Figure 1.

The effect of pharmacological calcium channel blockade on rat MC proliferation and apoptosis in vitro. A: Representative images of BrdU incorporation in primary rat MC co-incubated with 400 μmol/L nickel chloride (NiCl₂). Top panel: No drug and 400 μmol/L NiCl₂ show all nuclei stained with Hoechst. Bottom panel: No drug and 400 μmol/L NiCl₂ show nuclei stained with anti-BrdU antibody. Graphical representation of six independent experiments showed a significant dose-dependent reduction in BrdU incorporation in rat MCs treated with NiCl₂. ^∗∗P < 0.01, ^∗∗∗P < 0.001 versus control cells. B, left panel: A significant reduction in cell number was seen in rat MCs incubated with increasing concentrations of TH1177 for 72 hours (n = 5). ^∗P < 0.05, ^∗∗∗P < 0.001. Right panel: BrdU incorporation was significantly reduced when these cells were exposed to TH1177 at ≥10 μmol/L (n = 3). ^∗P < 0.05 versus vehicle-treated control cells. C: Dose-response curve demonstrated an ED₅₀ of 15 μmol/L for the anti-proliferative action of TH1177 in these conditions. D: Co-incubation of rat MCs with verapamil, an LCCB, had no effect on cell number over 72 hours. E: Representative images of nuclei stained with Hoechst. An example of the apoptotic nucleus is indicated by the arrow in the positive control image. The graph represents four independent experiments demonstrating that nickel (concentration of ≤400 μmol/L) and TH1177 (concentration of ≤20 μmol/L) caused no increase in the proportion of apoptotic nuclei seen compared to vehicle-treated cells.