携精氨酸-甘氨酸-天冬氨酸肽序列靶向超声微泡结合对比超声评价肿瘤新生血管

Evaluation of tumor angiogenesis using microbubbles conjugated with RGD peptides and contrast enhanced ultrasound

目的 探讨携精氨酸-甘氨酸-天冬氨酸（RGD）肽序列靶向超声微泡结合对比超声评价肿瘤新生血管的可行性.方法 应用声振法制备普通脂质微泡（MB）,通过共价偶联的方法,将RGD肽序列连接在MB外壳,制备成靶向超声微泡（MBRGD）.6只接种人肝癌HepG2细胞14 d的裸鼠（模型组）和6只未接种HepG2细胞的正常裸鼠（对照组）,经尾静脉弹丸式注射等量MB和MBRGD,两次注射间隔30 min.注射10 min后,分别行肿瘤和骨骼肌的对比超声检查,测量肿瘤和骨骼肌的第1帧对比超声图像的声强度（VI）,并对肿瘤组织和骨骼肌组织进行抗αvβ3-整合免疫组化检测.结果 模型组注射MBRGD和MB后,肿瘤第1帧对比超声图像分别显示显著及轻度的超声显影,VI值分别为17.03±3.58和5.33±1.71,注射MBRGD后的VI值是注射MB的3.18倍（P＜0.05）.对照组注射MBRGD和MB后,骨骼肌第1帧对比超声图像均无明显超声显影,VI值分别为3.10±0.48和2.90±0.29,两者差异无统计学意义（P＞0.05）.免疫组化检测显示,αvβ3-整合素在肿瘤新生血管内皮高度表达,而骨骼肌组织血管内皮却未见明显表达.结论 应用MBRGD行对比超声检查可有效评价肿瘤的新生血管,将有助于肿瘤性质的早期判断.

Objective To assess the feasibility of usage of microbubbles conjugated with RGD peptides and contrast enhanced ultrasound （CEU） in detection of tumor angiogenesis. Methods Lipid microbubbles （MB） were prepared, and the RGD peptides were covalently conjugated to the lipid shell of MB （MBRGD）. Six nude mice with tumor created by dorsal inoculation of HepG2 tumor cells were used as the test group. Six nude mice without tumor were served as the control group. 10 minutes after bolus injection of MB and MBRGD randomly （30 min interval） via a tail vein catheter, CEU was performed on the tumors of the test group and the thigh skeletal muscles of control group. The video intensity （VI） of tumors and the skeletal muscles were measured. The tumors and the skeletal muscles were harvested for immunohistochemical examination. Results Only a slight contrast enhancement of the tumor was seen with MB, and the VI was 5.33 ± 1.71. While a remarkable enhancement of the tumor was observed after injection of MBRGD. The VI was up to 17.03 ± 3.58, 3.18 folds higher as compared with that obtained by injection of MB （P 〈 0. 05）. As expected, there were no obvious contrast enhancement of the skeletal muscles with both MBRGD and MB. There was a high expression of αvβ3-integrin in tumor neovascular endothelium, however, no apparent expression of αvβ3-integrin was observed in the skeletal muscle vascular endothelium. Conclusion CEU with MBRGD can be used to effectively evaluate the angiogenesis of tumors,and it may greatly contribute to the early judgement of the nature of tumor.