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.

中文翻译：

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炎症肠壁的体内可视化和表征：靶向微泡在评估 NF-κB 表达中的探索

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。我们旨在开发靶向 NF-κB 的微泡 (MBs) 并进行分子对比增强超声 (CEUS) 以量化 IBD 小鼠体内肠壁上 NF-κB 的表达。在这项研究中，通过连接负载生物素的 NF-κB 抗体和负载抗生物素蛋白的 MB 来制造靶向 NF-κB 的 MB。NF-κB 靶向 MBs 呈透明圆形气泡，平均直径为 1.03/μm±0.01。通过流式细胞术、Western blot和免疫荧光等体外实验验证了靶向MBs与炎性细胞的特异性结合，揭示了靶向MBs与炎性细胞的特异性结合。随后，对患有化学性结肠炎的小鼠进行 NF-κB 靶向 CEUS 成像，并对峰强度 (PI) 和达峰时间 (TTP) 进行量化。进一步实施了病理和免疫组化（IHC）检查。对于目标 CEUS 组，观察到快速增强后缓慢下降。IBD 小鼠靶 CEUS 的 PI 显着高于 IBD 小鼠的非靶 CEUS、健康对照组和治疗 IBD 小鼠的靶 CEUS (34835%[13379–73492%] VS 437%[236–901 %]、130%[79–231%]、528%[274–779%]、p <0.0001)，根据 NF-κB 表达的 IHC 结果。治疗小鼠的目标 CEUS 的 TTP 显着高于未治疗小鼠（35.7s [18.1-49.5s] VS 8.3s [4.2-12.5s]， p <0.0001）。因此，我们建议以 NF-κB 为靶点的 CEUS 可以准确检测和量化 IBD 肠壁上 NF-κB 的表达，从而能够评估肠道炎症。