Effects Of Tiamulin On Proliferation And Apoptosis Of Human Lung Adenocarcinoma A549 Cell

Lung cancer has a high mortality rate among cancer patients,and due to its extremely high incidence,high metastatic potential,and heterogeneity,only 2%-20%of patients survive for 5 years from diagnosis.The recurrence of toxic side effects and drug resistance of anti-tumor drugs has put tremendous pressure on the research and development of new drugs.Therefore,it is urgent to find new anti-lung cancer drugs with good efficacy.In recent years,veterinary antibiotics have emerged in the field of cancer treatment,and their good R&D base and broad application prospects have brought new hope to the development of new anti-tumor drugs.Tiamulin,a truncated siderophore antibiotic,as one of the top ten veterinary drugs in the world,has shown some potential in cancer treatment.Therefore,the anti-lung cancer ability of Tiamulin was investigated in the hope of providing new and more reliable ideas for the development of anti-cancer drugs.In this experiment,the veterinary antibiotic Tiamulin was selected as the test drug to investigate whether it has anti-lung cancer activity and to study its preliminary mechanism of action on human lung adenocarcinoma A549 cells.Firstly,the effect of Tiamulin on the proliferation of A549 cells in vitro was analyzed from the changes in cell viability and number.Cell scratch test and Transwell chamber test were used to detect the changes in cell migration ability after the treatment of Tiamulin,and the effect of Tiamulin on cell proliferation was verified on the other hand.Meanwhile,combined with the tumor inhibition experiment in vivo to confirme the effect of tiamulin on the proliferation and growth of A549 cells.Secondly,the morphological changes of A549 cells after treatment with Tiamulin were observed by morphological method.Annexin V-FITC/PI double staining was used to detect and observe the apoptosis of the cells,and the ROS content was detected by reactive oxygen species kit to analyze the relationship between drugs and cell apoptosis.Finally,the activity of apoptosis-related caspases was detected,and the effect of caspase inhibitors on cell viability was detected.The expression of apoptosis-related proteins was detected to analyze the mechanism of Tiamulin inducing A549 cell apoptosis.The results showed that Tiamulin showed the ability to inhibit the proliferation of A549 cells in vitro and in vivo.Moreover,it can effectively induce apoptosis in A549 cells.Compared to controls,Tiamulin inhibited the survival of A549 cells in vitro,reduced the number of viable cells and colony formation,and had less toxic effects on human normal fibroblastic NHLF cells;cell scratch assays and Transwell chambers demonstrated the ability of tiamulin to inhibit horizontal and vertical migration of cells.At the same time,tiamulin slowed down the growth of transplanted lung cancer tumors in nude mice.Morphological assays directly observed that the cells showed typical apoptotic features such as chromatin fixation,breakage,convergence and apoptotic vesicles after treatment with thaumatin;intracellular ROS level and apoptosis rate increased significantly with increasing drug concentration;caspase enzyme activity assay and western blot results showed that caspase-3/-4/-8/-9enzymes were activated after Tiamulin treatment and had a significant effect on the apoptosis of A549 cells.The results of caspase activity assay and western blot showed that caspase-3/-4/-8/-9 enzymes were activated and inhibited cell proliferation,and the expression of pro-apoptotic proteins Bax and Cytc were up-regulated,while the expression of anti-apoptotic proteins Bcl-2 and XIAP were down-regulated,and PARP cleavage and Akt activity were decreased.It was shown that Tiamulin could efficiently induce apoptosis,indicating its cancer inhibitory activity.Conclusions:1.Tiamulin exhibited the ability to inhibit A549 cell proliferation in vitro and in vivo,and it was less toxic to human normal lung fibroblasts;2.Tiamulin can induce the accumulation of reactive oxygen species in A549 cells and promote the onset of apoptosis;3.Tiamulin can induce a variety of caspase enzymes involved in apoptosis of A549 cells,and its apoptotic process may be related to caspase-dependent mitochondrial pathway and PI3K/Akt pathway.