The Inhibitive Effect Of Extract From Selaginella Doederleinii Hieron On Ovary Cancer Cells

Objective:Selaginella doederleinii Hieron is a chinese traditional herbs which has been identified to exert anticancer function and usually used to treat nasopharyngeal darcinoma, gastric cancer and liver cancer. However,few research has reported that it has relationship with ovary cancer.In this study,different kinds of extract were isolated from the Selaginella doederleinii Hieron by three reagents with different hydrophobicity. These extract were added to the ovary cancer OVCAR-3 cells. The inhibitive effects of these extract on OVCAR-3 cells were investigated by In-vitro-Cytotoxicity-Test,phases of apoptosis analysis and observing ultrastructure of cells. Mean while the invasive ability of OVCAR-3cells treated with extract was tested and the synthesis changes of EGFR protein was tested. All these trials were putting effort to identify if the Selaginella doederleinii Hieron has positive effect on ovary cancer and provide some evidences for utilizing this herb on ovary cancer.Methods:1 Dry Selaginella doederleinii Hieron were grinded into powder and paste was gained after extracted by ethanol under high tempreture. This paste was further extracted by petroleum ether, ethyl acetate and n-butanol successively.2 Three kinds of extract were added to the ovary cancer OVCAR-3 cells which cultured in vitro. The inhibitive effects of these extract were tested through In-vitro-cytotoxicity-test using MTT; cells combined with Annexin V-FITC and/or PI to analyse different phases of apoptosis using Flow Cytometry; ultrastructures of cells were observed through the Transmission Electron Microscope. Then the invasive ability of OVCAR-3 cells was investigated by using Transwell Plate covered with Matrigel and synthesischanges of the EGFR protein were tested by Immunofluorescence technique using antibody specifically combinated to EGFR protein.Results:1 Three kinds of extract from Selaginella doederleinii Hieron were obtained successfully.2 When the concentration of extract mounted up to 50 μg/mL or more and cells treated long to 48 h or longer, extract obtained by petroleum ether and ethyl acetate could reduce the OD490 value of cultured cells, which meant to have fine inhibitive effect on OVCAR-3 cells; when the concentration of extract mounted up to 100 μg/mL and cells treated long to 48 h, extract obtained by petroleum ether had higher inhibition than extract obtained by ethyl acetate apparently. The inhibitive effect of these two kinds of extract rose with the concentration increased and time lasted longer. However, extract obtained by n-butanol had little impact on cells at any concentration in any time. Extract obtained by petroleum ether could destroy the structure of endoplasmic reticulum and made the endoplasmic reticulum swelled; Extract obtained by ethyl acetate could destroy the structure of mitochondria and broke down the cristae in the inner mitochondria and also could break down the membrance of nucleus and nucleolus.3 Three kinds of extract could reduce the number of cells which transferred through the Matrigel covered onto the Transwell Plate. Extract obtained by petroleum ether had the finest reductive effect and orderly followed by extract obtained by acetate ethyl and extract obtained by n-butanol.4 Extract obtained by petroleum ether and acetate ethyl could reduce the intensity of green fluorescence on the membrance of cells which meant the express of EGFR protein reduced. Extract obtained by petroleum ether had higher reductive effect than the extract obtained by acetate ethyl.Conclusions:1 Active substances could be isolated from Selaginella doederleinii Hieron toinhibite the growth of ovary cancer OVCAR-3 cells.2 Different substances have different targets when implement their function.Extract obtained by petroleum ether can swelled the endoplasmic reticulum while extract obtained by ethyl acetate can break down the cristae in the inner mitochondria and nucleus; These two kinds of extract also can down regulate the synthesis of EGFR protein and reduce the invasive ability of the cells.3 Extract obtained by n-butanol hardly could inhibite the growth ofOVCAR-3 cells and reduce the express of EGFR protein but can reduce their invasive ability to some extent.