Department of Radiology, Translational Research in Ultrasound Theranostics (TRUST) Program, University of Texas Southwestern Medical Center , 5323 Harry Hines Boulevard, Dallas, Texas 75390-8514, United States.
Department of Radiology, University of California, San Diego , La Jolla, California 92093, United States.
ACS Appl Mater Interfaces. 2017 Nov 1;9(43):37587-37596. doi: 10.1021/acsami.7b10592. Epub 2017 Oct 20.
Acute deep vein thrombosis (DVT) is the formation of a blood clot in the deep veins of the body that can lead to fatal pulmonary embolism. Acute DVT is difficult to distinguish from chronic DVT by ultrasound (US), the imaging modality of choice, and is therefore treated aggressively with anticoagulants, which can lead to internal bleeding. Here we demonstrate that conjugating perfluorobutane-filled (PFB-filled) microbubbles (MBs) with thrombin-sensitive activatable cell-penetrating peptides (ACPPs) could lead to the development of contrast agents that detect acute thrombosis with US imaging. Successful conjugation of ACPP to PFB-filled MBs was confirmed by fluorescence microscopy and flow cytometry. Fluorescein-labeled ACPP was used to evaluate the efficiency of thrombin-triggered cleavage by measuring the mean fluorescence intensity of ACPP-labeled MBs (ACPP-MBs) before and after incubation at 37 °C with thrombin. Lastly, control MBs and ACPP-MBs were infused through a tube containing a clot, and US contrast enhancement was measured with or without the presence of a thrombin inhibitor after washing the clot with saline. With thrombin activity, 91.7 ± 14.2% of the signal was retained after ACPP-MB infusion and washing, whereas only 16.7 ± 4% of the signal was retained when infusing ACPP-MBs in the presence of hirudin, a potent thrombin inhibitor.
急性深静脉血栓形成（DVT）是指身体深部静脉中形成的血栓，可导致致命性肺栓塞。超声（US）是首选的成像方式，很难将急性 DVT 与慢性 DVT 区分开来，因此需要积极使用抗凝剂治疗，这可能导致内出血。在这里，我们证明了将全氟丁烷填充（PFB 填充）微泡（MB）与凝血酶敏感的可激活细胞穿透肽（ACPP）缀合，可以开发出用于 US 成像检测急性血栓形成的造影剂。通过荧光显微镜和流式细胞术证实了 ACPP 与 PFB 填充 MB 的成功缀合。使用荧光素标记的 ACPP 通过测量在 37°C 与凝血酶孵育前后 ACPP 标记的 MB（ACPP-MB）的平均荧光强度来评估凝血酶触发切割的效率。最后，在使用盐水冲洗凝块后，在含有凝块的管中注入对照 MB 和 ACPP-MB，并测量 US 对比增强情况，有无凝血酶抑制剂。在存在凝血酶活性的情况下，在 ACPP-MB 输注和冲洗后保留了 91.7±14.2%的信号，而在存在凝血酶抑制剂水蛭素的情况下输注 ACPP-MB 时仅保留了 16.7±4%的信号。
