The Mechanism Of SIRT3 Inhibitor 3-TYP Increases Chemosensitivity Of Non-small Cell Lung Cancer By Modulating Oxidative Stress

Cisplatin is a first-line chemotherapy drug for the treatment of non-small cell lung cancer,but in the later stages of treatment,most patients will develop resistance to it.How to increase the sensitivity of cisplatin is an urgent problem in the medical community.Previous research reported that chemotherapy drugs including cisplatin can increase the production of ROS in tumor cells to drive oxidative stress and induce tumor cell apoptosis.People are looking forward to increasing the level of intracellular oxidative stress in as many ways as possible.To the maximum effect of tumor treatment.It is worth noting that SIRT3,as a deacetylase,plays an important role in the genesis and development of tumors.It can participate in tumor cells by deacetylating key proteins including superoxide dismutase in mitochondria.Oxidative stress,energy metabolism and apoptotic processes may provide a new direction for solving the problem of drug resistance in tumor cells.Therefore,exploring the relationship between SIRT3,oxidative stress and cisplatin resistance may provide an idea for the treatment of malignant tumors including but not limited to non-small cell lung cancer.In this study,non-small cell lung cancer patient tissues and NCI-H1299 cells were used as the research object.From the perspective of oxidative stress,the effect of SIRT3 inhibitor 3-TYP combined with cisplatin on tumor cell apoptosis and its mechanism were investigated.Optimizing chemotherapy for malignant tumors provides theoretical support.Methods: I.Detection of SIRT3 expression in tissues and cells of patients with non-small cell lung cancer Immunohistochemical staining was used to detect the expression of SIRT3 in tumor tissues of 84 patients with non-small cell lung cancer;Western-blot was used to detect the expression of SIRT3 in non-small cell lung cancer NCI-H1299 cells.II.Detection of SIRT3 inhibitor 3-TYP on the sensitivity and mechanism of cisplatin chemotherapy in non-small cell lung cancer NCI-H1299 cells Experimental grouping: control(CON)group,3-TYP group,cisplatin(DDP)group,3-TYP + DDP group 1.The MTT method was used to detect the effects of single-plus SIRT3 inhibitor 3-TYP,single-add DDP,and two drugs combined on the survival rate of non-small cell lung cancer cells NCI-H1299 cells and determine the optimal concentration of 3-TYP combined with DDP.2.Annexin V-FITC/ PI staining flow cytometry was used to detect the apoptosis of each group;Western-blot was used to detect the expression of apoptosis-related proteins in each group.3.DCFH-DA staining and flow cytometry were used to detect the ROS levels of cells in each group.4.SOD kit were used to detect SOD activity of cells in each group.5.JC-1 staining flow cytometry was used to detect the changes of mitochondrial membrane potential in each group of cells.6.Western-blot was used to detect the expression of SOD2 protein in each group of cells.Results: I.High expression of SIRT3 in tissues and cells of patients with non-small cell lung cancer Immunohistochemical staining showed that compared with normal lung tissue,SIRT3 expression was increased in non-small cell lung cancer tissues(P <0.05);Western-blot results showed that compared with normal lung epithelial cells,non-small cell lung cancer NCI-H1299 cells increased SIRT3 protein expression(P <0.01).II.SIRT3 inhibitor 3-TYP enhances chemotherapy sensitivity by increasing the level of oxidative stress in non-small cell lung cancer NCI-H1299 cells 1.MTT test results showed that compared with DDP group,3-TYP and DDP combined group can significantly inhibit the survival rate of non-small cell lung cancer NCI-H1299,and the optimal concentrations of 3-TYP and DDP are 5u M and 2.5ug / ml,respectively.2.Apoptosis test results showed that compared with the CON group,the apoptosis rate of the 3-TYP group had no significant change,and the apoptosis rate of the DDP group has increased.Compared with the DDP group,3-TYP and DDP combined increased of apoptosis rate in the group was more obvious(P <0.05);Western-blot results showed that the cell apoptosis rate in the Compared with the 3-TYP and DDP combined group,the expression of anti-apoptotic protein Bcl-2 decreased and the expression of pro-apoptotic proteins Bax and Cleaved-caspase3 increased(P <0.05).3.DCFH-DA staining flow cytometry results showed,the 3-TYP group and the DDP group had an increase in ROS content;compared with the DDP group,the 3-TYP and DDP combined group had a more increased ROS content.Obviously(P <0.05).4.The SOD kit test results showed that compared with the CON group,the SOD activity of the 3-TYP group and the DDP group decreased;compared with the DDP group,the SOD activity of the 3-TYP and DDP combined group decreased more significantly(P < 0.05).5.The results of JC-1 staining flow cytometry showed,the mitochondrial membrane potential decreased in the 3-TYP group and the DDP group(P <0.05);compared with the DDP group,the 3-TYP and The mitochondrial membrane potential of the DDP plus group was more significant(P <0.05).6.Western-blot test results showed that compared with the CON group,the expression of SOD2 protein was reduced in the 3-TYP group and the DDP group;compared with the DDP group,the expression of SOD2 protein in the 3-TYP and DDP combined group was significantly reduced(P <0.05).Conclusions:The combined use of SIRT3 inhibitor 3-TYP and cisplatin increased the oxidative stress level of non-small cell lung cancer NCI-H1299 cells and promoted their mitochondrial apoptosis,indicating that SIRT3 inhibitor 3-TYP enhanced non-small cell lung cancer NCI-H1299 cells are sensitive to cisplatin chemotherapy.