Acta Biomaterialia ( IF 9.4 ) Pub Date : 2020-05-13 , DOI: 10.1016/j.actbio.2020.04.023 Viktoriya Sokolova 1 , Goodwell Nzou 2 , Selina B van der Meer 1 , Tatjana Ruks 1 , Marc Heggen 3 , Kateryna Loza 1 , Nina Hagemann 4 , Florian Murke 5 , Bernd Giebel 5 , Dirk M Hermann 4 , Anthony J Atala 2 , Matthias Epple 1
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The neurovascular unit (NVU) is a complex functional and anatomical structure composed of endothelial cells and their blood-brain barrier (BBB) forming tight junctions. It represents an efficient barrier for molecules and drugs. However, it also prevents a targeted transport for the treatment of cerebral diseases. The uptake of ultrasmall nanoparticles as potential drug delivery agents was studied in a three-dimensional co-culture cell model (3D spheroid) composed of primary human cells (astrocytes, pericytes, endothelial cells). Multicellular 3D spheroids show reproducible NVU features and functions. The spheroid core is composed mainly of astrocytes, covered with pericytes, while brain endothelial cells form the surface layer, establishing the NVU that regulates the transport of molecules. After 120 h cultivation, the cells self-assemble into a 350 µm spheroid as shown by confocal laser scanning microscopy. The passage of different types of fluorescent ultrasmall gold nanoparticles (core diameter 2 nm) both into the spheroid and into three constituting cell types was studied by confocal laser scanning microscopy. Three kinds of covalently fluorophore-conjugated gold nanoparticles were used: One with fluorescein (FAM), one with Cy3, and one with the peptide CGGpTPAAK-5,6-FAM-NH2. In 2D cell co-culture experiments, it was found that all three kinds of nanoparticles readily entered all three cell types. FAM- and Cy3-labelled nanoparticles were able to enter the cell nucleus as well. The three dissolved dyes alone were not taken up by any cell type. A similar situation evolved with 3D spheroids: The three kinds of nanoparticles entered the spheroid, but the dissolved dyes did not. The presence of a functional blood-brain barrier was demonstrated by adding histamine to the spheroids. In that case, the blood-brain barrier opened, and dissolved dyes like a FITC-labelled antibody and FITC alone entered the spheroid. In summary, our results qualify ultrasmall gold nanoparticles as suitable carriers for imaging or drug delivery into brain cells (sometimes including the nucleus), brain cell spheroids, and probably also into the brain.
Statement of significance
3D brain spheroid model and its permeability by ultrasmall gold nanoparticles.
We demonstrate that ultrasmall gold nanoparticles can easily penetrate the constituting cells and sometimes even enter the cell nucleus. They can also enter the interior of the blood-brain barrier model. In contrast, small molecules like fluorescing dyes are not able to do that. Thus, ultrasmall gold nanoparticles can serve as carriers of drugs or for imaging inside the brain.
中文翻译：
在体外3D脑球模型中，超小金纳米颗粒（2 nm）可以穿透并进入细胞核。
神经血管单位（NVU）是一个复杂的功能和解剖结构，由内皮细胞及其血脑屏障（BBB）形成紧密连接组成。它代表了分子和药物的有效屏障。但是，它也阻止了针对脑疾病的靶向转运。在由原代人细胞（星形细胞，周细胞，内皮细胞）组成的三维共培养细胞模型（3D球体）中研究了超小纳米颗粒作为潜在药物的吸收。多细胞3D球体具有可重现的NVU特征和功能。球体核心主要由星形胶质细胞组成，覆盖有周细胞，而脑内皮细胞形成表面层，从而建立了调节分子运输的NVU。培养120小时后，共聚焦激光扫描显微镜显示，细胞可自组装成350 µm的球状体。通过共聚焦激光扫描显微镜研究了不同类型的荧光超小金纳米粒子（核心直径2 nm）既进入球体又进入三种构成细胞类型。使用了三种共价共轭荧光团的金纳米颗粒：一种带有荧光素（FAM），一种带有Cy3，另一种带有肽CGGpTPAAK-5,6-FAM-NH2。在2D细胞共培养实验中，发现所有三种纳米粒子都容易进入所有三种细胞类型。FAM和Cy3标记的纳米颗粒也能够进入细胞核。单独的三种溶解的染料不会被任何细胞类型吸收。3D球体也发生了类似的情况：三种纳米粒子进入了球体，但溶解的染料却没有。功能性血脑屏障的存在通过向球体中添加组胺来证明。在那种情况下，血脑屏障打开，溶解的染料（例如FITC标记的抗体和FITC）单独进入球体。总而言之，我们的研究结果证明超小金纳米颗粒可作为成像或药物递送到脑细胞（有时包括细胞核），脑细胞球体，
重要声明
超小型金纳米粒子的3D脑球模型及其渗透性。
我们证明，超小金纳米粒子可以轻松穿透组成细胞，有时甚至进入细胞核。他们还可以进入血脑屏障模型的内部。相反，像荧光染料这样的小分子则无法做到这一点。因此，超小金纳米颗粒可以用作药物的载体或在大脑内部成像。
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