OBJECTIVE: To explore the multi-component synergistic mechanism of Zuogui Wan(左归丸, ZGW) in treating postmenopausal osteoporosis(PMOP). METHODS: The main components and target genes of ZGW were screened via the Trad...OBJECTIVE: To explore the multi-component synergistic mechanism of Zuogui Wan(左归丸, ZGW) in treating postmenopausal osteoporosis(PMOP). METHODS: The main components and target genes of ZGW were screened via the Traditional Chinese Medicine Systems Pharmacology(TCMSP). In addition, the target gene sets of PMOP were derived from the Gene Cards and Online Mendelian Inheritance in Man databases. The search tool for recurring instances of neighbouring genes(STRING) 11.0 software was used to analyze the interaction among intersecting genes. Cytoscape 3.6.1 software and the Matthews correlation coefficient(MCC) algorithm were used to screen the core genes. Fifty Sprague-Dawley female rats were randomly divided into the sham-operated(Sham) group and the four ovariectomized(OVX) subgroups. Rats subjected to Sham or OVX were administered with the vehicle(OVX, 1 m L water/100 g weight), 17β-estradiol(E2, 50 μg·kg-1·d-1), and lyophilized powder of ZGW at a low dose of 2.3(ZGW-L) and high dose of 4.6(ZGW-H) g·kg-1·d-1 for three months. The bone density and bone strength were assessed using dual-energy X-ray and three-point bending tests, respectively. Furthermore, enzyme-linked immunosorbent assay, Hematoxylin-eosin staining, and western blot analysis were used to determine the potential pharmacological mechanisms of action of ZGW in PMOP. RESULTS: A total of 117 active compounds of ZGW were screened from the TCMSP. Furthermore, 108 intersecting genes of drugs and diseases were identified. Using STRING software and the MCC algorithm, ten core genes, including C-X-C chemokine living 8(CXCL8), C-C chemokine receptor type 2(CCR2), alpha-2a active receptor(ADRA2A), melatonin receptor type 1B(MTNR1B), and amyloid-beta A4 protein(APP), were identified. The anti-osteoporosis regulation network of ZGW was constructed using the Cytoscape software. The animal experiments demonstrated that ZGW groups significantly reduced the serum levels of β-C-terminal telopeptide of type I collagen(β-CTX) and increased serum levels of bone-specific alkaline phosphatase(BALP)(P < 0.05, P < 0.01). The OVX group exhibited a significant decrease in bone mineral density and bone strength compared with the Sham group(P < 0.01). Moreover, treatment with ZGW resulted in increased trabecular thickness, improved arrangement of trabecular structure, and reduced empty bone lacunae. Furthermore, treatment with ZGW significantly increased the protein expression of CXCL8, ADRA2A, and CCR2(P < 0.05, P < 0.01), and significantly decreased the protein expression of Runx2(P < 0.01). Furthermore, the ZGW and E2 groups demonstrated significantly increased BMD(P < 0.05, P < 0.01), improved bone strength(P < 0.05, P < 0.01), reduced expression of CXCL8, ADRA2A, and CCR2, and increased runt-related transcription factor 2 levels in bone tissue(P < 0.05, P < 0.01) compared with the OVX group. However, there were no significant differences in MTNR1B and APP expression among the groups. CONCLUSION: ZGW shows synergistic mechanisms in PMOP through multiple components, targets, and pathways.展开更多
基金the National Natural Science Foundation of China:Research on Mechanism of Zuogui Wan in Treating Postmenopausal Osteoporosis by Regulating Feed-forward Loop of Oxytocin/Oxytocin Receptor Based on Transcriptome so as to Explain the Theory of “All Marrows Dominated by Brain”(No. 82104730)Based on Protein Kinase Cθ/Nuclear Factor Kappa-B Signal Transduction Regulation,Shexiang Huangqi Compound Dropping Pills Regulate the Migration and Differentiation Mechanism of Mesenchymal Stem Cells Through the Blood-brain Barrier after Cerebral Ischemia (No. 81974564)+1 种基金The 72nd Batch of China Postdoctoral Science Foundation (2022M721065)Central Plains Talent Program-science and Technology Innovation Leading Talent Project (224200510027)。
文摘OBJECTIVE: To explore the multi-component synergistic mechanism of Zuogui Wan(左归丸, ZGW) in treating postmenopausal osteoporosis(PMOP). METHODS: The main components and target genes of ZGW were screened via the Traditional Chinese Medicine Systems Pharmacology(TCMSP). In addition, the target gene sets of PMOP were derived from the Gene Cards and Online Mendelian Inheritance in Man databases. The search tool for recurring instances of neighbouring genes(STRING) 11.0 software was used to analyze the interaction among intersecting genes. Cytoscape 3.6.1 software and the Matthews correlation coefficient(MCC) algorithm were used to screen the core genes. Fifty Sprague-Dawley female rats were randomly divided into the sham-operated(Sham) group and the four ovariectomized(OVX) subgroups. Rats subjected to Sham or OVX were administered with the vehicle(OVX, 1 m L water/100 g weight), 17β-estradiol(E2, 50 μg·kg-1·d-1), and lyophilized powder of ZGW at a low dose of 2.3(ZGW-L) and high dose of 4.6(ZGW-H) g·kg-1·d-1 for three months. The bone density and bone strength were assessed using dual-energy X-ray and three-point bending tests, respectively. Furthermore, enzyme-linked immunosorbent assay, Hematoxylin-eosin staining, and western blot analysis were used to determine the potential pharmacological mechanisms of action of ZGW in PMOP. RESULTS: A total of 117 active compounds of ZGW were screened from the TCMSP. Furthermore, 108 intersecting genes of drugs and diseases were identified. Using STRING software and the MCC algorithm, ten core genes, including C-X-C chemokine living 8(CXCL8), C-C chemokine receptor type 2(CCR2), alpha-2a active receptor(ADRA2A), melatonin receptor type 1B(MTNR1B), and amyloid-beta A4 protein(APP), were identified. The anti-osteoporosis regulation network of ZGW was constructed using the Cytoscape software. The animal experiments demonstrated that ZGW groups significantly reduced the serum levels of β-C-terminal telopeptide of type I collagen(β-CTX) and increased serum levels of bone-specific alkaline phosphatase(BALP)(P < 0.05, P < 0.01). The OVX group exhibited a significant decrease in bone mineral density and bone strength compared with the Sham group(P < 0.01). Moreover, treatment with ZGW resulted in increased trabecular thickness, improved arrangement of trabecular structure, and reduced empty bone lacunae. Furthermore, treatment with ZGW significantly increased the protein expression of CXCL8, ADRA2A, and CCR2(P < 0.05, P < 0.01), and significantly decreased the protein expression of Runx2(P < 0.01). Furthermore, the ZGW and E2 groups demonstrated significantly increased BMD(P < 0.05, P < 0.01), improved bone strength(P < 0.05, P < 0.01), reduced expression of CXCL8, ADRA2A, and CCR2, and increased runt-related transcription factor 2 levels in bone tissue(P < 0.05, P < 0.01) compared with the OVX group. However, there were no significant differences in MTNR1B and APP expression among the groups. CONCLUSION: ZGW shows synergistic mechanisms in PMOP through multiple components, targets, and pathways.
