title: Knockdown of hypoxia-inducible factor-1 alpha reduces proliferation, induces apoptosis and attenuates the aggressive phenotype of retinoblastoma WERI-Rb-1 cells under hypoxic conditions.
abstract: BACKGROUND
  Hypoxia-inducible factor-1 alpha (HIF-1α) plays a critical role in tumor cell adaption to hypoxia by inducing the transcription of numerous genes. The role of HIF-1α in malignant retinoblastoma remains unclear. We analyzed the role of HIF-1α in WERI-Rb-1 retinoblastoma cells under hypoxic conditions.
  
  
  METHODS
  CoCl2 (125 mmol/L) was added to the culture media to mimic hypoxia. HIF-1α was silenced using siRNA. Gene and protein expression were measured by semi-quantitative RT-PCR and Western blotting. Cell cycle and apoptosis were analyzed by flow cytometry. Cell proliferation, adhesion and invasion were assayed using MTT, Transwell invasion, and cell adhesion assays respectively.
  
  
  RESULTS
  Hypoxia significantly upregulated HIF-1α protein expression and the HIF-1α target genes VEGF, GLUT1, and Survivin mRNA. HIF-1α mRNA expression was not affected by hypoxia. Transfection of the siRNA expression plasmid pRNAT-CMV3.2/Neo-HIF-1α silenced HIF-1α by approximately 80% in hypoxic WERI-Rb-1 cells. The knockdown of HIF-1α under hypoxic conditions downregulated VEGF, GLUT1, and Survivin mRNA. It also inhibited proliferation, promoted apoptosis, induced the G0/G1 phase cell cycle arrest, and reduced the adhesion and invasion of WERI-Rb-1 cells.
  
  
  CONCLUSIONS
  HIF-1α plays a major role in the survival and aggressive phenotype of retinoblastoma cells under hypoxic conditions. Targeting HIF-1α may be a promising therapeutic strategy for human malignant retinoblastoma.
authors.affiliation: Department of Ophthalmology, the First Affiliated Hospital, Zhengzhou University, Zhengzhou, China. No.1 Jianshe East Road, Zhengzhou, 450052, China; .phone: 008613673666718; zhuyuxt@163.com.
authors.forename: Tian
authors.initials: T
authors.lastname: Xia
authors.forename: Hao
authors.initials: H
authors.lastname: Cheng
authors.forename: Yu
authors.initials: Y
authors.lastname: Zhu
mesh_terms: D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D002448:Cell Adhesion / Q000187:drug effects / Q000235:genetics; D002453:Cell Cycle / Q000187:drug effects / Q000235:genetics; D015687:Cell Hypoxia / Q000187:drug effects / Q000235:genetics; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D055785:Gene Knockdown Techniques; D051272:Glucose Transporter Type 1 / Q000235:genetics / Q000378:metabolism; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism*; D051034:Inhibitor of Apoptosis Proteins / Q000235:genetics / Q000378:metabolism; D009361:Neoplasm Invasiveness; D010641:Phenotype; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012175:Retinoblastoma / Q000235:genetics / Q000473:pathology*; D000077022:Survivin; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000378:metabolism
chemical_list: C107489:BIRC5 protein, human; D051272:Glucose Transporter Type 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D051034:Inhibitor of Apoptosis Proteins; D012333:RNA, Messenger; C495677:SLC2A1 protein, human; D000077022:Survivin; D042461:Vascular Endothelial Growth Factor A
keywords: HIF-1α; knockdown of HIF-1α; retinoblastoma; siRNA