The application of rhubarb concoctions in traditional Chinese medicine and its compounds, processing methods, pharmacology, toxicology and clinical research.

Objective: This study reviews the development of rhubarb processing and the current status of pharmacological research. We summarized the effects of different processing methods on the active compounds, pharmacological effects, and toxicity of rhubarb, as well as the clinical application of different concoctions, providing reference for further pharmacological research and clinical application of rhubarb. Methods: A comprehensive literature review was conducted using databases such as Pubmed, Embase, National Science and Technology Library, Web of science, CNKI, China Science and Technology Journal Database, SinoMed, and the Pharmacopoeia of the People's Republic of China. Search terms included "rhubarb", "raw rhubarb", "wine rhubarb", "cooked rhubarb", "rhubarb charcoal", "herbal processing", "compounds", "pharmacological effects", "inflammation", "gastrointestinal bleeding", and "tumor". Results: Historical records of rhubarb processing date back to the Han Dynasty, with continual innovations. Currently, the types of rhubarb used in traditional Chinese medicine have stabilized to three species: Rheum palmatum L., Rheum tanguticum Maxim.ex Balf. and Rheum officinale Baill. Common concoctions include raw rhubarb, wine rhubarb, cooked rhubarb and rhubarb charcoal. The active compounds of rhubarb are known to defecation, exhibit antibacterial and anti-inflammatory properties, regulate coagulation, protect the digestive system, and possess anti-tumor activities. Guided by Chinese medicine theory, the use of different rhubarb concoctions can enhance specific effects such as purgation to eliminate accumulation, clearing heat and toxins, cooling blood to stop hemorrhages, activating blood circulation to remove blood stasis, and inducing dampness to descend jaundice, thereby effectively treating various diseases. The therapeutic impact of these concoctions on diseases reflects not only in the changes to the active compounds of rhubarb but also in the formulations of traditional Chinese medicine. Processing has also shown advantages in reducing toxicity. Conclusion: Different processing methods alter the active compounds of rhubarb, thereby enhancing its various pharmacological effects and meeting the therapeutic needs of diverse diseases. Selecting an appropriate processing method based on the patient's specific conditions can maximize its pharmacological properties and improve clinical outcomes.

mtDNA regulates cGAS-STING signaling pathway in adenomyosis.

In various hyperproliferative disorders, damaged mitochondria can release mitochondrial DNA (mtDNA) into the cytoplasm, activating the cGAS-STING signaling pathway and subsequent immune imbalances. Our previous research has demonstrated that hypoxia plays a role in the development of adenomyosis (AM) by inducing mitochondrial dysfunction. However, the precise involvement of the cGAS-STING signaling pathway and mtDNA in AM remains unclear. Therefore, this study aims to investigate the relationship between mtDNA secretion, changes in the cGAS-STING signaling pathway, and the abnormal cellular proliferation observed in AM. We found the cGAS, STING, TBK1, p-TBK1, IRF3, and p-IRF3 proteins levels were significantly elevated in the tissues of patients with AM compared to the control group. Additionally, there was an increase in the expression of the pro-inflammatory cytokines IL-6 and IFN-α in the AM tissues. Hypoxia-induced an increase in the proliferation and migration abilities of endometrial stromal cells (ESCs), accompanied by the activation of the cGAS-STING signaling pathway and elevated levels of IFN-α. Furthermore, hypoxia promoted the leakage of mtDNA into the cytoplasm in AM ESCs, and the deletion of mtDNA reduced the activation of the cGAS-STING pathway. Moreover, knockdown of the STING gene inhibited the expression of TBK1, p-TBK1, IRF3, and p-IRF3 and suppressed the secretion of the inflammatory cytokines IL-6 and IFN-α. Furthermore, the migration and invasion abilities of AM ESCs were significantly diminished after STING knockdown. These findings provide valuable insights into the role of mtDNA release and the cGAS-STING signaling pathway in the pathogenesis of AM.

Role of Pink1-mediated mitophagy in adenomyosis.

Abstract Background: Recent studies indicate that endometrial hypoxia plays a critical role in adenomyosis (AM) development. Mitochondria are extremely sensitive to hypoxic damage, which can result in both morphological and functional impairment. Mitophagy is a crucial mechanism for preserving mitochondrial quality by selectively removing damaged mitochondria, thus ensuring the proper functioning of the entire mitochondrial network. In response to hypoxia, PINK1 is activated as a regulator of mitophagy, but its role in AM requires further study. Objective: To explore the potential mechanism of mitophagy mediated by PINK1 in the pathogenesis of AM. Method: The study compared PINK1, Parkin, OPTIN, P62, and NDP52 protein expression levels in patients with or without AM using clinical specimens and an AM mouse model. Pathological changes were compared using HE staining. Immunofluorescence and western blot were used to detect protein expression levels. Endometrial stromal cells (ESC) were isolated and examined for mitophagy, protein expression level, and cell invasion ability. Results: Both the endometrial tissue from patients with AM and AM ESC displayed an upregulation of protein levels for PINK1, Parkin, OPTIN, P62, and NDP52 when compared with the control group. Then, HE staining confirmed the successful establishment of the AM mouse model. Moreover, the ultrastructural analysis using transmission electron microscopy revealed that AM mice's endometrial glandular epithelial and stromal cells had exhibited swollen, deformed, and reduced mitochondria along with an increase in the number of lysosomes and mitochondrial autophagosomes. The protein levels of PINK1, Parkin, OPTIN, P62, and NDP52 in uterine tissue from AM mice were noticeably increased, accompanied by a considerable upregulation of ROS levels compared to the control group. In addition, cells in the AM group showed remarkably elevated mitophagy and invasion potentials compared to the control group. In contrast, the cell invasion ability decreased following PINK1 knockdown using the RNA interference technique. Conclusion: The high levels of PINK1-mediated mitophagy have been found in AM. The upregulation in mitophagy contributes to mitochondrial damage, which may result in the abnormal invasion characteristic of AM.

Network pharmacology prediction and experimental verification of Rhubarb-Peach Kernel promoting apoptosis in endometriosis.

BACKGROUND: "Rhubarb-Peach Kernel" herb pair (RP) one of the most frequently used drug pairs, has been used in traditional medicine in China to treat inflammation and diseases associated with pain. Although it is widely used clinically and has a remarkable curative effect, the mechanism of RP treatment for endometriosis (EMs) remains unclear due to its complicated components. The aim of this study was to investigate the anti-endometriosis effect of RP, with emphasis on apoptosis via network pharmacology prediction, molecular docking and experimental verification. METHODS: The related ingredients and targets of RP in treating EMs were screened out using Traditional Chinese Medicine Systems Pharmacology (TCMSP), Tool for Molecular mechanism of Traditional Chinese Medicine (BATMAN-TCM), and GeneCards database. The data of the protein-protein interaction (PPI) network was obtained by the Search Tool for the Retrieval of Interaction Gene/Proteins (STRING) Database. The Metascape database was adopt for Kyoto encyclopedia of genes and genomes (KEGG) enrichment analysis. After that, the molecular docking of the main active ingredients and apoptosis targets was performed. Finally, the pro-apoptotic effect of RP was verified in hEM15a cells. RESULTS: A total of 32 RP compounds were collected. Forty-two matching targets were picked out as the correlative targets of RP in treating EMs. Among these, 18 hub targets including P53, CASP3 were recognized by the PPI network. KEGG enrichment analysis discovered that the regulation of apoptosis was one of the potential mechanisms of RP against EMs. Anthraquinone compounds, flavonoids, and triterpenes in RP were identified as crucial active ingredients, involved in the pro-apoptotic effect, which were confirmed subsequently by molecular docking. Additionally, it was verified that RP treatment promoted apoptosis and inhibited the proliferation of EMs cells (assessed by MTT and Flow cytometry). Moreover, the induction of apoptosis in treated EMs cells may be due to the regulation of apoptosis-related protein expression, including P53, BAX, and CASP3. CONCLUSIONS: The results of our study demonstrated that RP may exert its therapeutic effects on EMs through the potential mechanism of promoting apoptosis. Anthraquinones, flavonoids and triterpenoids are the possible pro-apoptotic components in RP.

Dahuang-Taoren, a botanical drug combination, ameliorates adenomyosis via inhibiting Rho GTPases.

Introduction: Dahuang-Taoren (DT) is a classic combination of botanical drugs applied to treat pain-related diseases in ancient China. Today, DT is frequently applied for dysmenorrhea of adenomyosis (AM) in the clinic. Growing evidence indicates Rho GTPases may play an essential role in AM progression. However, the potential mechanism of DT on Rho GTPases in AM remains unclear. Methods: The expressions of Rho GTPases in the patients with AM were evaluated. Further, pituitary transplantation-induced AM mice and the primary AM endometrial stromal cells (AMESCs) were subjected to DT intervention. Results: The results revealed that the expressions of Rho GTPases were significantly upregulated in both AM patients and AM mice. The DT could reduce pathological infiltration, relieve hyperalgesia, and alleviate cytoskeleton remodeling in AM mice. Besides, the migration and invasion of AMESCs were markedly inhibited after exposure to DT. Discussion: These effects may be linked to the decreased Rho GTPases expression. The results may offer a novel explanation of DT against AM.

An insight into gut microbiota and metabolites in the mice with adenomyosis.

Background: Adenomyosis (AM) is a benign uterine disease characterized pathologically by the invasion of endometrial tissue into the myometrium. The pathogenesis of AM is still far from clear. Although the gut microbiome and metabolomics are thought to contribute to a variety of diseases, the role of them in AM has not been revealed. Objective: To investigate changes in the gut microbiota and derived metabolites in AM mice. Method: Female ICR mice were randomly assigned to AM and control groups, and pituitary transplantation was employed to perform AM modeling. Then, the fecal samples were obtained for microbial (16S rRNA gene sequencing) and metabolomic (liquid chromatography mass spectrometry, LC-MS) analysis. Result: The results of gut microbiota analysis showed that the intestinal microbiota composition of AM mice was altered. The ratio of Firmicutes/Bacteroidetes and the relative abundance of Lactobacillus in AM group increased compared with the control group. Sixty differential expressed metabolites were identified in intestinal metabolites, mainly involved in steroid hormone biosynthesis, cysteine and methionine metabolism, and alanine, aspartate, and glutamate metabolism. Further, correlation analysis verified that L-methionine and L-cystine were negatively correlated with Bacteroides and positively correlated with Desulfovibrio. The Pregnenolone, Androsterone glucuronide, and Testosterone glucuronide were negatively correlated with Unidentified_Ruminococcaceae and Alistipes, whereas they positively correlated with Bacteroides. Conclusion: AM mice have a unique gut microbiome and intestinal metabolites.

Integrating Network Pharmacology and Experimental Validation Deciphers the Mechanism of Guizhi Fuling Wan against Adenomyosis.

AIM: This study aimed to predict the key targets and endocrine mechanisms of Guizhi Fuling Wan (GZFLW) in treating adenomyosis (AM) through network pharmacology, molecular docking, and animal experiment verification. METHODS: The related ingredients and targets of GZFLW in treating AM were screened out using TCMSP, BATMAN-TCM, SwissTargetPrediction, and PubChem Database. Then, the protein-protein interaction (PPI) analysis and the network of compound-hub targets were constructed. At the same time, the key targets were uploaded to the Metascape Database for KEGG pathway enrichment analysis. After that, the molecular docking technology of the main active components and hub targets was performed. Furthermore, animal experiments were used to verify the results of network pharmacology analysis. RESULTS: A total of 55 active ingredients of GZFLW and 44 overlapping targets of GZFLW in treating AM were obtained. After screening, 25 hub targets were collected, including ESR1, EGF, and EGFR. Then, the KEGG pathway enrichment analysis results indicated that the endocrine therapeutic mechanism of GZFLW against AM is mainly associated with the estrogen signaling pathway, endocrine resistance, and an EGFR tyrosine kinase signaling pathway. Then, molecular docking showed that the significant compounds of GZFLW had a strong binding ability with ERα and EGFR. More importantly, the animal experiments confirmed that the GZFLW could downregulate the abnormal infiltration of the endometrial epithelium into the myometrium and had no interference with the normal sexual cycle. This effect may be directly related to intervening the local estrogen signaling pathway of the endometrial myometrial interface (EMI). It may also be associated with the myometrium cells' estrogen resistance via GPER/EGFR signaling pathway. CONCLUSION: The endocrine mechanism of GZFLW in treating AM was explored based on network pharmacology, molecular docking, and animal experiments, which provided a theoretical basis for the clinical application of GZFLW.

Computational Prediction of Antiangiogenesis Synergistic Mechanisms of Total Saponins of Panax japonicus Against Rheumatoid Arthritis.

Objective: To investigate the anti-angiogenesis mechanisms and key targets of total saponins of Panax japonicus (TSPJ) in the treatment of rheumatoid arthritis (RA). Methods: RStudio3.6.1 software was used to obtain differentially expressed genes (DEGs) by analyzing the differences in gene expression in the synovial tissue of RA and to predict the potential targets of active compounds from TSPJ by the PharmMapper and SwissTargetPrediction databases. We evaluated the overlapping genes by intersectional analysis of DEGs and drug targets. Based on the overlapping genes, we used Cytoscape 3.7.2 software to construct a protein-protein interactions (PPI) network and applied Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis to determine the mechanisms of the treatment. Finally, the correlations with angiogenesis-related genes were explored. Collagen-induced arthritis (CIA) model was established and treated with different doses of TSPJ. The manifestations of CIA were determined by evaluation of arthritis index and histology score. Serum levels of vascular endothelial growth factor (VEGF) and the hypoxia-inducible factor 1 (HIF-1) were tested by ELISA. The mRNA levels of IL-1ß and IL-17A were detected by real time-quantitative PCR. Results: Altogether, 2670 DEGs were obtained by differential analysis, and 371 drug targets were predicted for four active components (Araloside A, Chikusetsusaponin IVa, Ginsenoside Rg2, and Ginsenoside Ro). A total of 52 overlapping genes were included in the PPI network and the KEGG analysis. However, only 41 genes in the PPI network had protein interactions. The results of the KEGG enrichment analysis were all related to angiogenesis, including VEGF and HIF-1 signaling pathways. Seven genes with negative correlations and 16 genes with positive correlations were obtained by correlational analysis of DEGs in the VEGF and HIF-1 signaling pathways. SRC proto-oncogene, nonreceptor tyrosine kinase (SRC), and the signal transducer and activator of transcription 3 (STAT 3) had a higher value of degree and showed a significant correlation in the pathways; they were regarded as key targets. Compared with the model group, TSPJ significantly relieved the symptoms and decreased the expression of VEGFA, HIF-1α, IL-1ß, and IL-17A in serum or spleens of CIA mice. Conclusion: In the current study, we found that antiangiogenesis is one of the effective strategies of TSPJ against RA; SRC and STAT 3 may be the key targets of TSPJ acting on the VEGF and HIF-1 signaling pathways, which will provide new insight into the treatment of RA by inhibiting inflammation and angiogenesis.

Rhein ameliorates adenomyosis by inhibiting NF-κB and ß-Catenin signaling pathway.

In the present study, we examined the effects of rhein on pituitary gland implantation-induced adenomyosis, an animal model which mimics human adenomyosis. Oral administration of rhein dose-dependently attenuated hyperplastic and hypertrophic myometrium and improved adenomyosis. The activation of NF-κB and ß-catenin signaling pathway was observed in the ectopic endometria. While, rhein dose dependently inhibited the expressions of p-p65, p-AKT and actived Rac1. As Rac1 activation controlled nuclear localization of ß-catenin during canonical Wnt signaling, we found that the degradation complex of ß-catenin was improved by rhein. In addition, ß-catenin nuclear translocation and its downstream genes were markedly suppressed by different doses of rhein. At the same time, decreased activation of epithelial-mesenchymal transition (EMT)-associated proteins including Snail and ZEB1 was detected in rhein-treated mice, indicating that the activation of Wnt signaling pathway was suppressed by rhein. The in vitro study verified a negative regulation of rhein on ß-catenin in stromal cells. Stimulation of IL-1ß significantly increased the nuclear translocation of ß-catenin and improved its target genes expressions. While, rhein remarkably abolished the enhancement in a dose dependent manner. Taken together, our results demonstrated the ability of rhein to inhibit Wnt/ß-catenin activation and its potential use in the treatment of adenomyosis and other abnormal activation of ß-catenin -associated diseases.