title: Comparison of gene expression profiles in primary and immortalized human pterygium fibroblast cells.
abstract: OBJECTIVE
  Pterygium is a fibrovascular growth on the ocular surface with corneal tissue destruction, matrix degradation and varying extents of chronic inflammation. To facilitate investigation of pterygium etiology, we immortalized pterygium fibroblast cells and profiled their global transcript levels compared to primary cultured cells.
  
  
  METHODS
  Fibroblast cells were cultured from surgically excised pterygium tissue using the explant method and propagated to passage number 2-4. We hypothesized that intervention with 3 critical molecular intermediates may be necessary to propage these cells. Primary fibroblast cells were immortalized sequentially by a retroviral construct containing the human telomerase reverse transcriptase gene and another retroviral expression vector expressing p53/p16 shRNAs. Primary and immortalized fibroblast cells were evaluated for differences in global gene transcript levels using an Agilent Genechip microarray.
  
  
  RESULTS
  Light microscopic morphology of immortalized cells was similar to primary pterygium fibroblast at passage 2-4. Telomerase reverse transcriptase was expressed, and p53 and p16 levels were reduced in immortalized pterygium fibroblast cells. There were 3308 significantly dysregulated genes showing at least 2 fold changes in transcript levels between immortalized and primary cultured cells (2005 genes were up-regulated and 1303 genes were down-regulated). Overall, 13.58% (95% CI: 13.08-14.10) of transcripts in immortalized cells were differentially expressed by at least 2 folds compared to primary cells.
  
  
  CONCLUSIONS
  Pterygium primary fibroblast cells were successfully immortalized to at least passage 11. Although a variety of genes are differentially expressed between immortalized and primary cells, only genes related to cell cycle are significantly changed, suggesting that the immortalized cells may be used as an in vitro model for pterygium pathology.
authors.affiliation: Ocular Surface Research Group, Singapore Eye Research Institute, Singapore.
authors.forename: Aihua
authors.initials: A
authors.lastname: Hou
authors.forename: P Mathijs
authors.initials: PM
authors.lastname: Voorhoeve
authors.forename: Wanwen
authors.initials: W
authors.lastname: Lan
authors.forename: Minqi
authors.initials: M
authors.lastname: Tin
authors.forename: Louis
authors.initials: L
authors.lastname: Tong
mesh_terms: D015153:Blotting, Western; D002460:Cell Line; D005347:Fibroblasts / Q000166:cytology*; D005786:Gene Expression Regulation; D057809:HEK293 Cells; D006801:Humans; D011625:Pterygium / Q000235:genetics* / Q000473:pathology; D060888:Real-Time Polymerase Chain Reaction; D046888:Tissue Array Analysis; D059467:Transcriptome
keywords: Eye; Immortalization; Pterygium fibroblast