Name: modulating shape of polyester based polymersomes using osmotic pressure
Uploaded: 2021-04-21T15:00:00
Description: Watch this Scientific Journal Video about Modulating Shape of Polyester Based Polymersomes using Osmotic Pressure at JoVE.com

该项目部分由国家卫生研究院项目5P20GM103499-19通过学生启动研究项目项目资助。这项工作也得到了克莱姆森的创意调查计划的部分支持。我们也感谢尼古拉斯·拉莫罗和奥恩·阿里谁最初致力于创建这个协议，发表他们的第一篇论文引用这里14。

1. 使用溶剂注射法形成球形聚合物
<ol><li>有机溶剂中聚酯溶解 注：一次只应溶解一个聚酯溶解在各自的有机溶剂中，形成聚合物体。<ol>
<li>溶解聚酯PEG-PLA或PEG-b-poly（乳酸-共甘油酸）（PEG-PLGA）在二甲基硫氧化物（DMSO）的浓度为1.5%的重量。具体来说，溶解 0.015 克选定的聚酯在 1 mL 的 DMSO （15 毫克/毫升）.。聚合物的完全溶解可能需要长达15分钟的涡流。</li>
	</ol>
</li>
	<li>当聚酯溶解在有机溶?...

<ol>
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自组装系统是出了名的无法控制。它们的最终属性，包括大小、形状和结构，是由所选的两栖动物的疏水特性和所选溶剂环境驱动的。嗜血块共聚物倾向于球形，从而最大限度地减少吉布的自由能量，并导致热力学平衡23，从而形成聚合物体。由于其平衡性质，聚合物体在拉长或改变形状方面具有更大的挑战性，因此，与固体纳米粒子相比，研究较少。溶剂驱动的方法已经与PEG-b...

形状调制是改善纳米粒子介质药物输送的一种相对较新和有效的方法。形态的变化不仅增加了粒子的表面积，反过来又增加了承载能力，而且对提高稳定性、循环时间、生物可用性、分子靶向和受控释放1具有全面的影响。聚合物体是该方法中焦点的纳米粒子，它倾向于热力学自我组装成球形，这已被证明在细胞吸收中不切实际，并且更容易在免疫系统中作为异物检测到。能够将结构拉长成一根蛋白石或一根棒子，将使药物携带者通过模仿原生细胞来躲避巨噬细胞，并更成功地将它们交付到他们期望的目标2、3、4、5、6、7。聚合物体的显著优点，包括有效载荷的膜绑定保护，膜的刺激反应，以及亲水药物和疏水药物8，9，10的双重封装，使他们在形状调制过程中保持药物输送的有力候选者。
调节聚合物体的形状有许多不同的方法，每种方法都有各自的优缺点。然而，这些方法大多分为两类：溶剂驱动和盐驱动的渗透压力变化11。这两种方法都旨在克服聚合物形成球形平衡形状后存在的弯曲能量。通过引入渗透压力梯度，聚合物体可以被迫弯曲成拉长结构，尽管强弯曲能量11，12。
基于溶剂的方法探索形状的变化灵感来自金和范赫斯特13的工作。他们将有机溶剂和水混合物中的聚合物塑料化，将有机溶剂捕获在囊泡膜中，并将水从囊芯中排出。最终，粒子的内部体积如此之低，以至于拉长了。虽然这种方法已显示出希望，但它缺乏实用性。此方法需要不同的溶剂为每个单独的聚合物骨干参与调制。因此，它并不广泛适用于促进形状的变化。相反，盐基方法是均匀的，并利用一个通用驱动因素，可以引入渗透压力到许多块聚合物为基础的聚合物体。
该项目采用拉莫罗等人引进的盐基方法。此协议涉及两轮透析。一种旨在通过去除在生产过程中可能困在双层中的有机溶剂来净化和凝固聚（乙二醇）-b-聚合物（乳酸）（PEG-PLA）聚合物，另一种则促进形状变化。第二个透析步骤引入了 50 mM NaCl 溶液，可创建渗透压力梯度以驱动形状变化。这种方法得到了萨尔瓦等人的支持，他们指出，溶液中的超音速应激会导致囊泡收缩15。此方法基于先前发布的方法14， 从 50-200 m NaCl 中查看两种不同的聚酯聚合物和各种盐梯度。聚酯由于其生物相容性和生物降解而使用。盐梯度对形状有不同的影响，具体取决于块共聚体骨干的疏水性。它可用于创建前列腺、棒和豆类细胞。之所以选择这种盐驱动的方法，是因为复制的便利性和实验的多功能性。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>15*45 vials screw thread w/cap attached</td>
			<td>Fisherbrand</td>
			<td>9609104000</td>
			<td></td>
		</tr>
		<tr>
			<td>Dimethyl Sulfoxide</td>
			<td>Fisher Chemical</td>
			<td>D128-1</td>
			<td></td>
		</tr>
		<tr>
			<td>Dimethyl Sulfoxide</td>
			<td>BDH</td>
			<td>BDH1115-1LP</td>
			<td></td>
		</tr>
		<tr>
			<td>Isoremp stirrers, hotplates, and stirring hotplates</td>
			<td>Fisher scientific</td>
			<td>CIC00008110V19</td>
			<td></td>
		</tr>
		<tr>
			<td>LEGATO 130 SYRINGE PUMP</td>
			<td>kd Scientific</td>
			<td>788130</td>
			<td></td>
		</tr>
		<tr>
			<td>PEG(1000)-b-PLA(5000), Diblock Polymer</td>
			<td>Polysciences Inc</td>
			<td>24381-1</td>
			<td>note the molecular weights when replicating</td>
		</tr>
		<tr>
			<td>Poly(ethylene glycol) (2000) Methyl ether-block-poly(lactide-co-glycolide) (4500)</td>
			<td>Sigma aldrich</td>
			<td>764825-1G</td>
			<td>note the molecular weights when replicating</td>
		</tr>
		<tr>
			<td>Single-Use Syringe/BD PrecisionGlide Needle combination, sterile, BD medical</td>
			<td>BD medical</td>
			<td>BD305620</td>
			<td>Tuberculin</td>
		</tr>
		<tr>
			<td>Sodium Chloride</td>
			<td>BDH</td>
			<td>BDH9286</td>
			<td></td>
		</tr>
		<tr>
			<td>Zetasizer Nano ZS</td>
			<td>Malvern</td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

modulating shape of polyester based polymersomes using osmotic pressure

聚合物体是膜结合的双层囊泡，由嗜血块共聚体制成，可以封装疏水和亲水有效载荷，用于药物输送应用。尽管聚合物体很有前途，但由于其球形，其应用有限，而细胞不易被细胞占据，固体纳米粒子科学家证明了这一点。本文描述了一种盐基方法，用于提高球形聚合物（乙二醇）（PEG）基聚合物的纵横比。这种方法可以拉长聚合物体，并最终通过在形成后透析中加入氯化钠来控制其最终形状。盐浓度可以多种多样，如本方法所述，基于用作聚合物和目标形状基础的块聚合物的疏水性。拉长的纳米粒子有可能更好地瞄准直径较大的血管中的内皮，如观察到边缘的静脉。本方案可利用拉长技术与聚合物的双载长循环效益配合，扩大治疗纳米粒子的应用。

表 2 在遵循协议步骤 1 时呈现预期结果。请注意，DMSO 在聚合物体形成中用作 PEG-PLA 和 PEG-PLGA 的溶剂。偏离这种溶剂是可能的，因为其他水错溶剂将溶解共聚物，但预计将改变结果。预计PDI将低于0.2，表明单分散聚合物体17的形成。请注意，水灵性增加会导致聚合物直径和 PDI 的偏差增加。如果运行协议时，聚合物直径与此表中报告的直径有显著差异：最典型的罪魁?...

Watch this Scientific Journal Video about Modulating Shape of Polyester Based Polymersomes using Osmotic Pressure at JoVE.com

Modulating Shape of Polyester Based Polymersomes using Osmotic Pressure

聚合物体是自组装的聚合囊泡，形成球形，以尽量减少吉布的自由能量。在药物输送方面，更拉长的结构是有益的。本协议建立了方法，以创建更多的棒状聚合物体，与拉长的纵横比，使用盐诱导渗透压力和减少内部囊泡体积。

使用渗透压力调节聚酯聚合物的形状

Polymersomes are membrane-bound, bilayer vesicles created from amphiphilic block copolymers that can encapsulate both hydrophobic and hydrophilic payloads ...

纳米粒子形状影响细胞吸收，但改变聚合物体形状的工作有限。此协议演示了如何拉长聚合物以增强药物输送应用。该技术提供了一个简单的方法，可以创建许多不同的聚合物形状，在药物输送，伪细胞，或其他未开发的领域的潜在应用。拉长聚合物可以同时提供疏水和亲水药物，更容易地输送到细胞中。这项技术可以帮助将药物穿梭于血脑屏障，应对重大的医学挑战。首先，通过15分钟的涡流将0.015克选定的聚酯溶解在一毫升的DMSO中，并设置溶剂注射装置。将启动板直接放在垂直注射器泵下方，然后将一个五毫升玻璃胆汁与一毫升二型电离水放在搅拌盘上的微型星条中。调整注射器泵的高度，使针尖完全浸入二型去离子水中，并将注射器泵的输液率设置为每分钟五微升。将有机溶剂和聚酯溶液绘制成 27 量表针，执行溶剂注射。将针头放入注射器泵中，确保其安全。调整推手块以触摸注射器柱塞的末端。启动搅拌盘，使水以每分钟 100 转的速度旋转，然后启动注射器泵。一旦注射器泵将有机溶剂和聚合物完全注入水中，取出搅拌棒并盖住玻璃杯。要通过动态光散射或 DLS 对聚合物体进行定性，请将一毫升含有少量有机溶剂和聚合物的水添加到一毫升的 cuvette 中。通过将 cuvette 放入系统并设置运行来执行 DLS。阅读聚合物强度加权直径和多分散性指数或 PDI。根据制造商提供的协议清洗 300 千达尔顿透析膜后，在透析装置的储液库中加入一毫升聚合物溶液。将透析装置放在 250 毫升烧甲器中，将 150 毫升的二型电离水放在搅拌盘上。将搅拌板设置为允许透析装置的轻柔运动的速度，并让它在一夜之间搅拌。透析完成后，从透析装置中提取一毫升聚合物溶液。根据最终所需的聚合物特性，使用 50、100 或 200 毫摩尔浓度的氯化钠创建 150 毫升所需的盐缓冲器，并将透析装置的负载保持在 150 毫升盐溶液中 18 小时进行搅拌。调整形状后，执行 DLS 测量。与精神聚合物相比，特别注意 PDI 测量，因为 PDI 的变化表明聚合物体的形状发生了有效的变化。使用适当的控制进行成像，特别是非形状调制聚合物，以确保该方法的成功。TEM 对 PEG-PLA 基聚合物的观察表明，整体球形结构具有较厚的外线，表明膜。聚合物体作为物理结构存在，SEM 具有像 PEG 外部层一样的刷子。在水中透析一小时，总平均直径相同，溶剂去除会降低聚合物直径。当使用更大的有机溶剂初始浓度时，预计直径会减少。在制造PEG-PLA聚合物的同时，PDI有望略有变化，这可能表明形状有所变化。当透析PEG-PLGA聚合物时，PDI的增加与所有探索的盐梯度相一致，导致PDI增加。使用 50 毫摩尔盐梯度导致形状变化时，无论聚酯疏水性如何，都应观察到类似的结果。同时，100和200毫摩尔氯化钠盐梯度呈现出随着聚酯水恐惧性增加而增加的三角洲PDI的直接趋势。PEG-PLA聚合物的TEM图像用5000万低氯化钠解析，显示用雾化细胞和拉长棒。随着盐浓度增加到100毫摩尔，观察到的棒形成数量增加，番茄细胞数量减少。对200毫摩尔聚合物体进行透析，形成更一致的前列腺，具有适度的纵横比。此协议中最具挑战性的部分是始终如一地制造聚合物体。因此，重要的是要实践这个程序。

Research

JoVE Journal

Bioengineering

Engineering Biological-Based Vascular Grafts Using a Pulsatile Bioreactor

基于生物工程，人造血管使用脉动生物反应器

Much effort has been devoted to develop and advance the methodology to regenerate functional small-diameter arterial bypasses. In the physiological ...

科研

生物工程

Shape Memory Polymers for Active Cell Culture

形状记忆聚合物主动细胞培养

Shape memory polymers (SMPs) are a class of "smart" materials that have the ability to change from a fixed, temporary shape to a pre-determined permanent ...

Microfluidic Fabrication of Polymeric and Biohybrid Fibers with Predesigned Size and Shape

预设计尺寸和形状的聚合物纤维和生物贿赂纤维的微流体制造

A &#8220;sheath&#8221; fluid passing through a microfluidic channel at low Reynolds number can be directed around another &#8220;core&#8221; stream and ...

Fabrication of a Bioactive, PCL-based "Self-fitting" Shape Memory Polymer Scaffold

生物活性，PCL为基础的“自我装配”形状记忆聚合物支架的制备

Tissue engineering has been explored as an alternative strategy for the treatment of critical-sized cranio-maxillofacial (CMF) bone defects. Essential to ...

Forming Giant-sized Polymersomes Using Gel-assisted Rehydration

形成用凝胶辅助补液巨型的聚合物囊泡

Polymer vesicles, or polymersomes, are being widely explored as synthetic analogs of lipid vesicles based on their stability, robustness, barrier ...

Ammonia Synthesis at Low Pressure

低压氨合成

Ammonia can be synthesized at low pressure by the use of an ammonia selective absorbent. The process can be driven with wind energy, available locally in ...

Pattern-based Search of Epigenomic Data Using GeNemo

利用 GeNemo 模式搜索表数据

Compared with the robust text-based search tools for genomic or RNA sequencing data, current methodologies for pattern-based searches of epigenomic and ...

Antimicrobial Peptides Produced by Selective Pressure Incorporation of Non-canonical Amino Acids

非规范氨基酸选择性压力合成抗菌肽的研究

Nature has a variety of possibilities to create new protein functions by modifying the sequence of the individual amino acid building blocks. However, all ...

Light-sheet Fluorescence Microscopy to Capture 4-Dimensional Images of the Effects of Modulating Shear Stress on the Developing Zebrafish Heart

The hemodynamic forces experienced by the heart influence cardiac development, especially trabeculation, which forms a network of branching outgrowths ...

A Rapid Image-based Bacterial Virulence Assay Using Amoeba

一种基于变形虫的快速图像细菌毒力检测方法

Traditional bacterial virulence assays involve prolonged exposure of bacteria over the course of several hours to host cells. During this time, bacteria ...