title: Proinflammatory cytokine-induced NF-kappaB activation in human mesangial cells is mediated through intracellular calcium but not ROS: effects of silymarin.
abstract: BACKGROUND
  It is not fully understood whether intracellular calcium and/or reactive oxygen species (ROS) are involved in nuclear factor-kappaB (NF-kappaB) activation by proinflammatory cytokines. Silymarin exhibits anti-inflammatory and antioxidant effects but the effect of silymarin in human mesangial cells is largely unknown.
  
  
  METHODS
  NF-kappaB binding activity was measured by electrophoretic mobility shift assay. Intracellular calcium was monitored by confocal microscopy using Fluo-3 and intracellular ROS production was determined by flow cytometry. Monocyte chemoattractant protein-1 (MCP-1) expression was measured by Northern blot analysis and ELISA.
  
  
  RESULTS
  NF-kappaB was activated within 30 min by tumor necrosis factor-alpha (TNF-alpha) or interleukin-1beta (IL-1beta). Intracellular ROS was not produced until 30 min and also antioxidants such as N-acetylcysteine and tiron had no effect on the TNF-alpha- or IL-1beta-induced NF-kappaB activation. Intracellular calcium was increased by TNF-alpha and IL-1beta. Furthermore, a calcium chelator, BAPTA-AM, attenuated the NF-kappaB activation. Silymarin dose-dependently inhibited the TNF-alpha- or IL-1beta-induced NF-kappaB activation and MCP-1 expression. Silymarin also inhibited TNF-alpha-induced intracellular calcium.
  
  
  CONCLUSIONS
  Induction of NF-kappaB within 30 min by TNF-alpha- and IL-1beta was mediated through intracellular calcium but not ROS. Silymarin inhibited TNF-alpha-induced calcium-dependent NF-kappaB activation irrespective of its antioxidant effect.
authors.affiliation: Department of Internal Medicine, Urology and Asan Institute for Life Sciences, College of Medicine, University of Ulsan, Seoul, Korea.
authors.forename: Jai Wong
authors.initials: JW
authors.lastname: Chang
authors.forename: Choung Soo
authors.initials: CS
authors.lastname: Kim
authors.forename: Soon Bae
authors.initials: SB
authors.lastname: Kim
authors.forename: Su Kil
authors.initials: SK
authors.lastname: Park
authors.forename: Jung Sik
authors.initials: JS
authors.lastname: Park
authors.forename: Sang Koo
authors.initials: SK
authors.lastname: Lee
mesh_terms: D000975:Antioxidants / Q000494:pharmacology*; D015152:Blotting, Northern; D002118:Calcium / Q000502:physiology*; D018929:Cell Culture Techniques; D024202:Electrophoretic Mobility Shift Assay; D006801:Humans; D007150:Immunohistochemistry; D007249:Inflammation; D007375:Interleukin-1 / Q000502:physiology; D050527:Mesangial Cells / Q000187:drug effects* / Q000502:physiology*; D016328:NF-kappa B / Q000378:metabolism*; D017382:Reactive Oxygen Species; D015398:Signal Transduction / Q000187:drug effects; D012838:Silymarin / Q000494:pharmacology*; D014409:Tumor Necrosis Factor-alpha / Q000502:physiology
chemical_list: D000975:Antioxidants; D007375:Interleukin-1; D016328:NF-kappa B; D017382:Reactive Oxygen Species; D012838:Silymarin; D014409:Tumor Necrosis Factor-alpha; D002118:Calcium