Effect of p38MAPK gene silencing on expression of oncogenes and apoptotic genes induced by PM(2.5) in hepatocytes

Abstract

Objective: To study the effect of p38 mitogen-activated protein kinase (MAPK) gene silencing on expression of apoptotic genes and oncogenes in hepatocytes treated with PM(2.5). Methods: From June to September 2019, according to the p38MAPK gene mRNA sequence provided by GenBank, three interfering sequences were designed and synthesized, ligated into PLVX-shRNA2-puro after annealing, and the recombinant lentiviral vector was transfected into L02 hepatocytes. The p38MAPK silencing cells were identified by real-time fluorescent quantitative PCR and western blotting. The normal L02 cells and p38MAPK silencing cells were treated with 50 μg/mL PM(2.5) water soluble solution, 10 μmol/L positive control Cr(6+), and a blank control group was set up, the treatment time was 24 h. The mRNA levels of oncogenes (c-fos, c-myc, k-ras) , tumor suppressor gene (p53) and apoptotic genes (Caspase-3, Caspase-8, Caspase-9) were detected by real-time PCR. The protein levels of oncogenes and apoptotic genes were detected by Western blotting. Results: The expression levels of p38MAPK mRNA and protein in p38MAPK gene silencing cells were significantly lower than those in L02 hepatocytes (P<0.05) , and the p38MAPK gene silencing cell line was successfully constructed. Compared with the blank control group, the expression levels of the oncogenes c-fos, c-myc, k-ras and the apoptosis genes Caspase-3, Caspase-8 and Caspase-9 increased, the expression level of tumor suppressor gene p53 decreased in the L02 hepatocyte group treated with PM(2.5) water soluble matter, and the differences were statistically significant (P<0.05) . Compared with the L02 hepatocytes group treated with PM(2.5) water soluble matter, the expression levels of the oncogenes c-fos, c-myc, k-ras and apoptosis genes Caspase-3, Caspase-8 and Caspase-9 decreased, the expression level of tumor suppressor gene p53 increased in the p38MAPK gene silencing cells group treated with PM(2.5) water soluble matter, and the differences were statistically significant (P<0.05) . Conclusion: PM(2.5) has effects on the expression of oncogenes, tumor suppressor genes and apoptotic genes in L02 hepatocytes, while p38MAPK gene silencing can inhibit the effects of PM(2.5) on L02 hepatocytes.