以基质金属蛋白酶-2为靶向的荧光共振能量转移探针制备及其对肝癌细胞的检测

Preparation of the fluorescence resonance energy transfer probe targeting matrix metalloproteinase-2 and its detection of hepatoma cells

目的构建以基质金属蛋白酶-2(MMP-2)为靶向的聚乙二醇(PEG)修饰的荧光共振能量转移探针,探讨其用于肝癌细胞成像检测的可行性。方法将荧光基团7-(N,N-二乙基氨基)香豆素-3-羧酸琥珀酰亚胺酯通过能够被MMP-2切割的多肽链(GPLGVRGKGG)连接淬灭基团4-[4-(二甲基氨基)苯偶氮]苯甲酸N-丁二酰亚胺酯,构建荧光共振能量转移(FRET)探针。以人肝癌7402细胞和人正常肝细胞L-O2为研究对象,通过细胞成像实验检测人肝癌7402细胞和L-O2细胞中MMP-2的表达水平。结果体外实验(MMP-2、MMP-1及其他物种对探针的荧光响应)显示了探针对MMP-2的靶向性;体内实验(细胞成像)结果显示,人肝癌7402细胞中MMP-2表达水平高于人正常肝细胞L-O2。结论该实验制备的探针可以检测人肝癌7402细胞和人正常肝细胞L-O2中MMP-2的表达,将有望用于肝癌的筛查、诊断和预后判断。

Objective To prepare the polyethylene glycol（ PEG） modified fluorescence resonance energy transfer probe targeting matrix metalloproteinase-2（ MMP-2） and to explore the feasibility of it in hepatoma carcinoma cell imaging. Methods The fluorescence resonance energy transfer probe targeting MMP-2 was prepared by coupling a fluorophore [7-（ diethylamino）-2-oxo-2H-chromene-3-succinimidyl ester] with a quencher{ 4-[4-（ dimethylamino） phenylazo]benzoic acid N-succinimidyl ester} via a polypeptide chain（ GPLGVRGKGG） that could be cleaved by MMP-2. Human hepatoma 7402 cells and normal human liver L-O2 cells were taken as research object and cell imaging was done to detect the MMP-2 expression levels in hepatoma cells. Results In vitro experiments（ the fluorescent probe responses for MMP-2,MMP-1 and other species） showed that the probe targeted MMP-2 and in vivo experiments（ cell imaging） results showed that the MMP-2 expression level in human hepatoma 7402 cells was higher than that of normal human liver L-O2 cells. Conclusion The MMP-2 probe could be used to examine the MMP-2 expression in human hepatoma 7402 cells and human normal liver L-O2 cells,which was expected to be used for hepatoma screening,diagnosis and prognosis.