Department of Ultrasound, Chinese Academy of Medical Sciences and Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing, China.
J Cell Mol Med. 2021 Sep;25(18):8973-8984. doi: 10.1111/jcmm.16858. Epub 2021 Aug 19.
NF-κB, a critical cytokine of inflammatory bowel diseases (IBD), is a viable marker to reflect the inflammatory activity of the intestine. We aimed to develop NF-κB-targeted microbubbles (MBs) and perform molecular contrast-enhanced ultrasound (CEUS) to quantify NF-κB expressions on the intestinal wall in IBD mice in vivo. In this study, NF-κB-targeted MBs were fabricated by connecting biotin-loaded NF-κB antibodies and avidin-loaded MBs. NF-κB-targeted MBs presented as transparent and round bubbles with an average diameter of 1.03/μm±0.01. The specific binding of targeted MBs and inflammatory cells was validated by in vitro experiments, including flow cytometry, Western blot and immunofluorescence, which revealed the specific binding of targeted MBs and inflammatory cells. Subsequently, NF-κB-targeted CEUS imaging was performed on mice with chemical-induced colitis, and the peak intensity (PI) and time-to-peak (TTP) were quantified. Pathological and immunohistochemical (IHC) examinations were further implemented. For the target CEUS group, fast enhancement followed by slow subsiding was observed. The PI of target CEUS of the IBD mice was significantly higher than that of non-target CEUS of the IBD mice, healthy controls and target CEUS of the treated IBD mice (34835%[13379-73492%] VS 437%[236-901%], 130%[79-231%], 528%[274-779%], p<0.0001), in accordance with the IHC results of NF-κB expressions. The TTP of target CEUS of the treated mice was significantly higher than that of untreated mice (35.7s [18.1-49.5s] VS 8.3s [4.2-12.5s], p<0.0001). Therefore, we suggested that NF-κB-targeted CEUS could accurately detect and quantify NF-κB expressions on the intestinal walls of IBD, enabling the evaluation of intestinal inflammation.
NF-κB 是炎症性肠病 (IBD) 的关键细胞因子，是反映肠道炎症活动的可行标志物。本研究旨在开发 NF-κB 靶向微泡 (MB)，并进行分子对比增强超声 (CEUS)，以定量体内 IBD 小鼠肠壁上的 NF-κB 表达。在这项研究中，NF-κB 靶向 MB 通过连接生物素化的 NF-κB 抗体和亲和素化的 MB 来制备。NF-κB 靶向 MB 呈透明圆形气泡，平均直径为 1.03/μm±0.01μm。通过包括流式细胞术、Western blot 和免疫荧光在内的体外实验验证了靶向 MB 与炎症细胞的特异性结合，结果表明靶向 MB 与炎症细胞的特异性结合。随后，对化学诱导结肠炎的小鼠进行 NF-κB 靶向 CEUS 成像，并对峰值强度 (PI) 和达峰时间 (TTP) 进行定量。进一步进行了病理和免疫组织化学 (IHC) 检查。对于目标 CEUS 组，观察到快速增强后缓慢消退。IBD 小鼠的目标 CEUS 的 PI 明显高于 IBD 小鼠的非目标 CEUS、健康对照和治疗后的 IBD 小鼠的目标 CEUS (34835%[13379-73492%] VS 437%[236-901%]、130%[79-231%]、528%[274-779%]，p<0.0001)，与 NF-κB 表达的 IHC 结果一致。治疗组小鼠的 TTP 明显高于未治疗组小鼠 (35.7s [18.1-49.5s] VS 8.3s [4.2-12.5s]，p<0.0001)。因此，我们认为 NF-κB 靶向 CEUS 可以准确检测和定量 IBD 小鼠肠壁上的 NF-κB 表达，从而评估肠道炎症。
