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Hongling Du,1– 3 Shunjie Zhang,1 Kezhu Yuan,4 Zhirui Yang,5 Mingquan Wu4 1Department of Pharmacy, West China Hospital, Sichuan University, Chengdu, Sichuan, People’s Republic of China; 2Department of Pharmacy, Sichuan Public Health Clinical Center, Chengdu, Sichuan, People’s Republic of China; 3State Key Laboratory of Southwestern Chinese Medicine Resources, Pharmacy School, Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan, People’s Republic of China; 4Department of Pharmacy, Sichuan Orthopedic Hospital, Chengdu, Sichuan, People’s Republic of China; 5Department of Nuclear Medicine, Chengdu Second People’s Hospital, Chengdu, Sichuan, People’s Republic of ChinaCorrespondence: Mingquan Wu, Department of Pharmacy, Sichuan Orthopedic Hospital, Chengdu, Sichuan, People’s Republic of China, Email 18782976939@163.comBackground: Rehmanniae Radix (RR) has received attention for its antithrombotic effect. However, few studies have independently explored the bioactive components responsible for its antithrombotic bioactivity and the potential mechanism. We aimed to reveal the antithrombotic mechanisms of RR by using metabolomics integrated with network pharmacology.Methods: A thrombosis model was established by intraperitoneal injection of type I carrageenan in rats, and antithrombotic function was evaluated at different doses of RR. Metabolomics was used to identify the differential metabolites in the serum. Network pharmacology was then applied to identify the potential targets for the antithrombotic activity of the RR. An integrated network of metabolomics and network pharmacology was constructed using Cytoscape. Finally, key targets were verified using molecular docking.Results: RR at 5.4 g/kg significantly alleviated the thrombosis. Thirteen potentially significant metabolites were involved in the therapeutic effects of RR against thrombosis, most of which were regulated for recovery after RR treatment. An integrated analysis of metabolomics and network pharmacology showed that the antithrombosis effect of RR was closely associated with the regulation of PLA2G2A, PTGS1, ALOX5, and CYP2C9. Molecular docking showed high affinity between the key targets and components of RR. We speculated that the components of RR, such as catalpol, ferulic acid methyl ester, and methyl 4-hydroxycinnamate, might act on key proteins, including PLA2G2A, PTGS1, and ALOX5, to exert antithrombosis effects.Conclusion: This study confirmed the antithrombotic effect of high-dose RR, revealed the antithrombotic mechanism and potential material basis, and laid the foundation for the antithrombotic clinical application of RR. Furthermore, it provides a successful case reference for screening natural herbal components and exploring their potential pharmacological mechanisms.Keywords: Rehmanniae radix, antithrombotic effects, metabolomics, network pharmacology

Min-Dong Du,1,2,* Kai-Yi He,3,* Si-Qi Fan,3 Jin-Yi Li,3 Jin-Fu Liu,3 Zi-Qiang Lei,3 Gang Qin3 1Department of Osteoarthrosis, Xing-An Jieshou Orthopedics Hospital, Guilin, People’s Republic of China; 2Department of Orthopaedic Trauma and Hand Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, People’s Republic of China; 3Department of Osteoarthrosis, The First Affiliated Hospital of Guangxi Traditional Chinese Medical University, Nanning, People’s Republic of China*These authors contributed equally to this workCorrespondence: Gang Qin, Department of Osteoarthrosis, The First Affiliated Hospital of Guangxi Traditional Chinese Medical University, 89-9 Dongge Road, Nanning, Guangxi, 530023, People’s Republic of China, Tel +86-771-5361264, Email qqvvcbgq6016@163.comBackground: Cyperus rotundus (CR) is widely used in traditional Chinese medicine to prevent and treat a variety of diseases. However, its functions and mechanism of action in osteoarthritis (OA) has not been elucidated. Here, a comprehensive strategy combining network pharmacology, molecular docking, molecular dynamics simulation and in vitro experiments was used to address this issue.Methods: The bioactive ingredients of CR were screened in TCMSP database, and the potential targets of these ingredients were obtained through Swiss Target Prediction database. Genes in OA pathogenesis were collected through GeneCards, OMIM and DisGeNET databases. Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis were performed using DAVID database. STRING database and Cytoscape 3.10 software were used to construct “component-target-pathway” network, and predict the core targets affected by CR. The binding affinity between bioactive components and the core targets was evaluated by molecular docking and molecular dynamics simulation. The therapeutic activity of kaempferol on chondrocytes in inflammatory conditions was verified by in vitro experiments.Results: Fifteen CR bioactive ingredients were obtained, targeting 192 OA-related genes. A series of biological processes, cell components, molecular functions and pathways were predicted to be modulated by CR components. The core targets of CR in OA treatment were AKT serine/threonine kinase 1 (AKT1), interleukin 1 beta (IL1B), SRC proto-oncogene, non-receptor tyrosine kinase (SRC), BCL2 apoptosis regulator (BCL2), signal transducer and activator of transcription 3 (STAT3), epidermal growth factor receptor (EGFR), hypoxia-inducible factor 1 subunit alpha (HIF1A), matrix metallopeptidase 9 (MMP9), estrogen receptor 1 (ESR1) and PPARG orthologs from vertebrates (PPARG), and the main bioactive ingredients of CR showed good binding affinity with these targets. In addition, kaempferol, one of the CR bioactive components, weakens the effects of IL-1β on the viability, apoptosis and inflammation of chondrocytes.Conclusion: Theoretically, CR has great potential to ameliorate the symptoms and progression of OA, via multiple components, multiple targets, and multiple downstream pathways. Keywords: osteoarthritis, Cyperus rotundus, network pharmacology, molecular docking

Jiayue Zhou,1,2 Yingxiang Wu,1,2 Zhiyan Lu,1,2 Yan Wang1,2 1Yunnan Provincial Key Laboratory of Entomological Biopharmaceutical R&D, Dali University, Dali, Yunnan Province, 671000, People’s Republic of China; 2National-Local Joint Engineering Research Center of Entomoceutics, Dali University, Dali, Yunnan Province, 671000, People’s Republic of ChinaCorrespondence: Yan Wang, Yunnan Provincial Key Laboratory of Entomological Biopharmaceutical R&D, Dali University, Dali, Yunnan Province, 671000, People’s Republic of China, Tel +86 0872-2257401, Email jessica9428@sina.comPurpose: Rodgersia sambucifolia Hemsl (also known as Yantuo) is a traditional Chinese medicine commonly utilized as a medicinal herb with its rhizomes, mainly used to regulate the immune function of the human body. However, relatively few studies have investigated its active components and potential mechanisms of action in vivo.Methods: First, the chemical composition in vitro was identified and analyzed using the UPLC-Q-TOF MS/MS technique. Cyclophosphamide (CTX) was then administered intraperitoneally to rats to establish an immunosuppression model. Physiological and biochemical parameters, organ indices, and histopathological findings were evaluated for efficacy. Subsequently, potential biomarkers in rat serum were identified using multivariate statistical analysis and enriched and topologized using online platforms such as MetaboAnalyst and KEGG to reveal the critical metabolic pathways and their roles in the immunomodulatory network. Finally, the integrated analysis of components in vivo and in vitro, along with metabolic pathways, was performed using network pharmacology and molecular docking technology to elucidate the mechanisms of their roles in organismal immunity.Results: A total of 28 chemical components in vitro were identified, while pharmacodynamic experiments confirmed the immunomodulatory effects of Yantuo, especially in the high-dose administration group. Metabolomics analysis showed that 37 potential immune-related biomarkers were identified in positive and negative ion modes, involving 16 metabolic pathways such as arginine biosynthesis, pyrimidine metabolism, and riboflavin metabolism. The results of network pharmacology and molecular docking indicated that Yantuo may affect 7-O-galloyl-catechin, Cynaroside, Quercetin-7-O-beta-D-glucopyranoside, and 1.6-bis-O-galloyl-beta-D-glucose through interactions with the immune system, with significant pathways of action including galactose metabolism, glycolysis/gluconeogenesis, pyrimidine metabolism, and riboflavin metabolism.Conclusion: In our experiments, we confirmed the organismal modulatory effect of Yantuo on immunocompromised rats, clarified the key components, target proteins, and pathways of its possible action, and provided possibilities for follow-up studies. Keywords: immunosuppression, Yantuo, metabolomics, network pharmacology, molecular docking

Yule Wang,* Bei Li,* Yingjuan Zhang,* Ruiling Lu, Qianzhuo Wang, Yue Gao Zhejiang Key Laboratory of Traditional Chinese Medicine for the Prevention and Treatment of Senile Chronic Diseases, Department of Geriatrics, Affiliated Hangzhou First People’s Hospital, School of Medicine, Westlake University, Hangzhou, People’s Republic of China*These authors contributed equally to this workCorrespondence: Yue Gao, Zhejiang Key Laboratory of Traditional Chinese Medicine for the Prevention and Treatment of Senile Chronic Diseases, Department of Geriatrics, Affiliated Hangzhou First People’s Hospital, School of Medicine, Westlake University, 261 huansha Road, Shangcheng District, Hangzhou, 310006, People’s Republic of China, Email gaoyue@hospital.westlake.edu.cnBackground: Acute lung injury (ALI) is a life-threatening clinical syndrome with high mortality. Currently, the safe and effective therapies for ALI patients are still limited. Qingfei Huoxue decoction (QFHXD) is a hospital agreement prescription for treating pulmonary diseases and displays a remarkable efficacy. However, the pharmacological effect of QFHXD on preventing lipopolysaccharide (LPS)-induced ALI has yet to be reported, let alone questions of potential molecular mechanisms and anti-ALI active substances.Methods: To answer the above-mentioned questions, histopathological observation and kit detection were performed to estimate the protective effect of QFHXD pretreatment against LPS-induced ALI. Based on comprehensive chemical profiling of QFHXD, a network pharmacology strategy and experimental validation were integrated to elucidate the underlying functional mechanisms. The potential anti-ALI active components were identified by molecular docking. The anti-ALI activity of narirutin and its anti-inflammatory mechanism were further validated using animal and molecular experiments.Results: Pretreatment with different doses of QFHXD effectively mitigated histopathological lesions and systemic inflammation caused by LPS stimulation. A detailed analysis of established compound-target-disease network revealed the strong correlation between anti-ALI action of QFHXD and inflammatory mechanisms. Compared with the model group, QFHXD intervention markedly restrained the abnormally increased transcription and protein levels of pro-inflammatory factors (TLR4, NF-κB, IL-6, IL-1β, and TNF-α) in lung tissues of ALI mice. The results of molecular docking highlighted the anti-ALI potential of narirutin targeting to TLR4 and NF-κB p65. In addition to the protective effect of narirutin on suppressing LPS-induced pathological changes, we found that narirutin pretreatment effectively normalized the disordered protein levels of above pro-inflammatory factors of ALI mice.Conclusion: These interesting findings indicate the beneficial effects of QFHXD and its active component narirutin against ALI partly via regulating TLR4/NF-κB mediated inflammation. This work contributes to the development of novel medications for ALI patients. Keywords: Qingfei Huoxue decoction, narirutin, LPS-induced acute lung injury, network pharmacology, TLR4/NF-κB pathway, inflammatory response

Changhong Xu,1,&ast; Yun Deng,1,&ast; Jiangwei Man,1,&ast; Huabin Wang,1 Tuanjie Che,2 Liyun Ding,3 Li Yang1 1Department of Urology, Institute of Urology, Gansu Province Clinical Research Center for Urinary System Disease, Lanzhou University Second Hospital, Lanzhou, Gansu, 730030, People’s Republic of China; 2Innovation Center of Functional Genomics and Molecular Diagnostics Technology of Gansu Province, Lanzhou, People’s Republic of China; 3School of Physical Science and Technology, Lanzhou University, Lanzhou, 730000, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Li Yang, Department of Urology, Institute of Urology, Gansu Province Clinical Research Center for Urinary System Disease, Lanzhou University Second Hospital, Lanzhou, Gansu, 730030, People’s Republic of China, Email ery_yangli@lzu.edu.cnObjective: This study investigates the targets, pathways, and mechanisms of Schisandrin B (Sch B) in alleviating renal ischemia-reperfusion injury (RIRI) using RNA sequencing and network pharmacology.Methods: The effects of Sch B on RIRI were assessed using hematoxylin-eosin (HE) and periodic acid-Schiff (PAS) staining, along with measurements of blood creatinine and urea nitrogen (BUN). Differential gene expression in mouse models treated with RIRI and Sch B+RIRI was analyzed through RNA-Seq. Key processes, targets, and pathways were examined using network pharmacology techniques. The antioxidant capacity of Sch B was evaluated using assays for reactive oxygen species (ROS), mitochondrial superoxide, and JC-1 membrane potential. Molecular docking was employed to verify the interactions between key targets and Sch B, and the expression of these targets and pathway was confirmed using qRT-PCR, Western blot, and immunofluorescence.Results: Sch B pre-treatment significantly reduced renal pathological damage, inflammatory response, and apoptosis in a mouse RIRI model. Pathological damage scores dropped from 4.33 ± 0.33 in the I/R group to 2.17 ± 0.17 and 1.5 ± 0.22 in Sch B-treated groups (p < 0.01). Creatinine and BUN levels were also reduced (from 144.6 ± 21.05 μmol/L and 53.51 ± 2.34 mg/dL to 50.44 ± 5.61 μmol/L and 17.18 ± 0.96 mg/dL, p < 0.05). Transcriptomic analysis identified four key targets (AKT1, ALB, ACE, CCL5) and the PI3K/AKT pathway. Experimental validation confirmed Sch B modulated these targets, reducing apoptosis and oxidative stress, and enhancing renal recovery.Conclusion: Sch B reduces oxidative stress, inflammation, and apoptosis by modulating key targets such as AKT1, ALB, ACE, and CCL5, while activating the PI3K/AKT pathway, leading to improved renal recovery in RIRI.Keywords: renal ischemia-reperfusion injury, schisandrin B, transcriptomics, network pharmacology, PI3K/AKT pathway

Abstract Background Nomilin is a limonoid compound known for its multiple biological activities, but its role in triple negative breast cancer (TNBC) remains unclear. This study aims to uncover the potential therapeutic effect of nomilin on TNBC and elucidate the specific mechanism of its action. Methods We employed weighted gene co-expression network analysis (WGCNA), differential expression analysis, and the GeneCards database to identify potential targets for TNBC. Simultaneously, we utilized the Swiss Target Prediction, ChEMBL, and STITCH databases to identify potential targets of nomilin. The core targets and mechanisms of nomilin against TNBC were predicted through protein-protein interaction (PPI) network analysis, molecular docking, and enrichment analysis. The results of the network pharmacology were corroborated by conducting experiments. Results A total of 17,204 TNBC targets were screened, and 301 potential targets of nomilin were identified. Through the PPI network, eight core targets of nomilin against TNBC were pinpointed, namely BCL2, Caspase3, CyclinD1, EGFR, HSP90AA1, KRAS, PARP1, and TNF. Molecular docking, molecular dynamics simulation and proteome microarray revealed that nomilin exhibits strong binding activity to these core proteins. Enrichment analysis results indicated that the anti-TNBC effect of nomilin is associated with PI3K/Akt pathway. In vitro and in vivo experiments have demonstrated that nomilin inhibits TNBC cell proliferation and migration while promoting cell apoptosis through the PI3K/Akt pathway. Conclusion For the first time, the research effectively discovered the objectives and mechanisms of nomilin in combating TNBC using network pharmacology, molecular docking, molecular dynamics simulation, proteome microarray and experimental confirmation, presenting a hopeful approach for treating TNBC. Graphical Abstract

Xi Huang,1,2,&ast; Yu Peng,1,2,&ast; Lingfei Lu,1,&ast; Liwen Gao,1,2 Shanshan Wu,1,2 Jiandong Lu,1 Xinhui Liu1 1Department of Nephrology, Shenzhen Traditional Chinese Medicine Hospital, Guangzhou University of Chinese Medicine, Shenzhen, Guangdong, People’s Republic of China; 2The Fourth Clinical Medical College, Guangzhou University of Chinese Medicine, Shenzhen, Guangdong, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Xinhui Liu, Department of Nephrology, Shenzhen Traditional Chinese Medicine Hospital, Guangzhou University of Chinese Medicine, Shenzhen, Guangdong, 518033, People’s Republic of China, Email liuxinhui0317@163.comBackground: Huangqi-Danshen decoction (HDD) is a Chinese medicinal herb pair with good efficacy in treating chronic kidney disease, but its mechanism needs to be clarified.Aim: To uncover the underlying mechanism of HDD antagonizing renal fibrosis through network pharmacology (NP) analysis and experimental validation.Materials and Methods: The chemical components of water extract of HDD were analyzed by combining the ultra-high performance liquid chromatography coupled with Q-Exactive mass spectrum analysis (UHPLC-QE-MS) and HERB database. NP was used to identify core common targets of HDD components and renal fibrosis. Subsequently, male C57BL/6 mice were divided into Sham, unilateral ureteral obstruction (UUO) and UUO+HDD groups. Renal function, histopathology, Western blotting, and immunohistochemistry analyses were used to evaluate the protective effect of HDD on UUO mice. The effects of HDD on signaling pathways were validated in both UUO mice and transforming growth factor-β 1 (TGF-β 1)-induced HK-2 cells.Results: By combining UHPLC-QE-MS analysis and HERB database, 25 components were screened in HDD extract. There were 270 intersection targets of the 25 components and renal fibrosis. Based on their scores in protein-protein interaction analysis and degree values in component-pathway-target triadic network, 6 core common targets of the 25 components and renal fibrosis were identified, namely phosphoinositide 3-kinase (PI3K), signal transducer and activator of transcription 3 (Stat3), non-receptor tyrosine kinase Src (Src), epidermal growth factor receptor (EGFR), matrix metalloproteinase 9 (MMP9), and MMP2. HDD ameliorated renal tubular damage and collagen deposition and downregulated fibrosis-related proteins expression in UUO mice. Furthermore, HDD was demonstrated to reduce PI3K, Stat3, Src, EGFR, and MMP2 expressions, and enhance MMP9 expression in the kidney of UUO mice and in TGF-β 1-induced HK-2 cells.Conclusion: HDD can alleviate renal fibrosis which may be related to regulating the expression of essential proteins in the epithelial-mesenchymal transition and extracellular matrix production/degradation signaling pathways. Keywords: renal fibrosis, chronic kidney disease, Huangqi-Danshen decoction, network pharmacology, signaling pathway

Haoyang Xin,1,&ast; Man Wang,2,&ast; Shuo Wang,3 Haiyan Han,3 Nan Jiang1 1Department of Nephrology and Rheumatology, Second Affiliated Hospital of Tianjin University of TCM, Tianjin, People’s Republic of China; 2Department of Pharmacy, Tianjin Forth Central Hospital, Tianjin, People’s Republic of China; 3Graduate School, Tianjin University of Traditional Chinese Medicine, Tianjin, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Nan Jiang, Department of Nephrology and Rheumatology, Second Affiliated Hospital of Tianjin University of TCM, No. 69, Zengchan Street, Hebei District, Tianjin, 300250, People’s Republic of China, Tel +86 18622997255, Email jingnan_jn@126.comObjective: This study aimed to explore the potential active components and therapeutic targets of Shentong Zhuyu Decoction (SZD) in the treatment of ankylosing spondylitis (AS) through network pharmacology and animal experiments.Methods: Targets for AS and related pathways were obtained by network pharmacology, and the pathways with the best binding affinity in molecular docking were verified by animal experiments.Results: The network pharmacology analysis found 248 chemical components, 1068 drug targets and 803 AS-related targets in SZD. After intersection targets analysis, 116 common drug–disease targets were obtained, and matrix metalloproteinase-9, nucleotide-binding oligomerization domain (NOD)-like receptor thermal protein domain associated protein 3 (NLRP3) and cytochrome P450 2D6 were identified as the key targets of SZD for the treatment of AS. A total of 2152 biological processes, 38 cellular component expressions and 150 molecular function terms were obtained in gene ontology enrichment analysis, and 153 Kyoto Encyclopedia of Genes and Genomes (KEGG) signalling pathways were obtained in KEGG enrichment analysis. Molecular docking analysis showed that the absolute binding energies of glypallichalcone and NLRP3, quercetin and NLRP3 and kaempferol and NLRP3 were > 7. Animal experiments showed that the expressions of NLRP3, Caspase-1, interleukin (IL)-1β and IL-18 were significantly increased in the model group and the treatment group compared with those in the blank group, and the expression levels in the treatment group were significantly decreased compared with those in the model group (p < 0.05).Conclusion: The active components in SZD, such as baicalin, quercetin, kaempferol and glypallichalcone, may reduce the expression of IL-1β and IL-18 via the NLRP3/Caspase-1 signalling pathway to inhibit the development and progression of inflammation and play a role in the treatment of AS.Keywords: Shentong Zhuyu Decoction, ankylosing spondylitis, network pharmacology, mechanism of action

Chunxia Yin,1,&ast; Taohua Lan,2,3,&ast; Yunshan Wu,4,5 Jing Cai,1 Haoxiang Li,1 Xiaolan Kuang,1 Lin Jiao,1 Xiaomin Ou,1 Hua Yang,2 Bo Liu,3– 5 Weihui Lu2,6,7 1The Second Clinical Medical College, Guangzhou University of Chinese Medicine, Guangzhou, People’s Republic of China; 2Department of Cardiology, Guangdong Provincial Hospital of Chinese Medicine, Guangzhou, People’s Republic of China; 3State Key Laboratory of Dampness Syndrome of Chinese Medicine, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou, People’s Republic of China; 4Guangzhou Key Laboratory of Chirality Research on Active Components of Traditional Chinese Medicine, Guangdong Provincial Hospital of Chinese Medicine, Guangzhou, People’s Republic of China; 5Guangdong Provincial Key Laboratory of Clinical Research on Traditional Chinese Medicine Syndrome, Guangzhou University of Chinese Medicine, Guangzhou, People’s Republic of China; 6State Key Laboratory of Traditional Chinese Medicine Syndrome, Guangdong Provincial Hospital of Chinese Medicine, Guangzhou University of Chinese Medicine, Guangzhou, People’s Republic of China; 7Chinese Medicine Guangdong Laboratory, Guangdong Hengqin, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Weihui Lu, Bo Liu, The Second Clinical Medical College, Guangzhou University of Chinese Medicine, Email weihui.lu@gzucm.edu.cn; doctliu@gzucm.edu.cnPurpose: This study was designed to evaluate the effect and mechanism of the Qushi Huatan (QSHT) decoction against coronary heart disease (CHD) through network pharmacology and experimental verification.Methods: In the present study, the active ingredients of the QSHT decoction were identified by ultra performance liquid chromatography/tandem mass spectrometry (UPLC/MS), then the potential ingredients and coronary heart disease targets were predicted using the SwissTarget Prediction database and the database of Genecards and OMIM database, respectively. A herb-compound-target network was constructed using Cytoscape. GO and KEGG enrichment analysis were performed using the ClusterProfiler data package of R software. Molecular docking was used to predict the core targets of QSHT against CHD. In addition, we used a myocardial infarction (MI) and high-fat diet ApoE−/− mice model to investigate the cardioprotective effects of QSHT. Western blotting and immunochemistry were used to verify the core targets and the signaling pathway.Results: A total of 68 active ingredients were found in the QSHT decoction. Network pharmacology indicated 28 targets and 147 signal pathways, including AKT1, HIF-1α, GSK-3β, TLR4 and NF-κB, those key targets were also verified by molecular docking. The results of GO and KEGG enrichment analysis showed that the targets of QSHT against CHD were largely associated with inflammatory and oxidative stress, and AKT/HIF-1α and TLR4/NF-κB pathways might be key functional pathways. In vivo, QSHT significantly improved cardiac function and attenuated fibrosis and inflammation. Furthermore, QSHT could significantly inhibit the expression of HIF-1α, TLR4, phosphorylation of AKT1, GSK-3β and NF-κB after MI in ApoE−/− mice.Conclusion: Based on network pharmacology, molecular docking and experimental verification, this study demonstrated that QSHT could improve cardiac function and attenuate cardiac fibrosis by regulating TLR4/NF-κB and AKT/HIF-1α signaling pathway in post- MI and high-fat diet ApoE−/− mice.Keywords: network pharmacology, molecular docking, Qushi Huatan decoction, coronary heart disease

Context: Breast cancer therapy currently presents several uncomfortable side effects in patients, including effects on non-malignant tissues, recurrence, and resistance, which restrict their utilization. Consequently, researchers have directed their attention toward studying plant-derived anticancer compounds that exhibit high efficacy and safety profiles. Eugenol, a major component found in clove plants, demonstrates promising potential as a therapeutic agent for both estrogen receptor-positive and estrogen receptor-negative breast cancer. Aims: To predict the target of eugenol in estrogen receptor–positive breast cancer using network pharmacology and molecular docking analyses. Methods: Network pharmacology analysis was performed using the Chemical Toxigenomic Database, STITCH, GeneCards, Cytoscape, Enrichr, and Stringdb. Subsequently, molecular docking was performed using protein targets obtained from the RCSB-PDB and analyzed using AutoDock software. Results: Network pharmacology study and molecular docking revealed the anticancer effect of eugenol against breast cancer estrogen receptor–positive, especially in cancer and apoptotic pathways, by acting on caspase-3 (CASP3), epidermal growth factor receptor (EGFR), and poly [ADP-ribose] polymerase 1 (PARP1) signaling pathways. The docking results between the protein targets and eugenol showed that eugenol has the strongest binding with CASP3 (ligand binding energy: -5.78 kcal/mol), followed by eugenol binding with EGFR (ligand binding energy: -5.58 kcal/mol), and eugenol binding with PARP1 (ligand binding energy: -5.58 kcal/mol). Conclusions: Eugenol is a potential candidate for breast cancer therapy, especially for apoptosis mediated by CASP3 in breast cancer luminal A.

AbstractContext Sepsis-induced acute lung injury (ALI) is associated with high morbidity and mortality. Rhodiola rosea L. (Crassulaceae) (RR) and its extracts have shown anti-inflammatory, antioxidant, immunomodulatory, and lung-protective effects.Objective This study elucidates the molecular mechanisms of RR against sepsis-induced ALI.Materials and methods The pivotal targets of RR against sepsis-induced ALI and underlying mechanisms were revealed by network pharmacology and molecular docking. Human umbilical vein endothelial cells (HUVECs) were stimulated by 1 μg/mL lipopolysaccharide for 0.5 h and treated with 6.3, 12.5, 25, 50, 100, and 200 μg/mL RR for 24 h. Then, the lipopolysaccharide-stimulated HUVECs were subjected to cell counting kit-8 (CCK-8), enzyme-linked immunosorbent, apoptosis, and Western blot analyses. C57BL/6 mice were divided into sham, model, low-dose (40 mg/kg), mid-dose (80 mg/kg), and high-dose (160 mg/kg) RR groups. The mouse model was constructed through caecal ligation and puncture, and histological, apoptosis, and Western blot analyses were performed for further validation.Results We identified six hub targets (MPO, HRAS, PPARG, FGF2, JUN, and IL6), and the PI3K-AKT pathway was the core pathway. CCK-8 assays showed that RR promoted the viability of the lipopolysaccharide-stimulated HUVECs [median effective dose (ED50) = 18.98 μg/mL]. Furthermore, RR inhibited inflammation, oxidative stress, cell apoptosis, and PI3K-AKT activation in lipopolysaccharide-stimulated HUVECs and ALI mice, which was consistent with the network pharmacology results.Discussion and conclusion This study provides foundational knowledge of the effective components, potential targets, and molecular mechanisms of RR against ALI, which could be critical for developing targeted therapeutic strategies for sepsis-induced ALI.

AbstractContext Qinggong Shoutao Wan (QGSTW) is a pill used as a traditional medicine to treat age-associated memory decline (AAMI). However, its potential mechanisms are unclear.Objective This study elucidates the possible mechanisms of QGSTW in treating AAMI.Materials and methods Network pharmacology and molecular docking approaches were utilized to identify the potential pathway by which QGSTW alleviates AAMI. C57BL/6J mice were divided randomly into control, model, and QGSTW groups. A mouse model of AAMI was established by d-galactose, and the pathways that QGSTW acts on to ameliorate AAMI were determined by ELISA, immunofluorescence staining and Western blotting after treatment with d-gal (100 mg/kg) and QGSTW (20 mL/kg) for 12 weeks.Results Network pharmacology demonstrated that the targets of the active components were significantly enriched in the cAMP signaling pathway. AKT1, FOS, GRIN2B, and GRIN1 were the core target proteins. QGSTW treatment increased the discrimination index from −16.92 ± 7.06 to 23.88 ± 15.94% in the novel location test and from −19.54 ± 5.71 to 17.55 ± 6.73% in the novel object recognition test. ELISA showed that QGSTW could increase the levels of cAMP. Western blot analysis revealed that QGSTW could upregulate the expression of PKA, CREB, c-Fos, GluN1, GluA1, CaMKII-α, and SYN. Immunostaining revealed that the expression of SYN was decreased in the CA1 and DG.Discussion and conclusions This study not only provides new insights into the mechanism of QGSTW in the treatment of AAMI but also provides important information and new research ideas for the discovery of traditional Chinese medicine compounds that can treat AAMI.

Yujie Li,1 Yadong Yin,2 Juan Xiong,3 Zhipeng Zhang,4 Linglong Li,3 Baoshun Zhang,4 Feng Zhang,3 Dehong Mao3 1College of Traditional Chinese Medicine, Chongqing Medical University, Chongqing, People’s Republic of China; 2Xijing Hospital, Air Force Medical University, Xi’an, People’s Republic of China; 3Department of Otorhinolaryngology, Yongchuan Chinese Medicine Hospital Affiliated to Chongqing Medical University, Chongqing, People’s Republic of China; 4College of Pharmaceutical Sciences, Southwest University, Chongqing, People’s Republic of ChinaCorrespondence: Feng Zhang; Dehong Mao, Email 164382044@163.com; 19m001@hospital.cqmu.edu.cnBackground: Yujiang Paidu Decoction (YJPD) has demonstrated clinical efficacy in the treatment of chronic rhinosinusitis. However, the effects and mechanisms of the YJPD on chronic rhinosinusitis with nasal polyps (CRSwNP) remain unclear.Purpose: This study aimed to elucidate the potential mechanism of action of YJPD in the treatment of CRSwNP based on network pharmacology, transcriptomics and experiments.Methods: A CRSwNP mouse model was established using ovalbumin (OVA) and staphylococcus aureus enterotoxin B (SEB) for 12 weeks and the human nasal epithelial cell (HNEpC) model was induced with IL-13 in vitro. Behavioral tests, scanning electron microscopy (SEM), micro-CT and pathological change of nasal tissues were observed to investigate the therapeutic effects of YJPD. Network pharmacology and transcriptomics were launched to explore the pharmacological mechanisms of YJPD in CRSwNP treatment. Finally, an ELISA, immunofluorescence, RT-qPCR, Western blotting and Tunel were performed for validation.Results: Different doses of YJPD intervention effectively alleviated rubbing and sneezing symptoms in CRSwNP mice. Additionally, YJPD significantly reduced abnormal serological markers, structural damage of the nasal mucosa, inflammatory cell infiltration, goblet cell increases, and inhibited OVA-specific IgE levels and the secretion of Th2 cytokines such as IL-4, IL-5, and IL-13. Moreover, transcriptomics and network pharmacology analyses indicated that YJPD may exert anti-inflammatory and anti-apoptotic effects by inhibiting the MAPK/AP-1 signaling pathway. The experimental findings supported this conclusion, which was further corroborated by similar results observed in IL13-induced HNEpCs in vitro.Conclusion: YJPD could alleviate inflammatory status and epithelial apoptosis by inhibiting aberrant activation of MAPK/AP-1 signaling pathway. This finding provides a strong basis for using YJPD as a potential treatment in CRSwNP. Keywords: Yujiang paidu decoction, chronic rhinosinusitis with nasal polyps, network pharmacology, transcriptomics, inflammation

Jingjing Wei,1,&ast; Zhaoyang Liu,1,2,&ast; Mingming Li,1 Lingyun Du,1 Xia Zhu,1 Yi Leng,1 Changyu Han,1 Qingqing Xu,1 Chunhong Zhang1 1Department of Dermato-Venereology, the Second Hospital, Cheeloo College of Medicine, Shandong University, Jinan, People’s Republic of China; 2Department of Dermato-Venereology, Tianjin Medical University General Hospital, Tianjin, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Chunhong Zhang, Department of Dermato-Venereology, the Second Hospital, Cheeloo College of Medicine, Shandong University, 247 Beiyuan Dajie Street, Jinan, People’s Republic of China, Tel +86-531-85875027, Fax +86-531-88962544, Email aliu133@163.comBackground: Psoriasis is an immune-mediated chronic inflammatory disease. Qingre Lishi Decoction (QRLSD) has achieved great clinical effect in the treatment of psoriasis. However, the potential bioactive components and the mechanisms are yet unclear.Aim: To analyze the serum parameters of rats fed with QRLSD, screen out the active components of QRLSD, and explore the potential targets and pathway of QRLSD in the treatment of psoriasis.Materials and Methods: The active components of serum containing QRLSD were analyzed using ultra-high performance liquid chromatography quadrupole-time-of-flight mass spectrometry (UPLC-Q-TOF/MS). The targets of QRLSD in the treatment of psoriasis were predicted by network pharmacology and molecular docking. In vitro experiments verified the underlying mechanism.Results: By UPLC-Q-TOF/MS, 15 prototype components and 22 metabolites were identified in serum containing QRLSD. Subsequently, 260 chemical composition targets and 218 psoriasis targets were overlapped to obtain 23 intersection targets, including LGALS3, TNF, F10, DPP4, EGFR, MAPK14, STAT3 and others. TNF, IL-10, GAPDH, STAT3, EGFR, ITGB1, LGALS3 genes were identified as potential drug targets in the PPI network analyzed by CytoHubba. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses indicated that QRLSD may improve psoriasis by regulating immune and inflammatory pathways, the cytokine mediated signal transduction pathways and other signaling pathways. Molecular docking results showed that the main active components of the serum containing QRLSD had higher affinities for TNF and LGALS3. In vitro experiments confirmed that QRLSD may decrease levels of inflammatory cytokines by suppressing the NF-κB signaling pathway activated by TNF-α in human keratinocytes.Conclusion: This study explores the potential compounds, targets and signaling pathways of QRLSD in the treatment of psoriasis, which will help clarify the efficacy and mechanism of QRLSD.Keywords: qingre lishi decoction, psoriasis, UPLC-Q-TOF/MS, network pharmacology, molecular docking, NF-κB signaling pathway

Peijie Zhou,&ast; Shangshang Yu,&ast; Xuan Wang,&ast; Xiaofei Zhang, Dongyan Guo, Chongbo Zhao, Jiangxue Cheng, Jing Wang, Jing Sun Department of Pharmaceutics, College of Pharmacy, The Key Laboratory of Basic and New Drug Research of Traditional Chinese Medicine, Shaanxi Provincial Engineering Technology Research Center for Traditional Chinese Medicine Decoction Pieces, Shaanxi University of Chinese Medicine, Xianyang, Shaanxi, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Jing Sun, Email ph.175@163.comBackground: Cerebral ischaemia-reperfusion injury (CIRI) could worsen the inflammatory response and oxidative stress in brain tissue. According to previous studies, ferulic acid methyl ester (FAME), as the extract with the strongest comprehensive activity in the traditional Chinese medicine Huang Hua oil dot herb, has significant anti-oxidative stress and neuroprotective functions, and can effectively alleviate CIRI, but its mechanism of action is still unclear.Methods: Firstly, the pharmacological effects of FAME were investigated by in vitro oxidative stress and inflammatory experiments. Secondly, evaluate the therapeutic effects of FAME in the treatment of CIRI by brain histopathological staining and cerebral infarct area by replicating the in vivo MACO model. Thirdly, RNA-Seq and network pharmacology were utilized to predict the possible targets and mechanisms of FAME for CIRI at the molecular level. Finally, the expression of key target proteins, as well as the key regulatory relationships were verified by molecular docking visualization, Western Blotting and immunohistochemistry.Results: The results of in vitro experiments concluded that FAME could significantly reduce the content of TNF-α, IL-1β and ROS, inhibiting COX-2 and iNOS protein expression in cells(p< 0.01). FAME was demonstrated to have anti-oxidative stress and anti-inflammatory effects. The results of in vivo experiments showed that after the administration of FAME, the area of cerebral infarction in rats with CIRI was reduced, the content of Bcl-2 and VEGF was increased(p< 0.05). Network pharmacology and RNA-Seq showed that the alleviation of CIRI by FAME may be through PI3K-AKT and HIF-1 signaling pathway. Enhanced expression of HIF-1α, VEGF, p-PI3K, p-AKT proteins in the brain tissues of rats in the FAME group was verified by molecular docking and Western Blotting.Conclusion: FAME possesses significant anti-inflammatory and anti-oxidative stress activities and alleviates CIRI through the PI3K/HIF-1α/VEGF signaling pathway. Keywords: ferulic acid methyl ester, cerebral ischemia-reperfusion injury, WGCNA, RNA-Seq, network pharmacology, molecular docking techniques

Yun Zhong,1– 3 Yufei Zhao,4 Xin Meng,2 Fan Wang,2 Lei Zhou1 1Department of Dermatology, the Third Affiliated Hospital, Sun Yat-Sen University, Guangzhou, People’s Republic of China; 2Department of Dermatology, Xiangya Hospital, Central South University, Changsha, People’s Republic of China; 3Department of Dermatology, Nanfang Hospital, Southern Medical University, Guangzhou, People’s Republic of China; 4Department of Gastrointestinal Surgery, Laboratory of Surgery, the Second Affiliated Hospital of Guangzhou Medical University, Guangzhou, People’s Republic of ChinaCorrespondence: Lei Zhou, Department of Dermatology, the Third Affiliated Hospital, Sun Yat-sen University, 600 Tianhe Road, Tianhe District, Guangzhou, Guangdong, 510630, People’s Republic of China, Tel +020-85253333, Fax +020-85253336, Email zhoulei53@mail.sysu.edu.cnBackground: Rosacea, a recurring dermatological disorder, demands effective therapeutic approaches. Traditional Chinese medicine, particularly Liangxue Siwu Decoction (LXSWD), has shown promise in managing inflammatory skin diseases, such as rosacea. This study focuses on uncovering LXSWD’s specific effects on the inflammatory symptoms of rosacea.Objective: Our research investigates LXSWD’s therapeutic effectiveness in rosacea treatment and delves into its underlying mechanisms.Methods: Network pharmacology was utilized to identify LXSWD’s key components and their targets in rosacea management, which were then validated by molecular docking. An in vivo rosacea-like model in LL-37-induced mice was developed, subdividing them into control, model, and LXSWD groups. The LXSWD group received oral administration (25.0 g/kg/day) for six days before model induction. Post-treatment evaluations included skin tissue analyses to verify our network pharmacology predictions.Results: Key active ingredients in LXSWD, such as quercetin, kaempferol, and luteolin, were identified alongside central target proteins like TNF and MMPs. Our molecular docking study confirmed the interactions between these ingredients and targets. Analyses through GO and KEGG pathways indicated LXSWD’s role in mitigating inflammation, particularly influencing the TNF and IL-17 pathways. LXSWD treatment in vivo markedly alleviated LL-37-induced symptoms in mice, showing a marked reduction in inflammatory cytokines (p < 0.05) and modulation of crucial genes (p < 0.05). These results, supported by immunofluorescence analysis and Western blot, underline the modulatory effects of LXSWD on MMPs, offering significant protection against rosacea’s inflammation alterations (p < 0.05).Conclusion: Integrating network pharmacology, molecular docking, and in vivo experiments, this study elucidates LXSWD’s potential mechanisms in rosacea treatment. It offers a novel theoretical framework for its clinical use in managing rosacea.Keywords: Liangxue siwu decoction, molecular docking, network pharmacology, rosacea, skin inflammation

Qingyang Liu,1,&ast; Jian Wang,2,&ast; Chunhui Ding,3 Ying Chu,1 Fengying Jiang,1 Yunxia Hu,4 Haifeng Li,2 Qiubo Wang1 1Department of Clinical Laboratory, Wuxi Ninth People’s Hospital Affiliated to Soochow University, Wuxi, Jiangsu, 214000, People’s Republic of China; 2Department of Orthopedics, Wuxi Ninth People’s Hospital Affiliated to Soochow University, Wuxi, Jiangsu, 214000, People’s Republic of China; 3Department of Pharmacy, Wuxi Ninth People’s Hospital Affiliated to Soochow University, Wuxi, Jiangsu, 214000, People’s Republic of China; 4Department of Rheumatology and Immunology, The Affiliated Wuxi People’s Hospital of Nanjing Medical University, Wuxi, Jiangsu, 214000, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Qiubo Wang, Department of Clinical Laboratory, Wuxi Ninth People’s Hospital Affiliated to Soochow University, No. 999 Liang Xi Road, Binhu District, Wuxi, Jiangsu, 214000, People’s Republic of China, Email wangqiubo2020@suda.edu.cn Haifeng Li, Department of orthopedics, Wuxi Ninth People’s Hospital Affiliated to Soochow University, No. 999 Liang Xi Road, Binhu District, Wuxi, Jiangsu, 214000, People’s Republic of China, Email lihaifeng2008@suda.edu.cnPurpose: Sinomenine (SIN) is commonly used in Traditional Chinese Medicine (TCM) as a respected remedy for rheumatoid arthritis (RA). Nevertheless, the therapeutic mechanism of SIN in RA remains incompletely understood. This study aimed to delve into the molecular mechanism of SIN in the treatment of RA.Methods: The potential targets of SIN were predicted using the TCMSP server, STITCH database, and SwissTarget Prediction. Differentially expressed genes (DEGs) in RA were obtained from the GEO database. Enrichment analyses and molecular docking were conducted to explore the potential mechanism of SIN in the treatment of RA. In vitro and in vivo studies were conducted to validate the intervention effects of SIN on rheumatoid arthritis, as determined through network pharmacology analyses.Results: A total of 39 potential targets associated with the therapeutic effects of SIN in RA were identified. Enrichment analysis revealed that these potential targets are primarily enriched in PI3K-Akt signaling pathway, and the molecular docking suggests that SIN may act on specific proteins in the pathway. Experimental results have shown that exposure to SIN inhibits cytokine secretion, promotes apoptosis, reduces metastasis and invasion, and blocks the activation of the PI3K-Akt signaling pathway in RA fibroblast-like synoviocytes (RA-FLS). Moreover, SIN treatment alleviated arthritis-related symptoms and regulated the differentiation of CD4+ T cells in the spleen of collagen-induced arthritis (CIA) mice.Conclusion: By utilizing network pharmacology, molecular modeling, and in vitro/in vivo validation, this study demonstrates that SIN can alleviate RA by inhibiting the PI3K-Akt signaling pathway. These findings enhance the understanding of the therapeutic mechanisms of SIN in RA, offering a stronger theoretical foundation for its future clinical application. Keywords: sinomenine, rheumatoid arthritis, network pharmacology, PI3K-Akt signaling pathway

Jia Song,1 Jiancheng Zhang,2 Jun Shi,2 Xuming Pan,2 Dayu Mo3 1Department of General Practice, The Second Affiliated Hospital of Zhejiang Chinese Medical University, Hangzhou, Zhejiang, People’s Republic of China; 2Department of Emergency, The Second Affiliated Hospital of Zhejiang Chinese Medical University, Hangzhou, Zhejiang, People’s Republic of China; 3Department of Education, The Second Affiliated Hospital of Zhejiang Chinese Medical University, Hangzhou, Zhejiang, People’s Republic of ChinaCorrespondence: Dayu Mo, Department of Education, The Second Affiliated Hospital of Zhejiang Chinese Medical University, No. 318 Chaowang Road, Hangzhou, Zhejiang, 310005, People’s Republic of China, Tel +86-13857197190, Email modayu@zcmu.edu.cnBackground: Breviscapine has been demonstrated to have beneficial effects in ameliorating acute lung injury (ALI), yet its potential therapeutic value and molecular mechanisms in sepsis-induced ALI remain unexplored.Methods: We utilized network pharmacology approach to identify the potential targets and mechanisms of breviscapine in treating sepsis-induced ALI. To construct a murine model of sepsis, we performed cecal ligation and puncture (CLP). Hematoxylin and eosin (HE) staining and enzyme-linked immunosorbent assay (ELISA) were employed to respectively determine the pathologic changes and levels of inflammatory factors. Neutrophil count and total protein level in bronchoalveolar lavage fluid (BALF) were detected by corresponding kit. Additionally, we utilized flow cytometry, immunofluorescence, Western blotting, and real-time reverse transcription PCR (qRT-PCR) to detect cell apoptosis, protein expression, and gene expression. Finally, we used ELISA kits to detect the activity of myeloperoxidase (MPO) and caspase-8 (CASP8).Results: Breviscapine was revealed to target 81 potential proteins in the treatment of sepsis-induced ALI, while CASP8 was the most important one as demonstrated by network analysis. In vivo experiments demonstrated that breviscapine effectively reduced the severity of sepsis-induced ALI and inflammation, and significantly suppressed neutrophil infiltration in the lung tissues of CLP mice and promoted neutrophil apoptosis in the peripheral blood. In vitro experiments revealed that lipopolysaccharide (LPS)-induced neutrophil apoptosis was inhibited, and the expression and activity of CASP8 were down-regulated. Breviscapine intervention markedly up-regulated the expression and activity of CASP8, consequently activating neutrophil apoptosis and inhibiting inflammatory response by activating the NF-κB signaling pathway.Conclusion: Breviscapine is remarkably effective in improving sepsis-induced ALI, and its mechanism of action may be to induce neutrophil apoptosis, inhibit inflammatory overreaction and reduce its infiltration in pulmonary tissues by up-regulating the expression and activity of CASP8.Keywords: Breviscapine, acute lung injury, sepsis, network pharmacology, neutrophil

Rui Shen,1,2,&ast; Kebin Cheng,3,&ast; Guanyi Li,1 Zhendong Pan,4 Xijier Qiaolongbatu,1 Yuting Wang,1 Cui Ma,1 Xucong Huang,1 Li Wang,1 Wenjing Li,1 Yuanyuan Wang,2 Lili Jing,1 Guorong Fan,2 Zhenghua Wu2 1School of Pharmaceutical Sciences, Shanghai Jiao Tong University, Shanghai, 200240, People’s Republic of China; 2Department of Clinical Pharmacy, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200080, People’s Republic of China; 3Department of Respiratory and Critical Care Medicine, Shanghai Pulmonary Hospital, School of Medicine, Tongji University, Shanghai, 200433, People’s Republic of China; 4Department of Clinical Pharmacy, Eye and ENT Hospital, Fudan University, Shanghai, 200031, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Zhenghua Wu, Department of Clinical Pharmacy, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, No. 85 Wujin Road, Shanghai, 200080, People’s Republic of China, Tel +86-159-0161-7923, Email wuzhenghua@sjtu.edu.cnPurpose: Alisma orientale (AO, Alisma orientale (Sam). Juzep) has been widely employed for the treatment of macular edema (ME) in traditional Chinese medicine due to its renowned water-relief properties. Nonetheless, the comprehensive investigation of AO in alleviating ME remained unexplored. This study aims to identify the active components of AO that target the eye and investigate its pharmacological effects and mechanisms on ME.Methods: The study commenced with UPLC-Triple-TOF/MS analysis to identify the primary constituents of AO. Zebrafish eye tissues were then analyzed after a five-day administration of AO to detect absorbed components and metabolites. Subsequently, network pharmacology, molecular docking, and molecular dynamics simulations were employed to predict the mechanisms of ME treatment via biological target pathways. In vivo experiments were conducted to corroborate the pharmacological actions and mechanisms.Results: A total of 7 compounds, consisting of 2 prototype ingredients and 5 metabolites (including isomers), were found to traverse the blood-eye barrier and localized within eye tissues. Network pharmacology results showed that AO played a role in the treatment of ME mainly by regulating the pathway network of PI3K-AKT and MAPK with TNF-α centered. Computational analyses suggested that 11-dehydro-16-oxo-24-deoxy-alisol A, a metabolite of alisol A, mitigates edema through TNF-α inhibition. Furthermore, zebrafish fundus confocal experiments and HE staining of eyes confirmed the attenuating effects of alisol A on fundus angiogenesis and ocular edema, representing the first report of AO’s ME-inhibitory effects.Conclusion: In this study, computational analyses with experimental validation were used to understand the biological activity and mechanism of alisol A in the treatment of ME. The findings shed light on the bioactive constituents and pharmacological actions of AO, offering valuable insights and a theoretical foundation for its clinical application in managing ME. Keywords: Alisma orientale, macular edema, network pharmacology, zebrafish, TNF-&#x03B1

Xuan Wang,1,&ast; Hanjie Yu,2,&ast; Yunheng Zhang,3,&ast; Xin Chang,1 Chengyi Liu,1 Xiaodong Wen,1 Feng Tian,1 Yi Li1 1Department of Foot and Ankle Surgery, Honghui Hospital, Xi’an Jiaotong University, Xi’an, Shaanxi, People’s Republic of China; 2Laboratory for Functional Glycomics, College of Life Sciences, Northwest University, Xi’an, Shaanxi, People’s Republic of China; 3The Second Clinical Medical College, Shaanxi University of Chinese Medicine, Xianyang, Shaanxi, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Yi Li, Email liyidoctor@163.comObjective: Osteoarthritis (OA) is a common degenerative disease worldwide. While curcumin has shown therapeutic effects on OA, its mechanism remains unknown. This study aimed to investigate the molecular mechanism of curcumin in treating OA through network pharmacology and both in vivo and in vitro experiments.Methods: Curcumin-related targets were obtained using the HERB and DrugBank databases. GeneCards and DisGeNET were used to build a target database for OA. The STRING database was employed to construct protein-protein interaction networks and analyze related protein interactions. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway and gene ontology enrichment analyses of core targets were performed using Metascape. In addition, Autodock software was utilized for molecular docking validation of curcumin and disease targets. Further validation of the main findings was conducted through in vitro and in vivo experiments. In the in vitro experiments, an inflammation model was constructed through nitric oxide donor (SNP) stimulation of chondrocytes. Subsequently, the regulatory effects of curcumin on core targets and signaling pathways were validated using Western blotting and immunofluorescence staining techniques. In the in vivo experiments, an OA model was established by performing medial meniscectomy on male Sprague–Dawley rats. The therapeutic effects were evaluated using enzyme-linked immunosorbent assays, histologic staining, and micro-computed tomography (micro-CT) techniques.Results: Core targets of curcumin relevant to OA therapy included tumor necrosis factor-alpha (TNF-α), interleukin (IL)-1β, IL-6, matrix metalloproteinase 9 (MMP-9), B-cell lymphoma 2 (BCL-2), and caspase-3. The major biological processes involved oxidative stress and apoptotic processes, among others. The p38 mitogen-activated protein kinase (p38/MAPK) pathway was identified as the most likely pathway involved. In vitro experiments showed that curcumin significantly reduced oxidative stress levels, inhibited the expression of inflammatory factors IL-6 and Cyclooxygenase-2 (COX-2) and downregulated the expression of MMP-9 and MMP-1. In addition, curcumin was found to regulate the expression of BCL-2 and caspase-3 through the p38/MAPK pathway, inhibiting chondrocyte apoptosis. In vivo animal experiments demonstrated that curcumin significantly reduced the expression of OA-related factors (IL-1, IL-6, and TNF-α). Histological analysis and micro-CT results revealed that curcumin treatment significantly increased cartilage thickness, improved cartilage morphology, structure, and function, inhibited cartilage degradation, and enhanced the resorption of subchondral bone in the knee joints of rats with OA.Conclusion: Curcumin regulates oxidative stress and maintains mitochondrial function, thereby protecting chondrocyte guard. In addition, curcumin attenuates the inflammatory response of chondrocytes by inhibiting the phosphorylation of P38MAPK, slowing down the breakdown of the extrachondral matrix while preventing apoptosis of chondrocytes. Additionally curcumin attenuated cartilage degradation and bone damage while helping to boost bone density.Keywords: osteoarthritis, curcumin, molecular mechanism, network pharmacology, p38/MAPK

Yanan Li,1,2,&ast; Jingyi Hu,1,2,&ast; Ryan Au,1– 3 Cheng Cheng,1,2 Feng Xu,1,2 Weiyang Li,1,2 Yuguang Wu,1,2 Yuan Cui,1,2 Lei Zhu,1,2 Hong Shen1,2 1Affiliated Hospital of Nanjing University of Chinese Medicine, Nanjing, 210029, People’s Republic of China; 2Jiangsu Provincial Hospital of Chinese Medicine, Nanjing, 210029, People’s Republic of China; 3Academy of Chinese Culture and Health Sciences, Oakland, CA, 94612, USA&ast;These authors contributed equally to this workCorrespondence: Hong Shen; Lei Zhu, Jiangsu Provincial Hospital of Chinese Medicine (Affiliated Hospital of Nanjing University of Chinese Medicine), 155 Hanzhong Road, Qinhuai District, Nanjing, Jiangsu Province, People’s Republic of China, Tel\Fax +86-025- 86617141, Email shenhong999@njucm.edu.cn; zhulei5100@njucm.edu.cnBackground: Qingchang Tongluo Decoction (QTF) is clinically used for the treatment of intestinal fibrosis in Crohn’s Disease (CD). However, the role of QTF in CD-associated fibrosis and its potential pharmacological mechanism remains unclear.Purpose: The objective of this study was to elucidate the potential mechanism of QTF in treating CD-associated fibrosis, employing a combination of bioinformatics approaches — encompassing network pharmacology and molecular docking — complemented by experimental validation.Methods: To investigate the material basis and potential protective mechanism of QTF, a network pharmacology analysis was conducted. The core components and targets of QTF underwent molecular docking analysis to corroborate the findings obtained from network pharmacology. In vitro, a colon fibrotic model was established by stimulating IEC-6 cells with 10 ng/mL of transforming growth factor(TGF-β 1). In vivo, an intestinal fibrosis model was induced in BALB/c mice by TNBS. The role of QTF in inhibiting the TGF-β 1/Smad signaling pathway was investigated through RT-qPCR, Western blotting, immunohistochemistry staining, and immunofluorescence staining.Results: Network pharmacology analysis revealed that QTF could exert its protective effect. Bioinformatics analysis suggested that Flavone and Isoflavone might be the key components of the study. Additionally, AKT1, IL-6, TNF, and VEGFA were identified as potential therapeutic targets. Furthermore, experimental validation and molecular docking were employed to corroborate the results obtained from network pharmacology. RT-qPCR, Immunofluorescence, and Western blotting results demonstrated that QTF significantly improved colon function and inhibited pathological intestinal fibrosis in vivo and in vitro.Conclusion: Through the application of network pharmacology, molecular docking, and experimental validation, QTF could be confirmed to inhibit the proliferation of intestinal fibroblasts associated with CD and reduce the expression of Collagen I and VEGFA. This effect is achieved through the attenuation of ECM accumulation, primarily via the inhibition of the TGF-β 1/Smad signaling pathway. Keywords: Qingchang Tongluo Decoction, Crohn’s Disease, intestinal fibrosis, network pharmacology, Pharmacology

Banglan Cai,1,2,&ast; Manman Qi,2,3,&ast; Xue Zhang,1,2 Denghai Zhang1– 3 1School of Basic Medicine, Ningxia Medical University, Yinchuan, People’s Republic of China; 2Shanghai Health Commission Key Laboratory of Artificial Intelligence (AI)-Based Management of Inflammation and Chronic Diseases, Shanghai Pudong Gongli Hospital, Shanghai, People’s Republic of China; 3School of Medicine, Shanghai University, Shanghai, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Denghai Zhang; Xue Zhang, Shanghai Health Commission Key Laboratory of Artificial Intelligence (AI)-Based Management of Inflammation and Chronic Diseases, Shanghai Pudong Gongli Hospital, Shanghai, 200135, People’s Republic of China, Tel +86-18916173857 ; +86-13524805404, Email Denghai_zhang@163.com; 2021021020@nxmu.edu.cnBackground: As a traditional Chinese medicine monomer derived from Tripterygium wilfordii Hook.f. with potential anticancer activity, celastrol can induce ferroptosis in hepatic stellate cells and inhibit their activation to alleviate liver fibrosis. Activation of ferroptosis can effectively inhibit Hepatocellular carcinoma (HCC). Whether celastrol inhibits HCC by inducing ferroptosis remains to be studied.Purpose: To explore the potential targets of celastrol against HCC through ferroptosis based on network pharmacology and to verify the anticancer effect of celastrol on HepG2 cells.Methods: We collected celastrol targets, HCC, and ferroptosis-related genes through online databases, and got their intersection targets. Subsequently, we obtained a protein-protein interaction (PPI) network, and performed gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis to gain key genes for further study. They were verified in vitro and were performed molecular docking. The changes in cell proliferation and ferroptosis characteristics of HepG2 cells after celastrol treatment were detected.Results: 31 core target genes were screened for PPI network and enrichment analysis. The most significantly related KEGG pathway was chemical carcinogenesis-reactive oxygen species. The mRNA and protein levels of GSTM1 were significantly decreased after celastrol treatment. Molecular docking demonstrated the interaction between celastrol and GSTM1. Ferroptosis was induced and cell proliferation was inhibited by celastrol in HCC cells.Conclusion: Celastrol induces ferroptosis in HCC via regulating GSTM1 expression and may serve as a novel therapeutic compound with clinical potential in HCC treatment. Keywords: network pharmacology, celastrol, hepatocellular carcinoma, ferroptosis, reactive oxygen species

Kaiyue Zhang, Chunnan Li, Peitong Wu, Xiaochen Gao, Xueqin Feng, Jiaming Shen, Nanxi Zhang, Xuesheng Hu, Shuo Wang, Hui Zhang, Jingwei Lv, Jiaming Sun Jilin Ginseng Academy, Changchun University of Chinese Medicine, Changchun, Jilin, People’s Republic of ChinaCorrespondence: Jiaming Sun; Jingwei Lv, Changchun University of Chinese Medicine, No. 1035, Boshuo Road, Nanguan District, Changchun, 130117, People’s Republic of China, Tel +0431-86763809, Fax + 8676 3968, Email Sun_jiaming2000@163.com; jingwei-lv@hotmail.comPurpose: Zhixiao Tang (ZXT), a traditional Chinese compound prescription, has been used clinically to treat pneumonia in China. However, the underlying mechanism of ZXT treatment in pneumonia is still unclear. The present study aimed to reveal the potential mechanism of ZXT in pneumonia using a strategy combining metabolomics and network pharmacology.Methods: Initially, the chemical compositions were identified by UPLC-QE-Orbitrap-MS, while the prediction of potential signal pathways was performed through network pharmacology. To assess the anti-inflammatory properties of ZXT in the context of pneumonia, models of 16HBE cells induced by LPS and zebrafish induced by CuSO4 were established to measure levels of inflammatory markers and apoptosis. Subsequently, the differential changes of endogenous metabolites in cells caused by ZXT were examined using metabolomics technology, and the molecular docking analysis of key targets was carried out using Autodock Vina software. Ultimately, the validation of the primary pathways and targets was conducted through quantitative RT-PCR and Western blot techniques.Results: A total of 75 compounds were identified through UPLC-QE-Orbitrap-MS analyses. Network pharmacological analysis shows that it plays an anti-inflammatory role in C-type lectin receptor signaling pathway. After ZXT intervention, the inflammatory factors and apoptosis in cells were significantly reduced. Metabonomics analysis showed that 18 metabolites changed significantly. Four key genes were identified, which exhibited partial compatibility with the findings of network pharmacology. Molecular docking analysis confirmed the substantial affinity of the primary targets for ZXT. Furthermore, ZXT exerted a suppressive effect on neutrophil migration, down-regulated the expression of pro-inflammatory cytokine genes, and inhibited the up-regulation of the Dectin-1/SYK/NF-κB signaling pathway. In vivo cell experiments also yielded consistent experimental outcomes.Conclusion: This study enhances comprehension of the pharmacological mechanism underlying ZXT’s efficacy in pneumonia treatment, thereby establishing a scholarly basis for future research and clinical utilization of ZXT in pneumonia management. Keywords: Zhixiao Tang, pneumonia, network pharmacology, integrated metabolomics, 16HBE, Zebrafish

Benchi Cai,&ast; Qitong Wang,&ast; Lifan Zhong, Fang Liu, Xinyu Wang, Tao Chen Hainan Affiliated Hospital of Hainan Medical University, Haikou, Hainan, 570311, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Tao Chen, Email chentao@hnyxy15.wecom.workBackground: Parkinson’s disease (PD) is the most prevalent movement disorder. Curcumin, a polyphenol with hydrophobic properties, has been proved against Parkinson. Our previous study suggested that curcumin’s effectiveness in treating Parkinson’s disease may be linked to the gut-brain axis, although the specific mechanism by which curcumin exerts neuroprotective effects in the brain remains unknown.Methods: The therapeutic efficacy of curcumin was evaluated using behavioral tests, immunofluorescence of tyrosine hydroxylase (TH). Network pharmacology and transcriptomics predicted the mechanisms of curcumin in PD. Activation of the phosphatidylinositol 3-kinase PI3K/AKT pathway was confirmed by quantitative polymerase chain reaction (qPCR) and immunofluorescence.Results: Curcumin restored the dyskinesia and dopaminergic neurons damage of MPTP-induced mice. Curcumin against Parkinson’s disease by regulating inflammation, oxidative stress, and aging. The mechanisms of these were associated with activation of PI3K / AKT pathway.Conclusion: In conclusion, the neuroprotective mechanisms of curcumin activate PI3K / AKT pathway in Parkinson’s disease was revealed by our study.Keywords: Parkinson’s disease, curcumin, network pharmacology, transcriptomics, PI3K / AKT pathway

Jilei Li,1,&ast; Wenchao Dan,2,&ast; Chenchen Zhang,3 Nian Liu,3 Yichong Wang,1 Jixiang Liu,3 Shengsheng Zhang1 1Digestive Disease Center, Beijing Hospital of Traditional Chinese Medicine, Capital Medical University, Beijing, 100010, People’s Republic of China; 2Department of Dermatology, Beijing Hospital of Traditional Chinese Medicine, Capital Medical University, Beijing, 100010, People’s Republic of China; 3Beijing University of Chinese Medicine, Beijing, 100029, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Shengsheng Zhang, Digestive Disease Center, Beijing Hospital of Traditional Chinese Medicine, Capital Medical University, Beijing, 100010, Tel +8613801088329, Email zhangshengsheng@bjzhongyi.comPurpose: This study aimed to delineate the molecular processes underlying the therapeutic effects of berberine on UC by employing network pharmacology tactics, molecular docking, and dynamic simulations supported by empirical validations both in vivo and in vitro.Patients and Methods: We systematically screened potential targets and relevant pathways affected by berberine for UC treatment from comprehensive databases, including GeneCards, DisGeNET, and GEO. Molecular docking and simulation protocols were used to assess the interaction stability between berberine and its principal targets. The predictions were validated using both a DSS-induced UC mouse model and a lipopolysaccharide (LPS)-stimulated NCM460 cellular inflammation model.Results: Network pharmacology analysis revealed the regulatory effect of the TLR4/NF-κB/HIF-1α pathway in the ameliorative action of berberine in UC. Docking and simulation studies predicted the high-affinity interactions of berberine with pivotal targets: TLR4, NF-κB, HIF-1α, and the HIF inhibitor KC7F2. Moreover, in vivo analyses demonstrated that berberine attenuates clinical severity, as reflected by decreased disease activity index (DAI) scores, reduced weight loss, and mitigated intestinal inflammation in DSS-challenged mice. These outcomes include suppression of the proinflammatory cytokines IL-6 and TNF-α and downregulation of TLR4/NF-κB/HIF-1α mRNA and protein levels. Correspondingly, in vitro findings indicate that berberine decreases cellular inflammatory injury and suppresses TLR4/NF-κB/HIF-1α signaling, with notable effectiveness similar to that of the HIF-1α inhibitor KC7F2.Conclusion: Through network pharmacology analysis and experimental substantiation, this study confirmed that berberine enhances UC treatment outcomes by inhibiting the TLR4/NF-κB/HIF-1α axis, thereby mitigating inflammatory reactions and improving colonic pathology. Keywords: berberine, ulcerative colitis, TLR4/NF-κB/HIF-1α, network pharmacology, molecular docking, experimental validation, inhibitor

Ziyu Li,1,&ast; Qiuyan Zhang,2,&ast; Yuhe Gao,3,&ast; Fang Wan,3 Yincang Wang,3 Bin Hou,2 Wenwen Cui,4,5 Yanan Wang,2 Wei Feng,2 Yunlong Hou4,5 1Suzhou TCM Hospital Affiliated to Nanjing University of Chinese Medicine, Suzhou, People’s Republic of China; 2New Drug Evaluation Center, Shijiazhuang Yiling Pharmaceutical Co., Ltd, Shijiazhuang, People’s Republic of China; 3Graduate School, Heilongjiang University of Chinese Medicine, Harbin, People’s Republic of China; 4National Key Laboratory for Innovation and Transformation of Luobing Theory, Shijiazhuang Yiling Pharmaceutical Co., Ltd, Shijiazhuang, People’s Republic of China; 5Key Laboratory of State Administration of TCM (Cardio-Cerebral VesselCollateral Disease), Shijiazhuang, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Yunlong Hou, Email houyunlonghrb@Hotmail.comBackground: The LuoBiTong (LBT) capsule, a novel traditional Chinese medicine formulation, is currently in Phase III clinical trials. Preliminary preclinical and Phase II clinical studies suggest its efficacy and safety in treating rheumatoid arthritis (RA). However, the underlying mechanisms of its action remain to be elucidated.This research aims to explore the effects and mechanisms of LBT in conjunction with a maintenance dose of methotrexate (M-MTX) on RA.Methods: A Collagen-Induced Arthritis (CIA) mouse model was used to evaluate the anti-RA effects of LBT combined with M-MTX. Assessments included foot swelling, arthritis scoring, serum inflammatory factor analysis, and histopathological examination of the foot. These effects were compared with those of high-dose MTX (H-MTX). Network pharmacology was employed to construct a compound-target network for RA, based on drug composition, to predict its potential mechanism of action. Flow cytometry, Western Blot, and immunohistochemical analyses in animal models identified multiple inflammatory pathways targeted by LBT to augment the anti-RA effects of MTX.Results: The study revealed that LBT combined with M-MTX significantly alleviated CIA-induced arthritis without adverse effects. The combination of LBT and M-MTX showed similar or superior efficacy in regulating macrophage polarization, NF-κB, MAPK signaling pathways, and in the suppression of TH-17 expression in proinflammatory cells. These findings suggest that LBT may exert a multi-pathway therapeutic effect in RA treatment. The predicted pharmacological targets and mechanisms align well with this hypothesis.Conclusion: LBT, when combined with MTX, enhances the anti-RA effect by targeting multiple inflammatory pathways, demonstrating significant therapeutic potential.Keywords: rheumatoid arthritis, traditional Chinese medicine, methotrexate, network pharmacology

Lina Du,1 Dezhi Yang,2 Lan Wu,1 Li Mei,1 Sarula Wu,1 Yasula Ba,1 Yongchang Bao,1 Rigugaqiqige Su,1 Lin Song1 1College of Mongolian Medical, Inner Mongolia Medical University, Hohhot, Inner Mongolia, People’s Republic of China; 2Innovative Mongolian Medical Engineering Research Center, Inner Mongolia International Mongolian Hospital, Hohhot, Inner Mongolia, People’s Republic of ChinaCorrespondence: Lin Song, Inner Mongolia Medical University, Tel +15849163188, Email songlinwpsword@163.comObjective: To explored the potential molecular mechanism of Sugemule-4 decoction (MMS-4D) in treating insomnia.Methods: DL-4-chlorophenylalanine (PCPA) + chronic unpredictable mild stress stimulation (CUMS) was used to induce an insomnia model in rats. After the model was successfully established, MMS-4D was intervened at low, medium, and high doses for 7 days. The open-field test (OFT) was used to preliminarily evaluate the efficacy. The potential mechanism of MMS-4D in treating insomnia was investigated using gut microbiota, serum metabolomics, and network pharmacology (NP). Experimental validation of the main components of the key pathways was carried out using ELISA and Western blot.Results: The weights of the insomnia-model rats were significantly raised (p ≤ 0.05), the total exercise distance in the OFT increased (p ≤ 0.05), the rest time shortened, and the number of standing times increased (p ≤ 0.05), after treatment with MMS-4D. Moreover, there was a substantial recovery in the 5-HT, DA, GABA, and Glu levels in the hypothalamus tissue and the 5-HT and GABA levels in the colon tissue of rats. The expression of DAT and DRD1 proteins in the hippocampus of insomnia rats reduced after drug treatment. MMS-4D may treat insomnia by regulating different crucial pathways including 5-HT -, DA -, GABA -, and Glu-mediated neuroactive light receiver interaction, cAMP signaling pathway, serotonergic, glutamatergic, dopaminergic, and GABAergic synapses.Conclusion: This study revealed that MMS-4D can improve the general state and behavioral changes of insomnia model rats. Its mechanism may be related to the reversal of abnormal pathways mediated by 5-HT, DA, GABA, and Glu, such as Serotonergic synapse, Dopaminergic synapse, Glutamatergic synapse, and GABAergic synapse. Keywords: Mongolian medicine sugemule-4 decoction, insomnia, metabolomics, gut microbiota, network pharmacology, molecular mechanism

Bingjie Ma,1 Pengyi Hou,2 Ran Liu,3 Aijun Cang,1 Limei Zhao4 1Department of Pharmacy, People’s Hospital of Liaoning Province, Shenyang, 110010, People’s Republic of China; 2SCIEX Analytical Instrument Trading Co, Shanghai, 200000, People’s Republic of China; 3School of Food and Drug, Shenzhen Polytechnic, Shenzhen, 518000, People’s Republic of China; 4Department of Pharmacy, Shengjing Hospital of China Medical University, Shenyang, 110010, People’s Republic of ChinaCorrespondence: Limei Zhao, Department of Pharmacy, Shengjing Hospital of China Medical University, Shenyang, 110010, People’s Republic of China, Email lmzhao19@163.comBackground: Chemotherapy-induced myelosuppression (CIM) is a common adverse reaction with a high incidence rate that seriously affects human health. Shengyu Decoction (SYD) is often used to treat CIM. However, its pharmacodynamic basis and therapeutic mechanisms remain unclear.Purpose: This study aimed to clarify the active components and mechanisms of SYD in CIM.Methods: LC-QTOF/MS was used to identify the absorbable components of SYD. A series of network pharmacology methods have been applied to explore hub targets and potential mechanisms. Molecular docking was used to identify the binding ability of potential active ingredients and hub targets. Finally, in vitro experiments were performed to validate these findings.Results: In this study, 33 absorbable prototype components were identified using LC-QTOF/MS. A total of 62 possible targets of SYD in myelosuppression were identified. KEGG pathway enrichment analyses showed that some signaling pathways such as PI3K-Akt and HIF-1 may be the mechanisms by which it functions. Among them, we verified the PI3K-Akt pathway. 6 Hub proteins were screened by Protein–protein interaction (PPI) network analysis. Molecular docking results showed that four absorbable components in SYD showed good binding with six Hub targets. The effectiveness of the four predicted compounds and the mechanism were verified in vitro. It has also been shown that the active component could promote the proliferation of bone marrow stromal cells (BMSCs) and block apoptosis of BMSCs, which may be related to the PI3K-Akt pathway. This result is consistent with the network pharmacology approach and molecular docking predictions.Conclusion: Our results provided not only the candidate active component of SYD, but also a new insights into mechanism of SYD in the treatment of CIM. Keywords: Shengyu decoction, SYD, myelosuppression, network pharmacology, molecular docking, active components, mechanism

Yue Lu,1– 3,&ast; Simin Pan,4,&ast; Wenzhen Li,5 Yao Qi,6,7 Li Li,1– 3 Yu-Hong Yan,1– 3 Jianan Wei,1– 3 Dan-Ni Yao,1– 3 Jingjing Wu,1– 3 Hao Deng,1– 3 Shuyan Ye,1– 3 Haiming Chen,1– 3 Qubo Chen,1– 3 Hengjun Gao,6,7 Ling Han,1– 3 Chuanjian Lu1– 3 1State Key Laboratory of Dampness Syndrome of Chinese Medicine, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine (Guangdong Provincial Hospital of Chinese Medicine), Guangzhou, People’s Republic of China; 2Guangdong Provincial Key Laboratory of Clinical Research on Traditional Chinese Medicine Syndrome, Guangzhou, People’s Republic of China; 3Guangdong-Hong Kong-Macau Joint Laboratory on Chinese Medicine and Immune Disease Research, Guangzhou University of Chinese Medicine, Guangzhou, People’s Republic of China; 4The Second Clinical College, Guangzhou University of Chinese Medicine, Guangzhou, People’s Republic of China; 5The Clinical College of Acupuncture Rehabilitation, Guangzhou University of Chinese Medicine, Guangzhou, People’s Republic of China; 6Shanghai Molecular Medicine Engineering Technology Research Center, Shanghai, People’s Republic of China; 7Shanghai National Engineering Research Center of Biochip, Shanghai, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Ling Han; Chuanjian Lu, Email linghan99@gzucm.edu.cn; lcj@gzucm.edu.cnBackground: Psoriasis is a widespread chronic, immune-mediated skin disease with frequent recurrences, and is extremely harmful to the physical and mental health of patients, causing enormous suffering and exerting considerable economic burdens on the health care system as a whole. In more than a decade of clinical use, the optimized formula of Yinxieling (PSORI-CM01) has consistently demonstrated its effectiveness for treating psoriasis. However, its underlying mechanism remains largely unexplored.Methods: The network pharmacology analysis was conducted to predict the mechanism and protective effect of PSORI-CM01 in treating psoriasis. Subsequently, we collected blood samples from 21 patients with psoriasis as part of a randomized, double-blind, and double-dummy clinical trial for microRNA expression profiling. Finally, it was experimentally confirmed that PSORI-CM01 improved psoriasis by regulating miR-20a-3p and miR-3184-3p expression.Results: As a result of the network pharmacology analysis, PSORI-CM01 improved psoriasis through the regulation of autophagy, cellular apoptosis, cellular proliferation, and anti-inflammatory processes. In the target-miRNA regulatory network, these key targets were mainly associated with the regulation of hsa-miR-20a-3p, hsa-miR-155-5p, has-miR-3184-3p, hsa-miR-328-3p and hsa-miR-124-3p. Based on the microRNA expression profiling results, the PSORI-CM01 treatment group exhibited five up-regulated genes and 16 down-regulated genes compared with the healthy control group. In particular, miR-20a-3p and miR-3184-3p were the primary differentially expressed microRNAs, and they were significantly enriched in the signaling pathways involving autophagy, apoptosis, proliferation, and anti-inflammation. Further experiments confirmed that PSORI-CM01 effectively regulates miR-20a-3p and miR-3184-3p, resulting in increased autophagy.Conclusion: We demonstrated by combining network pharmacology and clinical studies of miRNA expression profiles in PBMCs that PSORI-CM01 effectively modulated miR-20a-3p and miR-3184-3p, leading to an increase in autophagy and a decrease in keratinocyte proliferation.Keywords: PSORI-CM01, Psoriasis vulgaris, MicroRNA expression profile, network pharmacology, autophagy

Background: Disorders of glutamate metabolism and excessive release participat in multiple neuronal pathologies including ischemic stroke (IS), Alzheimer's disease (AD), or Parkinson's disease (PD). Recently, herbal medicines have been widely used and have shown satisfactory results in the treatment of neurological disorders. Gastrodin is a traditional Chinese medicine (TCM) used for the treatment of nerve injuries, spinal cord injuries, and some central nervous system diseases as well. This research examines the neuroprotective effects of Gastrodin against glutamate-induced neurotoxicity in neuronal cells. Methods: The HERB database was used to explore the active ingredients and target genes of Gastrodia Elata. The STRING database and Cytoscape software were used to screen and construct the Protein-Protein Interaction (PPI). Furthermore, we used molecular docking to predict the potential targets of Gastrodin. The effects of Gastrodin were revealed by western blot, calcium imaging, membrane clamp, CCK8 and flow cytometry. Neuronal oxidative stress and damage were assessed by measuring malondialdehyde (MDA) levels and superoxide dismutase (SOD) activity. Neuronal morphology was examined using Golgi-Cox staining. Finally, animal behavior was examined using novel object recognition and fear conditioning tests. Results: We have obtained 22 components such as TM10, TM17, TM25 (Gastrodin), and 281 targets such as AKT, EGFR, and CDK1 through network pharmacology. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses revealed these genes were significantly enriched in protein phosphorylation, protein serine/threonine/tyrosine kinase activity, apoptosis and HIF-1 signaling pathways, etc. A higher affinity between Gastrodin and AKT was revealed by PPI analysis and molecular docking. Further, Gastrodin significantly inhibited Ca2+ influxes and excitatory synaptic transmission in cortical neurons. In addition, Gastrodin effectively alleviated neuron apoptosis, oxidative stress and damage. Conclusion: Gastrodin has neuroprotective effects against glutamate-induced neurotoxicity.

In this study, sixteen compounds were isolated from the branches of Mitragyna diversifolia, including twelve triterpenes (1–12), a phenolic compound (13), and three flavonoids (14–16). Among them, compounds 1–7, and 10–16 were reported for the first time from this plant. Compounds 7, 14, and 15 exhibited significant inhibitory activities against α-glucosidase, with IC50 values of 18.48 ± 2.74, 12.14 ± 1.58 and 35.77 ± 4.52 µM, respectively. Furthermore, the inhibitory kinetics of α-glucosidase revealed that all fractions, active compounds 7, 14, and 15 belong to the mix inhibition type. In molecular docking, the analysis showed that compounds 13, 14, 15, and 16 possessed superior binding capacities with α-glucosidase (−8.3, −9.6, −9.9, and −9.2 kcal/mol, respectively). The results of the glucose uptake experiment indicated that only compound 14 showed a significant promotion effect on the glucose uptake rate of 3T3-L1 adipocytes (P < 0.05). Meanwhile, compounds 13, 14, 15, and 16 possessed potent antioxidant abilities with DPPH, ABTS, and FRAP. In DNA and protein oxidative damage assays, compound 15 had a stronger effect than the positive control Vc. The network-based pharmacological analysis platform was used to predict the diabetes-related target proteins of active compounds 7, 13, 14, 15, and 16, and two candidate targets (ALB and PPARG) related to their therapeutic effects on diabetes were identified.

BackgroundPolycystic ovary syndrome (PCOS) is a heterogeneous gynecological endocrine disorder linked to immunity. Cangfu Daotan Decoction (CFDT), a classic Chinese medicine prescription, is particularly effective in treating PCOS, specifically in patients with obesity; however, its specific mechanism remains unclear.MethodsPart 1: Peripheral blood mononuclear cells were collected on egg retrieval day from obese and normal-weight patients with PCOS and healthy women undergoing in vitro fertilization (IVF)-embryo transfer. Next, scRNA-seq was performed to screen the key genes of bese patients with PCOS. Part 2: Active ingredients of CFDT and obesity-related PCOS targets were identified based on public databases, and the binding ability between the active ingredients and targets was analyzed. Part 3: This part was a monocentric, randomized controlled trial. The obese women with PCOS were randomized to CFDT (6 packets/day) or placebo, and the healthy women were included in the blank control group (43 cases per group). The clinical manifestations and laboratory outcomes among the three groups were compared.ResultsBased on the scRNA-seq data from Part 1, CYLD, ARPC3, CXCR4, RORA, JUN, FGL2, ZEB2, GNLY, FTL, SMAD3, IL7R, KIR2DL1, CTSD, BTG2, CCL5, HLA, RETN, CTSZ, and NCF2 were potential key genes associated with obese PCOS were identified. The proportions of T, B, and natural killer cells were higher in patients with PCOS compared to healthy women, with even higher proportions observed in obese patients with PCOS. Gene ontology and the Kyoto encyclopedia of genes and genomes analysis depicted that the differentially expressed genes were related to immune regulation pathways. Network pharmacology analysis identified that the key active components in CFDT were quercetin, carvacrol, β-sitosterol, cholesterol, and nobiletin, and TP53, AKT1, STAT3, JUN, SRC, etc. were the core targets. The core targets and their enrichment pathways overlapped with those in Part 1. Clinical trials in Part 3 found that CFDT reduced the dosage of gonadotropins use in patients with PCOS, increased the number of high-quality embryos, and improved the ongoing pregnancy rate.ConclusionCFDT can improve the immune microenvironment of patients to some extent, reduce their economic burden, and enhance IVF outcomes. The improvement in the immune microenvironment in obese patients with PCOS may be linked to targets such as JUN and AKT.

IntroductionBreast cancer is the most common cancer worldwide, the leading cause of cancer death in women, and the fifth leading cause of cancer death. Triple negative breast cancer (TNBC), with high metastasis and mortality rates, is the most challenging subtype in breast cancer treatment. There is an urgent need to develop anti-TNBC drugs with significant efficacy, low side effects and good availability. In early drug screening, blestriarene C was found to have inhibitory effects on TNBC cells. In this article, we further explore the mechanisms associated with blestriarene C for breast cancer.MethodsIn this article, we take the approach of network pharmacology combined with in vivo and in vitro experiments. Network pharmacology analysis was used to predict the active components in Baiji, and to investigate the hub targets and related mechanisms of BC in TNBC treatment. The mechanism of anti-TNBC in vitro was evaluated by CCK-8 assay, cell apoptosis and cell cycle assays, wound healing assay, WB assay, and molecular docking analysis. The inhibition effect in vivo was test in subcutaneous tumor models established in mice.ResultsThrough network pharmacology analysis and experiments, we screened out BC as the main active ingredient, and found that BC could inhibit the Ras/ERK/c-Fos signaling pathway while downregulating the expression of HSP90AA1 and upregulating the expression of PTGS2, thereby promoting apoptosis, causing S-phase cycle arrest, and inhibiting the proliferation and migration of BT549 cells. The in vivo results illustrated that BC inhibited the growth of TNBC tumors and has a high safety profile. By integrating network pharmacology with in vitro and in vivo experiments, this study demonstrated that BC inhibited the proliferation and migration of TNBC cells by inhibiting the Ras/ERK/c-Fos signaling pathway, promoting apoptosis, and causing S-phase cycle arrest.DiscussionThis study provides new evidence for the use of BC as a novel drug for TNBC treatment.

BackgroundSkin photoaging induced by ultraviolet A (UVA) and ultraviolet B (UVB) radiation manifests as skin roughness, desquamation, pigmentation, and wrinkle formation. Current treatments, such as sunscreen, hormones, and antioxidants, have limitations and side effects. Traditional Chinese Medicine Hibisci Mutabilis Folium (HMF), or Mu-Fu-Rong-Ye in Chinese name, refers to the dried leaves of the plant Hibiscus mutabilis L., which belongs to the Malvaceae family. It has been used traditionally to treat acute mastitis, parotitis, neurodermatitis, burns. The reported activities of HMF include anti-inflammatory and anti-oxidant effects. However, the therapeutic potential of HMF in preventing and treating UV-induced skin photoaging remains unexplored.ObjectiveThis study aimed to investigate the protective effects of HMF extract (EHMF) against UV-induced skin photoaging and the underlying mechanisms of action, by using network pharmacology and experimental verification.MethodsNetwork pharmacology was employed to identify the effective chemical components of EHMF. Potential targets were identified via PPI network analysis. Representative compounds were characterized using UPLC-MS/MS. In vitro validation involved assessing HaCaT cell viability, observing live/dead cell staining through fluorescence microscopy, and measuring inflammatory factors using ELISA. For in vivo validation, a UV-induced skin photoaging mice model was treated transdermally with EHMF or Methotrexate daily for 7 days. Dermatitis severity, skin morphology, and collagen fiber pathology were evaluated. Inflammatory cytokine and protein expression in dorsal skin lesions was confirmed using Elisa Kits, Western blot and immunohistochemistry.ResultsA total of 22 active ingredients of EHMF were identified. GO enrichment and KEGG pathway analyses revealed a focus on inflammatory signaling pathways. In vitro experiments showed that EHMF significantly reduced UV-induced inflammatory factors in HaCaT cells and improved cell survival rates. In vivo, EHMF alleviated back skin lesions in UV-exposed mice, reducing epidermal and dermal thickening and pathological inflammatory cell infiltration. It also decreased abnormal MMP-9 expression and collagen fiber proliferation, along with levels of inflammatory factors like TNF-α, IL-6, IL-17, and EGFR. Western blot and immunohistochemistry results indicated that the over-activation of the AKT-STAT3 signaling pathway was inhibited.ConclusionEHMF effectively reduced UV-induced skin damage, inflammation, and wrinkles, providing strong support for its clinical application as a dermatological agent.

Muscular dystrophies encompass a heterogeneous group of rare neuromuscular diseases characterized by progressive muscle degeneration and weakness. Among these, Duchenne muscular dystrophy (DMD) stands out as one of the most severe forms. The present study employs an integrative approach combining network pharmacology, quantitative structure-activity relationship (QSAR) modeling, molecular dynamics (MD) simulations, and free energy calculations to identify potential therapeutic targets and natural compounds for DMD. Upon analyzing the GSE38417 dataset, it was found that individuals with DMD exhibited 290 upregulated differentially expressed genes (DEGs) compared to healthy controls. By utilizing gene ontology (GO) and protein-protein interaction (PPI) network analysis, this study provides insights into the functional roles of the identified DEGs, identifying ten hub genes that play a critical role in the pathology of DMD. These key genes include DMD, TTN, PLEC, DTNA, PKP2, SLC24A, FBXO32, SNTA1, SMAD3, and NOS1. Furthermore, through the use of ligand-based pharmacophore modeling and virtual screening, three natural compounds were identified as potential inhibitors. Among these, compounds 3874518 and 12314417 have demonstrated significant promise as an inhibitor of the SMAD3 protein, a crucial factor in the fibrotic and inflammatory mechanisms associated with DMD. The therapeutic potential of the compounds was further supported by molecular dynamics simulation and Molecular Mechanics/Generalized Born Surface Area (MM/GBSA) analysis. These findings suggest that the compounds are viable candidates for experimental validation against DMD.

Aims: Diabetic nephropathy (DN) is a major complication of diabetes mellitus (DM) without curative interventions currently. Huperzine A (Hup A), a natural alkaloid, has demonstrated significant hypoglycemic and anti-inflammatory effects. We aim to investigate the protective effects of Hup A on DN and explore the underlying mechanisms Methods: We applied STZ induced diabetic rats as DN model and leveraged combination analysis of the transcriptome, metabolome, microbiome, and network pharmacology (NP). The total effect of Hup A on DN was detected (i.e. urine protein, renal tissue structure) and the differential genes were further verified at the level of diabetic patients, db/db mice and cells. Clinical data and small interfering RNA (siRNA)-Apoe were adopted. Results: Hup A alleviated kidney injury in DN rats. Transcriptomics data and Western blot indicated that the improvement in DN was primarily associated with Apoe and Apoc2. Additionally, metabolomics data demonstrated that DN-induced lipid metabolism disruption was regulated by Hup A, potentially involving sphingosine. Hup A also enriched microbial diversity and ameliorated DN-induced microbiota imbalance. Spearman's correlation analysis demonstrated significant associations among the transcriptome, metabolome, and microbiome. Apoe level was positively correlated with clinical biomarkers in DN patients. Si-Apoe also played protective role in podocytes. NP analysis also suggested that Hup A may treat DN by modulating lipid metabolism, microbial homeostasis, and apoptosis, further validating our findings. Conclusions: Collectively, we provide the first evidence of the therapeutic effect of Hup A on DN, indicating that Hup A is a potential drug for the prevention and treatment of DN.

Sarcopenic obesity (SO) is a metabolic disorder with increasing prevalence. It is characterized by a reduction in skeletal muscle mass and strength. Resveratrol (RSV) is one of the most frequently used herbs in the treatment of skeletal muscle atrophy. However, the precise mechanism of the action of RSV in SO remains unclear. The objective of this study was to examine the pharmacological mechanism of RSV in the context of SO through the lens of network pharmacology, to validate these findings through in vivo experimentation. A list of potential RSV targets was compiled by retrieving the data from multiple databases. This list was then cross-referenced with a list of potential targets related to SO. The intersections of RSV- and SO-related targets were analyzed using Venn diagrams. To identify the core genes, a protein-protein interaction (PPI) network of the intersection targets was constructed and subsequently analyzed. Molecular docking was used to predict RSV binding to its core targets. A high-fat diet was used to induce SO in mice. These findings indicated that RSV may prevent SO by acting on 11 targets. Among these, interleukin-6 (IL-6), C-reactive protein (CRP), and tumor necrosis factor (TNF) are considered core targets. The Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment results indicated that the anti-SO effect of RSV was predominantly linked to metabolic disease-related pathways, including those associated with nonalcoholic fatty liver disease. The anti-inflammatory effects of RSV were confirmed in vivo in an SO mouse model. This study contributes to a more comprehensive understanding of the key mechanisms of the action of RSV against SO and provides new possibilities for drug development in the pathological process of SO.

Background: In the early stages of diabetic kidney disease (DKD), the pathogenesis involves abnormal angiogenesis in the glomerulus. Hirudin, as a natural specific inhibitor of thrombin, has been shown in previous studies to inhibit the migration of various tumor endothelial cells and abnormal angiogenesis. However, its role in DKD remains unclear. Methods: The effects of hirudin in DKD were studied using spontaneous type 2 diabetic db/db mice (which develop kidney damage at 8 weeks). Network pharmacology was utilized to identify relevant targets. An in vitro high glucose model was established using mouse glomerular endothelial cells (MGECs) to investigate the effects of hirudin on the migration and angiogenic capacity of MGECs. Results: Hirudin can ameliorate kidney damage in db/db mice. Network pharmacology suggests its potential association with the VEGFA/VEGFR2 pathway. Western blot and immunohistochemistry demonstrated elevated protein expression levels of VEGFA, VEGFR2, AQP1, and CD31 in db/db mice, while hirudin treatment reduced their expression. In the MGECs high glucose model, hirudin may reverse the enhanced migration and angiogenic capacity of MGECs in a high glucose environment by altering the expression of VEGFA, VEGFR2, AQP1, and CD31. Moreover, the drug effect gradually increases with higher concentrations of hirudin. Conclusions: This study suggests that hirudin can improve early-stage diabetic kidney disease kidney damage by inhibiting the migration and angiogenesis of glomerular endothelial cells, thereby further expanding the application scope of hirudin. Additionally, the study found increased expression of AQP1 in DKD, providing a new perspective for further research on the potential pathogenesis of DKD.

IntroductionHyperlipidemia is a condition characterized by abnormal levels of lipids and lipoproteins in the plasma, posing significant health risks. Ferulic acid (FA) is an organic acid with therapeutic properties for diabetes and hyperlipidemia.MethodsTo explore biomarkers for FA treatment of hyperlipidemia and elucidate the mechanisms of lipid-lowering-related changes in metabolic pathways by metabolomics and network pharmacology. Initially, a hyperlipidemic mouse model induced by triton WR-1339 was established to evaluate the therapeutic effects of FA. Subsequently, serum metabolomics was utilized to identify differential metabolites, and metabolic pathway analysis was performed using MetaboAnalyst 6.0. Thirdly, network pharmacology was employed to identify potential targets of FA for hyperlipidemia. Finally, the compound-target-metabolite (C-T-M) network obtained core targets and validated them with molecular docking.ResultsBiochemical analysis and histological examination showed that FA had lipid-lowering effects on hyperlipidemic mice. It identified 31 potential biomarkers for FA against hyperlipidemia by metabolomics involving lipid and amino acid metabolism. Lipid and atherosclerosis signaling pathways were identified as the key signaling pathways of FA against hyperlipidemia by KEGG analysis. Conjoint analysis showed that FA against hyperlipidemia was associated with 18 core targets and six biomarkers. Molecular docking results showed that FA has a high binding affinity to these core targets.DiscussionThrough the synergy of network pharmacology and metabolomics, this study provides insights into how FA regulates endogenous metabolites, underscoring its promise as a treatment for hyperlipidemia.

Pulmonary fibrosis (PF) is a chronic progressive end-stage lung disease. However, the mechanisms underlying the progression of this disease remain elusive. Presently, clinically employed drugs are scarce for the treatment of PF. Hence, there is an urgent need for developing novel drugs to address such diseases. Our study found for the first time that a natural source of Prismatomeris connata Y. Z. Ruan (Huang Gen, HG) ethyl acetate extract (HG-2) had a significant anti-PF effect by inhibiting the expression of the transforming growth factor beta 1/suppressor of mothers against decapentaplegic (TGF-β1/Smad) pathway. Network pharmacological analysis suggested that HG-2 had effects on tyrosine kinase phosphorylation, cellular response to reactive oxygen species, and extracellular matrix (ECM) disassembly. Moreover, mass spectrometry imaging (MSI) was used to visualize the heterogeneous distribution of endogenous metabolites in lung tissue and reveal the anti-PF metabolic mechanism of HG-2, which was related to arginine biosynthesis and alanine, asparate and glutamate metabolism, the downregulation of arachidonic acid metabolism, and the upregulation of glycerophospholipid metabolism. In conclusion, we elaborated on the relationship between metabolite distribution and the progression of PF, constructed the regulatory metabolic network of HG-2, and discovered the multi-target therapeutic effect of HG-2, which might be conducive to the development of new drugs for PF.

BackgroundType 2 diabetes mellitus is a metabolic disease categorized by hyperglycemia, resistance to insulin, and ß-cell dysfunction. Around the globe, approximately 422 million people have diabetes, out of which 1.5 million die annually. In spite of innovative advancements in the treatment of diabetes, no biological drug has been known to successfully cure and avert its progression. Thereupon, natural drugs derived from plants are emerging as a novel therapeutic strategy to combat diseases like diabetes.ObjectiveThe current study aims to investigate the antidiabetic potential of natural compounds of Oryza sativa L. indica (black rice) in disease treatment.MethodsAntioxidant activity and alpha amylase assays were performed to evaluate the therapeutic potential of the extract of Oryza sativa L. indica. Gas chromatography–mass spectrometry (GC–MS) was used for identification of constituents from the ethanol extract. ADMET profiling (absorption, distribution, metabolism, excretion, and toxicity), network pharmacology, and molecular dynamics simulation were employed in order to uncover the active ingredients and their therapeutic targets in O. sativa L. indica against type 2 diabetes mellitus.ResultsGC–MS of the plant extract provided a list of 184 compounds. Lipinski filter and toxicity parameters screened out 18 compounds. The topological parameters of the protein–protein interaction (PPI) were used to shortlist the nine key proteins (STAT3, HSP90AA1, AKT1, SRC, ESR1, MAPK1, NFKB1, EP300, and CREBBP) in the type 2 diabetes mellitus pathways. Later, molecular docking analysis and simulations showed that C14 (1H-purine-8-propanoic acid, .alpha.-amino-2, 3, 6, 7-tetrahydro-1,3,7-trimethyl-2,6-dioxo-) and C18 (cyclohexane-carboxamide, N-furfuryl) bind with AKT1 and ESR1 with a binding energy of 8.1, 6.9, 7.3, and 7.2 kcal/mol, respectively. RMSD (root-mean-square deviation) and RMSF (root-mean-square fluctuation) values for AKT1 and ESR1 have shown very little fluctuation, indicating that proteins were stabilized after ligand docking.ConclusionThis study suggests therapeutic drug candidates against AKT1 and ESR1 to treat type 2 diabetes mellitus. However, further wet-lab analysis is required to discover the best remedy for type 2 diabetes mellitus.

BackgroundFFBZL is composed of three herbs: Scutellaria barbata D. Don (SBD), Astragali Radix (AR), and Ligusticum chuanxiong Hort (CX). FFBZL has been reported to be effective in the treatment of oral squamous cell carcinoma (OSCC). However, the molecular mechanism involved remains unclear. Based on network pharmacology combined with bioinformatics and molecular docking, the effect and molecular mechanism of action of FFBZL in treating OSCC were explored.Materials and methodsThis study employed an integrated approach using various databases and literature sources to identify the effective components of FFBZL, with a specific emphasis on screening active ingredients that align with traditional Chinese medicine principles. The TCMSP, ETCM, and SymMap databases were utilized to collect information on the active constituents and targets of FFBZL, while the PharmMapper database was used to predict targets. Key components were selected based on the degree value of the ‘active component−target’ network. Transcriptome data for OSCC samples were obtained from the TCGA and GEO databases. Differential gene expression analysis was conducted to identify targets associated with OSCC, and these targets were subsequently aligned with targets of the effective components of FFBZL to identify common targets. Subsequently, the STRING database was utilized to construct a protein‒protein interaction (PPI) network of these common targets, which was subsequently visualized using Cytoscape. Next, 71 targets were rescreened using the PPI network, and GO and KEGG enrichment analyses were performed; the PI3K-Akt signaling pathway was the top-ranking pathway related to cell apoptosis. Next, the expression of 19 genes enriched in the PI3K-Akt signaling pathway was analyzed using OSCC transcriptome data from the TCGA and GEO databases. The targets were subsequently mapped to the PI3K-Akt signaling pathway using the KEGG database, and the GSEA algorithm was used to assess the overall expression trend of the genes in this pathway. The 71 common targets were subsequently imported into the STRING database and visualized using Cytoscape. The DEGREE and MCC algorithms were used to select the corresponding targets within the PPI network. The intersection of these targets and the 19 targets mapped to the PI3K-Akt signaling pathway led to the identification of 6 key targets associated with cell apoptosis: GSK3B, PIK3CA, FN1, MET, SPP1, and MAPK3. Subsequently, the UALCAN database was utilized to analyze the expression levels and survival associations of the key genes related to cell apoptosis, and the transcriptome data from the GEO database were used to assess the correlations among the 6 key genes. Finally, molecular docking studies were conducted to explore the relationships between these targets and the active components with predicted associations.ResultsThis study identified six key components of FFBZL (quercetin, wogonin, carthamidin, scutellarein, senkyunolide K and astragalosidei: astragaloside I) as well as 820 potential target genes of these components. Intersection of these targets with those related to OSCC yielded 151 common targets. GO and KEGG enrichment analyses revealed that most of the top-ranked functions and pathways were associated with apoptosis, with the PI3K-Akt signaling pathway playing a critical role. Transcriptome analysis of data from the TCGA and GEO databases indicated that the genes enriched in the PI3K-Akt signaling pathway were strongly upregulated, and the GSEA algorithm indicated an overall upregulation trend for the PI3K-Akt signaling pathway. By intersecting the targets with the 19 genes mapped to the PI3K-Akt signaling pathway using the DEGREE and MCC algorithms, six key targets related to cell apoptosis were identified. The mRNA and protein expression levels of most these targets in head and neck squamous cell carcinoma were higher than those in normal tissues. Survival analysis revealed that low expression of SPP1 and FN1 was associated with increased patient survival time. Additionally, the molecular docking results indicated strong binding potential between the six identified key components and the six key targets.

BackgroundProstate cancer (PCa) is the most common non-cutaneous malignancy in men globally. Sappan lignum, which exists in the heartwood of Caesalpinia sappan L., has antitumor effects; however, its exact mechanism of action remains unclear. This study elucidated the underlying mechanisms of Sappan lignum in PCa through network pharmacology approaches and molecular docking techniques. Moreover, the therapeutic effects of Sappan lignum on PCa were verified through in vitro experiments.MethodsThe constituent ingredients of Sappan lignum were retrieved from the HERB database. Active plant-derived compounds of Sappan lignum were screened based on gastrointestinal absorption and gastric drug properties. Disease targets for PCa were screened using unpaired and paired case datasets from the Gene Expression Omnibus. Intersection targets were used for gene ontology and Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis. Core targets were identified through topological analysis parameters and their clinical relevance was validated through The Cancer Genome Atlas database. The affinity between the phytochemicals of Sappan lignum and core proteins was verified using the molecular docking technique. Validation experiments confirmed the significant potential of Sappan lignum in treating PCa.ResultsTwenty-one plant-derived compounds of Sappan lignum and 821 differentially expressed genes associated with PCa were collected. Among 32 intersection targets, 8 were screened according to topological parameters. KEGG analysis indicated that the antitumor effects of Sappan lignum on PCa were primarily associated with the p53 pathway. The molecular docking technique demonstrated a strong affinity between 3-deoxysappanchalcone (3-DSC) and core proteins, particularly cyclin B1 (CCNB1). CCNB1 expression correlated with clinicopathological features in patients with PCa. Experimental results revealed that 3-DSC exhibited anti-proliferative, anti-migratory, and pro-apoptotic effects on 22RV1 and DU145 cells while also causing G2/M phase cell cycle arrest, potentially through modulating the p53/p21/CDC2/CCNB1 pathway.ConclusionThis research highlights the promising therapeutic potential of Sappan lignum in treating PCa, with a particular focus on targeting the p53 pathway.

IntroductionYouhua Kuijie prescription (YHKJ) is a hospital preparation that is composed of nine kinds of herbs. Sulfasalazine (SASP) is widely used as a first-line clinical treatment for UC. Traditional Chinese medicine and Western medicine have their own advantages in the treatment of UC, and the mechanism of YHKJ combined with SASP in the treatment of UC needs to be investigated.MethodsIn this study, the therapeutic mechanism of YHKJ combined with SASP in the treatment of UC was predicted by network pharmacology and molecular docking. The chemical components and related targets of YHKJ were obtained from the TCMSP database. The chemical structure of SASP was obtained from the PubChem server, and related targets of SASP molecules were identified using the PharmMapper database. UC-related targets were obtained from the DisGeNET, GeneCards, OMIM, TTD, DrugBank and PharmGkb databases.ResultsIn total, 197 shared targets were identified by constructing a Venn diagram. PPI network data obtained from the STRING database were imported into Cytoscape to visualize the “drug-disease” target network, and STAT3 was selected as the core target by topological analysis. Gene Ontology revealed the biological functions of target genes, and KEGG analysis revealed that the core target STAT3 was differentially expressed in Th17 cells and the JAK-STAT signaling pathway. Thus, the core target STAT3 was subjected to molecular docking with the top 10 components, including nine YHKJ components (quercetin, luteolin, ursolic acid, daidzein, kaempferol, wogonin, myricetin, formononetin, indirubin) and SASP (C18H14N4O5S). The molecular docking results showed that STAT3 had favorable binding with the nine YHKJ components and SASP; STAT3 had the strongest binding with ursolic acid (−10.26 kcal/mol), followed by SASP (−8.54 kcal/mol). Qualitative analysis of the chemical constituents of YHKJ by HPLC revealed that sitosterol, ursolic acid, myricetin, daidzein, quercetin, kaempferol and formononetin were the main components. Additional experiments verified that YHKJ combined with SASP inhibited activation of the IL-6/JAK2/STAT3 pathway and alleviated inflammation in UC model rats.DiscussionOur results showed that seven chemical components in YHKJ cooperate with SASP to interfere with activation of the IL-6/JAK2/STAT3 pathway, thus playing a role in the treatment of UC.

BackgroundYangxue Xifeng Decoction (YXD) has been utilized in clinical settings for the treatment of Tourette Syndrome (TS). However, the action mechanism of YXD needs further research.MethodsThe ingredients and targets of YXD were identified via database searches and then constructed an active ingredient-target network using Cytoscape. Pathway enrichment analysis was performed via Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG). The core genes were determined by LASSO regression and SVM algorithm. Additionally, we analyzed the immune infiltration. The signaling pathways associated with core genes were investigated through KEGG and GO. We predicted the transcription factors using “RcisTarge”.Results127 active ingredients of YXD and 255 targets were obtained. TNF and the IL-17 signaling pathway were the main pathways. OPRM1 and VIM were screened out as core genes, which were associated with the immune infiltration. The signaling pathways involved in OPRM1 and VIM were enriched. Furthermore, remarkable correlation was found between OPRM1 and VIM levels and other TS-related genes such as MAPT and MAPT.ConclusionOPRM1 and MAPT, and the signaling pathways are associated with TS. YXD exerts its therapeutic TS through multi-component and multi-targets including immune infiltration.

Introduction: Dendrobium nobile Lindl alkaloids, or DNLA for short, are the most active ingredients found in D.nobile, a top grade plant in Shen Nong Ben Cao Jing, with an extensive history of medicinal use in Chinese traditional medicine (TCM) as a multifunctional therapeutic agent. Recent evidence has emerged linking the neuroprotective and anti-aging effects of DNLA to their involvement in promoting autophagy of toxic amyloid-β (Aβ) plaques and modulation of key enzymes involved in the hyperphosphorylation of Tau proteins. Although amyloid buildup and the aggregation of Tau proteins are central to the onset of Alzheimer's disease (AD), evidence on how DNLA relate to other overlooked dysregulated AD-associated pathways is still lacking. Methods: We intend on deciphering the underlying mechanisms driving the anti-AD effect of DNLA, using a combination of network analysis based on differentially expressed genes found in AD patients, target fishing, centrality analysis, enrichment analysis and hub genes identification. Results: In total, 2069 genes were found differentially expressed in SRP181886 and a PPI network constructed with common targets between DNLA and AD. Five hub genes were identified having a discriminatory power greater than 0.7; HTR2A, GRIN2B, GABRA1, HTR2C, GRIN2A, with the former being the top bottleneck node in the network. Enrichment analysis found that DNLA exert an anti-AD effect through the regulation of the calcium signaling pathway and the serotonergic system, by modulating key receptors implicated in excitatory/inhibitory neurotransmission. Additionally, DNLA were found to modulate two subunits of NMDA receptor involved in the release of pro-inflammatory cytokines, underlying the possible involvement of DNLA in neuroinflammation. Discussion: This further emphasizes the therapeutic value of D.nobile and the multi-target, multi-pathway potential of DNLA to counteract the deleterious effects of calcium dysregulation and excitatory toxicity in AD, while providing evidence-based rationale behind the traditional use of D. nobile in TCM.

Da Chuanxiong Formula (DCXF) is a traditional herbal prescription used for pain management. It consists of Chuanxiong Rhizoma (CR) and Gastrodiae Rhizoma (GR). Despite its long history of use, the underlying therapeutic mechanism of DCXF remains insufficiently understood. Therefore, in this study, key target genes were obtained through network pharmacology research methods and molecular docking techniques, including transient receptor potential vanilloid 1 (TRPV1), adenosine A2a receptor (ADORA2A), nuclear receptor subfamily 3 group C member 1 (NR3C1), and protein kinase C beta (PRKCB). Molecular dynamics simulations demonstrated the favorable binding between all four key genes and their corresponding compounds. Notably, chronic constriction injury (CCI) treatment resulted in a significant decrease in mechanical threshold and thermal latency period for rat foot contraction, which was ameliorated upon administration of DCXF. Furthermore, real-time quantitative reverse transcription PCR (RT-qPCR) and western blot (WB) analyses indicated an upregulation of TRPV1, ADORA2A, NR3C1, and PRKCB expression in the rat dorsal root ganglion following CCI, which was attenuated by treatment with DCXF. The expressions of inflammatory factors, including tumor necrosis factor-α (TNF-α), interleukin 1 beta (IL-1β), and interleukin 6 (IL-6), in the rat dorsal root ganglion were assessed using ELISA, confirming consistent trends with the aforementioned findings. The results of this study offer a promising theoretical foundation for the utilization of DCXF in the treatment of neuropathic pain (NP).

Lung cancer ranks as the 2nd most common cancer globally. It’s the most prevalent cancer in men and the 2nd most common in women. The prominent events in EGFR-mutated non-small-cell lung cancer (NSCLC) include the emergence of the L858R mutation within EGFR exon 21. Despite the promising efficacy of EGFR inhibitors in managing lung cancer, the development of acquired resistance poses a significant hurdle. In the current investigation, we focused on the screening of two phytochemicals, namely Dehydrocostus lactone and Mokkolactone, derived from the Saussurea lappa plant, as potential inhibitors targeting EGFR L858R mutant lung cancer. The chloroform and ethanol extract of the plant demonstrated anti-proliferative activity through the Resazurin chemosensitivity assay, exhibiting an IC50 value of 37.90 ± 0.29 µg/ml with selectivity index 2.4. Through a GC–MS study, we identified 11 phytochemicals for further insilico analysis. These compounds underwent ADMET assessment followed by drug likeliness analysis before being subjected to molecular docking against EGFR L858R, identified through protein–protein interaction network analysis. All phytochemicals exhibited binding energy scores ranging from −6.9 to −8.1 kcal/mol. Dehydrocostus lactone and Mokkolactone were specifically identified for their binding profile. Findings from 100 ns molecular dynamics simulations demonstrated their enhanced stability compared to the reference ligand DJK. This was evident in the root mean square deviation (RMSD) values, ranging from 0.23 ± 0.01 nm to 0.30 ± 0.05 nm, the radius of gyration values, from 1.71 ± 0.01 nm to 1.72 ± 0.01 nm, and the solvent accessible surface area values, from 155.39 ± 2.40 nm2 to 159.32 ± 2.14 nm2. Additionally, favourable characteristics were observed in terms of hydrogen bonding, principal component analysis, and free energy landscape analysis. Examination of their electronic structure via density functional theory revealed efficient properties, with the highest occupied molecular orbital-least unoccupied molecular orbital energy gap values ranging from −3.984 eV to −6.547 eV. Further, in vivo analysis is required to gain a more comprehensive understanding and efficacy of these identified phytochemicals against lung cancer.

Yihua Fan,1– 3,&ast; Wei Liu,1,2,&ast; Yue Jin,1,2,&ast; Hang Lu,1,2 Chunliu Liu,1,2 Aihua Wang,1,2 Qingxiang Gu,1,2 Yuxiu Ka1,2 1Department of Rheumatism and Immunity, First Teaching Hospital of Tianjin University of Traditional Chinese Medicine, Tianjin, People’s Republic of China; 2National Clinical Research Center for Chinese Medicine Acupuncture and Moxibustion, Tianjin, People’s Republic of China; 3Department of Rheumatism and Immunity, Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan Province, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Wei Liu, Email fengshiliuwei@163.comBackground: Acute gouty arthritis (AGA) is characterized by the accumulation of monosodium urate crystals within the joints, leading to inflammation and severe pain. Western medicine treatments have limitations in addressing this condition. Previous studies have shown the efficacy of Qinpi Tongfeng formula (QPTFF) in treating AGA, but further investigation is needed to understand its mechanism of action.Methods: We used ultra-high-performance liquid chromatography tandem Q-Exactive Orbitrap high-resolution mass spectrometry (UHPLC-Q-Orbitrap-MS) to identify compounds in QPTFF. Target proteins regulated by these compounds were obtained from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform, Chemistry Database, and Swiss Target Prediction Database. AGA-related targets were searched and screened from various databases, including Genecards, PharmGKB, Drugbank, etc. Intersection targets of QPTFF and AGA were analyzed for protein–protein interaction networks, GO function enrichment, and KEGG pathway enrichment. We then verified QPTFF’s mechanism of action using an AGA rat model, assessing pathological changes via H&E staining and target expression via ELISA, RT-qPCR, and Western blot.Results: UHPLC-Q-Orbitrap-MS identified 207 compounds in QPTFF, with 55 selected through network pharmacology. Of 589 compound-regulated targets and 1204 AGA-related targets, 183 potential targets were implicated in QPTFF’s treatment of AGA. Main target proteins included IL-1β, NFKBIA, IL-6, TNF, CXCL8, and MMP9, with the IL-17 signaling pathway primarily regulated by QPTFF. Experimental results showed that medium and high doses of QPTFF significantly reduced serum inflammatory factors and MMP-9 expression, and inhibited IL-17A, IL-6, IKK-β, and NF-κB p65 mRNA and protein expression in AGA rats compared to the model group.Conclusion: Key targets of QPTFF include IL-1β, NFKBIA, IL-6, TNF-α, CXCL8, and MMP9. QPTFF effectively alleviates joint inflammation in AGA rats, with high doses demonstrating no liver or kidney toxicity. Its anti-inflammatory mechanism in treating AGA involves the IL-17A/NF-κB p65 signaling pathway.Keywords: acute gouty arthritis, Qinpi Tongfeng formula, network pharmacology, UHPLC-Q-Orbitrap-MS, IL-17 signaling pathway