Longitudinal bioluminescence imaging of the dynamics of Doxorubicin induced apoptosis.
Longitudinal bioluminescence imaging of the dynamics of Doxorubicin induced apoptosis.
复制标题
阿霉素诱导细胞凋亡动力学的纵向生物发光成像
作者：
Niu G;Zhu L;Ho DN;Zhang F;Gao H;Quan Q;Hida N;Ozawa T;Liu G;Chen X
目的：大多数化疗药物主要通过诱导细胞凋亡引起肿瘤细胞死亡。在体内非侵入性成像细胞凋亡的能力可以显着受益于临床前和临床评价的化疗药物靶向细胞凋亡途径。本研究的目的是可视化的动态凋亡过程中的时间生物发光成像（BLI）使用凋亡特异性生物发光报告基因。研究方法：UM-SCC-22 B人头颈部鳞状细胞癌细胞和4 T1鼠乳腺癌细胞均用胱天蛋白酶-3特异性环萤火虫荧光素酶报告基因（pcFluc-DEVD）进行遗传修饰。用Annexin V染色和BLI检测不同浓度阿霉素诱导的细胞凋亡。在多柔比星或Doxil治疗后的不同时间点，在异种移植肿瘤模型中进行纵向BLI，以评估细胞凋亡。成像后，使用TUNEL染色在冷冻肿瘤切片中评估凋亡细胞中的DNA片段化。结果如下：在pcFluc-DEVD转染的UM-SCC-22 B和4 T1细胞中，阿霉素诱导的剂量和时间依赖性凋亡通过BLI可视化和定量。通过胱天蛋白酶活性测定和GloTM荧光素酶测定证实胱天蛋白酶-3活化。在22 B-pcFluc-DEVD异种移植肿瘤中，一剂阿霉素治疗早在治疗后24小时就诱导BLI强度急剧增加。在前3天观察到持续的信号增加，并且来自离体TUNEL染色的荧光信号与BLI成像结果一致。长期成像显示，BLI信号持续增加，并在用一剂Doxil治疗后约12天达到最大值。结论：以pcFluc-DEVD为报告基因的凋亡BLI有助于以实时方式确定凋亡过程的动力学，这为药物开发和治疗反应监测提供了独特的工具。
Objectives: Most chemotherapy agents cause tumor cell death primarily by the induction of apoptosis. The ability to noninvasively image apoptosis in vivo could dramatically benefit pre-clinical and clinical evaluation of chemotherapeutics targeting the apoptotic pathway. This study aims to visualize the dynamics of apoptotic process with temporal bioluminescence imaging (BLI) using an apoptosis specific bioluminescence reporter gene. Methods: Both UM-SCC-22B human head and neck squamous carcinoma cells and 4T1 murine breast cancer cells were genetically modified with a caspase-3 specific cyclic firefly luciferase reporter gene (pcFluc-DEVD). Apoptosis induced by different concentrations of doxorubicin in the transfected cells was evaluated by both annexin V staining and BLI. Longitudinal BLI was performed in xenografted tumor models at different time points after doxorubicin or Doxil treatment, to evaluate apoptosis. After imaging, DNA fragmentation in apoptotic cells was assessed in frozen tumor sections using TUNEL staining. Results: Dose- and time-dependent apoptosis induced by doxorubicin in pcFluc-DEVD transfected UM-SCC-22B and 4T1 cells was visualized and quantified by BLI. Caspase-3 activation was confirmed by both caspase activity assay and GloTM luciferase assay. One dose of doxorubicin treatment induced a dramatic increase in BLI intensity as early as 24 h after treatment in 22B-pcFluc-DEVD xenografted tumors. Sustained signal increase was observed for the first 3 days and the fluorescent signal from ex vivo TUNEL staining was consistent with BLI imaging results. Long-term imaging revealed that BLI signal consistently increased and reached a maximum at around day 12 after the treatment with one dose of Doxil. Conclusions: BLI of apoptosis with pcFluc-DEVD as a reporter gene facilitates the determination of kinetics of the apoptotic process in a real-time manner, which provides a unique tool for drug development and therapy response monitoring.
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影响因子：
11.2
作者：
Medarova Z;Rashkovetsky L;Pantazopoulos P;Moore A
通讯作者：
Moore A
DOI：
10.1513/pats.200901-001aw
发表时间：
2009-08-15
期刊：
Proceedings of the American Thoracic Society
影响因子：
--
作者：
Blankenberg, Francis G
通讯作者：
Blankenberg, Francis G
影响因子：
11.5
作者：
Lee, Kuei C.;Hamstra, Daniel A.;Rehemtulla, Alnawaz
通讯作者：
Rehemtulla, Alnawaz
影响因子：
--
作者：
Antczak C;Takagi T;Ramirez CN;Radu C;Djaballah H
通讯作者：
Djaballah H
影响因子：
2.6
作者：
Meijer, AE;Jernberg, ARM;Edgren, MR
通讯作者：
Edgren, MR