Acute thrombotic events play a major role in various cardiovascular diseases. Therefore, direct thrombus imaging can be proved beneficial for early diagnosis and prompt therapy of thrombosis. Our study investigated targeted dual‐modality cyclic arginine‐glycine‐aspartic micro bubbles (cRGD‐MBs) for direct imaging of thrombi by fluorescence and ultrasound.
cRGD‐MBs were prepared by mechanical vibration and chemical chelation methods.
Coulter counter analysis demonstrated that the cRGD‐MBs were well dispersed, with diameters ranging from 1 to 3 μm. They emitted bright red fluorescence under an excitation wavelength of 660 nm. In vivo fluorescence and ultrasound imaging revealed that cRGD‐MBs accumulated at the site of thrombus in the carotid artery with significant fluorescence and ultrasonic signal.
This study showed that novel microbubble cRGD‐MBs were successfully synthesized, and that these could potentially be used as contrast agents for immediate diagnosis of acute thrombus in vivo.
Lopez AD, Mathers CD, Ezzati M, Jamison DT, Murray CJ Global and regional burden of disease and risk factors, 2001: systematic analysis of population health data. Lancet. 2006;367(9524):1747–57. https://doi.org/10.1016/S0140-6736(06)68770-9
Furie B, Furie BC Mechanisms of thrombus formation. N Engl J Med. 2008;359(9):938–49. https://doi.org/10.1056/NEJMra0801082
Nighoghossian N, Derex L, Douek P The vulnerable carotid artery plaque: current imaging methods and new perspectives. Stroke. 2005;36(12):2764–72. https://doi.org/10.1161/01.STR.0000190895.51934.43
Qiao R, Qiao H, Zhang Y, Wang Y, Chi C, Tian J, et al Molecular imaging of vulnerable atherosclerotic plaques in vivo with osteopontin‐specific upconversion nanoprobes. ACS Nano. 2017;11(2):1816–25. https://doi.org/10.1021/acsnano.6b07842
Wang Y, Zhang Y, Wang Z, Zhang J, Qiao RR, Xu M, et al Optical/MRI dual‐modality imaging of M1 macrophage polarization in atherosclerotic plaque with MARCO‐targeted upconversion luminescence probe. Biomaterials. 2019;219:119378. https://doi.org/10.1016/j.biomaterials.2019.119378
Schutt EG, Klein DH, Mattrey RM, Riess JG Injectable microbubbles as contrast agents for diagnostic ultrasound imaging: the key role of perfluorochemicals. Angew Chem Int Ed Engl. 2003;42(28):3218–35. https://doi.org/10.1002/anie.200200550
Sanz J, Fayad ZA Imaging of atherosclerotic cardiovascular disease. Nature. 2008;451(7181):953–7. https://doi.org/10.1038/nature06803
Vyas SP, Vaidya B Targeted delivery of thrombolytic agents: role of integrin receptors. Expet Opin Drug Deliv. 2009;6(5):499–508. https://doi.org/10.1517/17425240902878002
Bennett JS Structure and function of the platelet integrin alphaIIbbeta3. J Clin Invest. 2005;115(12):3363–9. https://doi.org/10.1172/JCI26989
Ma YQ, Qin J, Plow EF Platelet integrin alpha(IIb)beta(3): activation mechanisms. J Thromb Haemostasis. 2007;5(7):1345–52. https://doi.org/10.1111/j.1538-7836.2007.02537
Kerrigan GN, Buchanan MR, Cade JF, Regoeczi E, Hirsh J Investigation of the mechanism of false positive 125I‐labelled fibrinogen scans. Br J Haematol. 1974;26(3):469–73. https://doi.org/10.1111/j.1365-2141.1974.tb00488
Springer TA, Wang JH The three‐dimensional structure of integrins and their ligands, and conformational regulation of cell adhesion. Adv Protein Chem. 2004;68:29–63. https://doi.org/10.1016/S0065-3233(04)68002-8
Huang G, Zhou Z, Srinivasan R, Penn MS, Kottke‐Marchant K, Marchant RE, et al Affinity manipulation of surface‐conjugated RGD peptide to modulate binding of liposomes to activated platelets. Biomaterials. 2008;29(11):1676–85. https://doi.org/10.1016/j.biomaterials.2007.12.015
Kononova O, Litvinov RI, Blokhin DS, Klochkov VV, Weisel JW, Bennett JS, et al Mechanistic basis for the binding of RGD‐ and AGDV‐peptides to the platelet integrin αIIbβ3. Biochemistry. 2017;56(13):1932–42. https://doi.org/10.1021/acs.biochem.6b01113
Luo BH, Carman CV, Springer TA Structural basis of integrin regulation and signaling. Annu Rev Immunol. 2007;25(1):619–47. https://doi.org/10.1146/annurev.immunol.25.022106.141618
Ziegler M, Hohmann JD, Searle AK, Abraham MK, Nandurkar HH, Wang X, et al A single‐chain antibody‐CD39 fusion protein targeting activated platelets protects from cardiac ischaemia/reperfusion injury. Eur Heart J. 2018;39(2):111–6. https://doi.org/10.1093/eurheartj/ehx218
Bonnard T, Tennant Z, Niego B, Kanojia R, Alt K, Jagdale S, et al Novel thrombolytic drug based on thrombin cleavable microplasminogen coupled to a single‐chain antibody specific for activated GPⅡb/Ⅲa. J Am Heart Assoc. 2017;6(2). https://doi.org/10.1161/JAHA.116.004535
Srinivasan R, Marchant RE, Gupta AS In vitro and in vivo platelet targeting by cyclic RGD‐modified liposomes. J Biomed Mater Res. 2010;93(3):1004–15. https://doi.org/10.1002/jbm.a.32549
Ntziachristos V, Bremer C, Weissleder R Fluorescence imaging with near‐infrared light: new technological advances that enable in vivo molecular imaging. Eur Radiol. 2003;13(1):195–208. https://doi.org/10.1007/s00330-002-1524-x
Frangioni JV In vivo near‐infrared fluorescence imaging. Curr Opin Chem Biol. 2003;7(5):626–34. https://doi.org/10.1016/j.cbpa.2003.08.007
Jin B, Lin M, Zong Y, Wan M, Xu F, Duan Z, et al. Microbubble embedded with upconversion nanoparticles as a bimodal contrast agent for fluorescence and ultrasound imaging. Nanotechnology. 2015;26(34):345601. https://doi.org/10.1088/0957-4484/26/34/345601
Wang X, Hagemeyer CE, Hohmann JD, Leitner E, Armstrong PC, Jia F, et al Novel single‐chain antibody‐targeted microbubbles for molecular ultrasound imaging of thrombosis: validation of a unique noninvasive method for rapid and sensitive detection of thrombi and monitoring of success or failure of thrombolysis in mice. Circulation. 2012;125(25):3117–26. https://doi.org/10.1161/CIRCULATIONAHA.111.030312
Wang X, Gkanatsas Y, Palasubramaniam J, Hohmann JD, Chen YC, Lim B, et al Thrombus‐targeted theranostic microbubbles: a new Technology towards concurrent rapid ultrasound diagnosis and bleeding‐free fibrinolytic treatment of thrombosis. Theranostics. 2016;6(5):726–38. https://doi.org/10.7150/thno.14514.eCollection2016
Maier A, Plaza‐Heck P, Meixner F, Guenther F, Kaufmann BA, Kramer M, et al A molecular intravascular ultrasound contrast agent allows detection of activated platelets on the surface of symptomatic human plaques. Atherosclerosis. 2017;267:68–77. https://doi.org/10.1016/j.atherosclerosis.2017.10.029
Jin Y, Ma X, Feng S, Liang X, Dai Z, Tian J, et al Hyaluronic acid modified tantalum oxide nanoparticles conjugating doxorubicin for targeted cancer theranostics. Bioconjugate Chem. 2015;26(12):2530–41. https://doi.org/10.1021/acs.bioconjchem.5b00551
Jin Y, Yang X, Tian J Targeted polypyrrole nanoparticles for the identification and treatment of hepatocellular carcinoma. Nanoscale. 2018;10(20):9594–601. https://doi.org/10.1039/c8nr02036a
Jin Y, Ma X, Zhang S, Meng H, Xu M, Yang X, et al A tantalum oxide‐based core/shell nanoparticle for triple‐modality image‐guided chemo‐thermal synergetic therapy of esophageal carcinoma. Cancer Lett. 2017;397:61–71. https://doi.org/10.1016/j.canlet.2017.03.030
Li S, Gou T, Wang Q, Chen M, Chen Z, Xu M, et al Ultrasound/optical dual‐modality imaging for evaluation of vulnerable atherosclerotic plaques with osteopontin targeted nanoparticles. Macromol Biosci. 2020;20(2):e1900279. https://doi.org/10.1002/mabi.201900279
Karasev MM, Stepanenko OV, Rumyantsev KA, Turoverov KK, Verkhusha VV. Near‐infrared fluorescent proteins and their applications. Biochemistry (Mosc). 2019;84((Suppl 1)):S32–50. https://doi.org/10.1134/S0006297919140037
Hong G, Antaris A, Dai H Near‐infrared fluorophores for biomedical imaging. Nat Biomed Eng. 2017;1:0010. https://doi.org/10.1038/s41551-016-0010
Stride E, Saffari N Microbubble ultrasound contrast agents: a review. Proc Inst Mech Eng H. 2003;217(6):429–47. https://doi.org/10.1243/09544110360729072
Qu E, Dai Z, Liang X, Qian Y, Wang S, Ke H, et al Detection and pathologic evaluation of sentinel lymph nodes in the VX2 tumor model using a novel ultrasound/near‐infrared dual‐modality contrast agent. Ultrasound Med Biol. 2015;41(7):1905–12. https://doi.org/10.1016/j.ultrasmedbio.2015.03.005
Pillai R, Marinelli ER, Fan H, Nanjappan P, Song B, von Wronski MA, et al A phospholipid‐PEG2000 conjugate of a vascular endothelial growth factor receptor 2 (VEGFR2)‐targeting heterodimer peptide for contrast‐enhanced ultrasound imaging of angiogenesis. Bioconjugate Chem. 2010;21(3):556–62. https://doi.org/10.1021/bc9005688
Hu G, Liu C, Liao Y, Yang L, Huang R, Wu J, et al Ultrasound molecular imaging of arterial thrombi with novel microbubbles modified by cyclic RGD in vitro and in vivo. Thromb Haemostasis. 2012;107(1):172–83. https://doi.org/10.1160/TH10-11-0701
Wu W, Wang Y, Shen S, Wu J, Guo S, Su L, et al In vivo ultrasound molecular imaging of inflammatory thrombosis in arteries with cyclic Arg‐Gly‐Asp‐modified microbubbles targeted to glycoprotein IIb/Ⅲa. Invest Radiol. 2013;48(11):803–12. https://doi.org/10.1097/RLI.0b013e318298652d
Tartis MS, Kruse DE, Zheng H, Zhang H, Kheirolomoom A, Marik J, et al Dynamic microPET imaging of ultrasound contrast agents and lipid delivery. J Contr Release. 2008;131(3):160–6. https://doi.org/10.1016/j.jconrel.2008.07.030
Sachs UJ, Nieswandt B In vivo thrombus formation in murine models. Circ Res. 2007;100(7):979–91. https://doi.org/10.1161/01.RES.0000261936.85776.5f
Farrehi PM, Ozaki CK, Carmeliet P, Fay WP Regulation of arterial thrombolysis by plasminogen activator inhibitor‐1 in mice. Circulation. 1998;97(10):1002–8. https://doi.org/10.1161/01.cir.97.10.1002