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《中华肺部疾病杂志》[ISSN:1006-6977/CN:61-1281/TN]

卷:

期数:

2024年06期

页码:

847-854

栏目:

论著

出版日期:

2024-12-25

文章信息/Info

Title:

In vitro investigation of cisplatin resistance reversal in lung cancer cells via chemo-photodynamic therapy utilizing a functionalized novel metal-organic framework

作者:

吕嘉文; 陈昕昕; 陆婉君; 吴冠楠; 顾晓凌; 吕镗烽; 王 栋; 宋 勇

210002 南京,南京大学医学院附属金陵医院呼吸与危重症医学科

Author(s):

Lv Jiawen;  Chen Xinxin;  Lu Wanjun;  Wu Guannan;  Gu Xiaoling;  Lv Tangfeng;  Wang Dong;  Song Yong.

Department of Respiratory Medicine, Jinling Hospital, Affiliated Hospital of Medical School, Nanjing University, Nanjing 210002, China.

关键词:

支气管肺癌;  金属有机框架;  纳米传输系统;  化学-光动力联合治疗;  顺铂耐药

Keywords:

Bronchogenic carcinoma;  Metal organic framework;  Nano transport system;  Chemo-photodynamic combined therapy;  Cisplatin resistance

分类号:

R734.2

DOI:

10.3877/cma.j.issn.1674-6902.2024.06.001

摘要:

目的 构建具有化学-光动力联合疗效的新型金属有机框架(metal-organic framework, MOF)纳米载药传输系统,改善肺癌细胞的顺铂耐药性。方法 以四羟基苯基卟啉为基础合成MOFs,利用超声乳化法负载顺铂,形成近红外照射下激活的载药纳米系统(Cisplatin@MOFs)。通过表征粒径、Zeta电位、单线氧检测实验等了解理化性质; 采用细胞活力检测试剂盒(cell counting kit-8, CCK-8)判断体外细胞毒性; 观察细胞增殖和细胞迁移功能判断联合抗肿瘤效应; 通过蛋白印迹实验(Western Blot)探索调控机制。结果 透射电子显微镜(transmission electron microscope, TEM)表征结果显示:Cisplatin@MOF呈多边球形结构,粒径为(431.33±21.47)nm; 激光动态光散射(dynamic light scattering, DLS)检测Cisplatin@MOF平均粒径为458 nm左右,分散系数(polymer dispersity index, PDI)为0.482; MOF和Cisplatin@MOF的Zeta电位值分别为(3.20±0.23)mV和( 6.63±0.81 )mV; 近红外照射后MOF纳米系统释放单线态氧。Cisplatin@MOF纳米给药系统的顺铂负载量为(41.73±2.66)%; 释放试验显示,Cisplatin@MOF在酸性条件下释放顺铂药物。体外细胞实验中,共聚焦显微镜观察显示肿瘤细胞摄取MOF纳米系统。细胞毒性实验显示,Cisplatin@MOF在光照条件下降低顺铂耐药型肺癌细胞(A549-DDP)的细胞活性; 细胞功能实验结果显示,Cisplatin@MOF在光照条件下抑制细胞增殖和细胞迁移能力。Cisplatin@MOF具有化学-光学联合抗肿瘤效应。蛋白印迹实验显示,光照条件下Cisplatin@MOF引起Bcl-2表达水平下降,Bax表达水平升高,Bcl-2/Bax信号通路可能参与调控机制。结论 多功能MOFs纳米载药传输系统是具有联合治疗疗效的新型抗肿瘤方案,可改善顺铂耐药,为高效肺癌靶向治疗手段。

Abstract:

Objective To overcome cisplatin resistance in lung cancer cells, a novel metal-organic framework(MOF)nanocarrier delivery system with chemo-photodynamic combined efficacy was developed. Methods MOFs were synthesized using the Tetrakis(4-hydroxyphenyl)porphyrin(THPP)and loaded with cisplatin through ultrasonic emulsification method to form a drug-loaded nanosystem(Cisplatin@MOFs), which were activated under the near-infrared irradiation. The physicochemical properties were characterized though the particle size, Zeta potential, and the singlet oxygen detection experiments. The particle size and surface zeta potential were measured by the dynamic light scattering(DLS)and the shape of samples was visualized by transmission electron microscopy(TEM). The singlet oxygen release was detected by the ROS indicator. The cisplatin release from Cisplatin@MOF nanoparticles were assessed by the dialysis diffusion method. In vitro cytotoxicity was observed by using a CCK8(cell counting kit-8)assay. The combined anti-tumor effect was investigated through the measurement of cell proliferation and cell migration. The proliferation was observed by Ki67 fluorescence staining and the cell migaration was assessed by the wound-healing experiment. Western Blot analysis was employed to explore the underlying regulatory mechanisms. Results Transmission electron microscopy(TEM)characterization showed that the Cisplatin@MOF exhibited a polygonal spherical structure with a particle size of(431.33±21.47)nm. The dynamic light scattering(DLS)measurements indicated an average size of approximately 458 nm for Cisplatin@MOF, with a polydispersity index(PDI)of 0.482. The Zeta potential values were(3.20±0.23)mV for MOF and(6.63±0.81)mV for Cisplatin@MOF, respectively. Under near-infrared irradiation, the MOF nanoparticles could release the singlet oxygen. The ROS generation ability of the MOFs was detected by the indicater(DPBF)and the results demonstrated the photodynamic protential. The loading capacity of cisplatin in the Cisplatin@MOF nanodrug delivery system was(41.73±2.66)%. Release tests showed that Cisplatin@MOF could release cisplatin under acidic conditions by a dialysis method. Then we used the confocal laser scanning microscopy(CLSM)to examine the cellular internalization of MOF nanoparticles. In vitro we conducted a cell viability assay(CCK8)to evaluate MOF's cytotoxicity in cisplatin-resistant lung cancer cells(A549-DDP). The results showed that compared to control and no-laser groups, a significant reduce in cell viability was observed in cells treated under light irradiation. Cisplatin@MOF under light irradiation exhibited the strongest damage on the cisplatin-resistant lung cancer cells. We used fluorescence immunostaining analysis of Ki67 marker to assess the cell proliferation. The results revealed that the lower Ki67 expression in the cells treated under light irradiation compared to the control and dark groups. Cisplatin@MOF under light irradiation showed the lowest Ki67 expression of cell proliferation. It was conducted that the cell scratch test to examine the impact of MOFs on cell migration. The results revealted that the cells receiving the later treatment demonstrated weak cell migration compared to the control and dark groups. Cisplatin@MOF under light irradiation exhibited the weakest cell migration compared to other rtreatmens. Thse results confrm a significant chemophotothermal combined antitumor effect on cisplatin-resistance tumor cells. Futhernore, we used the Western Blot assays to ascess the protein levels of Bcl-2/Bax signaling pathway. Cells treated with light irradiation inhibited significantly lower levels of Bcl-2 expresion compared to control and no-laser groups. And the protein levels of Bax were significantly increased in the cells from the laser groups compared to the control and dark groups. These results suggested Bcl-2/Bax signaling pathway's potential as the ksy mechanism for combined antitumor effects. Conclusion In summary, we developed a multifunctional MOFs-based drug delivery system that combines chemotherapy and PDT for a novel strategy. This system effectively overcomes cisplatin resistance and demonstrates combined therapeutic efficacy through chemo-photodynamic therapy. These findings suggest it could potentially serve as a more efficient targeted therapy for lung cancer.

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备注/Memo

备注/Memo:

基金项目: 东部战区总医院院管课题(22JCYYZD12)
东部战区总医院院管课题(2023JCYJDZ079)
通信作者: 王 栋, Email: doyle_wd@126.com
宋 勇, Email: yong_song6310@yahoo.com

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更新日期/Last Update: 2024-12-25