Ursolic Acid Inhibits Proliferation And Induces Apoptosis In Human Uterine Cervical Cancer Cell HELA

The cervical cancer is the most familiar gynecology malignant tumor. More than 20 milion women died of cervical cancer around the world every year.In the developing countries, the cervical cancer is the major gynecology tumor of all kindsnewfound casesin China, its mortality rate has the No. 1 potential of total cancer mortality rate.The apoptosis is the process of programmed cell death (PCD) that may occur in multicellular organisms. The process of apoptosis is controlled by a diverse range of cell signals, which may originate either extracellularly (extrinsic inducers) or intracellularly (intrinsic inducers). The apoptosis abnormality has important impact on many malignant tumor's occurrence and developments , therefore,studies foucus on theory and technique with optionally induced tumor apoptosisbecome one of the main strategies that cures a malignant tumor.The ursolic acid is pentacyclic triterpene acid, which has a far-ranging living creature activity, the anti- tumor activity is the pharmacology action of its most. Ursolic acid has relatively large resource, good prospects of development and application. And, as a kind of anti- tumor medicine, compared to other chemotherapy medicine, ursolic acid has poison secondary acting the low characteristicswhich means ithas a potential to prevent and cure tumor. The rsearch of its anti- tumor cell will be the important step of the anti-cancer ursolic acid series medicine research and developments anddirection and hot spot of future development , the ursolic acid hasa chance to become a kind of efficiently low poison new medicine on anti-tumor.Thr objects of this study are to discuss the inhitory effects and induce apoptosis of ursolic acid (UA) on human uterine cervical cancer cell HELA.Methods: 1.The concentrations of UA used in cell culture system were 0,20, 40,80μmol/L andthe termination points of cell culture are 24,48,72hours.Cell proliferation of UA on HELA cell was determined by MTT assay. Use 0.25% Parenzyme to digest single layer development cell, and then use it to develop unicellular suspension. Regulate the cell concentration to the 1.0x105/m1, pr. add 200μl unicellular suspension into 96 hole boards. Develop cell, until 80% cells inside the board present to remit to match a status, and then suck out development liquid, add no serum DMEMfor synchronous 24 hours , then add bear tartaric acid to make concentrationof the mixture for 0, 20, 40,and 80μmol/L, each one establishes 4 complex holes. Placing 96 hole boards in theculture boxto develop24, 48 and the 72 h respectively. After that, add MTT 20μl of 5 g/L, keep developing another 4 h, lightly and slowly suck away to develop inside the hole up pure, every hole joins DMSO 150μl, in take off a color to shake the osc 10 mins of bed postpose to mark the instrument 490 nm at the Mao a wavelength to measure to absorb a lintensity of light value(A).Calculating proliferation to repress a rate(IR%)=(1-the transaction set absorb the lintensity of light value/matched control to absorb a lintensity of light value)X 100%, pass rectilinear writing recorder regression equation computing IC50. Repeat 4 time of each experiment, and the take the average of the results.2.Cell apoptosis rate were analysed by flow cytometry.Using the streaming type cell techanque Annexin the V-FITC/PI cell apoptosis dye a law to directly determine the apoptosis of cell dead. For HELA cell in logarithm vegatation period, use 0.25% Parenzyme digest cell in a short time and develop a liquid hang cell again, adjust untill the cell concentrationis 4.0x105/m1, Be inoculated to 6 hole boards amid, add bear tartaric acid of 20μmol/L, react 24 h, 48h and 72 h respectively.After, click try the step of a box instruction in the book to carry on dyeing, immediately progress streaming type cell instrument examination, experiment again 3 times take it to be worth.3.Apoptosis was detected by morphological examination.Take HELA cell in logarithm vegetation period at long sight, use 0.25% Parenzyme to digest cell and develop a liquid hang cell again, adjust to go together with cell of concentration for the 4.0x105/m1, Be inoculated to 6 hole boards amid, join eventually a concentration for the ursolic acid of 0 and20μmol/L, act a 48 h, apply PI to carry on dyeing, Be placed to a fluorescence microscope to down observe the condition of apoptosis.4. Data were analyzed by SPSS16.0 Statistics software.Results:1.MTT assay showed that UA had a moderate anti-proliferation effect on HELA cells in a time-dependant and dose-dependant manner.2. The results of Flow Cytometry (FCM) represented that the apoptosis rate of HELA cells treated with20μmo/L UA for 24,48,72 hours were (6.64±1.01)%,(13.71±1.01)%,(18.60±2.51)%(P<0.05)。3.The morphologic changes of HELA includes red fluorescence appearance.Conclusion:1.UA had a moderate anti-proliferation effect on HELA cells in a time-dependant and dose-dependant manner.2.20μmo/L UA had a moderate apoptosis effect on HELA cells in a time-dependant manner.UA shows obvious inhibitory effects on HELA cells. The effects may be related to induce apoptosis.