Name: the assembly and application of 'shear rings': a novel endothelial model for orbital, unidirectional and periodic fluid flow and shear stress
Uploaded: 2016-10-31T14:00:00
Description: Watch this Scientific Journal Video about The Assembly and Application of 'Shear Rings': A Novel Endothelial Model for Orbital, Unidirectional and Periodic Fluid Flow and Shear Stress at JoVE.com

作者要感谢克里斯托弗先生阮亚伦亨特和什里夫波特的Jumpstart，SMART和Biostart培训计划的援助以及生物物理，什里夫波特，LA的路易斯安那百年学院中文系。

1.直径为150mm剪切戒指建设（图1） 注：剪切环可以被构造通过改变内外培养皿大小，导致与不同的总表面积，细胞产量的设备和开发的剪切力范围以创建许多不同的尺寸。本报告描述了150毫米的菜具有内部100mm培养皿为98 平方厘米（ 图2）的总表面积相结合。 <img alt="图1" src="/files/ftp_upload/54632/54632fig1.jpg" /> ...

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对暴露的内皮细胞剪切剪切环系统的建设是一个简单的方法来进行剪切应力的研究。然而，有一些是获得卓越的剪切戒指和更好的结果的关键几步之遥。一个完整的密封应在内和外环，以防止介质从泄漏可能创建样品之间不一致的剪切应力之间进行。如果完整的密封不进行，二氯甲烷最小量应通过在内圈的孔被加入到内和外碟用移液管之间的边缘。轻轻转动该环应该允许二氯甲烷以形成一个完整?...

流体剪切应力已经显示出调节内皮基因方案1 -通过顺式调节元件6的活化，组蛋白乙酰转移酶活7和剪切应力响应元件（SSRE）8 5。通过调节组织因子9和组织型纤溶酶原激活剂（tPA）10表达的剪切应力的影响对凝血内皮的贡献。剪应力也影响通过调节PDGF-B的合成和反应8血管生成11和血管重塑的控制权。内皮衍生的血管活性介质肾上腺髓质素，内皮素-1，尾加压素II和松弛素也被剪切机12调节。内皮型一氧化氮合成酶的生产和一氧化氮产生的转录都是剪切依赖性10。剪切还控制内皮的ICAM-1的表达13。流动引起的剪切应力，因此可以powerfuLLY影响种类繁多的内皮细胞反应。重要的是，血管脉动现在也出现在血管性痴呆14的正常血管老化和形式的病理生理学中发挥重要作用，甚至可能导致其它神经变性疾病，如多发性硬化15。 静脉和动脉内皮细胞固有地暴露于不同的血流动力学流动模式在体内 ，许多不同的内皮细胞的表型可以显示出16。取决于幅度和流量的周期性，对内皮细胞的作用可以包括炎性细胞活化和细胞凋亡，其可反映在基因或蛋白质表达17,18变化。研究内皮细胞对剪切现象，因此仍然被困难复杂， 在能够充分产生这样的剪切模式体外模型制作。 许多不同的experimental协议已被开发应用的流体剪切应力内皮细胞单层。一个最常用的系统是平行板流动室，它创建室19内均匀的层流- 21。蠕动泵通常连接到创建的周期性流，这可以概括通常在体内 22的许多地点找到的流动特性。另一种常见的建立使用&#39;锥板“的模式，其中，流体剪切应力是由锥体23的旋转速度确定的。两个系统，和其他安排类似于它们，可能是很乏味的设置和需要，可以是相对昂贵并且不可访问的许多实验室的组件。 这些当前模型的另一个主要限制是，可以同时进行，每一个相对低的表面积复制研究的数量相对较少。这增加了时间和共<html>这种方法的mplexity。因此，诱导单向和周期性剪切的理想模型可能之一，可以很容易地设置研究重复的较高的数值，每个具有相对较大的表面积。此外，上述模式需要一个相当复杂的设置，其可以是成本高昂的许多用户。可使用的基本材料实验室产生的流体剪切扰动模型可能有几个优点。 施加单向的，周期性的剪切应力的一种简单和非常经济的方法涉及的圆形菜定轨振荡器上24的位置。这个协议是很简单的，并且可以按比例增加，以实现高的数字研究复制，每个具有相对较大的表面面积，根据需要。但是，设在盘的中心的细胞暴露于不同的流动模式比沿周细胞，产生在相同的培养皿混合蜂窝的表型应答。 _content“&gt;在当前的报告中，我们通过限制流量描述”剪切戒指&#39;，我们的模型创建单向和定期剪应力的建设和使用，设计的剪切环有效限制“混合”蜂窝剪切引起的表型通过内圈的位置的圆形培养皿至周边内途径。剪切环的结构和操作是简单和经济的，并且可以很容易地扩展使用广泛使用的组织培养供应以适应广泛范围的轨道摇床中。这模型可以在内皮细胞实验应用于生理和病理水平之内提供单向和周期性流动模式。

<table border="0" cellpadding="0" cellspacing="0" width="694">
	<tbody>
		<tr height="42">
			<td height="42" style="height:42px;width:263px;">100 x 20 mm plastic tissue culture dish</td>
			<td style="width:119px;">Corning</td>
			<td align="right" style="width:119px;">430167</td>
			<td style="width:195px;">The dishes must be polystyrene</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:263px;">150 x 25 mm plastic tissue culture dish</td>
			<td style="width:119px;">Corning</td>
			<td align="right" style="width:119px;">430599</td>
			<td>The dishes must be polystyrene</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">150 mm glass petri dish&#160;</td>
			<td style="width:119px;">Fisher</td>
			<td style="width:119px;">3160150BO</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">15ml polystyrene tissue culture plastic tubes</td>
			<td>Falcon</td>
			<td align="right">352099</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:263px;">Methylene chloride</td>
			<td style="width:119px;">Sigma-Aldrich</td>
			<td style="width:119px;">D65100</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:263px;">silicone rubber sealant</td>
			<td style="width:119px;">DAP</td>
			<td align="right" style="width:119px;">7079808641</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:263px;">ethanol</td>
			<td style="width:119px;">Decon</td>
			<td align="right" style="width:119px;">2701</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:263px;">3 mL transfer pipette</td>
			<td style="width:119px;">Becton-Dickinson</td>
			<td align="right" style="width:119px;">357524</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:263px;">printer paper</td>
			<td style="width:119px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:263px;">scissors</td>
			<td style="width:119px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:263px;">gloves</td>
			<td style="width:119px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:263px;">rotary tool and set</td>
			<td style="width:119px;">Dremel</td>
			<td style="width:119px;">4000-6/50</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:263px;">rotary tool cutting head</td>
			<td>Dremel</td>
			<td>EZ476</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:263px;">rotary tool drill head</td>
			<td style="width:119px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:263px;">distilled water</td>
			<td style="width:119px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:263px;">orbital shaker</td>
			<td style="width:119px;">VWR</td>
			<td style="width:119px;">57018-754</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:263px;">incubator</td>
			<td style="width:119px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Rat retinal microvascular endothelial cells</td>
			<td>Cell Biologics</td>
			<td>RA-6065</td>
			<td></td>
		</tr>
	</tbody>
</table>

the assembly and application of 'shear rings': a novel endothelial model for orbital, unidirectional and periodic fluid flow and shear stress

通过诱导适应性以及在血管内皮表型和基因表达的病理变化，从正常水平，并在血管生理学和病理生理学的血管流体剪切中发挥重要作用的型态偏差。特别是，周期性的，单向流动引起的剪应力的适应不良效果可以触发各种几个血管细胞类型，尤其是血管内皮细胞的影响。而现在来自不同解剖起源的内皮细胞已经培养的，深入的其对流体剪切反应受到阻碍剪切模型的相对复杂性分析（ 例如，平行平板流动腔，锥板流速模型）。虽然这些都代表着优秀的方法，这种模式在技术上是复杂和缺点，包括相对漫长和复杂的设置时间，低表面积，泵和增压常常需要密封胶和密封垫要求受苦，创建对BOT的挑战^ h维护不育和无法运行多个实验。然而，如果流动和剪切的更高的吞吐量模型可用，对血管内皮的剪切响应更大的进步，在分子水平上尤其周期性剪切研究，可能会更迅速地先进。这里，我们描述了构造和使用剪切环：一种新颖的，简单到组装，和便宜的组织培养模型具有相对大的表面面积，可以轻松地允许在单向的，周期性的剪切应力在研究大量的实验重复的内皮细胞。

在这里，我们提出从两个hCMEC / D3脑内皮细胞和大鼠视网膜微血管内皮细胞单层，在剪切环培养代表性的结果。 允许hCMEC / D3脑内皮细胞单层生长在完整的EBM至汇合后，剪切环被放置在定轨摇床上72小时。从步骤3.5使用等式，计算出的最大剪应力<img alt="公式6" src="/files/ftp_upload/54632/54632eq6.jpg" />约为2.8达因/厘米2（与参数<i...

Watch this Scientific Journal Video about The Assembly and Application of 'Shear Rings': A Novel Endothelial Model for Orbital, Unidirectional and Periodic Fluid Flow and Shear Stress at JoVE.com

The Assembly and Application of 'Shear Rings': A Novel Endothelial Model for Orbital, Unidirectional and Periodic Fluid Flow and Shear Stress

Different levels and patterns of fluid shear are known to modulate endothelial gene expression, phenotype and susceptibility to disease. We discuss the assembly and use of 'shear rings': a model that produces unidirectional, periodic shear stress patterns. Shear rings are simple to assemble, economical and can produce high cell yields.

大会和“剪切环”中的应用：一种新型内皮模型轨道，单向和定期流体流动和剪应力

Deviations from normal levels and patterns of vascular fluid shear play important roles in vascular physiology and pathophysiology by inducing adaptive as ...

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Internal organs such as the  heart, brain, and gut develop left-right (LR) asymmetries that are critical for  their normal functions1. Motile cilia are ...

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