Hongwei Gao1, Naixin Kang2, Chao Hu2, Ziyu Zhang2, Qiongming Xu2, Yanli Liu3, Shilin Yang1. 1. College of Pharmaceutical Science, Soochow University, Suzhou 215123, China; College of Pharmacy, Guangxi University of Chinese Medicine, Nanning 530000, China. 2. College of Pharmaceutical Science, Soochow University, Suzhou 215123, China. 3. College of Pharmaceutical Science, Soochow University, Suzhou 215123, China. Electronic address: liuyanli@suda.edu.cn.
Abstract
BACKGOUND: Ginsenoside Rb1, the main active constituent of Panax ginseng, displays significant anti-inflammatory activity, although the mechanism has not been clearly unraveled. In this study, Rb1's mechanism of anti-inflammatory effects were investigated. METHODS: The flow cytometry and enzyme-linked immunosorbent assay (ELISA) were empolyed to detect pro-inflammatory cytokines release. The related protein and gene expression was investigated by western blotting and qRT-PCR. The dimerization of TLR4 was measured by co-immunoprecipitation and molecular docking assays. Cellular thermal shift assay was used for the determination of the binding of Rb1 and TLR4. For animal moldels, LPS- or cantharidin-induced acute kidney injury, LPS-induced septic death, and dimethyl benzene-induced ear edema were employed to investigate Rb1's anti-inflammatory activity in vivo. RESULTS: Rb1 significantly decreased inflammatory cytokines release in LPS-stimulated RAW264.7 cells and BMDMs, as well as COX-2 and iNOS amounts. Rb1 reduced LPS-associated calcium influx, ROS production, and NO generation. The NF-κB and MAPK axes participated in Rb1's anti-inflammatory effects. Molecular docking simulation indicated Rb1 bound to TLR4 to prevent TLR4 dimerization, as confirmed by co-immunoprecipitation and cellular thermal shift assay. Furthermore, MyD88 recruitment and TAK1 expression were altered by reduced TLR4 dimerization, indicating the TLR4-MyD88-NF-κB/MAPK pathways contributed to Rb1's anti-inflammatory process. In animal models, Rb1 markedly alleviated LPS- or cantharidin-induced acute kidney injury, rescued LPS-induced septic mice from death, and inhibited dimethyl benzene-induced mouse ear edema. CONCLUSION: Overall, these findings demonstrate Rb1 exhibits marked anti-inflammatory effects, suggesting Rb1 represents an optimal molecule for treating inflammatory diseases.
BACKGOUND: Ginsenoside Rb1, the main active constituent of Panax ginseng, displays significant anti-inflammatory activity, although the mechanism has not been clearly unraveled. In this study, Rb1's mechanism of anti-inflammatory effects were investigated. METHODS: The flow cytometry and enzyme-linked immunosorbent assay (ELISA) were empolyed to detect pro-inflammatory cytokines release. The related protein and gene expression was investigated by western blotting and qRT-PCR. The dimerization of TLR4 was measured by co-immunoprecipitation and molecular docking assays. Cellular thermal shift assay was used for the determination of the binding of Rb1 and TLR4. For animal moldels, LPS- or cantharidin-induced acute kidney injury, LPS-induced septic death, and dimethyl benzene-induced ear edema were employed to investigate Rb1's anti-inflammatory activity in vivo. RESULTS: Rb1 significantly decreased inflammatory cytokines release in LPS-stimulated RAW264.7 cells and BMDMs, as well as COX-2 and iNOS amounts. Rb1 reduced LPS-associated calcium influx, ROS production, and NO generation. The NF-κB and MAPK axes participated in Rb1's anti-inflammatory effects. Molecular docking simulation indicated Rb1 bound to TLR4 to prevent TLR4 dimerization, as confirmed by co-immunoprecipitation and cellular thermal shift assay. Furthermore, MyD88 recruitment and TAK1 expression were altered by reduced TLR4 dimerization, indicating the TLR4-MyD88-NF-κB/MAPK pathways contributed to Rb1's anti-inflammatory process. In animal models, Rb1 markedly alleviated LPS- or cantharidin-induced acute kidney injury, rescued LPS-induced septic mice from death, and inhibited dimethyl benzene-induced mouse ear edema. CONCLUSION: Overall, these findings demonstrate Rb1 exhibits marked anti-inflammatory effects, suggesting Rb1 represents an optimal molecule for treating inflammatory diseases.
Authors: Muhammad Naeem; Muhammad Omer Iqbal; Humaira Khan; Muhammad Masood Ahmed; Muhammad Farooq; Muhammad Moeen Aadil; Mohamad Ikhwan Jamaludin; Abu Hazafa; Wan-Chi Tsai Journal: Molecules Date: 2022-05-25 Impact factor: 4.927