Name: new application of an atmospheric pressure plasma jet as a neuro-protective agent against glucose deprivation-induced injury of sh-sy5y cells
Uploaded: 2017-10-09T15:00:00
Description: Watch this Scientific Journal Video about New Application of an Atmospheric Pressure Plasma Jet as a Neuro-protective Agent Against Glucose Deprivation-induced Injury of SH-SY5Y Cells at JoVE.com

这项工作得到了北京神经外科研究所 (2014-11)、中国国家自然科学基金 (Nos. 11475019 和 81271286) 和北京自然科学基金会 (No. 7152027) 的创新基金会的支持。

1. APPJ 设备的准备 警告: 请在使用前查阅所有相关的材料安全数据表 (MSDS)。在执行所有的和 #160 时, 请使用适当的安全做法; 实验, 包括使用通风罩和个人防护设备 (安全眼镜、防护手套、实验室大衣、 等 )。该协议要求标准的细胞处理技术 (杀菌, 细胞恢复, 细胞传代, 细胞冻结, 细胞染色, 等 ). 
<ol> <li> 选择一个内径为 1 mm 的石英管, 外部直径为 3 mm. 使用抛光?...

<ol>
	<li>Yang, S. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Alternative+mitochondrial+electron+transfer+for+the+treatment+of+neurodegenerative+diseases+and+cancers:+Methylene+blue+connects+the+dots.">Alternative mitochondrial electron transfer for the treatment of neurodegenerative diseases and cancers: Methylene blue connects the dots.</a> Prog. Neurobiol. , (2015).</li><li>Bhat, A. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Oxidative+stress,+mitochondrial+dysfunction+and+neurodegenerative+diseases;+a+mechanistic+insight.">Oxidative stress, mitochondrial dysfunction and neurodegenerative diseases; a mechanistic insight.</a> Biomed. Pharmacother. 74, 101-110 (2015).</li><li>Bullon, P., Newman, H. 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Cell Longev. 2016 (22), 1245049 (2016).</li><li>Contestabile, A., Ciani, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Role+of+nitric+oxide+in+the+regulation+of+neuronal+proliferation,+survival+and+differentiation.">Role of nitric oxide in the regulation of neuronal proliferation, survival and differentiation.</a> Neurochemistry International. 45 (6), 903-914 (2004).</li><li>Yan, X., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Protective+effect+of+atmospheric+pressure+plasma+on+oxidative+stress-induced+neuronal+injuries:+an+in+vitro+study.">Protective effect of atmospheric pressure plasma on oxidative stress-induced neuronal injuries: an in vitro study.</a> J. Phys. D: Appl. Phys. 50 (9), 095401 (2017).</li><li>Xie, H. R., Hu, L. S., Li, G. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=SH-SY5Y+human+neuroblastoma+cell+line:+in+vitro+cell+model+of+dopaminergic+neurons+in+Parkinson's+disease.">SH-SY5Y human neuroblastoma cell line: in vitro cell model of dopaminergic neurons in Parkinson's disease.</a> Chin. Med. J. 123 (8), 1086-1092 (2010).</li><li>Tsutsui, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Treatment+of+cardiac+disease+by+inhalation+of+atmospheric+pressure+plasma.">Treatment of cardiac disease by inhalation of atmospheric pressure plasma.</a> Japanese Journal of Applied Physics. 53 (6), 060309 (2014).</li><li>Graves, D. 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SH-SY5Y 细胞是人神经母细胞瘤衍生的干细胞系, 并广泛应用作为一个适当的细胞模型, 为体外神经系统毒性或脑保护的研究12。SH-SY5Y 细胞系对葡萄糖缺乏状态敏感。1小时葡萄糖缺乏后, 细胞活力降低到近 50%, 这是研究药效学的最佳细胞生存条件。此外, CCK-8 试剂对细胞没有细胞毒性, 在 APPJ 治疗后的葡萄糖剥夺条件下, CCK-8 试剂孵育4小时, 以检测细胞的活力。在目前的研究中...

在正常的生理条件下, 成年大脑几乎完全使用葡萄糖作为能量代谢的基质。人脑只占身体重量的 2%, 但在身体的总葡萄糖中消耗了大约25% 的1。众所周知, 葡萄糖代谢失调是缺血性中风和各种神经退行性疾病 (包括阿尔茨海默氏病 (AD)、亨廷顿氏病 (HD) 和帕金森病 (PD) 的主要病理变化之一2,3。葡萄糖的缺乏和糖的摄取受损或氧化磷酸化可以直接影响 ATP 的生产, 并进一步诱发神经细胞死亡, 这可能会增加神经元功能障碍的风险, 这表明维持细胞活力或延缓葡萄糖剥夺后的细胞损伤可能是治疗这些疾病的合理方法。通过葡萄糖调制对神经保护作用的研究, 着重于抗炎剂、离子通道调制器、自由基清道夫、神经营养因子、等。然而, 这些神经保护方法的翻译从长凳到临床实践没有成功4。
常压等离子体射流 (APPJs) 是一种新型的大气低温气体放电技术, 近年来引起了众多学科研究者的关注。APPJs 已用于几十年的各种生物医学应用, 如癌症细胞治疗, 细菌灭活, 凝血, 伤口愈合, 口服药物,等5,6, 由于其多种类型的排放活性氮种类 (RNS) 和活性氧 (ROS) (图 1)7。以前的等离子体生物医学应用主要集中在细菌、细胞和组织的氧化和/或 nitrative 压力上8。然而, APPJ 也可能是一把 "双刃剑", 因为 RNS 和 ROS 是重要的细胞内信号分子与许多生理学和病理生理学过程9。一氧化二氮 (no) 控制着广泛的生物过程, 在人体, 特别是中枢神经系统中扮演着双重角色。低水平的 NO 已经显示了他们的神经保护活动的体外和在体内通过多个信号通路10。我们先前的研究首次报告, 氦 APPJ 诱导 NO 的产生参与了 APPJ 抗氧化应激损伤的神经保护作用11。然而, 未报告 APPJs 对其他损伤的影响。因此, 本研究的目的是提供一个体外治疗方案, 关于氦 APPJ 在葡萄糖剥夺诱导的 SH-SY5Y 细胞损伤中的应用。与以往的研究不同, 我们的协议使用低剂量等离子治疗的神经保护应用, 而不会造成过多的血浆损伤, 表明 APPJ 治疗可能被用作一种新型的 "无供药物"为将来的研究, 甚至临床翻译。本协议也被建议作为细胞应用的其他细胞类型有不同的损伤, 但 APPJ 治疗条件应调整和治疗剂量必须相对较低。

<table width="911">
	<tbody>
		<tr height="20">
			<td height="20" style="width:216px;height:20px;">SH-SY5Y cell line</td>
			<td style="width:325px;">China Center for Type Culture Collection</td>
			<td style="width:135px;">3111C0001CCC000026</td>
			<td style="width:235px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="width:216px;height:21px;">RPMI 1640 medium</td>
			<td style="width:325px;">Thermo Scientific&#160;</td>
			<td style="width:135px;">21875091</td>
			<td>stored at 4 &#176;C</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:216px;height:21px;">RPMI 1640 medium no glucose</td>
			<td style="width:325px;">Thermo Scientific</td>
			<td style="width:135px;">11879020</td>
			<td>stored at 4 &#176;C</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:216px;height:21px;">fetal calf serum</td>
			<td style="width:325px;">Thermo Scientific</td>
			<td style="width:135px;">16000044</td>
			<td style="width:235px;">stored at -20 &#176;C</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:216px;height:21px;">tripsin-EDTA solution</td>
			<td style="width:325px;">Solarbio</td>
			<td style="width:135px;">T1300</td>
			<td>stored at 4 &#176;C</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:216px;height:21px;">96 wells plate</td>
			<td style="width:325px;">corning</td>
			<td style="width:135px;">3599</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="width:216px;height:21px;">Cell Counting Kit-8 (CCK-8)</td>
			<td style="width:325px;">Dojindo Laboratories</td>
			<td style="width:135px;">CK04</td>
			<td>stored at 4 &#176;C</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:216px;height:21px;">microplate reader</td>
			<td style="width:325px;">Tecan</td>
			<td style="width:135px;">M200 Pro</td>
			<td>for measuring the absorbance at 450 nm</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:216px;height:21px;">High &#8211; voltage Power Amplifier</td>
			<td style="width:325px;">Trek</td>
			<td style="width:135px;">PD06087</td>
			<td>for amplifing the power</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:216px;height:21px;">Function Signal Generator</td>
			<td style="width:325px;">MaZe Electronics Science&#38;Technology</td>
			<td style="width:135px;">AT30120</td>
			<td>for providing the specific signal</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:216px;height:21px;">High &#8211; Voltage Probe</td>
			<td style="width:325px;">Tektronix</td>
			<td style="width:135px;">P6015A</td>
			<td>for detecting high voltage</td>
		</tr>
		<tr height="21">
			<td height="21" style="width:216px;height:21px;">Digital Oscilloscope</td>
			<td style="width:325px;">Tektronix</td>
			<td style="width:135px;">DPO4104B</td>
			<td>for displaying the signal</td>
		</tr>
	</tbody>
</table>

new application of an atmospheric pressure plasma jet as a neuro-protective agent against glucose deprivation-induced injury of sh-sy5y cells

大气压力等离子体射流 (APPJ) 近年来引起了许多研究人员的关注, 因为它的排放包括多种类型的活性氮 (RNS) 和活性氧物种 (ROS)。我们先前的研究表明 APPJ 对氧化应激损伤的细胞作用。本研究的目的是提供一个详细的体外治疗方案, 关于神经保护应用的氦 APPJs 对葡萄糖剥夺诱导损伤 SH-SY5Y 细胞。SH-SY5Y 的人神经母细胞瘤的干细胞系维持在 RPMI 1640 培养基补充15% 胎小牛血清。在 APPJ 治疗前, 培养基被转化为 RPMI 1640, 无葡萄糖。在一个细胞培养器 1 h 孵化后, 细胞活力是确定使用细胞计数套件8。结果表明, 与葡萄糖剥夺组相比, APPJ 治疗的细胞表现出明显增加的细胞活力, 以剂量依赖性的方式, 8 s/良好观察作为最佳剂量。同时, 氦流动对葡萄糖剥夺诱导的细胞损伤无影响。我们的研究结果表明, APPJ 可能被用作治疗中枢神经系统中与葡萄糖缺乏相关的疾病的一种方法。本协议也可作为其他不同损伤细胞的细胞应用, 但细胞培养和 APPJ 处理条件应重新调整, 治疗剂量必须相对较低。

数据被表示为至少三独立实验的平均± SD。采用方差分析法对组结果进行方差统计。所有的分析都是用统计分析软件棱镜和 p 和 #60 进行的; 0.05 是统计意义的门槛。
CCK-8 孵育4小时后测定细胞活力。如图 3所示, 与对照组相比, 葡萄糖剥夺使 SH-SY5Y 细胞的生存能力降低到44.1 ± 2.6% (通常在 RPMI 1640 介质中含有 15% FBS)。APPJ 治疗显著增加了细胞生存能力的剂量?...

Watch this Scientific Journal Video about New Application of an Atmospheric Pressure Plasma Jet as a Neuro-protective Agent Against Glucose Deprivation-induced Injury of SH-SY5Y Cells at JoVE.com

New Application of an Atmospheric Pressure Plasma Jet as a Neuro-protective Agent Against Glucose Deprivation-induced Injury of SH-SY5Y Cells

常压等离子体射流作为神经保护剂抗葡萄糖剥夺诱导 SH-SY5Y 细胞损伤的新应用

小剂量大气压等离子体治疗糖剥夺性 SH-SY5Y 损伤的神经保护应用协议

The atmospheric pressure plasma jet (APPJ) has attracted the attention of many researchers from multiple disciplines in recent years because its emissions ...

Research

JoVE Journal

Engineering

Synthesis and Microdiffraction at Extreme Pressures and Temperatures

在极端的压力和温度的合成和Microdiffraction

High pressure compounds and polymorphs are investigated for a broad range of purposes such as determine structures and processes of deep planetary ...

科研

工程学

Visualization of High Speed Liquid Jet Impaction on a Moving Surface

高速喷液嵌塞对移动表面的可视化

Two apparatuses for examining liquid jet impingement on a high-speed moving surface are described: an air cannon device (for examining surface speeds ...

How to Ignite an Atmospheric Pressure Microwave Plasma Torch without Any Additional Igniters

如何点燃常压微波等离子体炬没有任何额外的点火器

This movie shows how an atmospheric pressure plasma torch can be ignited by microwave power with no additional igniters. After ignition of the plasma, a ...

Blast Quantification Using Hopkinson Pressure Bars

高炉利用定量霍普金森杆

Near-field blast load measurement presents an issue to many sensor types as they must endure very aggressive environments and be able to measure pressures ...

Three-dimensional Particle Tracking Velocimetry for Turbulence Applications: Case of a Jet Flow

湍流应用三维粒子跟踪测速：一喷流病例

3D-PTV is a quantitative flow measurement technique that aims to track the Lagrangian paths of a set of particles in three dimensions using stereoscopic ...

Applying X-ray Imaging Crystal Spectroscopy for Use as a High Temperature Plasma Diagnostic

运用X射线成像光谱水晶的使用为高温等离子体诊断

X-ray spectra provide a wealth of information on high temperature plasmas; for example electron temperature and density can be inferred from line ...

An Atmospheric Pressure Plasma Setup to Investigate the Reactive Species Formation

大气压等离子体设置调查活性物种形成

Non-thermal atmospheric pressure (&#39;cold&#39;) plasmas have received increased attention in recent years due to their significant biomedical potential. ...

Atmospheric Pressure Fabrication of Large-Sized Single-Layer Rectangular SnSe Flakes

大型单层矩形 SnSe 片的常压制备

Tin selenide (SnSe) belongs to the family of layered metal chalcogenide materials with a buckled structure like phosphorene, and has shown potential for ...

Magnet Assisted Composite Manufacturing: A Flexible New Technique for Achieving High Consolidation Pressure in Vacuum Bag/Lay-Up Processes

磁铁辅助复合制造: 真空袋/糊化过程中实现高固结压力的一种柔性新技术

This work demonstrates a protocol to improve the quality of composite laminates fabricated by wet lay-up vacuum bag processes using the recently developed ...

Nerve Stem Cell Differentiation by a One-step Cold Atmospheric Plasma Treatment In Vitro

Nerve Stem Cell Differentiation by a One-step Cold Atmospheric Plasma Treatment In Vitro

一步冷常压等离子体体外处理神经干细胞分化

As the development of physical plasma technology, cold atmospheric plasmas (CAPs) have been widely investigated in decontamination, cancer treatment, ...