630 nm激光诱导的血卟啉衍生物联合顺铂对人肺腺癌A549细胞增殖抑制及化疗增敏作用的影响

Effects of 630 nm Laser-Induced Hematoporphyrin Derivatives Combined with Cisplatin on the Proliferation Inhibition and Chemotherapy Sensitization of Human Lung Adenocarcinoma A549 Cells

摘要: 目的：探讨630 nm激光诱导的血卟啉衍生物(Hematoporphyrin Derivative-Photodynamic Therapy HPD-PDT)与顺铂(Cisplatin DDP)单独或联合应用对人肺腺癌A549细胞株的增殖抑制作用及HPD-PDT对DDP化疗的增敏作用。方法：将对数生长期的A549细胞分为HPD-PDT组、DDP组、联合组(DDP + HPD-PDT组)和对照组。CCK-8法检测各组细胞的增殖抑制率，Hoechst33342染色、annexinV-FITC/PI双染法检测细胞的凋亡，RT-PCR检测各组Bcl-2、Bax、cleaved-PARP及Caspase-9 mRNA的表达。结果：CCK8结果显示：对照组、DDP组、HPD-PDT组、DDP + HPD-PDT组A549细胞成活率分别为100%，(88.56 ± 3.57)%，(84.61 ± 3.18)%、(74.34 ± 3.54)%。各用药组对人肺腺癌A549细胞株均有增殖抑制作用，尤其以联合组抑制作用最强；组间比较(除HPD-PDT组和DDP组)有统计学意义(均P < 0.05)；Hoechst33342染色显示：HPD-PDT组，DDP组，DDP + HPD-PDT组均可见明显的凋亡细胞，以DDP + HPD-PDT组凋亡细胞数最多；流式细胞术检测结果显示：对照组、DDP组、HPD-PDT组、DDP + HPD-PDT组细胞早中期细胞凋亡率分别为(6.38 ± 0.36)%、(13.68 ± 0.67)%、(15.72 ± 0.56)%、(22.94 ± 0.55)%。各用药组与对照组比较，联合用药组与单独用药组比较，细胞凋亡率增加，差异均有统计学意义(均P < 0.05)，DDP + HPD-PDT组凋亡细胞数最多；RT-PCR显示：HPD-PDT和DDP作用细胞后可以上调Bax和PARPmRNA及下调Caspas9、Bcl-2mRNA的表达，组间比较(除HPD-PDT和DDP组)有统计学意义(均P < 0.05)。结论：光动力治疗联合化疗对肺腺癌A549细胞的杀伤效果最明显。血卟啉衍生物介导的光动力治疗可增强A549细胞对顺铂化疗的敏感性。

Abstract: Objective: To investigate the proliferation inhibitory effect of 630 nm laser-induced hematoporphyrin derivative photodynamic therapy (HPD-PDT) and cisplatin (DDP) alone or in combination on human lung adenocarcinoma A549 cell line and the sensitization effect of HPD-PDT on DDP chemotherapy. Methods: A549 cells in logarithmic growth phase were divided into HPD-PDT group, DDP group, combined group (DDP and HPD-PDT group) and control group. The proliferation inhibition rate of cells in each group was detected by CCK-8 method, and the apoptosis of cells was detected by Hoechst33342 staining and annexin V-FITC/PI double staining method, and the expression levels of Bcl-2, Bax, PARP and caspase-9 mRNA were detected by RT-PCR. Results: The results of CCK8 showed that the survival rate of A549 cells in the control group, DDP group, HPD-PDT group, DDP and HPD-PDT group was 100%, (88.56 ± 3.57)%, (84.61 ± 3.18)%, (74.34 ± 3.54)% respectively. Each group had a proliferation inhibitory effect on the human lung adenocarcinoma A549 cell line, especially the combined group had the strongest inhibitory effect; The comparison between the groups (except the HPD-PDT group and the DDP group) was statistically significant (both P < 0.05). Hoechst33342 staining showed that HPD-PDT group, DDP group, DDP and HPD-PDT group had obvious apoptotic cells. The results of flow cytometry showed that the early and mid-term cell apoptosis rates of the control group, DDP group, HPD-PDT group, DDP and HPD-PDT group were (6.38 ± 0.36)%, (13.68 ± 0.67)%, (15.72 ± 0.56)%, (22.94 ± 0.55)% respectively. The apoptosis rate between each medication group with the control group or the combination medication group with the single medication group was increased, and the difference was statistically significant (P < 0.05), and the number of apoptotic cells in the DDP and HPD-PDT group was the largest; RT-PCR showed that HPD-PDT and DDP can up-regulate the expression of Bax, PARPmRNA and down-regulate the level of caspase-9 and Bcl-2mRNA after acting on cells. The comparison between groups (except HPD-PDT group and DDP group) was statistically significant (P < 0.05). Conclusion: The killing effect of photodynamic therapy and chemotherapy on lung adenocarcinoma A549 cells is the most obvious, and hematoporphyrin derivative-mediated photodynamic therapy might enhance the sensitivity of A549 cells to cisplatin chemotherapy.