystine,prostaglandin G2,styrene,glycine,and taurine were downregulated.Among them,metabolites of intestinal microbiota such as acetic acid and propionic acid were highly expressed after medication.It is speculated that Guilu Erxian Gum may participate in regulating the content of intestinal microbiota and prevent PMOP;3.Tryptophan metabolism,galactose metabolism,alanine,aspartic acid and glutamic acid metabolism,phenylalanine metabolism,histidine metabolism and other metabolic pathways may be the main action pathway of Guilu Erxian Gum in preventing PMOP.The prevention of PMOP by Guilu Erxian Gum may be related to amino acid metabolism and glucose metabolism. |
PDF Full Text Request