Name: differentiated mouse adipocytes in primary culture: a model of insulin resistance
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我们感谢丹尼尔·蒙德拉贡、安东尼奥·普拉多、费尔南多·洛佩斯-巴雷拉、马丁·加西亚-塞尔文、亚历杭德拉·卡斯蒂利亚和玛丽亚·安东尼埃塔·卡尔巴霍的技术援助，感谢杰西卡·冈萨雷斯·诺里斯对手稿的批判性编辑。该议定书得到了墨西哥国家科学和技术委员会（CONACYT）、教育部门研究基金赠款284771（致Y.M.）的支持。

所有啮齿动物实验均由墨西哥国立自治大学神经生物学研究所生物伦理学委员会批准，协议编号075。
1.小鼠脂肪细胞前体细胞的分离
<ol><li>对8-10周龄的雄性C57BL / 6小鼠实施安乐死（例如，通过CO2吸入和随后的宫颈脱位）（每次分离四只动物）。用70%乙醇摩擦对小鼠进行消毒，并在处死后立即获得脂肪组织。 注意：啮齿动物安乐死后，立即分离脂肪组?...

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本文提供了一种研究胰岛素抵抗的方法，该方法使用用TNF-α处理的培养物中的原代脂肪细胞。该模型的优点是原代脂肪细胞可以在规定的条件下长时间培养，并严格控制细胞环境因素26。测定持续时间为15-20天，尽管实验之间可能发生分化脂肪细胞百分比的变化。原代脂肪细胞比细胞系具有优势，因为它们在培养中没有持续扩增，并且更紧密地捕获了它们来源的组织多样性<sup clas...

胰岛素是由胰岛β细胞产生的合成代谢激素，是葡萄糖和脂质代谢的关键调节因子。在其众多功能中，胰岛素调节葡萄糖摄取、糖原合成、糖异生、蛋白质合成、脂肪生成和脂肪分解1。胰岛素与其受体（IR）相互作用后的初始分子信号是IR2的内在酪氨酸蛋白激酶活性的激活，导致其自身磷酸化3和随后称为胰岛素受体底物（IRS）的蛋白质家族的激活，其与衔接蛋白结合导致级联蛋白激酶的激活4.胰岛素激活两种主要的信号通路：磷脂酰肌醇3激酶（PI3K）蛋白激酶B（AKT）和Ras-丝裂原活化蛋白激酶（MAPK）。前者构成一个主要的分支点或节点4，5，用于激活参与各种生理功能的众多下游效应子，包括调节燃料稳态，而后者调节细胞生长和分化4，6。胰岛素作用最终取决于细胞类型和生理环境7。
主要的胰岛素反应代谢组织之一是脂肪组织。白色脂肪组织是人类和啮齿动物中最丰富的脂肪类型，分布在皮下脂肪（皮肤和肌肉之间）和内脏脂肪（腹腔器官周围）中。鉴于其体积大，脂肪细胞或脂肪细胞是脂肪组织中最丰富的细胞类型。这些脂肪细胞可以是棕色/米色（产热）、粉红色（乳腺）和白色8，9。白色脂肪细胞以甘油三酯的形式保持体内的主要能量储备，这是一种胰岛素依赖性过程。胰岛素促进葡萄糖转运和脂肪生成，同时抑制脂肪分解或脂质分解7，10。它还有助于前脂肪细胞分化为脂肪细胞 - 成熟的脂肪储存细胞11。
当正常胰岛素水平产生减弱的生物反应时，就会发生胰岛素抵抗，从而导致代偿性高胰岛素血症12。胰岛素抵抗是一种与超重和肥胖相关的疾病5，当它们结合在一起时会导致2型糖尿病（T2D）和其他代谢性疾病13。高胰岛素血症补偿外周组织中的胰岛素抵抗，以维持正常的血糖水平14。然而，最终的β细胞丢失或衰竭，以及胰岛素抵抗加剧，导致血糖水平升高，符合T2D5。因此，胰岛素抵抗和高胰岛素血症可导致肥胖衍生代谢疾病的发展15。此外，肥胖可能导致慢性低度局部炎症，促进脂肪组织中的胰岛素抵抗15，16，17。此外，肥胖引起的脂肪组织改变，如纤维化、炎症以及血管生成和脂肪生成减少，导致脂联素血清水平（一种胰岛素增敏剂）降低，纤溶酶原激活剂抑制剂 1 （PAI-1）、游离脂肪酸和外泌体等因子分泌增加进入血液，加剧胰岛素抵抗17。
胰岛素抵抗的许多方面仍然未知。已经开发了体外和体内模型来研究介导主要靶组织（包括脂肪组织）中胰岛素抵抗的机制。体外模型的优点是研究人员可以更好地控制环境条件，并且可以评估特定细胞类型的胰岛素抵抗。特别是，脂肪细胞前体细胞（APC）具有供体组织的个体表型，这可能比脂肪细胞系更好地反映生理学。体外诱导胰岛素抵抗的一个主要因素是肿瘤坏死因子-α（TNF-α）。TNF-α是由脂肪组织中的脂肪细胞和巨噬细胞分泌的促炎细胞因子18。虽然适当的脂肪组织重塑和扩增是必需的19，但长期暴露于TNF-α会在体内脂肪组织和体外脂肪细胞中诱导胰岛素抵抗20。几种细胞类型的慢性TNF-α处理导致IR和IRS-1的丝氨酸磷酸化增加，从而促进酪氨酸磷酸化减少21。IRS-1在丝氨酸残基上的磷酸化增加抑制IR酪氨酸激酶活性，可能是慢性TNF-α治疗损害胰岛素作用的关键机制之一22，23。TNF-α激活涉及核因子ĸB激酶β（IKKβ）和c-Jun N末端激酶（JNK）的丝氨酸/苏氨酸激酶抑制剂的途径24。JNK诱导复杂的促炎转录程序，但也直接磷酸化IRS-16。
了解胰岛素抵抗的发病机制对于指导未来针对T2D的疗法的开发变得越来越重要。APC已被证明是研究脂肪细胞生物学的优秀模型，包括对胰岛素的敏感性和抵抗力，以及独立于全身环境鉴定脂肪细胞的内在特性。APC可以很容易地从不同的脂肪库获得，并在适当的条件下分化成成熟的脂肪细胞。使用这种方法，可以评估对脂肪细胞中胰岛素抵抗/敏感性的直接影响。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>1. Isolation mouse adipocyte precursor cells</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>ACK lysing buffer&#160;</td>
			<td>LONZA</td>
			<td>10-548E</td>
			<td></td>
		</tr>
		<tr>
			<td>Anti-Biotin Microbeads&#160;</td>
			<td>Miltenyi</td>
			<td>130-090-485</td>
			<td></td>
		</tr>
		<tr>
			<td>Anti-CD31</td>
			<td>eBioscience</td>
			<td>13-0311-85</td>
			<td></td>
		</tr>
		<tr>
			<td>AutoMACS Pro Separator</td>
			<td>Miltenyi</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Basement membrane matrix (matrigel)</td>
			<td>Corning</td>
			<td>354234</td>
			<td></td>
		</tr>
		<tr>
			<td>bFGF</td>
			<td>Sigma</td>
			<td>F0291</td>
			<td>Growth factor</td>
		</tr>
		<tr>
			<td>BSA</td>
			<td>Equitech-Bio, Inc.</td>
			<td>BAC63-1000</td>
			<td></td>
		</tr>
		<tr>
			<td>CD45 Monoclonal Antibody (30-F11) - Biotin&#160;</td>
			<td>eBioscience</td>
			<td>13-0451-85</td>
			<td></td>
		</tr>
		<tr>
			<td>Collagenase, Type 1</td>
			<td>Worthington Biochem</td>
			<td>LS004197</td>
			<td></td>
		</tr>
		<tr>
			<td>Dexamethasone</td>
			<td>Sigma</td>
			<td>D1756</td>
			<td></td>
		</tr>
		<tr>
			<td>DMEM</td>
			<td>GIBCO</td>
			<td>12800017</td>
			<td></td>
		</tr>
		<tr>
			<td>DMEM low glucose</td>
			<td>GIBCO</td>
			<td>31600-034</td>
			<td></td>
		</tr>
		<tr>
			<td>EGF</td>
			<td>Peprotech</td>
			<td>315-09</td>
			<td>Growth factor</td>
		</tr>
		<tr>
			<td>FBS</td>
			<td>GIBCO</td>
			<td>26140-079</td>
			<td></td>
		</tr>
		<tr>
			<td>ITS mix</td>
			<td>Sigma</td>
			<td>I3146</td>
			<td></td>
		</tr>
		<tr>
			<td>L-ascorbic acid 2-phosphate</td>
			<td>Sigma</td>
			<td>A8960</td>
			<td></td>
		</tr>
		<tr>
			<td>LIF</td>
			<td>Millipore</td>
			<td>ESG1107</td>
			<td>Growth factor</td>
		</tr>
		<tr>
			<td>Linoleic acid-albumin</td>
			<td>Sigma</td>
			<td>L9530</td>
			<td></td>
		</tr>
		<tr>
			<td>MCDB 201 medium</td>
			<td>Sigma</td>
			<td>M6770</td>
			<td></td>
		</tr>
		<tr>
			<td>Normocin</td>
			<td>InvivoGen</td>
			<td>ant-nr-2</td>
			<td></td>
		</tr>
		<tr>
			<td>PDGF-BB&#160;</td>
			<td>Peprotech</td>
			<td>315-18</td>
			<td>Growth factor</td>
		</tr>
		<tr>
			<td>Peniciline-Streptomycine</td>
			<td>BioWest</td>
			<td>L0022-100</td>
			<td></td>
		</tr>
		<tr>
			<td>Pre-Separation Filters (70 &#181;m)</td>
			<td>Miltenyi</td>
			<td>130-095-823</td>
			<td></td>
		</tr>
		<tr>
			<td>Purified Rat Anti-Mouse CD16 / CD32&#160;</td>
			<td>BD Pharmingen</td>
			<td>553142</td>
			<td></td>
		</tr>
		<tr>
			<td>Trypsin-EDTA&#160;</td>
			<td>GIBCO</td>
			<td>25300062</td>
			<td></td>
		</tr>
		<tr>
			<td>2. Adipocyte differentiation and insulin resistance induction</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>3-Isobutyl-1-methylxanthine [IBMX]</td>
			<td>Sigma</td>
			<td>I5879</td>
			<td>Differentiation cocktail</td>
		</tr>
		<tr>
			<td>BMP4</td>
			<td>R&#38;D Systems</td>
			<td>5020-BP-010</td>
			<td>Differentiation cocktail</td>
		</tr>
		<tr>
			<td>Dexamethasone</td>
			<td>Sigma</td>
			<td>D1756</td>
			<td>Differentiation cocktail</td>
		</tr>
		<tr>
			<td>Insulin</td>
			<td>Sigma</td>
			<td>I9278</td>
			<td></td>
		</tr>
		<tr>
			<td>Rosiglitazone</td>
			<td>Cayman</td>
			<td>71742</td>
			<td>Differentiation cocktail</td>
		</tr>
		<tr>
			<td>TNF&#945;</td>
			<td>R&#38;D Systems</td>
			<td>210-TA-005&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>3. Evaluation of insulin signaling pathway by western blot</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Anti-beta tubulin antibody</td>
			<td>Abcam</td>
			<td>ab6046</td>
			<td></td>
		</tr>
		<tr>
			<td>Bromophenol blue</td>
			<td>BioRad</td>
			<td>161-0404</td>
			<td>Laemmli buffer</td>
		</tr>
		<tr>
			<td>EDTA</td>
			<td>Sigma</td>
			<td>E5134</td>
			<td>RIPA buffer</td>
		</tr>
		<tr>
			<td>EGTA</td>
			<td>Sigma</td>
			<td>E4378</td>
			<td>RIPA buffer</td>
		</tr>
		<tr>
			<td>FluorChem E system</td>
			<td>ProteinSimple</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Glycerol</td>
			<td>Sigma</td>
			<td>G6279</td>
			<td>Laemmli buffer</td>
		</tr>
		<tr>
			<td>Glycine</td>
			<td>Sigma</td>
			<td>G7126</td>
			<td>Running and Transfer buffer</td>
		</tr>
		<tr>
			<td>Igepal</td>
			<td>Sigma</td>
			<td>I3021</td>
			<td>RIPA buffer</td>
		</tr>
		<tr>
			<td>2- mercaptoethanol</td>
			<td>Sigma</td>
			<td>M3148</td>
			<td>Laemmli buffer</td>
		</tr>
		<tr>
			<td>Methanol</td>
			<td>JT Baker</td>
			<td>907007</td>
			<td>Transfer buffer</td>
		</tr>
		<tr>
			<td>NaCl</td>
			<td>JT Baker</td>
			<td>3624-05</td>
			<td>TBS-T</td>
		</tr>
		<tr>
			<td>NaF</td>
			<td>Sigma</td>
			<td>77F-0379</td>
			<td>RIPA buffer</td>
		</tr>
		<tr>
			<td>NaOH&#160;</td>
			<td>JT Baker</td>
			<td>3722-19</td>
			<td></td>
		</tr>
		<tr>
			<td>Na4P2O7</td>
			<td>Sigma</td>
			<td>114F-0762</td>
			<td>RIPA buffer</td>
		</tr>
		<tr>
			<td>Na3VO4</td>
			<td>Sigma</td>
			<td>S6508</td>
			<td>RIPA buffer</td>
		</tr>
		<tr>
			<td>Nitrocellulose membrane&#160;</td>
			<td>BioRad</td>
			<td>1620112</td>
			<td></td>
		</tr>
		<tr>
			<td>Nonfat dry milk</td>
			<td>BioRad</td>
			<td>1706404</td>
			<td>Blocking solution</td>
		</tr>
		<tr>
			<td>Prestained protein standard&#160;</td>
			<td>BioRad</td>
			<td>1610395</td>
			<td></td>
		</tr>
		<tr>
			<td>Protease inhibitor cocktail&#160;</td>
			<td>Sigma</td>
			<td>P8340-5ML</td>
			<td></td>
		</tr>
		<tr>
			<td>Peroxidase AffiniPure Donkey Anti-Rabbit IgG (H+L)&#160;</td>
			<td>Jackson ImmunoResearch</td>
			<td>711-035-132</td>
			<td></td>
		</tr>
		<tr>
			<td>Phospho- Insulin Receptor &#946;&#160;</td>
			<td>Cell signaling</td>
			<td>3024</td>
			<td></td>
		</tr>
		<tr>
			<td>Phospho-Akt (Ser473) Antibody</td>
			<td>Cell signaling</td>
			<td>9271</td>
			<td></td>
		</tr>
		<tr>
			<td>Phospho-IRS1 (Tyr608) antibody</td>
			<td>Millipore</td>
			<td>9432</td>
			<td></td>
		</tr>
		<tr>
			<td>Saccharose&#160;</td>
			<td>JT Baker</td>
			<td>407205</td>
			<td>RIPA buffer</td>
		</tr>
		<tr>
			<td>SDS</td>
			<td>BioRad</td>
			<td>1610302</td>
			<td>Running and laemmli buffer</td>
		</tr>
		<tr>
			<td>SuperSignal West Pico PLUS Chemiluminescent Substrate</td>
			<td>Thermo Scientific</td>
			<td>34577</td>
			<td></td>
		</tr>
		<tr>
			<td>Tris-base</td>
			<td>Promega</td>
			<td>H5135</td>
			<td>Running, transfer and laemmli buffer</td>
		</tr>
		<tr>
			<td>Tris-HCl</td>
			<td>JT Baker</td>
			<td>4103-02</td>
			<td>RIPA buffer - TBS</td>
		</tr>
		<tr>
			<td>Tween 20</td>
			<td>Sigma</td>
			<td>P1379</td>
			<td>TBS-T</td>
		</tr>
	</tbody>
</table>

differentiated mouse adipocytes in primary culture: a model of insulin resistance

胰岛素抵抗是胰岛素对其靶细胞的影响降低，通常源于胰岛素受体信号传导的减少。胰岛素抵抗导致2型糖尿病（T2D）和其他肥胖衍生疾病的发展，这些疾病在世界范围内患病率很高。因此，了解胰岛素抵抗的潜在机制非常重要。几种模型已被用于研究 体内 和 体外胰岛素抵抗;原代脂肪细胞是研究胰岛素抵抗机制和鉴定抵消这种情况的分子和胰岛素增敏药物分子靶标的一种有吸引力的选择。在这里，我们使用用肿瘤坏死因子-α（TNF-α）处理的培养物中的原代脂肪细胞建立了胰岛素抵抗模型。
通过磁细胞分离技术从胶原酶消化的小鼠皮下脂肪组织中分离出的脂肪细胞前体细胞（APC）分化为原代脂肪细胞。然后通过TNF-α治疗诱导胰岛素抵抗，TNF-是一种促炎细胞因子，可减少胰岛素信号级联反应成员的酪氨酸磷酸化/激活。通过蛋白质印迹定量胰岛素受体 （IR）、胰岛素受体底物 （IRS-1） 和蛋白激酶 B （AKT） 磷酸化降低。该方法为研究脂肪组织中介导胰岛素抵抗的机制提供了极好的工具。

在过去的几年中，肥胖和T2D患病率的增加促使人们强烈寻找介导脂肪组织中胰岛素抵抗的机制。通过此处描述的方案，APC可以分化为成熟的脂肪细胞，以评估胰岛素抵抗和敏感性。一旦APC达到汇合，则需要10天才能完成分化为成熟脂肪细胞及其TNF-α介导的胰岛素抵抗诱导（图1）。
APCs显示出成纤维细胞样形态，其特征在于其扁平和细长的形状以及它们在?...

Watch this Scientific Journal Video about Differentiated Mouse Adipocytes in Primary Culture: A Model of Insulin Resistance at JoVE.com

Differentiated Mouse Adipocytes in Primary Culture: A Model of Insulin Resistance

该协议描述了从皮下脂肪中分离小鼠前脂肪细胞，它们分化为成熟脂肪细胞以及诱导胰岛素抵抗。通过蛋白质印迹的胰岛素信号通路成员的磷酸化/激活来评估胰岛素作用。该方法可以直接测定原代脂肪细胞的胰岛素抵抗/敏感性。

原代培养中分化的小鼠脂肪细胞：胰岛素抵抗模型

Insulin resistance is a reduced effect of insulin on its target cells, usually derived from decreased insulin receptor signaling. Insulin resistance ...

可以在原代脂肪细胞中评估胰岛素抵抗，从而可以在不同的生理病理学背景下研究供体，例如瘦与肥胖，或来自不同脂肪沉积物的细胞。原代脂肪细胞保留了许多内在特性，可以在规定的条件下长时间培养，并严格控制细胞环境因素。要开始分化过程，细胞必须处于80%汇合。如果分化从较低或较高的汇合处开始，细胞将分化较少或失去分化能力。牺牲动物后，用70%乙醇摩擦对小鼠进行消毒。解剖每只小鼠的腹股沟皮下脂肪组织，并将其收集在15毫升锥形管中，其中含有15毫升冰上的一型胶原酶溶液。用无菌手术剪刀将脂肪组织切成小块，并通过在 37 摄氏度的轨道振荡器中以 150 RPM 的速度与 1 型胶原酶溶液一起孵育 30 分钟来消化样品。每 10 分钟检查一次消化，以确保其正常工作，并防止过度消化。使用200微米网状注射器过滤以消除未被胶原酶消化的组织，并将过滤器通过管的边缘以将尽可能多的细胞排入溶液中。加入 15 毫升冷 DMEM 1%BSA 以停止消化，并在 400 摄氏度下以 400 G 离心 10 分钟。吸出含有成熟脂肪细胞的顶层和大部分液体层。向其中加入20毫升冷PBS 2%FBS，并重悬沉淀。再次离心五分钟后，通过吸出上层除去上清液以消除剩余的脂肪细胞和脂肪。将沉淀重悬于一毫升ACK裂解缓冲液中。在冰上孵育五分钟。加入10毫升PBS 2%FBS并混合。离心五分钟并吸出上清液后，将沉淀重悬于200微升抗FC溶液中。在冰上孵育五分钟，然后将细胞悬液转移到适合磁性细胞分离器的预冷架的五毫升管中。离心5分钟并除去上清液后，加入200微升CD 31单克隆抗体生物素和CD 45单克隆抗体生物素的混合物。充分混合，在冰上孵育15分钟。然后加入400微升PBS 2%FBS，并在4摄氏度下再次以400G离心5分钟。吸出上清液并将其在100微升抗生物素微珠中在冰上孵育15分钟。加入 400 微升 PBS 2%FBS，并在 400 摄氏度下再次以 400 G 离心五分钟，如前所示。弃去上清液后，将沉淀重悬于350微升PBS 2%FBS中。要使用 70 微米预分离过滤器从单细胞悬浮液中去除细胞聚集体或大颗粒，请用 100 微升 PBS 2%FBS 激活过滤器。将细胞悬液通过过滤器，并将其收集在干净的管中。然后用100微升PBS 2%FBS清洗过滤器。要使用负分离策略对细胞进行磁性分离，请将样品放置在冷藏架的位置A中，将两个空管放在位置B和C处以回收未标记和标记的细胞。然后将洗涤缓冲液和电泳缓冲液装入相应的瓶子中。在分离部分中，选择要分离的样品数量并选择耗尽方案。按运行并开始分离。在程序结束时，恢复未标记的单元格。接下来，通过添加 400 微升 2.5% 基底膜基质覆盖每个孔的整个表面，用基底膜基质涂覆 12 孔板。取出多余的溶液，让板在层流罩内干燥至少一小时。离心五分钟并弃去上清液后，将沉淀重悬于500微升生长培养基中。将脂肪细胞前体细胞或APCs接种在先前用2.5%基底膜基质包被的12孔板的一个孔中。在37%二氧化碳气氛中以37摄氏度孵育，每48小时更换一次培养基，直到细胞达到80%汇合。完全除去培养基后，用350微升PBS 2%FBS洗涤细胞。用350微升0.05%tripsin EDTA在37摄氏度下收获细胞两分钟，并加入两毫升生长培养基。将细胞收集在新的50毫升锥形管中，并以400 G离心五分钟。将APC传代到先前涂有2.5%基底膜基质的12孔板中，并在37摄氏度下用5%二氧化碳孵育。每48小时更换一次培养基，直到细胞达到80%汇合。在80%汇合时，吸出任何生长培养基，并用500微升分化培养基代替，每孔中含有3.3纳摩尔骨形态发生蛋白。48小时后，用每孔500微升分化培养基和分化混合物替换培养基。72小时后取出培养基，并在500微升新鲜分化培养基中加入100纳摩尔胰岛素。48小时后，用每毫升4毫微克的肿瘤坏死因子α或TNF α诱导胰岛素抵抗。吸出任何分化培养基，并用 500 微升含有 TNF α 的简单培养基 2%FBS 代替。孵育24小时后，在简单的培养基0%FBS中加入每毫升TNF α4纳克。24 小时后，通过加入 100 纳摩尔胰岛素激活胰岛素信号通路，并在 37% 二氧化碳气氛中在 57% 的气氛中孵育 15 分钟，如前所述。孵育后，除去培养基并用500微升PBS洗涤，包括没有胰岛素处理的对照孔。提取蛋白质并通过蛋白质印迹测量胰岛素信号标志物的磷酸化。该图显示了在12孔培养板中诱导分化之前皮下脂肪细胞前体细胞在80%汇合，以及诱导分化7天后的原代脂肪细胞。TNF α在皮下原代脂肪细胞中诱导的胰岛素抵抗通过胰岛素诱导的胰岛素受体、胰岛素受体底物1和蛋白激酶B或AKT磷酸化减少来证明。用抗磷酸化胰岛素受体、抗磷酸化胰岛素受体底物一、抗磷酸化AKT探测膜，抗β微管蛋白作为上样对照。通过化学发光检测对信号进行可视化。此处显示了代表性印迹和三个独立实验的定量结果。在尝试此程序时，需要注意的一件事是，TNF α诱导胰岛素抵抗是2%FBS持续24小时，最后24小时使用0%FBS。

Research

JoVE Journal

Biology

Primary Culture and Plasmid Electroporation of the Murine Organ of Corti. 

原代培养及质粒电穿孔尔蒂小鼠器官。

In all mammals, the sensory epithelium for audition is located along the spiraling organ of Corti that resides within the conch shaped cochlea of the ...

科研

生物学

In vivo Imaging and Therapeutic Treatments in an Orthotopic Mouse Model of Ovarian Cancer

In vivo Imaging and Therapeutic Treatments in an Orthotopic Mouse Model of Ovarian Cancer

活体成像和治疗卵巢癌的原位小鼠模型

Human cancer and response to therapy is better represented in orthotopic animal models.  This paper describes the development of an orthotopic mouse model ...

Isolation and Primary Culture of Rat Hepatic Cells

大鼠肝细胞的分离和小学文化

Primary hepatocyte culture is a valuable tool that has been extensively used in basic research of liver function, disease, pathophysiology, pharmacology ...

Isolation and Culture of Mouse Primary Pancreatic Acinar Cells

小鼠原代胰腺腺泡细胞的分离和培养

This protocol permits rapid isolation (in less than 1 hr) of murine pancreatic acini, making it possible to maintain them in culture for more than one ...

A Video Protocol of Retroviral Infection in Primary Intestinal Organoid Culture

逆转录病毒感染的原发性肠化培养的视频协议

Lgr5-positive stem cells can be supplemented with the essential growth factors Egf, Noggin, and R-Spondin, which allows us to culture ever-expanding ...

Isolation and Primary Culture of Mouse Aortic Endothelial Cells

隔离和鼠标主动脉内皮细胞原代培养

The vascular endothelium is essential to normal vascular homeostasis. Its dysfunction participates in various cardiovascular disorders. The mouse is an ...

Validation of a Mouse Model to Disrupt LINC Complexes in a Cell-specific Manner

小鼠模型的验证扰乱LINC配合物在细胞特异性的方式

Nuclear migration and anchorage within developing and adult tissues relies heavily upon large macromolecular protein assemblies called LInkers of the ...

Glucose Uptake Measurement and Response to Insulin Stimulation in In Vitro Cultured Human Primary Myotubes

Glucose Uptake Measurement and Response to Insulin Stimulation in In Vitro Cultured Human Primary Myotubes

葡萄糖摄取测量和对胰岛素刺激的反应<em&gt;体外</em&gt;培养的人类初级肌管

Skeletal muscle is the largest glucose deposit in mammals and largely contributes to glucose homeostasis. Assessment of insulin sensitivity of muscle ...

Isolation and Culture of Primary Mouse Keratinocytes from Neonatal and Adult Mouse Skin

来自新生儿和成年鼠皮肤的原代小鼠角化细胞的分离和培养

The keratinocyte (KC) is the predominant cell type in the epidermis, the outermost layer of the skin. Epidermal KCs play a critical role in providing skin ...

A Tissue Culture Model of Estrogen-producing Primary Bovine Granulosa Cells

雌激素原代牛颗粒细胞的组织培养模型

Ovarian granulosa cells (GC) are the major source of estradiol synthesis. Induced by the preovulatory luteinizing hormone (LH) surge, cells of the theca ...