| Study objective:Colorectal cancer is associated with genetic and environmental risk factors and is one of the most common life-threatening cancers.Surgery,radiotherapy,and chemotherapy are common treatment modalities for colorectal cancer.Chemotherapy includes the use of fluorouracil-based drugs or platinum-based drugs,etc.However,the high cost of chemotherapy and the tendency of patients with colorectal cancer to develop resistance to chemotherapy drugs severely limit the therapeutic effect of colorectal cancer patients.Therefore,there is an urgent need to develop a new,effective drug for colorectal cancer treatment with few side effects.Currently,structurally rich natural products are gradually becoming a hot spot for antitumor research as an important source of molecularly targeted antitumor drugs.Linalool,a natural product,is a monoterpene alcohol with antiviral,anti-inflammatory and antibacterial properties found in basil,laurel,jasmine,mahogany and lavender.Previous reports have shown that it has anti-cancer effects in breast and liver cancers,but its role and mechanism in colorectal cancer are far from being clearly elucidated.This project will investigate the anti-colorectal effects and mechanisms of linalool in human colorectal cancer SW480 cells,and provide new ideas for the design and development of novel molecular targeting drugs against tumors.Study methods:1.The cell morphology was observed microscopically after treatment of SW480 colorectal cancer cells with different concentrations of linalool,and the effect of linalool on cell proliferation was detected using the CCK-8 method.2.SW480 cells were treated with different concentrations of linalool and subjected to nuclear staining,membrane staining and DCFH-DA probe,and the extent of apoptosis was examined by fluorescence microscope and flow cytometry,and the cell cycle and intracellular ROS levels were measured by flow cytometry.3.The cycle-related proteins(p21 and Cyclin D1),apoptosis-related proteins(Cleave d-PARP),autophagy-related proteins(Beclin-1 and LC3B)and AKT/GSK-3β pathwa y-related proteins were detected in SW480 cells treated with different concentrations of linalool by western blot,respectively.4.After pretreatment of SW480 cells with ROS scavenger(NAC)and autophagy inhibitor(3-MA),respectively,the effect of linalool stimulation on cell viability was detected by CCK-8 method,and the expression of AKT/GSK-3β pathway-related proteins after NAC treatment was detected by WB.Study results:1.The CCK-8 results showed that linalool drug significantly inhibited the viability of SW480 cells in a significant concentration-dependent manner.Morphological observations also showed an increase in the number of SW480 cell death with an increase in linalool concentration.2.By flow cytometry,we can conclude that the percentage of G0/G1 phase in SW480 cells treated with different concentrations of linalool increases with increasing concentration,and WB results show that increasing linalool concentration can up-regulate p21 protein expression and down-regulate Cyclin D1 protein expression.3.The results of DAPI nuclear staining of SW480 cells treated with different concentrations of linalool showed that the number of intracellular fluorescence points gradually increased and intensified with the increase of linalool concentration.By flow cytometry,we found that the number of apoptosis in SW480 cells increased with increasing drug concentration,and WB assay revealed that Cleaved-PARP protein expression was up-regulated with increasing linalool concentration.4.We examined the ROS levels in SW480 cells treated with different concentrations of linalool drug by flow cytometry,and the results showed that the ROS levels in SW480 cells were positively correlated with linalool concentrations.The inhibitory effect of linalool on SW480 cell viability was attenuated by NAC after adding ROS scavenger treatment.5.The results of the AKT/GSK-3β pathway protein assay indicated that an increase in linalool concentration was able to decrease the expression of both p-AKT and p-GSK-3βproteins;meanwhile,the expression of p-AKT and p-GSK-3β proteins was increased after the addition of NAC compared to the single drug group.6.The assay of autophagy-related proteins in SW480 cells showed that linalool was able to upregulate the expression of LC3B-II as well as Beclin-1 protein.The results of CCK-8assay showed that the autophagy inhibitor 3-MA was able to reduce the inhibitory effect of the drug linalool on SW480 cell viability.Study conclusions:This study showed that linalool could induce SW480 cells to block in G0/G1 phase to inhibit cell proliferation,and linalool could induce apoptosis and inhibit AKT/GSK-3βpathway by increasing the level of ROS in SW480 cells to achieve its anti-tumor effect,and also induce autophagy in SW480 cells.This study illustrates that linalool can be used as a potential antitumor drug and provide a new direction for the development of new drugs for colorectal cancer. |
PDF Full Text Request