title: Inhibition of aldose reductase prevents lipopolysaccharide-induced inflammatory response in human lens epithelial cells.
abstract: OBJECTIVE
  Bacterial infections are one of the major causes of human eye disease. Because the bacterial endotoxin lipopolysaccharide (LPS) is known to cause cytotoxicity through oxidative stress and an earlier study has shown that aldose reductase (AR) mediates oxidative stress signals, the purpose of this study was to investigate the anti-inflammatory effects of AR inhibition on LPS-induced activation of NF-kappaB-dependent signals in human lens epithelial cells (HLECs).
  
  
  METHODS
  Growth-arrested HLECs were cultured without or with AR inhibitors or transfected with an AR small interfering (si)RNA. Subsequently, the cells were stimulated with LPS (1-10 mug/mL) for 24 hours. The cell viability was assessed by cell counts and MTT assay, and apoptosis was measured by nucleosomal degradation. Electrophoretic mobility gel shift assays were performed to determine the activation of NF-kappaB and AP1. The levels of nitric oxide, MMP-2, MMP-9, Cox-2, and TNF-alpha were measured by using specific ELISA kits. Western blot analysis was performed to determine the cleavage of poly(ADP-ribose) polymerase (PARP) and the activation of PKC and mitogen-activated protein kinase (MAPK).
  
  
  RESULTS
  Bacterial LPS caused apoptosis of HLECs. Inhibition of AR by two structurally unrelated inhibitors, sorbinil and tolrestat, or ablation by AR siRNA prevented the LPS-induced apoptosis, activation of caspase-3 and cleavage of PARP protein. Inhibition of AR in HLECs also prevented the LPS-induced activation of redox-sensitive transcription factors such as NF-kappaB and AP1 and their downstream signals that lead to expression of Cox-2, MMP-2, MMP-9, and TNF-alpha proteins. In addition, inhibition of AR prevented LPS-induced activation of protein kinases upstream to NF-kappaB activation such as PKC and MAPK in HLECs.
  
  
  CONCLUSIONS
  The results indicate that AR mediates the bacterial endotoxin signaling that could damage HLECs by regulating the signals that activate the redox-sensitive transcription factor NF-kappaB and cause inflammation. Thus, inhibition of AR could be a therapeutic target for Gram-negative bacterial infection-induced visual complications.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Texas Medical Branch, Galveston, Texas 77555-0647, USA.
authors.forename: Agnieszka
authors.initials: A
authors.lastname: Pladzyk
authors.forename: Aramati B M
authors.initials: AB
authors.lastname: Reddy
authors.forename: Umesh C S
authors.initials: UC
authors.lastname: Yadav
authors.forename: Ravinder
authors.initials: R
authors.lastname: Tammali
authors.forename: Kota V
authors.initials: KV
authors.lastname: Ramana
authors.forename: Satish K
authors.initials: SK
authors.lastname: Srivastava
mesh_terms: D000449:Aldehyde Reductase / Q000037:antagonists & inhibitors*; D015153:Blotting, Western; D053148:Caspase 3 / Q000378:metabolism; D024161:Collagen Type XI / Q000378:metabolism; D051546:Cyclooxygenase 2 / Q000378:metabolism; D015232:Dinoprostone / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004797:Enzyme-Linked Immunosorbent Assay; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D048289:Imidazolidines / Q000494:pharmacology; D007223:Infant; D007908:Lens, Crystalline / Q000187:drug effects* / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology*; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D009281:Naphthalenes / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
chemical_list: C499319:COL11A2 protein, human; D024161:Collagen Type XI; D004791:Enzyme Inhibitors; D048289:Imidazolidines; D008070:Lipopolysaccharides; D016328:NF-kappa B; D009281:Naphthalenes; D014409:Tumor Necrosis Factor-alpha; C040550:tolrestat; D009569:Nitric Oxide; D000449:Aldehyde Reductase; D051546:Cyclooxygenase 2; D020929:Mitogen-Activated Protein Kinase Kinases; D053148:Caspase 3; D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9; C026411:sorbinil; D015232:Dinoprostone