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客戶在外泌體相關(guān)研究中采用我司巰基磁珠發(fā)表高分論文(IF=27)

來源:生物磁珠專家 2022-9-27 22:21:27??????點(diǎn)擊:



Chen, C.Sun, M.Wang, J.Su, L.Lin, J., & Yan, X. (2021). Active cargo loading into extracellular vesicles: Highlights the heterogeneous encapsulation behaviourJournal of Extracellular Vesicles10,e12163. https://doi.org/10.1002/jev2.12163

Active cargo loading into extracellular vesicles: Highlights the heterogeneous encapsulation behaviour

Chaoxiang Chen,Mengdi Sun,Jialin Wang,Liyun Su,Junjie Lin,Xiaomei Yan
First published: 31 October 2021 https://doi.org/10.1002/jev2.12163 

Abstract

Extracellular vesicles (EVs) have demonstrated unique advantages in serving as nanocarriers for drug delivery, yet the cargo encapsulation efficiency is far from expectation, especially for hydrophilic chemotherapeutic drugs. Besides, the intrinsic heterogeneity of EVs renders it difficult to evaluate drug encapsulation behaviour. Inspired by the active drug loading strategy of liposomal nanomedicines, here we report the development of a method, named “Sonication and Extrusion-assisted Active Loading” (SEAL), for effective and stable drug encapsulation of EVs. Using doxorubicin-loaded milk-derived EVs (Dox-mEVs) as the model system, sonication was applied to temporarily permeabilize the membrane, facilitating the influx of ammonium sulfate solution into the lumen to establish the transmembrane ion gradient essential for active loading. Along with extrusion to downsize large mEVs, homogenize particle size and reshape the nonspherical or multilamellar vesicles, SEAL showed around 10-fold enhancement of drug encapsulation efficiency compared with passive loading. Single-particle analysis by nano-flow cytometry was further employed to reveal the heterogeneous encapsulation behaviour of Dox-mEVs which would otherwise be overlooked by bulk-based approaches. Correlation analysis between doxorubicin auto-fluorescence and the fluorescence of a lipophilic dye DiD suggested that only the lipid-enclosed particles were actively loadable. Meanwhile, immunofluorescence analysis revealed that more than 85% of the casein positive particles was doxorubicin free. These findings further inspired the development of the lipid-probe- and immuno-mediated magnetic isolation techniques to selectively remove the contaminants of non-lipid enclosed particles and casein assemblies, respectively. Finally, the intracellular assessments confirmed the superior performance of SEAL-prepared mEV formulations, and demonstrated the impact of encapsulation heterogeneity on therapeutic outcome. The as-developed cargo-loading approach and nano-flow cytometry-based characterization method will provide an instructive insight in the development of EV-based delivery systems.


細(xì)胞外囊泡(EVs)在作為藥物遞送納米載體方面表現(xiàn)出獨(dú)特的優(yōu)勢(shì),但貨物包封效率遠(yuǎn)未達(dá)到預(yù)期,特別是對(duì)于親水性化療藥物。此外,電動(dòng)汽車的內(nèi)在異質(zhì)性使得評(píng)估藥物封裝行為變得困難。受脂質(zhì)體納米藥物的主動(dòng)載藥策略的啟發(fā),我們?cè)诖藞?bào)告了一種名為“超聲和擠出輔助主動(dòng)加載”(SEAL) 的方法的開發(fā),該方法用于對(duì) EV 進(jìn)行有效和穩(wěn)定的藥物封裝。使用載有阿霉素的牛奶衍生 EVs (Dox-mEVs) 作為模型系統(tǒng),應(yīng)用超聲處理暫時(shí)透化膜,促進(jìn)硫酸銨溶液流入管腔,從而建立主動(dòng)加載所必需的跨膜離子梯度。除了通過擠壓來縮小大型 mEV、使粒徑均勻化和重塑非球形或多層囊泡外,與被動(dòng)加載相比,SEAL 的藥物封裝效率提高了約 10 倍。通過納米流式細(xì)胞術(shù)進(jìn)行的單粒子分析被進(jìn)一步用于揭示 Dox-mEV 的異質(zhì)封裝行為,否則這些行為會(huì)被基于體積的方法所忽視。多柔比星自發(fā)熒光與親脂性染料 DiD 熒光之間的相關(guān)性分析表明,只有脂質(zhì)包裹的顆粒是可主動(dòng)加載的。同時(shí),免疫熒光分析顯示,85% 以上的酪蛋白陽性顆粒不含阿霉素。這些發(fā)現(xiàn)進(jìn)一步激發(fā)了脂質(zhì)探針和免疫介導(dǎo)的磁分離技術(shù)的發(fā)展,以分別選擇性地去除非脂質(zhì)封閉顆粒和酪蛋白組件的污染物。最后,細(xì)胞內(nèi)評(píng)估證實(shí)了 SEAL 制備的 mEV 制劑的卓越性能,并證明了封裝異質(zhì)性對(duì)治療結(jié)果的影響。所開發(fā)的貨物裝載方法和基于納米流式細(xì)胞儀的表征方法將為基于 EV 的遞送系統(tǒng)的開發(fā)提供有益的見解。


客戶在該研究中使用我司生產(chǎn)的巰基磁珠偶聯(lián)抗體。

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