DNAJC19 Regulates Autophagy Via PI3K/AKT Signaling Pathway In The Radiosensitivity Of NSCLC

Research background and purpose:Radiation therapy is one of the main treatments for advanced non-small cell lung cancer(NSCLC).because of the widespread radioresistance,the efficacy of radiotherapy is limited,and it is also one of the important reasons for the failure of treatment in patients with NSCLC.Autophagy is involved in radioresistance.In previous studies,we used sh RNA to specifically inhibit the expression of DNAJC19 protein,and RNA-seq sequencing found that PI3K/AKT autophagy-related signaling pathways were involved in molecular events in NSCLC cell A549.Therefore,it is speculated that DNAJC19 modulates autophagy through PI3K/AKT signaling and may be an important mechanism affecting the radiosensitivity of human NSCLC.In this study,we intend to explore the potential mechanism of DNAJC19/PI3K/AKT participation in autophagy and thus affect radiosensitivity,in order to provide a new target for radiosensitization of NSCLC.Materials and Methods:(1)We explored the relationship between DNAJC19 and autophagy-related proteins and clinicopathological features through immunohistochemistry(2)Cytology function experiments(1)We used Transwell,cell cloning experiments,CCK-8,western blot,and flow cytometry to investigate the effects of radiotherapy on cell migration,proliferation,viability,autophagy,and apoptosis.(2)We used Transwell,cell cloning experiments,CCK-8,western blot,and flow cytometry to investigate the effects of Si RNA-DNAJC19 combined with autophagy inhibitors on cell migration,proliferation,viability,autophagy and apoptosis were observed(3)We used Transwell,cell cloning experiments,CCK-8,western blot,and flow cytometry to investigate the effects on cell migration,proliferation,viability,autophagy and apoptosis were observed using si RNA-DNAJC19 combined with autophagy inhibitors plus radiotherapy(4)We used western blot,to investigate the mechanism of DNAJC19 regulating autophagy through the PI3K/AKT signaling pathway(3)Statistical methodsWe used SPSS 11.0,Graph Pad Prism 8,and R for data processing and analysis,The survival curve was plotted using the Kaplan Meier method,and the difference in survival time of patients was analyzed by the Log-rank method.Experimental data are expressed as mean ± standard error(Mean±SEM),comparisons between the means of two samples are tested using independent samples t-test,and Fisher’s exact test,spearman,and chi-square test analysis are used to compare between-group variables.P<0.05 is considered statistically significant.Results:(1).We retrospectively analysed 50 cases of NSCLC.The results of immunohistochemistry showed that 34 cases(68%)had high expression of DNAJC19 and 16 cases(32%)had low expression.Meaningfully,patients with high expression of DNAJC19 survived shorter than those with low expression of DNAJC19,with an OS of35.5 months,statistically significant difference(P=0.01).The results of immunohistochemistry showed that 17 cases(34%)had high expression of LC3 and 33cases(66%)had low expression.Patients with high expression of LC3 survived shorter than those with low expression of LC3,with an OS of 30 months,statistically significant difference(P=0.02).(2).Radiotherapy significantly inhibits tumor cell migration,proliferation and cell viability,and promotes tumor cell apoptosis,DNAJC19 and increased expression of autophagy protein.(3)Knocking down DNAJC19 in A549 cells or using autophagy inhibitors can induce apoptosis by inhibiting clonal formation and cell activity.(4).Under radiation,we used si RNA-DNAJC19 or autophagy inhibitors,which can enhance the inhibitory effect of radiation on tumor cell migration,proliferation and cell viability,and the effect is stronger when combined with si RNA-DNAJC19 and autophagy inhibitors.(5).Western blot showed that si RNA-DNAJC19 or autophagy inhibitors inhibited the expression of PI3 K,AKT and DNAJC19 proteins.(6)Radiation acting on A549 cells,combined with si RNA-DNAJC19 or autophagy inhibitors,reverses the promotion of the expression of P3 K,AKT and DNAJC19 proteins.Conclusions:High expression of DNAJC19 and LC3 proteins affects the prognosis of NSCLC patients.Si RNA-DNAJC19 participated in the regulation of autophagy in lung cancer cells by inhibiting the PI3K/AKT signaling pathway.si RNA-DNAJC19 combined with autophagy inhibitors can significantly increase the radiotoxicity of radiation to A549 cells,providing a new target for NSCLC radiotherapy resistance.