Figure 6.

Astrocytes sequester cytoplasmic calcium from tumor cells through GJC. (A) Temporal analysis of cytoplasmic calcium (relative Fura-2 fluorescence) and DNA fragmentation (propidium iodide analysis) in A375P cells incubated with paclitaxel (10 ng/ml). Data are expressed relative to the initial (t = 0 hour) values. (B) A375P cells were preincubated with the indicated concentrations of BAPTA-AM before incubation with paclitaxel (10 ng/ml), and the apoptotic index was determined as described earlier. (C) RFP-expressing astrocytes were cocultured with tumor cells and incubated with paclitaxel (10 ng/ml) for 12 hours in the absence or presence of CBX. The cells were then labeledwith Fluo-3AMand harvested, and cytoplasmic calciumlevels in the tumor cells (RFP-negative)were determined by flow cytometry. Data are presented relative to untreated control tumor cells. Note that inhibition of GJC between tumor cells and the astrocytes with CBX (100 µM) resulted in higher cytoplasmic calcium levels compared with paclitaxel-treated tumor cells alone. (D) Proposed working model for astrocytes' protection of tumor cell apoptosis. When tumor cells cultured alonewere treatedwith chemotherapeutic drugs, an accumulation of cytoplasmic calcium (green circles) led to tumor apoptosis. Coculture of tumor cells with astrocytes, however, resulted in GJC channels between these cells that served as conduits for sequestration and, hence, depletion of calcium and other proapoptotic molecules from the tumor cytoplasm. Alternatively, astrocyte-derived survival factors and signals could be transferred to tumor cells, thereby reducing tumor calcium overload.