The Growth-inhibiting And Apoptosis-inducing Effects Of Oridonin On Human Gastric Cancer Drug-resistant Cell Line SGC7901/VCR

Background Rabdosia rubescens have been used to treat gastric carcinoma for more than half a century . Nowadays , Oridonin have been extracted from Rabdosia rubescens and proved to be a very effective anti-tumor reagent. Recently , experimental data showed that Oridonin could inhibit the growth of many malignant cells , such as leukemic cells, breast cancer cells and non-small cell lung cancer cells. However, its effect on gastric carcinoma drug-resistant cells hasn't been demonstrated.Aim This experiment was intent to evaluate the growth-inhibiting and apoptosis-inducing effects of Oridonin on human gastric cancer drug-resistant cell line SGC7901/VCR cultured in vitro.Methods1. Determination of the effective concentration of Oridonin on SGC7901/VCR.The cells were cultured in vitro and treated with various concentrations of Oridonin for 24 h. The growth inhibitory rates were measured by MTT test, and theIC50 was determined.2. Growth-inhibiting effect of Oridonin at low concentration on SGC7901/VCR cells.The cells were treated with various concentrations of Oridonin (O/zmolL^lO^mollA 20/^mol-L'1) for 24 h , washed with PBS and fixed with 4% paraformaldehyde . After stained with Giemsa , the morphological changes of the treated cells were observed under microscope.3. Apoptosis-inducing effect of Oridonin on SGC7901/VCR cells .(1) After treatment with 40^molL': of Oridonin , the morphological changes were observed every an hour until apoptotic bodies appeared. After the appearance of apoptotic bodies , the cells were observed further under phase contrast microscope.(2) From the time of apoptotic bodies appearing , the cells were stained with Trypan blue and Giemsa respectively to observe the morphology of cells and apoptotic bodies.(3) From the time of apoptotic bodies appearing, the activity of succinate dehydrogenase of the cells was detected by MTT staining to reflect the function of mitochondria.(4)The DNA Ladder test was used to demonstrate the apoptosis-inducing effect of Oridonin on SGC7901/VCR cells further .(5) The range of concentration of Oridonin inducing the apoptosis of gastric cells was determined by the appearance of apoptotic bodies .4. Cell-killing effect of Oridonin on SGC7901 /VCR cellsThe cell necrosis was observed to indicate the cell-killing effect of Oridonin at high concentration on SGC7901/VCR cells.Results1. MTT test: Oridonin could inhibit the growth of human gastric cancer cell line SGC7901/VCR efficiently in a dose-dependent manner in vitro. At the concentration of 75/imol-L'1, the growth inhibitory rates could reach to 90%. The IC50 was about 55/^molL"1.2. Treated with various concentration of Oridonin (QamolL*1 > 20,umol-L~1) for 24 h, SGC7901/VCR cells changed from polygonal and short shuttle to round in shape . Some of the cells floated from the culture plate. In the specimens stained with Giemsa, the shrinked nucleus and condensed chromatin could be found.3. Apoptosis of SGC7901/VCR cells could be induced by Oridonin at certain concentration. After treatment with Oridonin (4Qamol-L"J) for about 4 h, the cells became round in shape, and the apoptotic bodies started to appear . Both the cells and apoptotic bodies couldn't be stained with Trypan Blue. In the Giemsa-stained specimens, condensed nucleus and marginated chromatin could be found. On the surface of apoptotic cells in the early stage in the specimens stained with MTT, the needle-like crystal could be found which might reflect the activity of succinate dehydrogenase of mitochondria. A distinct DNA Ladder occurred in the sample treated with 40lwmol-L"1 Oridonin. From SS^amolL"1 to 65/miol-L"1 Oridonin, apoptosis of gastric cancer cells SGC7901/VCR could be induced determined by the appearing of apoptotic bodies.4. Necrosis of SGC7901/VCR cells could be caused by 75/jmolL'1 to 105/imolL"1 Oridonin.Conclusions1. Oridonin could inhibit the growth of human gastric cancer drug-resistant cell line SGC7901/VCR efficiently in a dose-dependent manner in vitro.2. 35/imol-L'1 to 65/onol-L"1 Oridonin could induce the apoptosis of SGC7901/VCR cells.3. 75^molL1 to 105/imolL"1 Oridonin could killed SGC7901/VCR cells directly .