Preventive effect of germanium dioxide on the inhibition of gap junctional intercellular communication by TPA

Gap junctional intercellular communication (GJIC) is thought to be essential for maintaining cellular homeostasis and growth control. In order to detect any protective agent against tumor formation, we examined the anticarcinogenic effect of a germanium dioxide (GeO2) using a model system of GJIC in F344 rat liver epithelial cells, named WB cells. 12-O-tetradecanoylphorbol-13-acetate (TPA), known as tumor promoters, inhibited GJIC in the epithelial cells as determined by the scrape loading/dye transfer (SL/DT) assay. And GeO2 recovered this inhibition of GJIC. Immunostaining of connexin 43 (Cx43) protein in WB cells indicated that TPA caused a loss of Cx43 protein from the cell membranes. However, GeO2 treatment showed re-appearance of Cx43 protein on the membrane. Reverse transcription-polymerase chain reaction (RT-PCR) and Western blots were analyzed to determine whether the test compounds might have altered the steady-state levels of gap junction mRNA and/or connexin protein levels or phosphorylation. The inhibition of GJIC by TPA in WB cells was correlated with the hyperphosphorylation of Cx43 as measured by mobility shifts of the western blot bands of Cx43. TPA induced hyperphosphorylation of Cx43 protein, while GeO2 appeared to partially block this hyperphosphorylation. Here, we showed that pre- and co-incubation with GeO2 in TPA-treated WB-cells abolished down-regulation of GJIC by TPA. These data suggest that GeO2 may inhibit tumor promotion by enhancing GJIC.