PMID- 32062328 OWN - NLM STAT- MEDLINE DCOM- 20200713 LR - 20200713 IS - 1618-095X (Electronic) IS - 0944-7113 (Linking) VI - 68 DP - 2020 Mar TI - Wogonoside alleviates colitis by improving intestinal epithelial barrier function via the MLCK/pMLC2 pathway. PG - 153179 LID - S0944-7113(20)30012-X [pii] LID - 10.1016/j.phymed.2020.153179 [doi] AB - BACKGROUND: Intestinal epithelial barrier dysfunction, which involves myosin light chain kinase (MLCK) activation, contributes to the occurrence and progression of inflammation in inflammatory bowel disease (IBD). Wogonoside helps maintain intestinal homeostasis in mice with dextran sulfate sodium (DSS)-induced colitis, but it is unclear whether it modulates intestinal barrier function. PURPOSE: Here, we demonstrate that wogonoside protects against intestinal barrier dysfunction in colitis via the MLCK/pMLC2 pathway both in vivo and in vitro. METHODS: Caco-2 cell monolayers treated with the proinflammatory cytokine TNF-alpha showed barrier dysfunction and were assessed in the absence and presence of wogonoside for various physiological, morphological, and biochemical parameters. Colitis was induced by 3% DSS in mice, which were used as an animal model to explore the pharmacodynamics of wogonoside. We detected MLCK/pMLC2 pathway proteins via western blot analysis, assessed the cytokines IL-13 and IFN-gamma via ELISA, tested bacterial translocation via fluorescence in situ hybridization (FISH) and a proper sampling of secondary lymphoid organs for bacterial culture. In addition, the docking affinity of wogonoside and MLCK was observed with DS2.5 software. RESULTS: Wogonoside alleviated the disruption of transepithelial electrical resistance (TER) in TNF-alpha exposured Caco-2 cell; FITC-dextran hyperpermeability; loss of the tight junction (TJ) proteins occludin, ZO-1 and claudin-1 in Caco-2 cell monolayers; and bacterial translocation in colitic mice. Moreover, wogonoside reduced the levels of the proinflammatory cytokines IL-13 and IFN-gamma to maintain intestinal immune homeostasis. Transmission electron microscopy (TEM) confirmed that wogonoside ameliorated the destruction of intestinal epithelial TJs. Wogonoside not only inhibited the cytoskeletal F-actin rearrangement induced by TNF-alpha, stabilized the cytoskeletal structure, suppressed MLCK protein expression, and reduced MLC2 phosphorylation. In addition, the results of molecular docking analysis showed that wogonoside had a high affinity for MLCK and formed hydrogen bonds with the amino acid residue LYS261 and pi bonds with LYS229. CONCLUSION: Collectively, our study indicates that wogonoside alleviates colitis by protecting against intestinal barrier dysfunction, and the potential mechanism may involve regulation of TJs via the MLCK/pMLC2 signaling pathway. Meanwhile, our study also explains the success of S. baicalensis in the treatment of ulcerative colitis (UC). CI - Copyright (c) 2020 Elsevier GmbH. All rights reserved. FAU - Huang, Shaowei AU - Huang S AD - School of Pharmaceutical Sciences, Guangzhou University of Chinese Medicine, Guangzhou, China. FAU - Fu, Yajun AU - Fu Y AD - School of Pharmaceutical Sciences, Guangzhou University of Chinese Medicine, Guangzhou, China. FAU - Xu, Bo AU - Xu B AD - School of Pharmaceutical Sciences, Guangzhou University of Chinese Medicine, Guangzhou, China. FAU - Liu, Chang AU - Liu C AD - School of Pharmaceutical Sciences, Guangzhou University of Chinese Medicine, Guangzhou, China. FAU - Wang, Qing AU - Wang Q AD - School of Pharmaceutical Sciences, Guangzhou University of Chinese Medicine, Guangzhou, China. FAU - Luo, Shuang AU - Luo S AD - School of Pharmaceutical Sciences, Guangzhou University of Chinese Medicine, Guangzhou, China. FAU - Nong, Feifei AU - Nong F AD - School of Pharmaceutical Sciences, Guangzhou University of Chinese Medicine, Guangzhou, China. FAU - Wang, Xiaojing AU - Wang X AD - School of Pharmaceutical Sciences, Guangzhou University of Chinese Medicine, Guangzhou, China. FAU - Huang, Songyu AU - Huang S AD - School of Pharmaceutical Sciences, Guangzhou University of Chinese Medicine, Guangzhou, China. FAU - Chen, Jinyan AU - Chen J AD - School of Basic Medicine, Guangzhou University of Chinese Medicine, Guangzhou, China. FAU - Zhou, Lian AU - Zhou L AD - School of Pharmaceutical Sciences, Guangzhou University of Chinese Medicine, Guangzhou, China. Electronic address: zl@gzucm.edu.cn. FAU - Luo, Xia AU - Luo X AD - School of Pharmaceutical Sciences, Guangzhou University of Chinese Medicine, Guangzhou, China. Electronic address: luoxia@gzucm.edu.cn. LA - eng PT - Journal Article DEP - 20200203 PL - Germany TA - Phytomedicine JT - Phytomedicine : international journal of phytotherapy and phytopharmacology JID - 9438794 RN - 0 (Flavanones) RN - 0 (Glucosides) RN - 0 (Myosin Light Chains) RN - 0 (Tight Junction Proteins) RN - 0 (myosin light chain 2) RN - 9042-14-2 (Dextran Sulfate) RN - EC 2.7.11.18 (Myosin-Light-Chain Kinase) RN - EC 3.6.1.- (Cardiac Myosins) RN - ETX4944Z3R (wogonoside) SB - IM MH - Animals MH - Caco-2 Cells MH - Cardiac Myosins/*metabolism MH - Colitis/chemically induced/*drug therapy/metabolism MH - Dextran Sulfate/toxicity MH - Flavanones/chemistry/*pharmacology MH - Glucosides/chemistry/*pharmacology MH - Humans MH - Intestinal Mucosa/metabolism MH - Male MH - Mice, Inbred C57BL MH - Molecular Docking Simulation MH - Myosin Light Chains/*metabolism MH - Myosin-Light-Chain Kinase/*metabolism MH - Phosphorylation MH - Tight Junction Proteins/metabolism OTO - NOTNLM OT - Colitis OT - Intestinal epithelial barrier OT - MLCK/pMLC2 pathway OT - Wogonoside COIS- Declaration of Competing Interest There are no conflicts of interest to declare. EDAT- 2020/02/18 06:00 MHDA- 2020/07/14 06:00 CRDT- 2020/02/17 06:00 PHST- 2019/05/06 00:00 [received] PHST- 2020/01/20 00:00 [revised] PHST- 2020/02/02 00:00 [accepted] PHST- 2020/02/18 06:00 [pubmed] PHST- 2020/07/14 06:00 [medline] PHST- 2020/02/17 06:00 [entrez] AID - S0944-7113(20)30012-X [pii] AID - 10.1016/j.phymed.2020.153179 [doi] PST - ppublish SO - Phytomedicine. 2020 Mar;68:153179. doi: 10.1016/j.phymed.2020.153179. Epub 2020 Feb 3.