Fig. 6.

Treatment with H₂O₂ increased apoptosis and cell death of lens fiber cells, and this increase was augmented in lens fiber cells with impairment of Cx50 and Cx46 hemichannels. (A) Primary chick lens cell cultures were infected with high-titer RCAS(A) retroviral vehicle (V) or recombinant RCAS(A) retroviruses containing Cx50, and Cx50 mutants E48K, P88S or H156N. After 10–11 days of cell culturing, cells were treated with 0.2 mM H₂O₂ for 4 h and analyzed by using the FITC–Annexin V Apoptosis Detection Kit with PI (BioLegend). At least five microphotographs of fluorescence fields were taken under a 20× microscope (Olympus) under phase-contrast or with FITC or Rhodamine filters. Scale bar: 100 μm. (B) Positive cell areas were quantified by means of ImageJ software and are shown as a percentage. The data are presented as the mean±s.e.m. (n=3). *P<0.05; **P<0.01; ***P<0.001.