Study On Mitochondrial Damage Effect Of Ampelopsin Sodium In Tumor Cells In Vitro

To make clear the anti-tumor targets and mechanisms of Amp-Na, the experiment studied the effects of ampelopsin sodium (Amp-Na) on proliferation of B16 and SPC-A-1 cells and on morphological and mitochondrial function changes of SPC-A-1.The tumor cells was divided in groups treated with Amp-Na alone or combined with Carboplatin. The inhibitory effect on proliferation of tumor cells was examined by MTT colorimetric assay. The changes of cell ultramicrostructure were observed under electron microscope. The changes of mitochondrial membrane potential and intracellular calcium concentration were measured by laser scanning confocal microscope (LSCM). The levels of NO was measured by nitrate reductase method and total ATPase activity was examined by inorganic phosphate in mitochondrial of tumor cells.Results showed that:1. The proliferation of SPC-A-1 cells was inhibited by Amp-Na 6.25～200μg/ml the in concentration-dependent manner. However, the proliferation of B16 cells was inhibited only in high concentration and synergistic effect of Amp-Na was not observed as combined with carboplatin.2. The electron microscope analysis showed morphological changes of SPC-A-1 cells induced by Amp-Na, such as vacuoles in cytoplasm, mitochondria swelling, nuclear membrane shrinkage, and chromatin margination, etc.3. LSCM analysis showed that mitochondrial membrane potential (ΔΨm) of SPC-A-1 cells was decreased after 30 min without concentration-effect relationship and 48 hours with an obvious concentration-effect relationship treated with Amp-Na. Amp-Na also increased intracellular calcium concentration after 30 min and 48 h concentration-independently. 4. Themitochondrial NO level in SPC-A-1 cells was increased with concentration-dependent manner,while the total ATPase activity was decreased by Amp-Na.The above results suggest that Amp-Na injuries mitochondria and induces apoptosis of tumor cells by mechanisms as decreasing mitochondrial membrane potential (ΔΨm) , inducing overloading of intracellular free calcium, increasing mitochondrial NO level, and decreasing total ATPase activity...