The State Key Laboratory Breeding Base of Basic Science of Stomatology (Hubei-MOST) & Key Laboratory of Oral Biomedicine Ministry of Education, School and Hospital of Stomatology, Wuhan University, Wuhan 430079, China.
Department of Oral and Maxillofacial Surgery, School and Hospital of Stomatology, Wuhan University, Wuhan 430079, China.
Anal Chem. 2023 Jan 17;95(2):1016-1026. doi: 10.1021/acs.analchem.2c03725. Epub 2022 Dec 19.
Small extracellular vesicles (sEVs) are heterogeneous membrane-bound vesicles that carry numerous bioactive molecules. Studies have reported that sEVs carrying PD-L1 on the surface could contribute to immunosuppression; however, the precise mechanisms are unclear. To fully dissect their mode of action, it requires qualified methods to specifically isolate natural PD-L1-positive sEVs from heterogeneous sEVs. This study reported an aptamer-assisted capture-and-release strategy for traceless isolation of PD-L1-positive sEVs. The PD-L1 aptamer-anchored magnetic microspheres enable the specific capture of PD-L1-positive sEVs. The traceless release of captured PD-L1-positive sEVs was triggered by competition of complementary oligonucleotides, endowing the obtained label-free PD-L1-positive sEVs with natural properties. Benefited from this traceless isolation strategy, the distinct molecule profiles in adhesion and immuno-regulation between PD-L1-positive and PD-L1-negative sEVs were revealed. Compared to PD-L1-negative sEVs, PD-L1-positive sEVs were much more concentrated in cadherin binding, accompanied by increased adhesion to lymphatic endothelial cells and T cells but decreased adhesion to the extracellular matrix. Moreover, PD-L1-positive sEVs could transfer their enriched immunosuppressive "synapse"-related proteins to antigen-presenting cells, thereby inducing a tolerogenic-like phenotype. In summary, the present work dissects the subpopulation signature and action mode of PD-L1-positive sEVs for the first time and provides a general approach to the traceless isolation of sEV subpopulations.
小细胞外囊泡 (sEVs) 是具有多种生物活性分子的异质膜结合囊泡。研究报道，表面带有 PD-L1 的 sEV 可导致免疫抑制；然而，确切的机制尚不清楚。为了充分剖析其作用模式，需要有合格的方法从异质 sEV 中特异性分离天然 PD-L1 阳性 sEV。本研究报道了一种适体辅助的捕获-释放策略，用于无痕迹分离 PD-L1 阳性 sEV。PD-L1 适体锚定的磁性微球能够特异性捕获 PD-L1 阳性 sEV。通过互补寡核苷酸的竞争触发捕获的 PD-L1 阳性 sEV 的无痕迹释放，赋予获得的无标记 PD-L1 阳性 sEV 天然特性。得益于这种无痕迹分离策略，揭示了 PD-L1 阳性和 PD-L1 阴性 sEV 之间在黏附和免疫调节方面的明显分子谱差异。与 PD-L1 阴性 sEV 相比，PD-L1 阳性 sEV 在钙黏蛋白结合中更为集中，伴随与淋巴管内皮细胞和 T 细胞的黏附增加，但与细胞外基质的黏附减少。此外，PD-L1 阳性 sEV 可以将其富含免疫抑制性“突触”相关蛋白转移到抗原呈递细胞，从而诱导耐受性样表型。总之，本研究首次剖析了 PD-L1 阳性 sEV 的亚群特征和作用模式，并为 sEV 亚群的无痕迹分离提供了一种通用方法。
