Study On The Inhibition Effect And Mechanism Of Action Of Brucea Javanica Oil Emulsion On Hela Cervical Cells

ObjectiveCervical cancer is a serious threat to women's health .It is one of the most common cancers in developing countries. Currently,its incidence and mortality rates are increasing.Doctors use mainly surgery combined with radiotherapy and chemotherapy treatment, but most of radiotherapy and chemotherapy drug has side effects, They seriously impact on the life quality of patients. So we need find a kind of better drug .Brucea javanica oil emulsion is the extracts of Brucea javanica mature fruit. The study found that it has a higher affinity for cancer cells, It can treat cancer, combined with radiotherapy and chemotherapy it can improve immunity, protect bone marrow, raise peripheral blood and so on. Its side effects are small so the clinical application is safe.At present it has been widely applied to treat lung cancer, metastatic cancer, liver cancer, esophageal cancer and other malignant tumor. At the same time it can be used for treatment of HPV-related flat wart, verruca vulgaris, condylomata acuminata and so on. But it has not been applied to the treatment of cervical cancer.Therefore, we do an experiment using Brucea javanica oil emulsion to observe its inhibition to the Hela cells, discuss the prospects of its application in the prevention of HPV-related cervical cancer.We carried out the following study: culture cell, plus Brucea javanica oil emulsionâ‘ use Papanicolaou stain to observed cell morphologyâ‘¡MTT assay to detect cell inhibition rateâ‘¢use flow cytometry to detect Hela cell cycle.MethodsCell Culture: Use conventional methods to culture Hela cells.The Observation Of Cells: After treatment with different concentrations of Brucea javanica, use an inverted microscope to observe the cell growth.Drugs for 48 hours, collect all the cells , add formaldehyde to fix for Papanicolaou stain, Observe that cell morphology changes.The Rate Of Inhibition Of Cell: Hela cells are transferred to 96-well plate, add different concentrations of drugs, incubate for 24 h, 72 h and 48h, add MTT and DMSO to each hole, measure the absorbance at 490nm by Meibiaoyi.Detect Changes In The Cell Cycle: add different concentrations of Brucea javanica oil emulsion, incubate for 48 h, collect all the Hela cells ,detect cell cycle by flow cytometry.ResultsCells Morphological Change: used inverted microscope to observe: With the increase of drug concentration and the extension of time, we saw that the density of cell growth decreased, round cells increased, and some cells fell off from bottles debris, the majority of cells showed different degrees of apoptosis and necrosis state. Papanicolaou stain Results: chromatin condensed and attached to a membrane surrounding,we could observe the typical apoptotic bodies.Test The Inhibition Rate of Drugs to Cell: Different concentrations of Brucea javanica oil emulsion effected on Hela cells for 24 h,48 h,72 h, we tested the apparent inhibitory. The same concentration drugs at different time points showed inhibition rates were significantly different, with the extension of time, the inhibition rate increased. at the same time, inhibition rate of different concentrations there was an obvious difference, with the concentration increasing the inhibitory rate increased.Brucea Javanica effected Cell Cycle: The different concentrations of Brucea javanica oil emulsion added for 48 h, the cells were analyzed by flow cytometry. Cell cycle changes can be seen with the drug concentration increased, The cell of G₀-G₁ phase declined, the percentage of cells in S phase and G₂-M phase significant increased. The experimental group were detected significantly apoptosis peak, with the drag concentration increasing apoptosis peak was greater.DiscussBrucea javanica oil emulsion has inhibition on the Hela cells. After 48 hours, the inhibition rate of experimental groups had been over 50%, after 72 hours,the inhibitory rate of almost were more than 80%. Brucea javanica oil emulsion could significantly inhibit the proliferation of Hela cells, with the increase of drug concentration and the extension of time the effect of inhibition enhanced.Tumor occurrence and cell apoptosis are related, tumor cell apoptosis induced by chemotherapeutic drugs is one of the mechanisms. The study found that Hela cells were added Brucea javanica, Papanicolaou stain showed obvious characteristics of apoptotic bodies. In addition, the results that were analyzed by flow cytometry showed obviously peak of apoptosis of the experimental group.The Brucea javanica oil emulsion inhibited Hela cells that may be the mechanism of drug-induced apoptosis.Experimental results showed that Brucea javanica oil emulsion had an impact on the cell cycle by flow cytometry. As the drug concentration increased, G₀-G₁ phase cells fell,cells ratio in S phase risen significantly and in G2-M phase cells increased. Brucea javanica oil emulsion mainly blocked Hela cells in S phase and G2-M phase, reduced tumor cell division, inhibited the proliferation of tumor cells and by interfering with the growth and metabolism induced apoptosis of tumor cells.Conclusion1,Brucea javanica oil emulsion has effected on Hela cells in shape, internal structure, the proliferation rate and the growth cycle it can inhibit cell proliferation, induce apoptosis and block cell growth cycle.2,Brucea javanica oil emulsion except for the treatment flat wart, verruca vulgaris, condyloma and laryngeal papilloma which caused by HPV. It also have a certain prospect to the HPV-related cervical cancer.