title: Downregulation of β-actin gene and human antigen R in human keratoconus.
abstract: OBJECTIVE
  The purpose of this study was to determine the expression levels and regulation of β-actin in the stroma of keratoconus (KC) and normal corneas.
  
  
  METHODS
  A total of 15 different human corneas from both KC and normal individuals were used for this study. Additionally, 3 Fuch's dystrophic corneas were also used. The β-actin gene expression was analyzed at the transcriptional and translational levels in the epithelium and stroma of the KC and normal corneas. The human antigen R (HuR) gene expression was analyzed by real-time PCR in the stroma of five KC and five normal corneas. The keratocytes from three normal and three KC corneas were cultured in the presence of serum, and the expression levels of β-actin and human antigen R (HuR) were analyzed by using confocal imaging in both normal and KC fibroblasts.
  
  
  RESULTS
  The expression of the β-actin gene was downregulated in the stroma of the six KC corneas but not in the stroma of six normal and Fuchs' dystrophic corneas. Immunofluorescence detection of β-actin showed that it was absent in the KC fibroblast. The real-time PCR analysis of the HuR gene showed a relative 4.7-fold lower expression in KC corneas relative to the normal corneas, which was further confirmed by the immunofluorescence detection of HuR in fibroblasts of KC corneas.
  
  
  CONCLUSIONS
  Although ubiquitous β-actins are essential for cell survival during early embryogenesis, the effects on various stages of development are not well understood. Our results show that β-actin is downregulated in the corneal stroma of patients with KC, which may be related to reduced levels of a stabilizing factor (HuR) for β-actin mRNA. We propose that loss of β-actin in the corneal stroma might be a triggering factor in the development of KC.
authors.affiliation: Department of Vision Sciences, University of Alabama at Birmingham, Birmingham, Alabama 35294-4390, USA.
authors.forename: Roy
authors.initials: R
authors.lastname: Joseph
authors.forename: Om P
authors.initials: OP
authors.lastname: Srivastava
authors.forename: Roswell R
authors.initials: RR
authors.lastname: Pfister
mesh_terms: D000199:Actins / Q000096:biosynthesis / Q000235:genetics*; D015153:Blotting, Western; D002478:Cells, Cultured; D003319:Corneal Stroma / Q000378:metabolism* / Q000473:pathology; D015536:Down-Regulation; D051959:ELAV Proteins / Q000096:biosynthesis / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D005347:Fibroblasts / Q000378:metabolism / Q000473:pathology; D005786:Gene Expression Regulation; D006801:Humans; D007150:Immunohistochemistry; D007640:Keratoconus / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D008297:Male; D018613:Microscopy, Confocal; D008875:Middle Aged; D012333:RNA, Messenger / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D000199:Actins; D051959:ELAV Proteins; D012333:RNA, Messenger