title: Growth and differentiation of human lens epithelial cells in vitro on matrix.
abstract: OBJECTIVE
  To characterize the growth and maturation of nonimmortalized human lens epithelial (HLE) cells grown in vitro.
  
  
  METHODS
  HLE cells, established from 18-week prenatal lenses, were maintained on bovine corneal endothelial (BCE) extracellular matrix (ECM) in medium supplemented with basic fibroblast growth factor (FGF-2). The identity, growth, and differentiation of the cultures were characterized by karyotyping, cell morphology, and growth kinetics studies, reverse transcription-polymerase chain reaction (RT-PCR), immunofluorescence, and Western blot analysis.
  
  
  RESULTS
  HLE cells had a male, human diploid (2N = 46) karyotype. The population-doubling time of exponentially growing cells was 24 hours. After 15 days in culture, cell morphology changed, and lentoid formation was evident. Reverse transcription-polymerase chain reaction (RT-PCR) indicated expression of alphaA- and betaB2-crystallin, fibroblast growth factor receptor 1 (FGFR1), and major intrinsic protein (MIP26) in exponential growth. Western analyses of protein extracts show positive expression of three immunologically distinct classes of crystallin proteins (alphaA-, alphaB-, and betaB2-crystallin) with time in culture. By Western blot analysis, expression of p57(KIP2), a known marker of terminally differentiated fiber cells, was detectable in exponential cultures, and levels increased after confluence. MIP26 and gamma-crystallin protein expression was detected in confluent cultures, by using immunofluorescence, but not in exponentially growing cells.
  
  
  CONCLUSIONS
  HLE cells can be maintained for up to 4 months on ECM derived from BCE cells in medium containing FGF-2. With time in culture, the cells demonstrate morphologic characteristics of, and express protein markers for, lens fiber cell differentiation. This in vitro model will be useful for investigations of radiation-induced cataractogenesis and other studies of lens toxicity.
authors.affiliation: Life Sciences Division, Lawrence Berkeley National Laboratory, Berkeley. Cellrex, San Francisco, USA. eablakely@lbl.gov
authors.forename: E A
authors.initials: EA
authors.lastname: Blakely
authors.forename: K A
authors.initials: KA
authors.lastname: Bjornstad
authors.forename: P Y
authors.initials: PY
authors.lastname: Chang
authors.forename: M P
authors.initials: MP
authors.lastname: McNamara
authors.forename: E
authors.initials: E
authors.lastname: Chang
authors.forename: G
authors.initials: G
authors.lastname: Aragon
authors.forename: S P
authors.initials: SP
authors.lastname: Lin
authors.forename: G
authors.initials: G
authors.lastname: Lui
authors.forename: J R
authors.initials: JR
authors.lastname: Polansky
mesh_terms: D000818:Animals; D020346:Aquaporins; D015153:Blotting, Western; D002417:Cattle; D002454:Cell Differentiation; D002478:Cells, Cultured; D003459:Crystallins / Q000235:genetics / Q000378:metabolism; D050761:Cyclin-Dependent Kinase Inhibitor p57; D004728:Endothelium, Corneal / Q000166:cytology; D004847:Epithelial Cells / Q000166:cytology* / Q000378:metabolism; D005109:Extracellular Matrix; D005136:Eye Proteins / Q000235:genetics / Q000378:metabolism; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D007621:Karyotyping; D007908:Lens, Crystalline / Q000166:cytology* / Q000378:metabolism; D008562:Membrane Glycoproteins; D008856:Microscopy, Fluorescence; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism; D051496:Receptor, Fibroblast Growth Factor, Type 1; D017468:Receptors, Fibroblast Growth Factor / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D020346:Aquaporins; C493964:CDKN1C protein, human; D003459:Crystallins; D050761:Cyclin-Dependent Kinase Inhibitor p57; D005136:Eye Proteins; D008562:Membrane Glycoproteins; D009687:Nuclear Proteins; D012333:RNA, Messenger; D017468:Receptors, Fibroblast Growth Factor; C030328:aquaporin 0; D016222:Fibroblast Growth Factor 2; C496348:FGFR1 protein, human; D020794:Receptor Protein-Tyrosine Kinases; D051496:Receptor, Fibroblast Growth Factor, Type 1