Isolation and characterization of extracellular vesicles in saliva of children with asthma.
Isolation and characterization of extracellular vesicles in saliva of children with asthma.
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DOI：
10.20517/evcna.2020.09
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发表时间：
2021
期刊：
影响因子：
--
通讯作者：
Baccarelli A
中科院分区：
文献类型：
--
作者：
Comfort N;Bloomquist TR;Shephard AP;Petty CR;Cunningham A;Hauptman M;Phipatanakul W;Baccarelli A
目的：证实哮喘儿童唾液中是否存在细胞外小泡(EV)，并描述分离的EV种群。使用ExoQuick-TC从180名参与者中分离出的CFS EV的汇集样本在下游分析中进行了检查。用透射电子显微镜(TEM)证实EVS的存在。采用纳米颗粒跟踪分析(NTA)和荧光单粒子干涉反射成像传感(SP-IRIS)技术对电动汽车进行了检测、计数和表型鉴定。蛋白质定量采用毛细管免疫分析法。TEM证实了不同大小的电动汽车的存在，表明PREP包含了不同类型的电动汽车。毛细管免疫分析证实了EV相关蛋白(CD9、CD63、CD81、ICAM-1和ANXA5)的存在，并表明细胞污染有限。正如其他人也报告的那样，电动汽车的规模和枚举在不同平台上存在差异。荧光NTA检测到模直径为~90 nm的粒子，而SP-IRIS报告的粒子大小为~55-60 nm，这与透射电子显微镜的结果更接近。与蛋白免疫分析结果一致的是，SP-IRIS荧光显示，这些EV大多为CD9和CD63阳性，CD81很少表达。可以使用一种高通量方法分离来自CFS的EV，这种方法可以扩展到大型流行病学研究。据我们所知，我们是第一个从哮喘患者身上鉴定CFS EV的人。CFS EVS作为潜在的哮喘新生物标志物的应用值得进一步研究，并为未来的研究开辟了一条新的研究途径。
To confirm the presence of extracellular vesicles (EVs) in cell-free saliva (CFS) of children with asthma and describe the isolated EV population. A pooled sample of CFS EVs isolated from 180 participants using ExoQuick-TC was examined in downstream analyses. Transmission electron microscopy (TEM) was used to confirm the presence of EVs. Nanoparticle tracking analysis (NTA) and single particle interferometric reflectance imaging sensing (SP-IRIS) with fluorescence were used for sizing, counting, and phenotyping of EVs. Capillary immunoassays were used for protein quantitation. TEM confirmed the presence of EVs of diverse sizes, indicating the prep contained a heterogeneous population of EVs. Capillary immunoassays confirmed the presence of EV-associated proteins (CD9, CD63, CD81, ICAM-1, and ANXA5) and indicated limited cellular contamination. As others have also reported, there were discrepancies in the EV sizing and enumeration across platforms. Fluorescent NTA detected particles with a mode diameter of ~90 nm, whereas SP-IRIS reported sizes of ~55–60 nm that more closely approximated the TEM results. Consistent with protein immunoassay results, SP-IRIS with fluorescence showed that the majority of these EVs were CD9- and CD63-positive, with little expression of CD81. EVs from CFS can be isolated using a high-throughput method that can be scaled to large epidemiological studies. To our knowledge, we are the first to characterize CFS EVs from patients with asthma. The use of CFS EVs as potential novel biomarkers in asthma warrants further investigation and opens a new avenue of research for future studies.
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影响因子：
16
作者：
Lötvall J;Hill AF;Hochberg F;Buzás EI;Di Vizio D;Gardiner C;Gho YS;Kurochkin IV;Mathivanan S;Quesenberry P;Sahoo S;Tahara H;Wauben MH;Witwer KW;Théry C
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影响因子：
3.7
作者：
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通讯作者：
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影响因子：
4.9
作者：
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通讯作者：
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影响因子：
16
作者：
Bachurski, Daniel;Schuldner, Maximiliane;von Strandmann, Elke Pogge
通讯作者：
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影响因子：
6.7
作者：
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通讯作者：
HYYPPA, T