title: Molecular mechanisms of rosmarinic acid from Salvia miltiorrhiza in acute lymphoblastic leukemia cells.
abstract: BACKGROUND
  Rosmarinic acid (RA), a major hydrosoluble bioactive compound found in the Chinese medicinal herb, Salvia miltiorrhiza Bunge, which has been used in traditional Chinese medicine to treat various diseases, including cancer. However, the mechanisms have not been fully elucidated.
  
  
  OBJECTIVE
  Guided by microarray hybridization and Ingenuity Pathway Analysis, we identified modes of action of rosmarinic acid (RA) isolated from S. miltiorrhiza on acute lymphoblastic leukemia cells.
  
  
  METHODS
  Microarray data were verified by independent methods: Real-time RT-PCR (mRNA expression), resazurin assay (cytotoxicity of RA towards parental CCRF-CEM, multidrug-resistant CEM/ADR5000 cells and normal lymphocytes), flow cytometry (cell cycle arrest, apoptosis, necroptosis, generation of reactive oxygen species (ROS), disruption of mitochondrial membrane potential (MMP)), single cell gel electrophoresis (DNA damage), molecular docking and gene promoter binding motif analysis (NFκB), Western blotting (nuclear NFκB translocation, PARP cleavage, caspase 3/7/9 expression), and fibronectin-based cell adhesion assay.
  
  
  RESULTS
  RA dose-dependently inhibited CCRF-CEM and CEM/ADR5000 cells, but caused less cytotoxicity towards normal lymphocytes. RA simultaneously induced apoptosis and necrosis, as shown by cell morphology and annexin V-PI assay. DNA damage was dose-dependently induced without ROS generation, which subsequently led to cell cycle arrest. RA-stimulated MMP dysfunction activated PARP-cleavage and caspase-independent apoptosis. In accordance with molecular docking and gene promoter binding motif analyses, p65 translocation from the cytosol to the nucleus was blocked by RA, indicating a mechanistic role of the NFκB pathway to explain RA's action. RA affected cellular movement as evaluated by ameliorating cell adhesion to fibronectin.
  
  
  CONCLUSIONS
  RA induced apoptosis and necrosis in a ROS-independent DNA damage and caspase-independent manner. These results may contribute to the rationale use of S. miltiorrhiza and RA in traditional medicine of leukemia.
authors.affiliation: Department of Pharmaceutical Biology, Institute of Pharmacy and Biochemistry, Johannes Gutenberg University, Staudinger Weg 5, 55128 Mainz, Germany.
authors.forename: Ching-Fen
authors.initials: CF
authors.lastname: Wu
authors.affiliation: Department of Pharmaceutical Biology, Institute of Pharmacy and Biochemistry, Johannes Gutenberg University, Staudinger Weg 5, 55128 Mainz, Germany.
authors.forename: Chunlan
authors.initials: C
authors.lastname: Hong
authors.affiliation: Working Group Cancer Genome Research, German Cancer Research Center (DKFZ) and German Cancer Consortium (DKTK), National Center for Tumor Diseases (NCT), Heidelberg, Germany.
authors.forename: Sabine M
authors.initials: SM
authors.lastname: Klauck
authors.affiliation: National Research Institute of Chinese Medicine, Taipei, Taiwan.
authors.forename: Yun-Lian
authors.initials: YL
authors.lastname: Lin
authors.affiliation: Department of Pharmaceutical Biology, Institute of Pharmacy and Biochemistry, Johannes Gutenberg University, Staudinger Weg 5, 55128 Mainz, Germany. Electronic address: efferth@uni-mainz.de.
authors.forename: Thomas
authors.initials: T
authors.lastname: Efferth
mesh_terms: D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D002448:Cell Adhesion / Q000187:drug effects; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002934:Cinnamates / Q000494:pharmacology*; D020552:Comet Assay; D004249:DNA Damage; D053630:Depsides / Q000494:pharmacology*; D018432:Drug Resistance, Multiple; D019008:Drug Resistance, Neoplasm; D020869:Gene Expression Profiling; D006801:Humans; D008214:Lymphocytes / Q000187:drug effects; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D062105:Molecular Docking Simulation; D016328:NF-kappa B / Q000378:metabolism; D054198:Precursor Cell Lymphoblastic Leukemia-Lymphoma / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D027561:Salvia miltiorrhiza; D000097123:Rosmarinic Acid
chemical_list: D000970:Antineoplastic Agents; D002934:Cinnamates; D053630:Depsides; D016328:NF-kappa B; D017382:Reactive Oxygen Species; D000097123:Rosmarinic Acid
keywords: Cell death; Multidrug resistance; NFκB; Pharmacogenomics