Curcuma Oil On Cultured A549 Cell Cycle And Cathepsin

ObjectivePulmonary fibrosis is a kind of diffuse lung disease caused by many reasons and has similar pathological process, and its pathology is characterized by alveolar structure destruction and diffuse pulmonary alveolitise in the early, and then induce over aggradation of Extracellular Matrix (ECM) and diffuse pulmonary fibrosis, leading to pulmonary dysfunction. Most of patients died of progressive and aggravating pulmonary respiratory failure due to pulmonary fibrosis.Up to now, the pathogenesis of pulmonary fibrosis has not been fully elucidated. In recent years, the incidence of pulmonary fibrosis increased year by year with the lack of effective treatment measures. Therefore, the key question in the medical science field is to find effective medicine for pulmonary fibrosis.Zedoary turmeric oil is the main active component of Zedoary, Guangxi Rhizoma Zedoariae and Turmeric rhizome. Studies suggested that Zedoary turmeric oil inhibit the proliferation of the cell by the effect of direct cytotoxicity, the process of cell apoptosis and the disordering of the nucleic acid metabolism of cells. In recent years, experiments in vitro indicated that Zedoary turmeric oil can inhibit the proliferation of human ovarian cancer cell (SKOV3) line, human nasopharyngeal carcinoma cell line (CNE-2), mouse liver cells line (HepA) and murine lung cancer cell line (Lewis). Several studies also.suggested that Zedoary turmeric oil and the microsphere of Zedoary turmeric oil inhibited the proliferation of human hepatocellular carcinoma cell (SMMC-7721) significantly, which was in the dose-dependent manner, and induced the cell apoptosis and blocked the cell cycle. However, the effect of Zedoary turmeric oil on human lung adenocarcinoma cell line A549 is still rarely reported.The cell line A549 is an epithelial lung tumor cell line with feathures of alveolar typeⅡepithelial cells. Alveolar epithelial cells have been shown to have regulatory functions in the maintenance of lung structure and function. Several studies indicated that alveolar epithelial cells are capable of expressing Cathepsins that participated in the process of extracellular matrix degradation and apoptosis.In this study, human lung adenocarcinoma cell line A549 was cultured in vitro and stimulated by Zedoary turmeric oil. Meanwhile the inhibition rate of the cell proliferation was tested, and the cell cycle and apoptosis rate were measured by the flow cytometry and the detection of the Cathepsins was accomplished by the Western blotting. According to the result of experiments, the effect of Zedoary turmeric oil on epithelial cells was explored and discussed. Method1 The inhibition rate of Zedoary turmeric oil on the proliferation of lung adenocarcinoma cell line A549 for 24 h,48 h,72 h were tested by MTT assay.2 The cell cycle of lung adenocarcinoma cell line A549 stimulated by Zedoary turmeric oil for 24 h was analyzed by flow cytometry and PI simple staining.3 The apoptosis and necrosis of lung adenocarcinoma cell line A549 stimulated by Zedoary turmeric oil for 24 h was tested by flow cytometry and Annexin V-FITC/PI double staining.4 The effect of Zedoary turmeric oil stimulated by Zedoary turmeric oil for 24 h on the level of Cathepsin D, K in lung adenocarcinoma cell line was tested.Result1 The A549 cells co-cultured with Zedoary turmeric oil for 24 h had the morphological change. There were a lot of granular material in the cytoplasm. The number of cells is growing obvious with the increasingly concentration of Zedoary turmeric oil.2 MTT assay indicated that the inhibition of Zedoary tunneric oil on the proliferation of lung adenocarcinoma cell line A549 increased significantly with the growing of hours and concentrations. The inhibition rate for 48 h is not significantly different from that for 72 h, but higher than the inhibition rate for 24 h.3 Further analysis by flow cytometry showed that Zedoary turmeric oil stimulating the A549 cells for 24 h can lead to G0/G1 phase arrest and blocked S phase entry, however G2/M phase had no significant change, in which A549 cells was stained by PI. Meanwhile cells in early apoptosis, late apoptosis and necrosis increased with the increase of the concentration of Zedoary turmeric oil, which was measured by flow cytometry and AnnexinⅤ-FITC/PI double staining, however the difference was not statistically significant among groups.4 The result of Western blotting indicated that normal cultured A549 cells can express Cathepsin D, K, while the level of Cathepsin D, K in the A549 cells co-cultured with different concentrations of Zedoary turmeric oil for 24 h was affected. The level of Cathepsin D and Cathepsin K were increasing with the growingly concentration of Zedoary turmeric oil.ConclusionZedoary turmeric oil can markedly inhibit the proliferation of lung adenocarcinoma cell line A549 in time-dependent and dose-dependent manners, and its role was achieved through preventing cell-cycle. Zedoary turmeric oil can increase the level of Cathepsins. 1 The study indicated that Zedoary turmeric oil reduce the degree of lung fibrosis, which due to the Epithelial-Mescnchymal Transition, through inhibiting epithelial cell proliferation.2 This study also indicated that Zedoary turmeric oil has the possibility of regulating the pulmonary matrix by increasing the level of Cathepsins.