Department of Ultrasound, China-Japan Friendship Hospital, Beijing, China.
National Center for Respiratory Medicine, National Clinical Research Center for Respiratory Diseases, Institute of Respiratory Medicine of Chinese Academy of Medical Sciences, Beijing, China.
Front Endocrinol (Lausanne). 2023 Mar 3;14:1052862. doi: 10.3389/fendo.2023.1052862. eCollection 2023.
Accurate diagnosis of highly aggressive papillary thyroid cancer (PTC) may greatly help avoid overdiagnosis and overtreatment of PTC. However, there is still a lack of a convenient and accurate method. Targeted microbubbles, an emerging ultrasound contrast agent, have the potential to accurately diagnose highly aggressive PTC.
To design and prepare a targeted microbubble for specific contrast-enhanced ultrasound (CEUS) imaging of highly invasive PTC.
Using β-galactoside-binding protein galectin-3 (Gal-3) overexpressed on the surface of highly invasive PTC cells as a target, C12 polypeptide (ANTPCGPYTHDCPVKR) with high affinity and specificity for Gal-3 was coupled to the surface of lipid microbubbles to prepare targeted microbubbles (Gal-3-C12@lipo MBs). The targeted microbubbles were prepared by thin-film hydration method and mechanical shaking method. The morphology, diameter, concentration and stability of microbubbles were investigated by fluorescence microscopy and an AccuSizer. The biosafety of microbubbles was studied using BCPAP cells through CCK8 assay. Confocal laser scanning microscope and flow cytometry were applied to research the cellular uptake of microbubbles to investigate the targeting ability to highly aggressive PTC. Finally, the specific contrast-enhanced ultrasound imaging of microbubbles in highly invasive PTC was validated on the mice bearing subcutaneous BCPAP tumor model a clinically ultrasound imaging system.
Gal-3-C12@lipo MBs were successfully prepared which showed a well-defined spherical morphology with an average diameter of 1.598 ± 0.848 μm. Gal-3-C12@lipo MBs showed good stability without rupture within 4 hours after preparation. At the cellular level, Gal-3-C12@lipo MBs exhibited favorable biosafety and superior targeting ability to BCPAP cells, with 2.8-fold higher cellular uptake than non-targeted lipid microbubbles (Lipo MBs). At the animal level, Gal-3-C12@lipo MBs significantly improved the quality of contrast-enhanced ultrasound imaging in highly invasive PTC, with an echo intensity of tumor significantly higher than that of Lipo MBs.
We designed and fabricated a novel targeted microbubble for the specific ultrasound imaging diagnosis of highly aggressive PTC. The targeted microbubbles have good stability, superior biosafety and high targeting specificity, which can significantly improve the tumor signal-to-noise ratio of highly invasive PTC, and have the potential to facilitate and accurately diagnose highly invasive PTC.
准确诊断侵袭性强的甲状腺乳头状癌（PTC）可能有助于避免 PTC 的过度诊断和过度治疗。然而，目前仍然缺乏一种便捷且准确的方法。靶向微泡，一种新兴的超声造影剂，具有准确诊断侵袭性强的 PTC 的潜力。
设计并制备一种针对侵袭性强的 PTC 的特定对比增强超声（CEUS）成像的靶向微泡。
利用高侵袭性 PTC 细胞表面过表达的β-半乳糖苷结合蛋白半乳糖凝集素-3（Gal-3）作为靶点，将对 Gal-3 具有高亲和力和特异性的 C12 多肽（ANTPCGPYTHDCPVKR）偶联到脂质微泡表面，制备靶向微泡（Gal-3-C12@lipo MBs）。采用薄膜水化法和机械振摇法制备靶向微泡。通过荧光显微镜和 AccuSizer 观察微泡的形态、直径、浓度和稳定性。通过 CCK8 检测研究微泡的生物安全性，采用 BCPAP 细胞。应用共聚焦激光扫描显微镜和流式细胞术研究微泡对高侵袭性 PTC 的细胞摄取，以研究其对高侵袭性 PTC 的靶向能力。最后，在皮下种植 BCPAP 肿瘤模型的小鼠上验证微泡的特异性对比增强超声成像，使用临床超声成像系统。
成功制备了 Gal-3-C12@lipo MBs，其具有良好的定义的球形形态，平均直径为 1.598±0.848μm。Gal-3-C12@lipo MBs 在制备后 4 小时内无破裂，表现出良好的稳定性。在细胞水平上，Gal-3-C12@lipo MBs 表现出良好的生物安全性和对 BCPAP 细胞的优越靶向能力，细胞摄取率比非靶向脂质微泡（Lipo MBs）高 2.8 倍。在动物水平上，Gal-3-C12@lipo MBs 显著提高了侵袭性 PTC 的对比增强超声成像质量，肿瘤回声强度明显高于 Lipo MBs。
我们设计并制备了一种新型的靶向微泡，用于侵袭性强的 PTC 的特定超声成像诊断。靶向微泡具有良好的稳定性、优越的生物安全性和高靶向特异性，可显著提高侵袭性 PTC 的肿瘤信噪比，有望促进和准确诊断侵袭性 PTC。
