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Jiahua Liang,1,* Yingjie Huang,1,* Zhexing Mai,1,* Qunzhang Zhan,1 Hengchen Lin,1 Yuxin Xie,1 Haihao Wang,1 Yan Liu,1 Chuanjin Luo2 1The First Clinical Medical College, Guangzhou University of Chinese Medicine, Guangzhou, Guangdong, People’s Republic of China; 2The Department of Cardiovascular Disease, Guangzhou University of Traditional Chinese Medicine First Affiliated Hospital, Guangzhou, Guangdong, People’s Republic of China*These authors contributed equally to this workCorrespondence: Chuanjin LuoThe Department of Cardiovascular Disease, Guangzhou University of Traditional Chinese Medicine First Affiliated Hospital, Jichang Road 16#, District Baiyun, Guangzhou, 510405, Guangdong, People’s Republic of ChinaEmail Gztcm1964@163.comBackground: This study used network pharmacology, molecular docking and experimental validation to assess the effects of Huanglian Jiedu Decoction (HLJDD) on atherosclerosis (AS).Methods: The components and targets of HLJDD were analyzed using the Traditional Chinese Medicine Systems Pharmacology database, and information on the genes associated with AS was retrieved from the GeneCards and OMIM platforms. Protein–protein interactions were analyzed using the STRING platform. A component–target–disease network was constructed using Cytoscape. GO and KEGG analyses were performed to identify molecular biological processes and signaling pathways, and the predictions were verified experimentally. Molecular docking was conducted with ChemOffice software, PyMOL software and Vina to verify the correlation of targets and compounds.Results: HLJDD contained 31 active compounds, with quercetin, kaempferol, moupinamide and 5-hydroxy-7-methoxy-2-(3,4,5-trimethoxyphenyl)chromone as the core compounds. The most important biotargets of HLJDD in AS were ICAM-1, CD31 and RAM-11. The molecular docking results showed that the molecular docking interaction energy between the 3 key targets and the 4 high-degree components were much less than − 5 kJ∙mol− 1. The experimental validation results showed that HLJDD might treat AS mainly by reducing TC, TG and LDL-C and increasing HDL-C, upregulating CD31 expression, reducing ICAM-1 and RAM-11 expression, and downregulating inflammatory factors, including CRP, IL-6 and TNF-α. These results support the network pharmacology data and demonstrate that HLJDD affects the expression of core genes and alters the leukocyte transendothelial migration signaling pathway.Conclusion: Based on network pharmacology and experimental validation, our study indicated that HLJDD exerted anti-AS effect through upregulating CD31 expression and reducing the expression of ICAM-1 and RAM-11. HLJDD may be a potential therapeutic drug to the prevention of AS.Keywords: Huanglian Jiedu Decoction, atherosclerosis, network pharmacology, molecular docking, experimental validation

Nuanxin capsule (NX), an in-hospital preparation of Guangdong Provincial Hospital of Chinese Medicine, has been used in heart failure (HF) treatment for 15 years, but its mechanism and protective effect have not been investigated. This study was aimed at exploring the mechanism and protective effect of NX on HF treatment via network pharmacology analysis and experimental validation. Network pharmacology analysis predicted that NX was involved in the regulation of response to apoptotic process and hypoxia via protecting cellular damage and mitochondrial dysfunction against chronic hypoxia. Its mechanism may be involved in the regulation of the PI3K-Akt signaling pathway, HIF-1 signaling pathway, AMPK signaling pathway, and MAPK signaling pathway. Experimental validation indicated that NX was capable of improving cellular viability, restoring cellular morphology, and suppressing cellular apoptosis cellular. NX also exerted cardioprotection by inhibiting mitochondrial membrane potential injury and protecting mitochondrial respiratory and energy metabolism in a chronic hypoxia cellular model, which was consistent with the results of network pharmacology prediction. In addition, the screened active compounds of NX did have a good binding with their key targets, indicating NX may exert protective effect through multicompounds and multitargets. In conclusion, NX had a protective effect on HF through cellular and mitochondrial protection against chronic hypoxia via multicompounds, multitargets, and multipathways, and its mechanism may be involved in modulating the PI3K-Akt signaling pathway, HIF-1 signaling pathway, AMPK signaling pathway, and MAPK signaling pathway.

Macrophage polarization plays a vital impact in triggering atherosclerosis (AS) progression and regression. Huang-Lian-Jie-Du Decoction (HLJDD), a famous traditional Chinese decoction, displays notable anti-inflammatory and lipid-lowering effects in different animal models. However, its effects and mechanisms on AS have not been clearly defined. We determined whether HLJDD attenuated atherosclerosis and plaques vulnerability by regulating macrophage polarization in ApoE−/− mice induced by high-fat diet (HFD). Furthermore, we investigated the effects of HLJDD on macrophage polarization in oxidized low-density lipoprotein (ox-LDL) induced RAW264.7 cells. For in vivo assay, compared with the model group, HLJDD ameliorated lipid metabolism, with significantly decreased levels of serum triglyceride, total cholesterol (CHOL), and lipid density lipoprotein. HLJDD suppressed serum tumor necrosis factor α (TNF-α) and IL-1β levels with increased serum IL-10 level, and inhibited mRNA level of NLRP3 inflammasome in carotid tissues. HLJDD enhanced carotid lesion stability by decreasing macrophage infiltration together with increased expression of collagen fibers and α-SMA. Moreover, HLJDD inhibited M1 macrophage polarization, which decreased the expression and mRNA levels of M1 markers [inducible nitric oxide synthase (iNOS) and CD86]. HLJDD enhanced alternatively activated macrophage (M2) activation, which increased the expression and mRNA levels of M2 markers (Arg-1 and CD163). For in vitro assay, HLJDD inhibited foam cell formation in RAW264.7 macrophages disturbed by ox-LDL. Besides, groups with ox-LDL plus HLJDD drug had a lower expression of CD86 and mRNA levels of iNOS, CD86, and IL-1β, but higher expression of CD163 and mRNA levels of Arg-1, CD163, and IL-10 than ox-LDL group. Collectively, our results revealed that HLJDD alleviated atherosclerosis and promoted plaque stability by suppressing M1 polarization and enhancing M2 polarization.

Drynariae Rhizoma (DR) has been demonstrated to be effective in promoting fracture healing in clinical use. In the study, we tried to predicate potential signaling pathways and active ingredients of DR via network pharmacology, uncover its regulation mechanism to improve large bone defects by in vivo and in vitro experiment. We total discovered 18 potential active ingredients such as flavonoids and 81 corresponding targets, in which mitogen-activated protein kinase (MAPK) signaling pathway has the highest correlation with bone defects in pathway and functional enrichment analysis. Therefore, we hypothesized that flavonoids in DR improve large bone defects by activating MAPK signaling pathway. Animal experiments were carried out and all rats randomly divided into TFDR low, medium, and high dosage group, model group and control group. 12 weeks after treatment, according to X-ray and Micro-CT, TFDR medium dosage group significantly promote new bone mineralization compared with other groups. The results of HE and Masson staining and in vitro ALP level of BMSC also demonstrated the formation of bone matrix and mineralization in the TFDR groups. Also, angiographic imaging suggested that flavonoids in DR promoting angiogenesis in the defect area. Consistently, TFDR significantly enhanced the expression of BMP-2, RUNX-2, VEGF, HIF-1 in large bone defect rats based on ELISA and Real-Time PCR. Overall, we not only discover the active ingredients of DR in this study, but also explained how flavonoids in DR regulating MAPK signaling pathway to improve large bone defects.