Name: activating autophagy by aerobic exercise in mice
Uploaded: 2017-02-03T14:00:00
Description: Watch this Scientific Journal Video about Activating Autophagy by Aerobic Exercise in Mice at JoVE.com

We thank the Northwestern University Mouse Histology and Phenotyping Laboratoryfor technical support and assistance, and Noboru Mizushima (University of Tokyo) for providing GFP-LC3 transgenic mice. A. R. and C. H. were supported by the startup funds from Northwestern University and the grant from National Institutes of Health (DK094980).

所有涉及动物的程序进行根据西北大学机构动物护理和使用委员会（IACUC）批准的指导进行。 1.鼠标型号<ol><li>使用8-12周龄小鼠的运动训练。以检测体内行使诱导自噬，用于成像研究和C57BL / 6小鼠进行生化分析的GFP-LC3转基因小鼠（C57BL / 6背景）。 </li></ol> 2.运动诱发的自噬<ol><li> 跑步机设置-强制运动 <ol><li>适应和培养在10°上坡开跑步机小鼠2天。在第...

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自噬是分解代谢过程提供能量和细胞质组件或受损细胞器溶酶体降解减少细胞毒性。自噬研究重要的是要了解细胞稳态的调节和应激反应的机制。新的模式和方法不断涌现的研究领域15，研究如何吞噬受损有助于众多病理过程16,17。 营养缺乏（饥饿）和药理学诱导常用的方法来诱导自噬在体外和体?...

自噬是一种进化上保守的降解途径，这是响应于各种胁迫条件如饥饿，缺氧1,2诱导。在自噬，双膜囊，称为自噬体，包括不必要的或损坏的亚细胞成分，并将它们运送到溶酶体降解3。基底自噬是在许多疾病，包括神经变性，肿瘤和2型糖尿病4，5，6被牵连于细胞功能和生物体的发展，和受损基底自噬是必不可少的。 最知名的生理自噬诱导剂是饥饿。然而，它有两个主要的限制。首先，饥饿需要很长的时间，以有效地诱导自噬在动物中， 例如 ，在小鼠中食物限制的48小时在大多数器官。第二，饥饿勉强诱导脑自噬，由于大脑中的相对稳定的营养供给。实际上，它也难以通过小分子诱导剂来检测自噬诱导，因为许多药物不能通过血脑屏障。因此，为了更好地分析自噬活化在疾病的发病机制的功能，我们最近发现，运动是诱导自噬在时间7，8，9短时间内更有效的生理方法。与饥饿相比，自噬是有效地跑步机上跑步一样快，30分钟所致。因此，运动是一种方便和有效的生理方法来研究细胞自噬机制在调解医疗福利和预防疾病。 有用于检测自噬活动的，包括LC3和p62的几种蛋白质的标志物。 LC3（微管相关蛋白1A / 1B-光速c海恩3）是胞质蛋白（LC3-I形式）是，在自噬诱导缀合的PE（磷脂酰乙醇胺）。 PE-脂化LC3（LC3-II的形式）被募集到autophagosomal膜，可用于当用GFP标记的可视化自噬。其从细胞溶质向点状在显微镜下自噬体结构易位是自噬诱导的指示。 P62是自噬底物（如泛素化蛋白质）货物的受体，并且被并入自噬体为好。因为蛋白质是与自噬体沿溶酶体降解，其水平可用于测量自噬通量。在这里，我们展示了如何使用这些标志通过有氧运动，包括强迫运动（跑步机）和自愿运动（跑步轮）引起不同的鼠标组织量化自噬。相同的过程也可以治疗其它诱导物后施加到自噬的体内测量。

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	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">Treadmill</td>
			<td style="width:204px;">Columbus Instruments</td>
			<td style="width:195px;">150-RM Exer 3/6</td>
			<td style="width:217px;"></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">Mouse running wheel</td>
			<td style="width:204px;">Super Pet</td>
			<td style="width:195px;">100079365</td>
			<td>diameter 11.4 cm</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">Odometer</td>
			<td style="width:204px;">Bell</td>
			<td style="width:195px;">DASHBOARD 100</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">Syringe pump</td>
			<td style="width:204px;">KD Scientific</td>
			<td style="width:195px;">KDS100</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:247px;">Fluorescence microscope</td>
			<td style="width:204px;">Nikon</td>
			<td style="width:195px;">Model: inverted microscope ECLIPSE</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">Cryostat</td>
			<td style="width:204px;">Leica</td>
			<td style="width:195px;">CM 1850UV</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:247px;">Homogenizer</td>
			<td style="width:204px;">IKA</td>
			<td style="width:195px;">003737001 / Model: T10 Basic S1</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:247px;">Chloroquine</td>
			<td style="width:204px;">CAYMAN CHEMICAL COMPANY</td>
			<td style="width:195px;">14194</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">Parafolmaldehye</td>
			<td style="width:204px;">SIGMA-ALDRICH</td>
			<td style="width:195px;">P6148</td>
			<td>Personal protection equipment required. This product may release formaldehyde gas, a chemical known to cause cancer</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">Mounting media</td>
			<td style="width:204px;">Vector Laboratories</td>
			<td style="width:195px;">H-1200</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">p62 antibody</td>
			<td style="width:204px;">BD Biosciences</td>
			<td style="width:195px;">610833</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">LC3 antibody</td>
			<td style="width:204px;">Novus Biologicals</td>
			<td style="width:195px;">NB100-2220</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">2X Laemmli Sample Buffer</td>
			<td style="width:204px;">Bio-Rad Laboratories</td>
			<td style="width:195px;">161-0737</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:247px;">ImageJ</td>
			<td style="width:204px;">NIH</td>
			<td style="width:195px;"></td>
			<td></td>
		</tr>
	</tbody>
</table>

activating autophagy by aerobic exercise in mice

Autophagy is a lysosomal degradation pathway essential for cell homeostasis, function and differentiation. Under stress conditions, autophagy is induced and targets various cargos, such as bulk cytosol, damaged organelles and misfolded proteins, for degradation in lysosomes. Resulting nutrient molecules are recycled back to the cytosol for new protein synthesis and ATP production. Upregulation of autophagy has beneficial effects against the pathogenesis of many diseases, and pharmacological and physiological strategies to activate autophagy have been reported. Aerobic exercise is recently identified as an efficient autophagy inducer in multiple organs in mice, including muscle, liver, heart and brain. Here we show procedures to induce autophagy in vivo by either forced treadmill exercise or voluntary wheel running. We also demonstrate microscopic and biochemical methods to quantitatively analyze autophagy levels in mouse tissues, using the marker proteins LC3 and p62 that are transported to and degraded in lysosomes along with autophagosomes.

本协议描述两种不同的方法来诱导自噬通过有氧运动小鼠组织：共90分钟由驯化两天进行一个多车道的跑步机锻炼强迫的;两个星期自愿行使对单只装一个用来跑轮。在每个运动方案，我们可以通过测量各个器官荧光显微镜和Western blot分析自噬通量。 我们用表达GFP标记的LC3当记者通过系统的锻炼1，监测自噬的?...

Watch this Scientific Journal Video about Activating Autophagy by Aerobic Exercise in Mice at JoVE.com

Activating Autophagy by Aerobic Exercise in Mice

自噬活化是在预防许多疾病有益。一种生理方法的诱导自噬体内是体育锻炼。在这里，我们将展示如何通过有氧运动来激活自噬和在小鼠体内测量细胞自噬水平。

小鼠中自噬激活有氧运动

Autophagy is a lysosomal degradation pathway essential for cell homeostasis, function and differentiation. Under stress conditions, autophagy is induced ...

Research

JoVE Journal

Biology

Agar-Block Microcosms for Controlled Plant Tissue Decomposition by Aerobic Fungi

为控制植物组织分解好氧菌的琼脂块缩影

The two principal methods for studying fungal biodegradation of lignocellulosic plant tissues were developed for wood preservative testing (soil-block; ...

科研

生物学

Determining the Contribution of the Energy Systems During Exercise

确定的能源系统，在运动的贡献

One of the most important aspects of the metabolic demand is the relative contribution of the energy systems to the total energy required for a given ...

Transient Expression of Proteins by Hydrodynamic Gene Delivery in Mice

蛋白通过流体动力基因传递在小鼠体内瞬时表达

Efficient expression of transgenes in vivo is of critical importance in studying gene function and developing treatments for diseases. Over the past ...

In Vivo 4-Dimensional Tracking of Hematopoietic Stem and Progenitor Cells in Adult Mouse Calvarial Bone Marrow

In Vivo 4-Dimensional Tracking of Hematopoietic Stem and Progenitor Cells in Adult Mouse Calvarial Bone Marrow

在造血干细胞和祖细胞的成年小鼠颅骨骨髓体内四维跟踪

Through a delicate balance between quiescence and proliferation, self renewal and production of differentiated progeny, hematopoietic stem cells (HSCs) ...

Analysis of Autophagy in Penicillium chrysogenum by Using Starvation Pads in Combination With Fluorescence Microscopy

Analysis of Autophagy in Penicillium chrysogenum by Using Starvation Pads in Combination With Fluorescence Microscopy

自噬的分析<em&gt;产黄青霉</em&gt;通过结合使用饥饿垫荧光显微镜

The study of cellular quality control systems has emerged as a highly dynamic and relevant field of contemporary research. It has become clear that cells ...

Fecal Glucocorticoid Analysis: Non-invasive Adrenal Monitoring in Equids

糖皮质激素粪分析：在马属动物无创监测肾上腺

Adrenal activity can be assessed in the equine species by analysis of feces for corticosterone metabolites. During a potentially aversive situation, ...

Study of Protein-protein Interactions in Autophagy Research

自噬研究中蛋白质-蛋白质相互作用的研究

Protein-protein interactions are important for understanding cellular signaling cascades and identifying novel pathway components and protein dynamics. ...

In Situ Immunofluorescent Staining of Autophagy in Muscle Stem Cells

In Situ Immunofluorescent Staining of Autophagy in Muscle Stem Cells

肌肉干细胞中自噬的原位免疫荧光染色

Increasing evidence points to autophagy as a crucial regulatory process to preserve tissue homeostasis. It is known that autophagy is involved in skeletal ...

The Lactate Dehydrogenase Sequestration Assay &#8212; A Simple and Reliable Method to Determine Bulk Autophagic Sequestration Activity in Mammalian Cells

The Lactate Dehydrogenase Sequestration Assay ÃƒÆ’Ã‚Â¢ÃƒÂ¢Ã¢â‚¬Å¡Ã‚Â¬ÃƒÂ¢Ã¢â€šÂ¬Ã‚Â A Simple and Reliable Method to Determine Bulk Autophagic Sequestration Activity in Mammalian Cells

乳酸脱氢酶螯合测定--一种简便、可靠的测定哺乳动物细胞中自噬性螯合活性的方法

Bulk autophagy is characterized by the sequestration of large portions of cytoplasm into double/multi-membrane structures termed autophagosomes. Here a ...

Measurement of Insulin- and Contraction-Stimulated Glucose Uptake in Isolated and Incubated Mature Skeletal Muscle from Mice

测量小鼠分离和孵育成熟骨骼肌中胰岛素和收缩刺激的葡萄糖摄取

Skeletal muscle is an insulin-responsive tissue and typically takes up most of the glucose that enters the blood after a meal. Moreover, it has been ...