Induction of Intracellular Ca(2+) and pH Changes in Sf9 Insect Cells by Rhodojaponin-III, A Natural Botanic Insecticide Isolated from Rhododendron molle

Many studies on intracellular calcium ([Ca(2+)](i)) and intracellular pH (pH(i)) have been carried out due to their importance in regulation of different cellular functions. However, most of the previous studies are focused on human or mammalian cells. The purpose of the present study was to characterize the effect of Rhodojaponin-III (R-III) on [Ca(2+)](i) and pH(i) and the proliferation of Sf9 cells. R-III strongly inhibited Sf9 cells proliferation with a time- and dose-dependent manner. Flow cytometry established that R-III interfered with Sf9 cells division and arrested them in G2/M. By using confocal scanning technique, effects of R-III on intracellular free calcium ([Ca(2+)](i)) and intracellular pH (pH(i)) in Sf9 cells were determined. R-III induced a significant dose-dependent (1, 10, 100, 200 μg/mL) increase in [Ca(2+)](i) and pH(i) of Sf9 cells in presence of Ca(2+)-containing solution (Hanks) and an irreversible decrease in the absence of extra cellular Ca(2+). We also found that both extra cellular Ca(2+) and intracellular Ca(2+) stores contributed to the increase of [Ca(2+)](i), because completely treating Sf9 cells with CdCl(2) (5 mM), a Ca(2+) channels blocker, R-III (100 μg/mL) induced a transient elevation of [Ca(2+)](i) in case of cells either in presence of Ca(2+) containing or Ca(2+) free solution. In these conditions, pH(i) showed similar changes with that of [Ca(2+)](i) on the whole. Accordingly, we supposed that there was a certain linkage for change of [Ca(2+)](i), cell cycle arrest, proliferation inhibition in Sf9 cells induced by R-III.