光动力疗法对喉癌HEp-2细胞迁移和侵袭的影响
发布时间：2019-02-23 19:56
【摘要】：目的：光动力疗法(Photodynamic Therapy, PDT)是借助某种特定的药物(称为光敏剂)进入患者体内后,富集于生长异常的组织(如肿瘤),光敏剂经一定波长光的辐照后发生光动力敏化反应而产生活性氧(reactive oxygen species, ROS),导致生物大分子光氧化失活,并由此造成细胞损伤以达到破坏目标病变组织的目的。我们通过本实验评估PDT对喉癌HEp-2细胞迁移与侵袭的影响,探讨PDT诱导的ROS在抑制喉癌HEp-2细胞迁移与侵袭中的作用机制。 方法：不同浓度的光敏剂9-羟基脱镁叶绿酸甲酯a(9-hydroxypheophorbide a,9-HPbD)与HEp-2细胞共培养6小时,随之以激光照射15分钟(波长664nm,能量密度2.0J/cm2),培养24小时后,MTT法测定Hep-2细胞的活性,确定亚致死剂量。利用还原型谷胱甘肽(GSH)抑制ROS的产生,将HEp-2细胞分为4组：正常对照组(无光动力疗法干预,无GSH干预)、光动力组(有光动力疗法干预,无GSH干预)、GSH组(无光动力疗法干预,有GSH干预)、光动力加GSH组(有光动力疗法干预,有GSH干预)。H2DCFDA探针检测细胞内活性氧水平；划痕试验、transwell小室侵袭试验检测亚致死剂量光动力疗法处理24h后HEp-2细胞的迁移与侵袭能力；Western blotting对比分析MEKl/2和ERK1/2的磷酸化水平及MMP-2和MMP-9的表达情况。 结果：9-HPbD介导的PDT对人喉癌HEp-2细胞具有明显杀伤效应,且与药物浓度有关。PDT诱导喉癌HEp-2细胞产生ROS,而GSH抑制PDT介导的ROS的产生。细胞划痕试验、transwell小室迁移和侵袭试验显示PDT抑制喉癌HEp-2细胞的迁移与侵袭能力,而GSH使PDT抑制喉癌HEp-2细胞的迁移与侵袭的能力下降。Western显示光动力抑制喉癌HEp-2细胞中MEK1/2和ERK1/2的磷酸化,下调喉癌HEp-2细胞MMP-2和MMP-9的表达,而GSH可以逆转这一结果。 结论：光动力产生的ROS抑制MEK1/2和ERKl/2的磷酸化,下调MMP-2和MMP-9的表达,进而抑制喉癌HEp-2细胞的迁移与侵袭。
[Abstract]:Objective: photodynamic therapy (Photodynamic Therapy, PDT) is the use of a specific drug (known as Guang Min) into the body of patients, enriched in abnormal growth tissue (such as tumor), The reactive oxygen species (reactive oxygen species, ROS),) produced by the photodynamic sensitization reaction of Guang Min after irradiation with a certain wavelength of light caused photooxidation inactivation of biomolecules and caused cell damage in order to destroy the target pathological tissues. We evaluated the effect of PDT on the migration and invasion of HEp-2 cells in laryngeal carcinoma and explored the mechanism of PDT induced ROS in inhibiting the migration and invasion of HEp-2 cells. Methods: HEp-2 cells were co-cultured with different concentrations of Guang Min (9-hydroxypheophorbide a 9-HPbD) for 6 hours and then irradiated with laser for 15 minutes (wavelength 664 nm, energy density 2.0J/cm2). After 24 hours of culture, the activity of Hep-2 cells was determined by MTT assay, and the sublethal dose was determined. Using reduced glutathione (GSH) to inhibit the production of ROS, HEp-2 cells were divided into four groups: normal control group (no photodynamic therapy intervention, no GSH intervention), photodynamic group (photodynamic therapy intervention, no GSH intervention). GSH group (no photodynamic therapy intervention, GSH intervention), photodynamic therapy plus GSH group (photodynamic therapy intervention, GSH intervention). H2DCFDA probe detection of intracellular reactive oxygen species; The migration and invasiveness of HEp-2 cells were detected by transwell chamber invasion test and sublethal photodynamic therapy for 24 hours. The phosphorylation levels of MEKl/2 and ERK1/2 and the expression of MMP-2 and MMP-9 were compared between MEKl/2 and ERK1/2 by; Western blotting. Results: 9-HPbD mediated PDT had a significant killing effect on human laryngeal cancer HEp-2 cells and was related to drug concentration. PDT induced ROS, production of HEp-2 cells and GSH inhibited PDT mediated ROS production. Cell scratch test, transwell chamber migration and invasion test showed that PDT inhibited the migration and invasion of laryngeal carcinoma HEp-2 cells. GSH decreased the ability of PDT to inhibit the migration and invasion of laryngeal carcinoma HEp-2 cells. Western showed photodynamic inhibition of MEK1/2 and ERK1/2 phosphorylation, and down-regulated the expression of MMP-2 and MMP-9 in HEp-2 cells. GSH can reverse this result. Conclusion: photodynamic ROS inhibits the phosphorylation of MEK1/2 and ERKl/2, down-regulates the expression of MMP-2 and MMP-9, and inhibits the migration and invasion of HEp-2 cells.
【学位授予单位】：复旦大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R739.65
本文编号：2429143
[Abstract]:Objective: photodynamic therapy (Photodynamic Therapy, PDT) is the use of a specific drug (known as Guang Min) into the body of patients, enriched in abnormal growth tissue (such as tumor), The reactive oxygen species (reactive oxygen species, ROS),) produced by the photodynamic sensitization reaction of Guang Min after irradiation with a certain wavelength of light caused photooxidation inactivation of biomolecules and caused cell damage in order to destroy the target pathological tissues. We evaluated the effect of PDT on the migration and invasion of HEp-2 cells in laryngeal carcinoma and explored the mechanism of PDT induced ROS in inhibiting the migration and invasion of HEp-2 cells. Methods: HEp-2 cells were co-cultured with different concentrations of Guang Min (9-hydroxypheophorbide a 9-HPbD) for 6 hours and then irradiated with laser for 15 minutes (wavelength 664 nm, energy density 2.0J/cm2). After 24 hours of culture, the activity of Hep-2 cells was determined by MTT assay, and the sublethal dose was determined. Using reduced glutathione (GSH) to inhibit the production of ROS, HEp-2 cells were divided into four groups: normal control group (no photodynamic therapy intervention, no GSH intervention), photodynamic group (photodynamic therapy intervention, no GSH intervention). GSH group (no photodynamic therapy intervention, GSH intervention), photodynamic therapy plus GSH group (photodynamic therapy intervention, GSH intervention). H2DCFDA probe detection of intracellular reactive oxygen species; The migration and invasiveness of HEp-2 cells were detected by transwell chamber invasion test and sublethal photodynamic therapy for 24 hours. The phosphorylation levels of MEKl/2 and ERK1/2 and the expression of MMP-2 and MMP-9 were compared between MEKl/2 and ERK1/2 by; Western blotting. Results: 9-HPbD mediated PDT had a significant killing effect on human laryngeal cancer HEp-2 cells and was related to drug concentration. PDT induced ROS, production of HEp-2 cells and GSH inhibited PDT mediated ROS production. Cell scratch test, transwell chamber migration and invasion test showed that PDT inhibited the migration and invasion of laryngeal carcinoma HEp-2 cells. GSH decreased the ability of PDT to inhibit the migration and invasion of laryngeal carcinoma HEp-2 cells. Western showed photodynamic inhibition of MEK1/2 and ERK1/2 phosphorylation, and down-regulated the expression of MMP-2 and MMP-9 in HEp-2 cells. GSH can reverse this result. Conclusion: photodynamic ROS inhibits the phosphorylation of MEK1/2 and ERKl/2, down-regulates the expression of MMP-2 and MMP-9, and inhibits the migration and invasion of HEp-2 cells.
【学位授予单位】：复旦大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R739.65
【参考文献】
相关期刊论文 前3条
1 罗国仪，孙振权，，王洪涛;染料激光和Ar￣＋激光进行光动力学治疗鼻咽癌108例疗效分析[J];癌症;1995年05期
2 杜波,王苹,杜宝东,郭晓峰;MMP-2和MMP-9在喉癌组织中的表达及其与预后的关系[J];吉林大学学报(医学版);2002年06期
3 李黎波,罗荣城,廖旺军,张鸣江,刘晓军,罗宇玲,缪景霞;PHOTOFRIN光动力治疗喉癌及改善发声功能的疗效[J];中国临床康复;2005年02期
本文编号：2429143
本文链接：https://www.wllwen.com/yixuelunwen/yank/2429143.html
最近更新
教材专著
