Department of Physics, Sapienza University of Rome, Piazzale Aldo Moro 2, I-00185 Roma, Italy.
International School for Advanced Studies (SISSA), I-34136 Trieste, Italy.
Molecules. 2021 Feb 8;26(4):887. doi: 10.3390/molecules26040887.
Extracellular vesicles are membrane-delimited structures, involved in several inter-cellular communication processes, both physiological and pathological, since they deliver complex biological cargo. Extracellular vesicles have been identified as possible biomarkers of several pathological diseases; thus, their characterization is fundamental in order to gain a deep understanding of their function and of the related processes. Traditional approaches for the characterization of the molecular content of the vesicles require a large quantity of sample, thereby providing an average molecular profile, while their heterogeneity is typically probed by non-optical microscopies that, however, lack the chemical sensitivity to provide information of the molecular cargo. Here, we perform a study of individual microvesicles, a subclass of extracellular vesicles generated by the outward budding of the plasma membrane, released by two cultures of glial cells under different stimuli, by applying a state-of-the-art infrared nanospectroscopy technique based on the coupling of an atomic force microscope and a pulsed laser, which combines the label-free chemical sensitivity of infrared spectroscopy with the nanometric resolution of atomic force microscopy. By correlating topographic, mechanical and spectroscopic information of individual microvesicles, we identified two main populations in both families of vesicles released by the two cell cultures. Subtle differences in terms of nucleic acid content among the two families of vesicles have been found by performing a fitting procedure of the main nucleic acid vibrational peaks in the 1000-1250 cm frequency range.
细胞外囊泡是一种膜限定结构，参与多种细胞间通讯过程，包括生理和病理过程，因为它们传递复杂的生物货物。细胞外囊泡已被确定为几种病理疾病的潜在生物标志物；因此，对其进行特征描述是至关重要的，以便深入了解其功能和相关过程。传统的方法来描述囊泡的分子内容需要大量的样本，从而提供平均的分子谱，而它们的异质性通常通过非光学显微镜来探测，然而，后者缺乏化学敏感性来提供分子货物的信息。在这里，我们通过应用一种基于原子力显微镜和脉冲激光耦合的最先进的红外纳米光谱技术，对两种不同刺激下的神经胶质细胞培养物释放的两种细胞外囊泡亚类——微囊泡进行了个体研究。这种技术结合了红外光谱的无标记化学敏感性和原子力显微镜的纳米分辨率。通过对单个微囊泡的形貌、力学和光谱信息进行相关分析，我们在两种细胞培养物释放的囊泡家族中都鉴定出了两个主要群体。通过对 1000-1250 cm 频率范围内主要核酸振动峰进行拟合处理，我们发现了两种囊泡家族在核酸含量方面的细微差异。
