Osteoporosis, as a globally prevalent chronic malady, exerts a profound impact on patients' quality of life and imposes substantial burdens upon both healthcare systems and socioeconomic structures[14]. The primary objective of this investigation is to gain a comprehensive understanding of the molecular mechanisms underpinning osteoporosis. Through a holistic analysis encompassing the Gene Expression Omnibus (GEO) database and the pharmaceutical agent Osteoking, this study has yielded novel insights into various facets of this pathological condition.
Primarily, employing data mining techniques on the GEO database facilitated the identification of a plethora of differentially expressed genes within the GSE35959 dataset, including both upregulated and downregulated genes. This outcome not only contributes novel insights into the genetic underpinnings of osteoporosis but also establishes a foundational basis for future investigations seeking innovative therapeutic targets. The discerned alterations in the gene expression profile of bone marrow mesenchymal stem cells under pathological conditions emphasize pivotal clues for comprehending the molecular regulatory mechanisms governing osteoporosis.
Subsequently, through an in-depth scrutiny of the Osteoking pharmaceutical, we have unveiled its potential therapeutic mechanisms against osteoporosis. The pharmaceutical composition of Osteoking encompasses diverse active constituents intricately linked to pertinent pathways and biological processes associated with osteoporosis. The construction of an active ingredient-disease target network and a protein-protein interaction network has identified specific components, such as GINSENG RADIX ET RHIZOMA and ASTRAGALI RADIX, playing crucial roles within the overall treatment network. This not only furnishes the molecular foundation for Osteoking's efficacy in osteoporosis treatment but also guides future endeavors towards the development of more precision-oriented therapeutic strategies. These findings carry profound clinical significance. Primarily, a nuanced understanding of the molecular mechanisms underlying osteoporosis facilitates enhanced prediction of disease risk among patients, enabling timely intervention and personalized treatment modalities. Secondly, an in-depth comprehension of Osteoking contributes to the optimization of its treatment protocols, thereby augmenting its efficacy among osteoporotic patients. This holds paramount value in enhancing patient quality of life and alleviating the burdens on healthcare resources.
Our identification of 3578 downregulated genes and 1204 upregulated genes within the GSE35959 dataset reflects crucial involvement of bone marrow mesenchymal stem cells in the onset of osteoporotic symptoms. The increased abundance of downregulated genes may be associated with diminished functionality of patients' bone marrow mesenchymal stem cells and a decline in the capacity for bone formation and repair. The emergence of upregulated genes suggests the activation of biological processes such as inflammation and cellular metabolism abnormalities, playing pivotal roles in the development of osteoporosis. The discovery of differentially expressed genes profoundly reveals the biological distinctions between osteoporotic patients and normal individuals. Under normal circumstances, the gene expression pattern of bone marrow mesenchymal stem cells typically maintains the structure and functionality of the skeleton. Among the upregulated genes, pathways involving inflammation, bone resorption, and lipid metabolism were implicated, closely aligning with known pathogenic mechanisms of osteoporosis. Simultaneously, the enrichment of downregulated genes pertained to bone marrow mesenchymal stem cell differentiation, bone cell formation, and bone matrix synthesis, suggesting that the abnormal functionality of bone marrow stem cells in osteoporotic patients may be a crucial link in the disease's progression. These findings bear significant clinical implications.
As a representative of traditional Chinese medicine formulations, the selection of Osteoking's pharmaceutical and active ingredients is well-considered, based on a profound understanding of the extensive history and widespread use of Chinese herbs in the treatment of osteoporosis. This rational selection not only reflects the accumulated knowledge of traditional medicine in the field of osteoporosis research but also provides robust support for exploring novel treatment strategies. We delved into the historical and current status of key components in Osteoking. Ingredients such as GINSENG RADIX ET RHIZOMA, ASTRAGALI RADIX, and CARTHAMI FLOS have long been considered in traditional Chinese medicine to have the efficacy of nourishing qi and blood, strengthening tendons and bones [15–20]. Through an analysis of the active ingredient-disease target network and topological analysis, we emphasize the crucial roles of components such as GINSENG RADIX ET RHIZOMA, ASTRAGALI RADIX, and CARTHAMI FLOS in the Osteoking network for treating osteoporosis. These components were found to have high degree values in the network, suggesting their potential significance throughout the treatment process. GINSENG RADIX ET RHIZOMA, as the component with the highest degree value in the network, has been widely used in traditional Chinese medicine for invigorating the body and nourishing qi and blood. ASTRAGALI RADIX and CARTHAMI FLOS are well-known in clinical practice for their properties of replenishing qi and promoting blood circulation [21, 22]. These components may impact the pathogenic mechanisms of osteoporosis through multiple pathways, including promoting bone cell proliferation, inhibiting bone resorption, and improving the functionality of bone marrow mesenchymal stem cells [23, 24]. Further comparison of our findings with existing literature reveals consistency in the selection of Osteoking's components with other research results [25, 26]. By corroborating our experimental results, we further consolidate the feasibility and potential efficacy of Osteoking in the treatment of osteoporosis.
The construction of protein-protein interaction networks is a crucial aspect in the investigation of the therapeutic mechanisms of osteoporosis. In our study, we successfully delineated the protein-protein interaction network of Osteoking's therapeutic effects on osteoporosis by integrating the analysis results from the GENEMANIA and STRING databases. This network not only provides a comprehensive perspective on understanding the mechanism of action of Osteoking but also underscores the significance of its core targets, such as PRKACA, PTGS2, and PRKCA. Firstly, it is essential to emphasize that the construction of protein-protein interaction networks is not merely a presentation of molecular-level interactions but also a grasp of potential key nodes throughout the entire therapeutic process. The high connectivity of core targets like PRKACA, PTGS2, and PRKCA in the network implies their potential critical roles in Osteoking's treatment of osteoporosis. PRKACA has been identified as significantly associated with skeletal disorders, with mutations in PRKACA being notably prevalent in individuals with skeletal defects. Furthermore, its correlation with the proliferation of osteoblasts underscores its pivotal role in bone health [27, 28]. PTGS2 is a key protein regulating inflammatory responses and cellular apoptosis, and its modulation in bone metabolism may be related to Osteoking's anti-inflammatory and pro-survival effects on bone cells [29–31]. PRKCA, a protein kinase C isoform, is intricately linked to crucial biological processes such as cell proliferation, differentiation, and apoptosis [32, 33]. The identification of these core targets holds promise for providing clues towards unraveling the molecular mechanisms underlying Osteoking's treatment of osteoporosis.
KEGG enrichment analysis represents a critical step in our study, as its results not only offer a series of enriched pathway mechanisms but also deepen our understanding of the potential effects of Osteoking on treating osteoporosis. In our investigation, pathways such as the cAMP signaling pathway and PI3K-AKT signaling pathway were found to occupy significant positions in the enrichment results. These pathways have been extensively studied in the pathophysiological processes of osteoporosis and are closely associated with the regulation of bone cell proliferation, differentiation, and metabolism. The cAMP signaling pathway plays a crucial role in bone formation by activating key molecules like cAMP response element-binding protein (CREB), regulating the expression of various genes, thus influencing the functionality of bone cells [34, 35]. On the other hand, the PI3K-AKT signaling pathway plays a pivotal role in regulating crucial biological processes such as cell survival, proliferation, and differentiation, exerting a significant impact on the activity of bone cells and bone formation [36, 37]. The discovery of these enriched pathway mechanisms provides crucial clues for explaining the effects of Osteoking in treating osteoporosis. Firstly, the enrichment of the cAMP signaling pathway may suggest that Osteoking modulates intracellular cAMP levels, affecting the expression of relevant genes, thereby mediating the regulation of bone cell proliferation, differentiation, and bone matrix generation. Secondly, the enrichment of the PI3K-AKT signaling pathway indicates that Osteoking may promote the survival and proliferation of bone cells by activating the PI3K-AKT signaling pathway, thereby aiding in the repair and regeneration of bone tissue.
Integrating active ingredients, disease targets, and osteoporosis-related pathways into a comprehensive network diagram is a crucial step in our study. This not only provides a deeper insight into the understanding of the treatment mechanism but also highlights the key roles of active ingredients such as Cetylic Acid, Sitosterol, and Kaempferol in the entire treatment network. Firstly, the integration of the active ingredient-disease target network emphasizes the potential mechanisms of action of these active ingredients in treating osteoporosis. Through Cytoscape visualization, we clearly observe the associations between Cetylic Acid, Sitosterol, and Kaempferol with disease targets like PRKACA, RXRA, PPARD. This underscores that these active ingredients may influence biological processes related to osteoporosis by regulating these key targets. Secondly, the integrated network diagram highlights the comprehensive roles of these active ingredients throughout the entire treatment network. Taking Cetylic Acid as an example, its close connection with PRKACA in the active ingredient-disease target network aligns with its crucial role in the cAMP signaling pathway implicated in KEGG enrichment analysis. At the same time, we tested the good binding ability of these targets and active ingredients through molecular docking. This multi-level association suggests that these active ingredients may exert comprehensive regulation on osteoporosis by modulating multiple pathways and targets.
Despite achieving encouraging results in this study, it is important to acknowledge some limitations. Firstly, the data primarily relies on public databases such as GEO, ETCM, GENEMANIA, and STRING. While these databases provide rich information, they are subject to inherent limitations, such as sample size and heterogeneity in experimental conditions. Future research can enhance the reliability of study results by incorporating more diverse samples and finer experimental designs. Secondly, although molecular docking results suggest that Kaempferol may play a crucial role in Osteoking's treatment, laboratory results are preliminary and limited to in vitro data. To comprehensively understand the clinical application potential of Kaempferol, more experiments, even validation in animal models, are needed. This will help determine the pharmacokinetics and toxicity characteristics of Kaempferol, providing support for its further development as a potential therapeutic drug. Future research directions should include in-depth investigations into other potential targets. While this study focused on key targets such as PRKACA, RXRA, and PPARD, osteoporosis is a complex disease involving multiple signaling pathways and biological processes. Exploring more targets, especially those closely related to the pathogenic mechanisms of osteoporosis, will help enhance the comprehensiveness and effectiveness of treatment.