BACKGROUND Antibody-mediated rejection (AMR) constitutes an important cause of cardiac allograft loss; however, all current therapeutic strategies represent systemic applications with unsatisfactory efficacy. Previously, we successfully non-invasively detected C4d, a specific marker for AMR diagnosis, in allografts using C4d-targeted microbubbles (MBC4d). In this study, we extended this approach by incorporating nitric oxide (NO), as high NO levels manifest immunosuppressive and anti-thrombotic effects. METHODS We designed novel MBC4d loaded with NO (NO-MBC4d). A rat model of AMR was established by pre-sensitization with skin transplantation. Contrast-enhanced ultrasound (CEUS) images were obtained and quantitatively analyzed following NO-MBC4d injection. Allograft survival and histologic features were analyzed to evaluate the therapeutic effect and underlying mechanism of NO-MBC4d toward AMR. RESULTS We successfully obtained CEUS images following NO-MBC4d injection and demonstrated that the ultrasound signal intensity of the myocardial area and clearance time of NO-MBC4d both increased with increased C4d grade, thereby realizing non-invasive diagnosis of AMR. Furthermore, allograft survival was significantly prolonged, and rejection was obviously attenuated following NO-MBC4d injection through significant suppression of thrombosis and reduction of inflammatory cell infiltrates. Overall, the therapeutic efficacy was significantly improved in the NO-MBC4d group compared with the control NO-MB group, demonstrating that precise treatment could significantly improve the therapeutic efficacy compared with that afforded by systemic applications. CONCLUSIONS This study presented a novel tool to provide simultaneous non-invasive diagnosis and precise treatment of AMR using NO-MBC4d CEUS imaging, which may be expected to provide a better option for recipients with AMR in clinic.

中文翻译：

---

使用装载有一氧化氮的C4d靶向微气泡精确治疗大鼠急性抗体介导的心脏异体移植排斥反应。

背景技术抗体介导的排斥反应（AMR）构成心脏同种异体移植物丢失的重要原因。然而，目前所有的治疗策略都代表了全身应用，疗效不尽人意。以前，我们使用靶向C4d的微泡（MBC4d）在同种异体移植中成功地非侵入性地检测了C4d，这是一种用于AMR诊断的特异性标记。在这项研究中，我们通过掺入一氧化氮（NO）扩展了这种方法，因为高NO水平显示出免疫抑制和抗血栓形成作用。方法我们设计了新型的NO加载的MBC4d（NO-MBC4d）。通过皮肤移植预敏化建立了AMR大鼠模型。注射NO-MBC4d后，获得了超声造影（CEUS）图像并进行了定量分析。分析同种异体移植物的存活和组织学特征，以评估NO-MBC4d对AMR的治疗作用和潜在机制。结果我们成功获得了NO-MBC4d注射后的CEUS图像，并证明心肌面积的超声信号强度和NO-MBC4d的清除时间均随C4d等级的增加而增加，从而实现了AMR的无创诊断。此外，通过显着抑制血栓形成和减少炎症细胞浸润，NO-MBC4d注射后，同种异体移植的存活时间显着延长，排斥反应明显减弱。总体而言，与对照组NO-MB组相比，NO-MBC4d组的疗效显着提高，这表明与全身应用相比，精确治疗可以显着提高治疗效果。结论本研究提出了一种新颖的工具，可使用NO-MBC4d CEUS成像同时进行非侵入性诊断和AMR的精确治疗，有望为临床接受AMR的患者提供更好的选择。