Fig. 7.

Cx50 hemichannels mediate uptake of GSH and protect cells against oxidative stress induced by H₂O₂. CEF cells were infected with high-titer recombinant RCAS(A) retroviruses containing Cx50 or the Cx50 H156N mutant. (A) Cells were treated with 1 mM GSH for various periods of time then rinsed and labeled with fluorescent Thioltracker™. The intracellular GSH level was quantified by determining the fluorescence intensity. (B,C) Cells were pretreated with 1 mM GSH (B) or 1 mM DTT (C) for 20 min, and in some groups GSH or DTT was removed (preGSH or preDTT) before the treatment with 1 mM H₂O₂. Cells under apoptosis were analyzed by using the Dead Cell Apoptosis Kit with FITC–Annexin V and PI (BioLegend, San Diego, CA). At least five microphotographs of fluorescence fields were taken under a 10× microscope (Olympus) with under phase-contrast or with FITC or Rhodamine filters. Positive cells were quantified by counting 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.