Network Pharmacology Study On Estrogenic-Like Effect Of Glycosides Of Cistanche Deserticola
Apr 22, 2025
Abstract
This study explores the potential mechanism of estrogenic-like effects of glycosides of Cistanche deserticola (GCs) based on network pharmacology combined with transcriptomic data. A total of 40 active ingredients of GCs with estrogenic effects were predicted using databases such as TCMSP, SymMap, ETCM, Stitch, and QSAR.
The target information on estrogenic deficiency diseases was retrieved from GeneCards and OMIM databases. Key targets were screened, and a PPI network was constructed. GO functional enrichment analysis and KEGG pathway enrichment analysis were conducted to identify key targets and pathways.
Combined with preliminary transcriptomic data, correlation analysis between network pharmacological prediction targets and KEGG pathway was performed to further reveal the mechanism of the estrogenic-like effect of GCs.
183 targets were screened, and 1,712 targets were selected for estrogen deficiency diseases.
24 core key targets were obtained.
GO function and KEGG pathway enrichment showed that GCs mainly act on biological processes such as:
Cytokine receptor binding
Estrogen receptor binding
Growth factor receptor binding
Nuclear receptor activity
Nuclear hormone receptor binding
Steroid hormone receptor activity
Hormone receptor binding
The regulatory pathways include:
Tumor necrosis factor (TNF) signaling pathway
Interleukin-17 (IL-17) signaling pathway
Hypoxia-inducible factor-1 (HIF-1) signaling pathway
Estrogen signaling pathway
Gene expression changes were detected in all 24 core targets except Mcl1 and Gcg. Differentially expressed genes included Ar, Casp8, Esr1, Fos, and Hmox1. A total of 68 KEGG pathways were identified, with the estrogen signaling pathway, cAMP signaling pathway, and TNF signaling pathway being the main pathways.
Conclusion
Glycosides of Cistanche deserticola can exert estrogen-like effects through a "multi-component, multi-target, multi-pathway" mechanism. This mechanism may be related to the regulation of:
Estrogen signaling pathway
cAMP signaling pathway
TNF signaling pathway
Natural Cistanche extract pills instead of Estromimetic
Keywords
Glycosides of Cistanche deserticola
Estromimetic
Network pharmacology
Transcriptomics
Joint analysis
Estrogen is an essential hormone in the development of the female reproductive system and secondary sexual characteristics, playing a crucial role in regulating women's fertility and gender traits [1]. After entering menopause, women experience a decline in ovarian function, leading to reduced estrogen production, which in turn causes a series of conditions such as osteoporosis, atherosclerosis, menopausal syndrome, and coronary heart disease [2]. Traditional estrogen replacement therapy can alleviate or reduce the symptoms caused by estrogen deficiency, but this treatment may also lead to potential side effects [3]. Studies have shown that long-term use of estrogen may result in excessive proliferation of the endometrium, uterine fibroids, and mammary gland hyperplasia, among other adverse effects. Hence, seeking an effective, safe, and practical treatment to improve low estrogen levels is of great significance and value [4].
Plant estrogens are substances capable of exerting estrogen-like activity. Their molecular structure is similar to that of mammalian estrogen, enabling them to specifically bind to estrogen receptors and produce mild estrogen-like effects [5]. Research indicates that plant estrogens have various pharmacological effects, including anti-osteoporosis and altering metabolic disorders [6].
Cistanche deserticola is the dried, fleshy stem with scaly leaves of the Orobanchaceae plant Cistanche deserticola Y. C. Ma. It has functions such as tonifying kidney yang, enriching essence and blood, and moistening the intestines to relieve constipation. It is commonly used to treat kidney yang deficiency, essence and blood deficiency, impotence, infertility, soreness and weakness of the waist and knees, and constipation caused by intestinal dryness [7]. Previous studies by the research group confirmed that total glycosides of Cistanche deserticola (GCs), with a content of greater than 60% of echinacoside, are the main active components responsible for its estrogen-like effects. GCs significantly increase the uterine coefficient and the height of endometrial epithelial cells in immature rats. Transcriptomic analysis results showed that GCs exert estrogen-like effects through 2,953 differentially expressed genes, of which 1,475 were upregulated and 1,478 were downregulated. These genes are mainly related to estrogen response, positive regulation of nitrogen compound metabolic processes, developmental processes, and biological process regulation. They primarily participate in the estrogen signaling pathway, cAMP signaling pathway, Rap1 signaling pathway, and MAPK signaling pathway [8-9].
Traditional Chinese medicine (TCM) network pharmacology can explore the mechanisms of TCM in disease treatment by screening TCM components, predicting the target proteins of active components, and analyzing the signaling pathways involved in these target proteins [10-12]. To further reveal the mechanism of the estrogen-like effects of GCs, this study employs bioinformatics analysis based on the group's previous transcriptomic data to uncover the possible mechanism by which total glycosides of Cistanche deserticola exert estrogen-like effects.
1. Experimental Databases and Software
TCMSP: http://lsp.nwu.edu.cn/tcmsp.php
SymMap: https://www.symmap.org
ETCM: http://www.nrc.ac.cn:9090/ETCM/
Stitch: http://stitch.embl.de/
QSAR: http://targetnet.scbdd.com/
PUBChem: https://pubchem.ncbi.nlm.nih.gov
UniProt: http://www.uniprot.org/
GeneCards: http://www.genecards.org/
OMIM: https://www.omim.org/
STRING: https://string-db.org/
KEGG: https://www.kegg.jp/
GO: http://geneontology.org/
R Language: Rx64 3.5.3 with the following packages:
VennDiagram
count.R
clusterProfiler
org.Hs.eg.db
symbol2id
Perl Script: addSymbol.pl
Cytoscape: Version 3.7.1
2. Experimental Methods
2.1 Network Pharmacology Prediction of GCs' Estrogen-like Mechanism
2.1.1 Prediction of Targets of GCs' Main Active Components
Using UPLC/Q-TOF-MS analysis, 40 compounds with estrogen-like pharmacological activity were identified [13]. These compounds were searched in TCMSP, SymMap, and ETCM databases to find potential gene targets for each compound. Additionally, compound SMILES structures were imported into Stitch and QSAR databases to collect targets. All collected targets were merged, and duplicates were removed to obtain potential target proteins.
2.1.2 Estrogen-like Target Database Information
The keywords "Estrogen deficiency" were used to search for relevant targets in the GeneCards and OMIM databases.
2.1.3 Construction of the "Active Compound - Core Target - Estrogen-like" Network
Using the VennDiagram package in R, the intersection of targets from both sources was plotted as a Venn diagram. Candidate protein interaction data were imported into Cytoscape v3.9.1, and core targets were identified. The corresponding compounds of the core targets were selected, and the "Compound - Core Target - Estrogen-like" network was visually analyzed and constructed using Cytoscape v3.9.1.
2.1.4 Construction of the PPI Network
Core target information was imported into the STRING database, selecting the species "Homo sapiens" and setting the confidence score to >0.7 to construct the protein-protein interaction (PPI) network. The count.R package in R was used to analyze and select the top 20 core genes with the highest connectivity among core target proteins, which were visualized.
2.1.5 GO Functional Enrichment Analysis and KEGG Pathway Enrichment Analysis
For core target genes, GO and KEGG database searches were performed using the R packages org.Hs.eg.db, symbol2id, and clusterProfiler. The top 20 pathways with corrected p-values were visualized as bar charts, analyzing the significant functional annotations and pathways enriched by the core targets of GCs' estrogen-like effects.
2.2 Integration with Transcriptomic Data
2.2.1 Correlation Analysis Between Predicted Targets and Transcriptomic Genes
Since the species in network pharmacology predictions is human, the targets were converted into homologous rat genes using HGNC via the "Tools-HCOP" function. The expression levels of differential genes were analyzed between the GCs group and the control group in transcriptomic sequencing [9].
2.2.2 Correlation Analysis Between Network Pharmacology Predictions and Transcriptomic KEGG Pathways
The core target genes predicted by network pharmacology (converted to rat homologous genes) were searched in the KEGG database using the R packages bioconductor and clusterProfiler along with the id2symbol.pl script. The species was restricted to "rno," and pathways with Padjust < 0.05 were screened. Venn analysis was conducted to identify overlapping pathways between KEGG pathways enriched through network pharmacology predictions and transcriptomic experiments related to GCs' estrogen-like effects.
