The study of killing effect and inducing apoptosis of 630-nm laser on lung adenocarcinoma A549 cells mediated by hematoporphyrin derivatives in vitro.

Abstract

To investigate the killing effect and inducing apoptosis of 630-nm laser mediated by hematoporphyrin derivatives (HPD) on human lung adenocarcinoma A549 cells. The human lung adenocarcinoma A549 cells were incubated at random with different concentrations of HPD (5, 10, 12, 15, 20μg/ml) for 4h and then illuminated by 630-nm laser with different energy densities (25, 50, 75, 100mW/cm2). And, meanwhile, the simple photosensitizer group, laser irradiation group, and blank control group were established. Then, CCK8, Hoechst 33258 staining, RT-PCR, and Western blot were employed. HPD-PDT proved no killing effect on the lung adenocarcinoma A549 cells with photosensitizer or laser irradiation alone. With the combination, the killing effect was obvious. CCK8 showed that the A549 cell viability in 15μg/ml and 20μg/ml HPD group as well as 50mW/cm2, 75mW/cm2, and 100mW/cm2 power density group decreased significantly compared with the control group. Hoechst 33258 staining showed that with the increase of HPD concentration, the cells presented chromatin fixation and hyperchromatic nuclei. The Annexin V-FITC/PI double staining was used to detect the apoptosis rate, and the difference was statistically significant. RT-PCR and Western blot showed that the expression of Caspase-3 and Bax were significantly up-regulated. However, the Bcl-2 and Survivin were significantly down-regulated in the HPD-PDT group, while those of the other three groups showed no significant changes. HPD-PDT has a significant effect on A549 cells. The mechanism of action may be related to the upregulation of the expression of Caspase-3, Bax, and downregulation of the expression of Bcl-2 and Survivin.