title: Antiproliferative and apoptotic effects of indomethacin on human retinoblastoma cell line Y79 and the involvement of β-catenin, nuclear factor-κB and Akt signaling pathways.
abstract: BACKGROUND
  To determine in vitro if indomethacin inhibits proliferation and induces apoptosis in human retinoblastoma cell line Y79, and to explore possibly involved signaling pathways.
  
  
  METHODS
  The human retinoblastoma cell line Y79 was cultured with indomethacin at various concentrations (0, 25, 50, 100, 200 and 400 µmol/l). The effect of indomethacin on cell proliferation and apoptosis was examined by the Cell Counting Kit-8 and TUNEL test, respectively. The mRNA level of survivin, β-catenin and Bcl-2 was detected by RT-PCR. The protein level of survivin was measured by ELISA. Western blot was used to analyze β-catenin, nuclear factor (NF)-κB/p65, phosphorylated Akt (pAkt) and total Akt (tAkt) expression in cultured cells.
  
  
  RESULTS
  Indomethacin treatment inhibits proliferation (at concentrations from 25 to 400 µmol/l) and induces apoptosis (at concentrations from 100 to 400 µmol/l) of human retinoblastoma cell line Y79 in a dose-dependent manner. RT-PCR showed that the mRNA expression of Bcl-2 (F = 20.497; p < 0.001) and of β-catenin (F = 14.835; p < 0.001) was significantly different among the treated groups. Survivin mRNA levels remained steady, but its protein levels decreased significantly as measured by ELISA (F = 67.633; p < 0.001). Western blot analysis showed a dose-dependent downregulation of β-catenin (F = 37.411; p < 0.001), NF-κB/p65 (F = 16.302; p < 0.001) and of pAkt (F = 27.700; p < 0.001) after indomethacin treatment, while tAkt protein expression was steady among the groups.
  
  
  CONCLUSIONS
  Treatment with indomethacin can potently suppress proliferation and induce apoptosis in the retinoblastoma Y79 cell line. Wnt/β-catenin, NF-κB and Akt/PKB pathways might be implicated in the process.
authors.affiliation: Eye Hospital, Wenzhou Medical College, Wenzhou, China.
authors.forename: Qinxiang
authors.initials: Q
authors.lastname: Zheng
authors.forename: Ying
authors.initials: Y
authors.lastname: Zhang
authors.forename: Yueping
authors.initials: Y
authors.lastname: Ren
authors.forename: Yijun
authors.initials: Y
authors.lastname: Wu
authors.forename: Shelley
authors.initials: S
authors.lastname: Yang
authors.forename: Yuanping
authors.initials: Y
authors.lastname: Zhang
authors.forename: Hao
authors.initials: H
authors.lastname: Chen
authors.forename: Wensheng
authors.initials: W
authors.lastname: Li
authors.forename: Yihua
authors.initials: Y
authors.lastname: Zhu
mesh_terms: D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D049109:Cell Proliferation / Q000187:drug effects*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D020287:In Situ Nick-End Labeling; D007213:Indomethacin / Q000494:pharmacology*; D051034:Inhibitor of Apoptosis Proteins / Q000235:genetics / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D019572:Retinal Neoplasms / Q000378:metabolism / Q000473:pathology*; D012175:Retinoblastoma / Q000378:metabolism / Q000473:pathology*; D000077022:Survivin; D051996:Transcription Factor RelA / Q000378:metabolism; D014407:Tumor Cells, Cultured; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism
chemical_list: D000894:Anti-Inflammatory Agents, Non-Steroidal; C107489:BIRC5 protein, human; C495270:CTNNB1 protein, human; D051034:Inhibitor of Apoptosis Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D000077022:Survivin; D051996:Transcription Factor RelA; D051176:beta Catenin; D007213:Indomethacin