Exploring the pharmacological mechanisms of the flower of Rhododendron molle in rheumatoid arthritis rats based on metabolomics integrated network pharmacology.

ETHNOPHARMACOLOGICAL RELEVANCE: As a traditional Chinese medicine, the flower of Rhododendron molle G. Don (RMF) is record in the Chinese pharmacopoeia, and is commonly utilized for treating rheumatoid arthritis (RA) in clinical practice. However, its precise mechanisms necessitate further exploration. AIM OF THE STUDY: To expound the effective components, targets, metabolites, and pathways participated in RMF's anti-RA effects by metabolomics integrated network pharmacology. MATERIALS AND METHODS: CIA rats were intragastric administered RMF for 2 weeks, following which the therapeutic effects were comprehensively evaluated. Serum metabolomics was adopted to investigate the differential metabolites (DEMs). UHPLC-Q-Exactive-MS method was applied to identify the components of RMF, and then network pharmacology was utilize to select the component-RA-targets. Molecular docking and Western blotting were utilized to validate the key targets. RESULTS: RA symptoms were alleviated by RMF through the inhibition secretion of pro-inflammatory factors IL-1ß, IL-6 and TNF-α, along with relief in bone destruction observed in CIA rats. Four targets, namely AKR1B1, TPH1, CYP1A1, and CYP1A2, were identified, along with their corresponding metabolites, namely D-glucose, D-mannose, L-tryptophan, 11-deoxycorticosterone, and 17α-hydroxyprogesterone. These were found to be involved in three key metabolic pathways: steroid hormone biosynthesis, tryptophan metabolism, and galactose metabolism. Additionally, five significant anti-RA active components were identified from RMF, including Rhodojaponin (Rj)-Ⅱ, Rj-Ⅲ, Rj-Ⅴ, Rj-Ⅵ, and quercetin. CONCLUSIONS: The anti-RA mechanisms of RMF were investigated in this study, focusing on active components, upstream targets, and downstream metabolites. These findings lay a foundation for the clinical practice and drug development of RMF.

Ganlu formula ethyl acetate extract (GLEE) blocked the development of experimental arthritis by inhibiting NLRP3 activation and reducing M1 type macrophage polarization.

ETHNOPHARMACOLOGICAL RELEVANCE: The Tibetan medicine Ganlu Formula, as a classic prescription, is widely used across the Qinghai-Tibet Plateau area of China, which has a significant effect on relieving the course of rheumatoid arthritis (RA). However, the active compounds and underlying mechanisms of Ganlu Formula in RA treatment remain largely unexplored. AIM OF THE STUDY: This study aimed to elucidate the active substances and potential mechanisms of the ethyl acetate extract of Ganlu Formula ethyl acetate extract (GLEE) in the treatment of RA. MATERIALS AND METHODS: Ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) was utilized to analyze and identify the chemical constituents within GLEE. Discovery Studio molecular virtual docking technology was utilized to dock the interaction of GLEE with inflammation-related pathway proteins. The GLEE gene library was obtained by transcriptome sequencing. Collagen-induced arthritic（CIA) rats were utilized to assess the antiarthritic efficacy of GLEE. Micro-CT imaging was employed to visualize the rat paw, and ultrasound imaging revealed knee joint effusion. Evaluation of synovial tissue pathological changes was conducted through hematoxylin-eosin staining and saffranine solid green staining, while immunohistochemical staining was employed to assess NLRP3 expression along with inflammatory markers. Immunofluorescence staining was utilized to identify M1 macrophages. RESULTS: Metabolomic analysis via UPLC-Q-TOF-MS identified 28 potentially bioactive compounds in GLEE, which interacted with the active sites of key proteins such as NLRP3, NF-κB, and STAT3 through hydrogen bonds, C-H bonds, and electrostatic attractions. In vitro analyses demonstrated that GLEE significantly attenuated NLRP3 inflammasome activation and inhibited the polarization of bone marrow-derived macrophages (BMDMs) towards the M1 phenotype. In vivo, GLEE not only prevented bone mineral density (BMD) loss but also reduced ankle swelling in CIA rats. Furthermore, it decreased the expression of the NLRP3 inflammasome and curtailed the release of inflammatory mediators within the knee joint. CONCLUSION: GLEE effectively mitigated inflammatory responses in both blood and knee synovial membranes of CIA rats, potentially through the down-regulation of the NLRP3/Caspase-1/IL-1ß signaling pathway and reduction in M1 macrophage polarization.

Chinese medicine Di-long (Pheretima vulgaris) and its active fraction exhibit anti-rheumatoid arthritis effects by inhibiting CXCL10/CXCR3 chemotaxis in synovium.

ETHNOPHARMACOLOGICAL RELEVANCE: Di-Long (Pheretima vulgaris) is a classic animal sourced traditional Chinese medicine. It has been used for the treatment of joint inflammation and arthralgia for over two thousand years due to its effects of Tong-Luo-Zhi-Tong (dredging collaterals and alleviating pain). Our previous study showed that Chinese medicine Di-Long has significant anti-rheumatoid arthritis (RA) effects. AIM OF THE STUDY: Considering Di-Long as a potential source of active compounds with specific anti-RA therapeutic effects, this research was to obtain the anti-RA target-specific active fraction from Di-Long extracts (DL), and to further explore the chemical basis and verify the anti-RA mechanism of this active fraction. MATERIALS AND METHODS: Transcriptomic was applied to obtain the main anti-RA targets of DL on human RA fibroblast-like synoviocytes (FLS) and validated by qPCR. The target-corresponding active fraction was isolated from DL by ethanol precipitation and gel chromatography, and analyzed by nanoliter chromatography-mass spectrometry. Anti-RA effects of this active fraction was investigated by collagen-induced arthritis (CIA) in mice, and anti-RA mechanisms were verified in cocultured model of rat FLS and peripheral blood lymphocytes. RESULTS: We confirmed that CXCL10/CXCR3 was the main anti-RA target of DL. The active fraction - A (2182 - 890 Da) was isolated from DL based on its CXCL10 inhibiting effects in RA-FLS. Fraction A contains 195 peptides (192 were newly discovered), 26 of which might be bioactive and were considered to be the chemical basis of its anti-RA effects. Fraction A significantly ameliorated the joint destruction and overall inflammation in CIA mice, and downregulated CXCR3 expression in mice joint. Fraction A inhibited the chemotaxis of Th-cells in rat peripheral blood lymphocytes towards the TNF-α-induced rat FLS through CXCL10/CXCR3 pathway. CONCLUSIONS: Our work indicated that active fraction from DL containing small peptides exhibits promising therapeutic effects for RA through inhibiting CXCL10/CXCR3 chemotaxis.

Network pharmacology combined with affinity ultrafiltration to elucidate the potential compounds of Shaoyao Gancao Fuzi Decoction for the treatment of rheumatoid arthritis.

ETHNOPHARMACOLOGICAL RELEVANCE: Shaoyao Gancao Fuzi Decoction (SGFD), has been employed for thousands of years in the treatment of rheumatoid arthritis (RA) with remarkable clinical efficacy. However, the material basis underlying the effectiveness of SGFD still remains unclear. AIM OF THE REVIEW: This study aims to elucidate the material basis of SGFD through the application of network pharmacology and biological affinity ultrafiltration. RESULTS: UPLC-Q-TOF-MS/MS was employed to characterize the components in SGFD, the identified 145 chemical components were mainly categorized into alkaloids, flavonoids, triterpenoids, and monoterpenoids according to the structures. Network pharmacology method was utilized to identify potential targets and signaling pathways of SGFD in the RA treatment, and the anti-inflammatory and anti-RA effects of SGFD were validated through in vivo and in vitro experiments. Moreover, as the significant node in the pharmacology network, TNF-α, a classical therapeutic target in RA, was subsequent employed to screen the interacting compounds in SGFD via affinity ultrafiltration screening method, 6 active molecules (i.e.,glycyrrhizic acid, paeoniflorin, formononetin, isoliquiritigenin, benzoyl mesaconitine, and glycyrrhetinic acid) were exhibited significant interactions. Finally, the significant anti-inflammatory and anti-TNF-α effects of these compounds were validated at the cellular level. CONCLUSIONS: In conclusion, this study comprehensively elucidates the pharmacodynamic material basis of SGFD, offering a practical reference model for the systematic investigation of traditional Chinese medicine formulas.

Anti-inflammatory and anti-rheumatic effects of Phoenix dactylifera L. (date palm) seed by controlling cytokines and inhibiting JAK1/STAT3 pathway on CFA-induced arthritis rat and its phytochemical profiling.

ETHNOPHARMACOLOGICAL RELEVANCE: Phoenix dactylifera L. (date palm) seed is widely used in Arabian traditional medicine to alleviate several health problems including inflammatory conditions. The herbal tea of date palm seed has been consumed by rheumatoid patients to relief their symptoms. AIM OF THE STUDY: The purpose of this study was to investigate the claimed beneficial use of P. dactylifera L. (Sewy variety) seed (PDS) in the treatment of rheumatoid arthritis (RA) and its mechanism of action as well as to study its phytoconstituents. MATERIALS AND METHODS: The anti-inflammatory and anti-oxidative properties of the non-polar and the polar extracts of PDS were studied using Complete Freund's adjuvant (CFA)-induced arthritis rat model. Paw edema, body weight, total nitrate/nitrite NOX content and cytokine markers were evaluated to monitor the progress of arthritis. Also, histological examination and thermal analysis were conducted. The phytoconstituent profiles of non-polar and polar extracts of PDS were investigated using liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). The multiple reactions monitoring mode (MRM) of liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) was used to quantify phenolic phytoconstituents in both extracts. RESULTS: According to the findings, the polar and non-polar PDS extracts kept body weight comparable to those of healthy individuals while considerably lowering paw swelling, edema, and neutrophil infiltration. It also reduced the levels of Nuclear Factor Kappa B (NF-κB), Tumor Necrosis Factor Alpha (TNF-α), Interleukin 22, Interleukin 23, Interferon (IFN), Interleukin 17, Interleukin 1ß, Interleukin 6, Interleukin 36, Janus Kinase 1 (JAK1), and Signal Transducer and Activator of Transcription 3 (STAT3). They also reduced the degenerative alterations caused by RA. Thermal research gave additional support for these findings. 83 phytoconstituents were identified in the non-polar PDS extract and 86 phytoconstituents were identified in the polar PDS extract. 74 of the identified phytoconstituents were common in both extracts. 33 phytoconstituents were identified here from P. dactylifera for the first time as far as we know. In MRM-LC-ESI-MS/MS analysis, the major phenolics in both extracts were chlorogenic acid, naringenin, and vanillin. Catechin was only detected in the non-polar PDS extract. On the other hand, apigenin, kaempferol, and hesperetin were only detected in the polar PDS extract. Generally, the polar PDS extract showed higher concentrations of the identified phenolics than the non-polar extract. CONCLUSIONS: The PDS extracts especially the non-polar extract showed significant anti-inflammatory and anti-oxidative properties in the CFA-induced arthritis rat model. PDS might be used to produce RA medicines.