神经纤毛蛋白-1靶向性分子探针在裸鼠异位脑胶质瘤分级诊断中的应用

Application of NRP-1 targeting molecular probe in grading diagnosis of heterotopic brain glioma in nude mice

目的观察神经纤毛蛋白-1(neuropilin-1,NRP-1)在不同级别脑胶质瘤细胞中的表达;通过磁共振成像(MRI)探讨新型分子探针(USPIO-PEG-tLyP-1)在裸鼠异位脑胶质瘤分级诊断中的应用价值。方法利用Western blot检测不同级别脑胶质瘤细胞系中NRP-1表达水平;利用碳二亚胺法合成USPIO-PEG-tLyP-1;建立U87-MG荷瘤鼠模型(U87-MG组)、CHG-5荷瘤鼠模型(CHG-5组)各10只,从两组中各选取移植瘤体积约0.6 cm3的荷瘤鼠6只,分别经尾静脉给予2 mg/kg分子探针,在0 h、6 h、12 h、24 h进行磁共振检测,计算出R2值;成像结束后处死荷瘤鼠,制作肿瘤组织切片,行普鲁士蓝染色检测铁颗粒含量,比较两组瘤组织与分子探针的结合能力。结果U87-MG、CHG-5细胞系中NRP-1表达水平明显高于人星形胶质细胞(HA),且NRP-1在U87-MG中的表达高于在CHG-5细胞中的表达(P<0.01)。MRI结果:注射分子探针前,两组瘤组织R2值比较,差异无统计学意义[U87-MG组:(10.35±0.52),CHG-5组:(9.86±0.43),t=1.779,P=0.106];注射分子探针后,与CHG-5组比较,U87-MG组瘤组织R2值较高[6 h:U87-MG组(11.63±0.85),CHG-5组(10.51±0.49),t=2.796,P=0.019;12 h:U87-MG组(14.23±0.68),CHG-5组(12.29±0.28),t=6.462,P=0.000;24 h: U87-MG组(13.36±0.92),CHG-5组(11.32±0.64),t=4.459,P=0.001]。普鲁士蓝染色结果:U87-MG组瘤组织蓝染颗粒明显多于CHG-5组,差异有统计学意义(P<0.01)。结论神经纤毛蛋白-1靶向性分子探针可以对不同恶性程度的裸鼠异位脑胶质瘤进行分级诊断。

Objective To observe the expression of neuropilin-1(NRP-1)in glioma cells of different grades, and evaluate the application value of a novel molecular probe(USPIO-PEG-tLyP-1)in the grading diagnosis of heterotopic glioma in nude mice by magnetic resonance imaging(MRI). Methods Expression levels of NRP-1 in glioma cell lines of different grades were detected by Western-Blot.USPIO-PEG-tLyP-1 was synthesized by carbon diimine method.The U87-MG tumor-bearing mice model(U87-MG group)and CHG-5 tumor-bearing mice model(CHG-5 group) were established with 10 mice in each group.Six tumor-bearing mice with a tumor volume about 0.6 cm3 were selected from each group, and they were given with 2mg/kg molecular probes via tail vein respectively and was detected by MRI at 0 h, 6 h, 12 h and 24 h, then R2 values were calculated.After the imaging, tumor-bearing mice were sacrificed, and tumor tissue sections were made.The iron particles in the sections was detected by Prussian blue staining.The binding ability of molecular probes and tumor tissues in the two groups was compared. Results The expression of NRP-1 in U87-MG and CHG-5 cell lines was significantly higher than that in HA.In addition, the expression of NRP-1 in U87-MG was higher than that in CHG-5 cell(P<0.01). MRI results showed that R2 values of tumor tissues in the two groups were compared, and the difference was not statistically significant before the injection of molecular probe(U87-MG group(10.35±0.52)vs CHG-5 group(9.86±0.43), t=1.779, P=0.106). The R2 value of tumor tissue in the U87-MG group was higher than that in the CHG-5 group after the injection of molecular probe(6 h: U87-MG group(11.63±0.85)vs CHG-5 group(10.51±0.49), t=2.796, P=0.019;12 h: U87-MG group(14.23±0.68)vs CHG-5 group(12.29±0.28), t=6.462, P=0.000;24 h: U87-MG group(13.36±0.92)vs CHG-5 group(11.32±0.64), t=4.459, P=0.001). The results of Prussian blue staining showed that there were significantly more blue staining particles in tumor tissues of the U87-MG group than that of the CHG-5 group, and the difference was statistically significant(P<0.01). Conclusion The NRP-1 targeted molecular probe can be used for grading diagnosis of high and low grade heterotopic brain glioma in nude mice.