Effect Of Tea Tree Oil Extract On The Growth Of B16F10 Melanoma Cell And Its Transplant Tumor In Mice

Objective:Tea tree oil,a natural mixture extracted from tea tree,has been reported to possess antibacterial,antifungal,antiviral,immune enhancement activities.Recent researches have shown that tea tree oil inhibited the growth of numerous tumor cells.The present study was set to investigate the activity of different tea tree oil extract formulation against the growth of B16F10 melanoma cell in vitro and its transplantable tumor model in vivo.This research aimed to screen out the best formula of the different formulas and investigate its anti-melanoma mechanism,which providing the basic data for using tea tree oil as a new anti-melanoma drug.Methods:In cell culture experiments in vitro,B16F10 melanoma cells were treated by 0.0125%-0.1%tea tree oil extract of six formulas,the inhibitory effect on the cell activity tested using MTT assay was to screen the best recipe of tea tree oil extract.The influence of formula 5 on cell cycle distribution and apoptosis were determined by flow cytometry,expression of apoptotic associated proteins such as cleaved caspase-3 and cleaved caspase-9 were detected via Western blot.In the mice transplanted melanoma model in vivo,B16F10 cells were subcutaneous injected toC57BL/6 mice.The mice were then intratumoral injected with PBS,formula 5(10%),cyclophosphamide respectivelly for 7 days.During the experiment,body weight and tumor diameter were determined each day.At the end of the experiment,tumors were stripped and weighted to check the inhibitory effect of tea tree oil extract on tumor growth.The tumor tissues were collected for HE staining analysis,Western blot and immunohistochemistry of cleaved caspase-3 and/or cleaved caspase-9.Results:In the cell culture experiments in vitro,MTT assay showed that all of tea tree oil extract formulas could effectively inhibit the growth of B16F10 melanoma cells with a significant dose-effect relationship,the inhibitory rate of the six formulas was 30%?75%at 0.025%concentration.formula 5 was appeared best potent inhibitory effect,followed by formula 2,and 4 formula better than other three formulas.The cells activity were decreased more than 60%at 0.0125%?0.05%of the formula 5.In the Flow cytometry and Western blot analysis,the cell cycle arrest and apoptosis were induced by formula 5,and the expression level of apoptotic associated proteins including cleaved caspase-3 and cleaved caspase-9 were significantly increased.In the experiments of mice transplanted melanoma model in vivo,the tumor growth was obviously inhibited at formula 5 treatment with.The growth of tumor cell and the structure of tumor tissue were affected by formula 5 in the HE staining analysis.The expression of cleaved caspase-3 and/or cleaved caspase-9 in tumor tissue were significantly increased in the Western blot and immunohistochemistry analysivs.Conclusion:Tea tree oil extract can effectiely inhibit the growth of B16F10 melanoma cells and their transplanted tumors in vitro and in vivo,in which the mitochondrial pathway apoptosis may be involved.