Charité - Universitätsmedizin Berlin, corporate member of Freie Universität Berlin, Humboldt-Universität zu Berlin, and Berlin Institute of Health, Charitéplatz 1, 10117, Berlin, Germany.
MR and CT Contrast Media Research, Bayer AG, Berlin, Germany.
Mol Imaging Biol. 2021 Jun;23(3):382-393. doi: 10.1007/s11307-020-01563-z. Epub 2020 Dec 7.
Contrast-enhanced magnetic resonance imaging (MRI) has the potential to replace angiographic evaluation of atherosclerosis. While studies have investigated contrast agent (CA) uptake in atherosclerotic plaques, exact CA spatial distribution on a microscale is elusive. The purpose of this study was to investigate the microdistribution of gadolinium (Gd)- and iron (Fe) oxide-based CA in atherosclerotic plaques of New Zealand White rabbits.
The study was performed as a post hoc analysis of archived tissue specimens obtained in a previous in vivo MRI study conducted to investigate signal changes induced by very small superparamagnetic iron oxide nanoparticles (VSOP) and Gd-BOPTA. For analytical discrimination from endogenous Fe, VSOP were doped with europium (Eu) resulting in Eu-VSOP. Formalin-fixed arterial specimens were cut into 5-μm serial sections and analyzed by immunohistochemistry (IHC: Movat's pentachrome, von Kossa, and Alcian blue (pH 1.0) staining, anti-smooth muscle cell actin (anti-SMA), and anti-rabbit macrophage (anti-RAM-11) immunostaining) and elemental microscopy with laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) and synchrotron radiation μX-ray fluorescence (SR-μXRF) spectroscopy. Elemental distribution maps of Fe, Eu, Gd, sulfur (S), phosphorus (P), and calcium (Ca) were investigated.
IHC characterized atherosclerotic plaque pathomorphology. Elemental microscopy showed S distribution to match the anatomy of arterial vessel wall layers, while P distribution corresponded well with cellular areas. LA-ICP-MS revealed Gd and Fe with a limit of detection of ~ 0.1 nmol/g and ~ 100 nmol/g, respectively. Eu-positive signal identified VSOP presence in the vessel wall and allowed the comparison of Eu-VSOP and endogenous Fe distribution in tissue sections. Extracellular matrix material correlated with Eu signal intensity, Fe concentration, and maximum Gd concentration. Eu-VSOP were confined to endothelium in early lesions but accumulated in cellular areas in advanced plaques. Gd distribution was homogeneous in healthy arteries but inhomogeneous in early and advanced plaques. SR-μXRF scans at 0.5 μm resolution revealed Gd hotspots with increased P and Ca concentrations at the intimomedial interface, and a size distribution ranging from a few micrometers to submicrometers.
Eu-VSOP and Gd have distinct spatial distributions in atherosclerotic plaques. While Eu-VSOP distribution is more cell-associated and might be used to monitor atherosclerotic plaque progression, Gd distribution indicates arterial calcification and might help in characterizing plaque vulnerability.
对比增强磁共振成像（MRI）有可能取代动脉粥样硬化的血管造影评估。虽然已经研究了动脉粥样斑块中对比剂（CA）的摄取，但微观尺度上 CA 的精确空间分布仍然难以捉摸。本研究的目的是研究新西兰白兔动脉粥样硬化斑块中钆（Gd）和氧化铁基 CA 的微观分布。
本研究是先前体内 MRI 研究的存档组织标本的事后分析，该研究旨在研究超小超顺磁氧化铁纳米颗粒（VSOP）和 Gd-BOPTA 诱导的信号变化。为了与内源性 Fe 进行分析区分，VSOP 被掺 Eu（Eu-VSOP）以形成 Eu-VSOP。福尔马林固定的动脉标本切成 5μm 的连续切片，通过免疫组织化学（IHC：Movat 五重染色、von Kossa 和 Alcian blue（pH 1.0）染色、抗平滑肌细胞肌动蛋白（抗-SMA）和抗兔巨噬细胞（抗-RAM-11）免疫染色）和元素显微镜（激光烧蚀电感耦合等离子体质谱法（LA-ICP-MS）和同步辐射 μX 射线荧光光谱法（SR-μXRF））进行分析。研究了 Fe、Eu、Gd、硫（S）、磷（P）和钙（Ca）的元素分布图谱。
IHC 描述了动脉粥样硬化斑块的病理形态学。元素显微镜显示 S 的分布与动脉壁层的解剖结构相匹配，而 P 的分布与细胞区域非常吻合。LA-ICP-MS 显示 Gd 和 Fe 的检测限分别约为 0.1nmol/g 和 100nmol/g。Eu 阳性信号识别出血管壁中的 VSOP 存在，并允许比较组织切片中的 Eu-VSOP 和内源性 Fe 分布。细胞外基质材料与 Eu 信号强度、Fe 浓度和最大 Gd 浓度相关。在早期病变中，Eu-VSOP 局限于内皮，但在晚期斑块中积累于细胞区。健康动脉中的 Gd 分布均匀，但在早期和晚期斑块中不均匀。分辨率为 0.5μm 的 SR-μXRF 扫描显示，在内膜中层界面处，Gd 热点处的 P 和 Ca 浓度增加，并且存在从几微米到亚微米的尺寸分布。
Eu-VSOP 和 Gd 在动脉粥样硬化斑块中有不同的空间分布。虽然 Eu-VSOP 的分布更与细胞相关，可能用于监测动脉粥样硬化斑块的进展，但 Gd 的分布表明动脉钙化，可能有助于表征斑块的脆弱性。
