Raizada Geetika, Guillouzouic Joan, Rouleau Alain, Lesniewska Eric, Le Ferrec Eric, Elie-Caille Céline, Boireau Wilfrid
Université Marie et Louis Pasteur, CNRS, Institut FEMTO-ST, 25030 Besançon, France.
Univ Rennes, Inserm, EHESP, Irset (Institut de Recherche en Santé Environnement et Travail), UMR_S 1085, 35000 Rennes, France.
Biosensors (Basel). 2025 Feb 11;15(2):103. doi: 10.3390/bios15020103.
Recent advances in the clinical extracellular vesicles (EVs) field highlight their potential as biomarkers for diverse diseases and therapeutic applications. This study provides an in-depth characterization of 10k EVs from human microvascular endothelial cells (HMEC-1) exposed to benzo[a]pyrene (B[a]P), a polycyclic aromatic hydrocarbon found in food and smoke. Given EVs' complexity, with numerous surface and cargo proteins, phenotyping remains challenging. Here, we introduce a multiplex biosensor, in µarray format, for profiling EVs from distinct cellular conditions, employing a multimodal approach that combines surface plasmon resonance imaging (SPRi) and in situ atomic force microscopy (AFM) to decipher EVs' biochemical and biophysical properties. SPRi experiments showed notable EV capture differences on ligands such as Anti-CD36, Anti-CD81, and Anti-ApoA between treated and control conditions, likely due to B[a]P exposure. A complementary AFM study and statistical analyses revealed size differences between EVs from treated and control samples, with ligands like Annexin-V, Anti-CD36, and Anti-VEGFR1 emerging as ligands specific to potential cytotoxicity biomarkers. Our findings suggest that B[a]P exposure may increase EV size and alter marker expression, indicating phenotypic shifts in EVs under cytotoxic stress. The original combination of SPRi and AFM reveals valuable data on the phenotypical and morphological heterogeneities of EV subsets linked to cytotoxic stresses and highlights the potential of EVs as specific toxicological markers.
临床细胞外囊泡（EVs）领域的最新进展凸显了其作为多种疾病生物标志物和治疗应用的潜力。本研究深入表征了来自人微血管内皮细胞（HMEC-1）且暴露于苯并[a]芘（B[a]P）的10k EVs，苯并[a]芘是一种存在于食物和烟雾中的多环芳烃。鉴于EVs的复杂性，其具有众多表面蛋白和 cargo 蛋白，对其进行表型分析仍然具有挑战性。在此，我们引入一种微阵列形式的多重生物传感器，用于分析来自不同细胞条件的EVs，采用一种多模态方法，该方法结合表面等离子体共振成像（SPRi）和原位原子力显微镜（AFM）来解读EVs的生化和生物物理特性。SPRi实验表明，在处理组和对照组条件下，Anti-CD36、Anti-CD81和Anti-ApoA等配体对EVs的捕获存在显著差异，这可能是由于B[a]P暴露所致。一项互补的AFM研究和统计分析揭示了处理组和对照组样品的EVs之间的大小差异，膜联蛋白-V、Anti-CD36和Anti-VEGFR1等配体成为潜在细胞毒性生物标志物的特异性配体。我们的研究结果表明，B[a]P暴露可能会增加EVs的大小并改变标志物表达，表明细胞毒性应激下EVs的表型发生了变化。SPRi和AFM的原始组合揭示了与细胞毒性应激相关的EV亚群的表型和形态异质性的有价值数据，并突出了EVs作为特定毒理学标志物的潜力。
