Name: spatial and temporal control of murine melanoma initiation from mutant melanocyte stem cells
Uploaded: 2019-06-07T14:45:00
Description: Watch this Scientific Journal Video about Spatial and Temporal Control of Murine Melanoma Initiation from Mutant Melanocyte Stem Cells at JoVE.com

这项工作得到了主管卫生事务助理国防部长办公室的支持, 该方案通过 W81XWH-1-0272 奖的同行审查癌症研究方案提供了支持。意见、解释、结论和建议是作者的意见、解释、结论和建议, 不一定得到国防部的认可。这项工作也得到了康奈尔大学干细胞计划向 a. c. white 提供的种子赠款的支持。H. Moon 得到了康奈尔大学脊椎动物基因组学研究中心学者计划的支持。

所有动物手术都是根据康奈尔大学动物护理和使用机构委员会 (IACUC) 进行的。
1. 准备工作
<ol><li>
收集12天产后小鼠的尾夹, 在95°c 下在0.05Naoh 的400Μl 中消化 1 h. 涡流, 并加入32μl 的 1 m Rris-hcl, pH 值7。根据杰克逊实验室提供的 PCR 协议, 对有兴趣基因型的小鼠进行基因型鉴定6。 - Tyrer-半形 (+/creer) - Lsl-brafV600e– 杂合 (+/ls...

<ol>
	<li>Wernli, K. J., Henrikson, N. B., Morrison, C. C., Nguyen, M., Pocobelli, G., Blasi, P. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Screening+for+skin+cancer+in+adults:+updated+evidence+report+and+systematic+review+for+the+US+preventive+services+task+force.">Screening for skin cancer in adults: updated evidence report and systematic review for the US preventive services task force.</a> The Journal of the American Medical Association. 316 (4), 436-447 (2016).</li><li>Siegel, R. L., Miller, K. D., Jemal, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cancer+statistics,+2018.">Cancer statistics, 2018.</a> CA: A Cancer Journal for Clinicians. 68 (1), 7-30 (2018).</li><li>White, A. C., Lowry, W. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Refining+the+role+for+adult+stem+cells+as+cancer+cells+of+origin.">Refining the role for adult stem cells as cancer cells of origin.</a> Trends in Cell Biology. 25 (1), 11-20 (2015).</li><li>Moon, H., Donahue, L. R., White, A. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Understanding+cancer+cells+of+origin+in+cutaneous+tumors.">Understanding cancer cells of origin in cutaneous tumors.</a> Cancer Stem Cells. , 263-284 (2016).</li><li>Blanpain, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tracing+the+cellular+origin+of+cancer.">Tracing the cellular origin of cancer.</a> Nature Cell Biology. 15 (2), 126-134 (2013).</li><li>Moon, H., Donahue, L. R., 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=Melanocyte+stem+cell+activation+and+translocation+initiate+cutaneous+melanoma+in+response+to+UV+exposure.">Melanocyte stem cell activation and translocation initiate cutaneous melanoma in response to UV exposure.</a> Cell Stem Cell. 21 (5), 665-678 (2017).</li><li>Moon, H., White, A. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+path+from+melanocyte+stem+cells+to+cutaneous+melanoma+illuminated+by+UVB.">A path from melanocyte stem cells to cutaneous melanoma illuminated by UVB.</a> Molecular &#38; Cellular Oncology. 5 (2), e1409864 (2018).</li><li>Rabbani, P., Takeo, M., 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=Coordinated+activation+of+Wnt+in+epithelial+and+melanocyte+stem+cells+initiates+pigmented+hair+regeneration.">Coordinated activation of Wnt in epithelial and melanocyte stem cells initiates pigmented hair regeneration.</a> Cell. 145 (6), 941-955 (2011).</li><li>Chou, W. C., Takeo, M., 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=Direct+migration+of+follicular+melanocyte+stem+cells+to+the+epidermis+after+wounding+or+UVB+irradiation+is+dependent+on+Mc1r+signaling.">Direct migration of follicular melanocyte stem cells to the epidermis after wounding or UVB irradiation is dependent on Mc1r signaling.</a> Nature Medicine. 19 (7), 924-929 (2013).</li><li>Keyes, B. E., Segal, J. P., 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=Nfatc1+orchestrates+aging+in+hair+follicle+stem+cells.">Nfatc1 orchestrates aging in hair follicle stem cells.</a> Proceedings of the National Academy of Sciences of the United States of America. 110 (51), (2013).</li><li>M&#252;ller-R&#246;ver, S., Handjiski, B., 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=A+comprehensive+guide+for+the+accurate+classification+of+murine+hair+follicles+in+distinct+hair+cycle+stages.">A comprehensive guide for the accurate classification of murine hair follicles in distinct hair cycle stages.</a> The Journal of Investigative Dermatology. 117 (1), 3-15 (2001).</li><li>Kastenmayer, R. J., Fain, M. A., Perdue, K. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+retrospective+study+of+idiopathic+ulcerative+dermatitis+in+mice+with+a+C57BL/6+background.">A retrospective study of idiopathic ulcerative dermatitis in mice with a C57BL/6 background.</a> Journal of the American Association for Laboratory Animal Science&#8239;: JAALAS. 45 (6), 8-12 (2006).</li><li>Vultur, A., Herlyn, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=SnapShot:+melanoma.">SnapShot: melanoma.</a> Cancer Cell. 23 (5), (2013).</li><li>Dankort, D., Curley, D. P., 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=Braf(V600E)+cooperates+with+Pten+loss+to+induce+metastatic+melanoma.">Braf(V600E) cooperates with Pten loss to induce metastatic melanoma.</a> Nature Genetics. 41 (5), 544-552 (2009).</li><li>Viros, A., Sanchez-Laorden, B., 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=Ultraviolet+radiation+accelerates+BRAF-driven+melanomagenesis+by+targeting+TP53.">Ultraviolet radiation accelerates BRAF-driven melanomagenesis by targeting TP53.</a> Nature. 511 (7510), 478-482 (2014).</li></ol>

在皮肤黑色素瘤肿瘤中经常发现的 hallmark 基因改变已经被很好地描述了13。最主要的驱动突变是BrafV600e,和基因工程小鼠模型的 braf v600e 介导黑色素瘤是由博森伯格组14产生, 并沉积在杰克逊实验室. 利用这个小鼠模型, 我们最近的研究证明了细胞激活的要求, 肿瘤易发的 Mcsc 显著启动braf V600e介导的黑色素瘤在背皮肤<sup...

黑色素瘤是黑色素瘤的恶性形式, 是皮肤中最具攻击性的癌症, 皮肤黑色素瘤占皮肤癌死亡的大多数1。在美国, 黑色素瘤通常被诊断出来;据预测, 在2018年估计的新癌症病例中, 它将是第5至第6种最常见的癌症类型2。此外, 虽然近几十年来癌症的总体发病率呈逐渐下降的趋势, 但在过去几十年中, 皮肤黑色素瘤发病率显示,2 男女的皮肤黑色素瘤发病率都在持续和迅速增加。
为了更有效地对抗皮肤黑色素瘤, 重要的是要清楚地了解黑色素瘤发展的早期事件, 从他们的起源细胞, 以更好地确定临床上有效的预防方法。现在人们知道, 在包括皮肤组织 3、4、5在内的许多不同类型的器官中, 成人干细胞作为癌症的细胞起源,可以显著促进肿瘤的形成。同样, 在小鼠背皮的成人黑细胞干细胞 (Mcsc) 可以作为黑色素瘤细胞的起源时, 异常激活拉斯拉夫和阿克特途径6。然而, 仅这些致癌突变并不能有效地诱导静止的 Mcsc 形成肿瘤, 当 Mcsc 在自然头发循环过程中变得活跃时, 黑色素瘤最终会发展起来。然而, 在这个协议中, 我们将描述的方法, 人为地诱导细胞激活易发生肿瘤的 mcsc, 从而促进黑色素瘤的启动精确控制 6,7。
通过这里描述的方法, 空间和时间控制的皮肤黑色素瘤开始从肿瘤易发的 Mcsc表达致癌 BrafV600e 连同 pten表达的损失, 可以成功地执行, 因为我们此前曾有报道。这种方法结合了先前的研究结果, 表明毛囊干细胞通过脱毛激活, 以及如何小鼠皮肤暴露于 UV-B 可以促进激活 Mcsc 居民的毛囊和这个细胞的移位亚群到卵泡间表皮 6,8,9,10。这些体内模型系统可以提供有关生理和环境变化如何改变易患黑色素瘤的 Mcsc 的细胞状态的宝贵信息, 进而导致皮肤中黑色素瘤的显著启动。

<table>
	<tbody>
		<tr>
			<td>Tamoxifen</td>
			<td>Sigma</td>
			<td>T5648-1G</td>
			<td>For systemic injection</td>
		</tr>
		<tr>
			<td>Tamoxifen</td>
			<td>Cayman Chemical</td>
			<td>13258</td>
			<td>For systemic injection</td>
		</tr>
		<tr>
			<td>Corn oil</td>
			<td>Sigma</td>
			<td>45-C8267-2.5L-EA</td>
			<td></td>
		</tr>
		<tr>
			<td>4OH-tamoxifen</td>
			<td>Sigma</td>
			<td>H7904-25MG</td>
			<td>For topical treatment</td>
		</tr>
		<tr>
			<td>26g 1/2&#34; needles</td>
			<td>various</td>
			<td></td>
			<td>Veterinary grade</td>
		</tr>
		<tr>
			<td>1 mL syringe</td>
			<td>various</td>
			<td></td>
			<td>Veterinary grade</td>
		</tr>
		<tr>
			<td>Pet hair trimmer</td>
			<td>Wahl</td>
			<td>09990-502</td>
			<td></td>
		</tr>
		<tr>
			<td>Hair removal cream</td>
			<td>Nair</td>
			<td>n/a</td>
			<td>Available at most drug stores</td>
		</tr>
		<tr>
			<td>Cotton swabs</td>
			<td>various</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Ultraviolet light bulb</td>
			<td>UVP</td>
			<td>95-0042-08</td>
			<td>model XX-15M midrange UV lamp</td>
		</tr>
		<tr>
			<td>200 proof ethanol</td>
			<td>various</td>
			<td></td>
			<td>pure ethanol</td>
		</tr>
		<tr>
			<td>Histoplast PE</td>
			<td>Fisher Scientific</td>
			<td>22900700</td>
			<td>paraffin pellets</td>
		</tr>
		<tr>
			<td>Neutral Buffered Formalin, 10%</td>
			<td>Sigma</td>
			<td>HT501128-4L</td>
			<td></td>
		</tr>
		<tr>
			<td>Clear-Rite 3</td>
			<td>Thermo Scientific</td>
			<td>6901</td>
			<td>xylene substitute</td>
		</tr>
		<tr>
			<td>O.C.T. Compound</td>
			<td>Thermo</td>
			<td>23730571</td>
			<td></td>
		</tr>
		<tr>
			<td>Tissue Cassette</td>
			<td>Sakura</td>
			<td>89199-430</td>
			<td>for FFPE processing</td>
		</tr>
		<tr>
			<td>Cryomolds</td>
			<td>Sakura</td>
			<td>4557</td>
			<td>25 x 20 mm</td>
		</tr>
		<tr>
			<td>FFPE metal mold</td>
			<td>Leica</td>
			<td>3803082</td>
			<td>24 x 24 mm</td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>various</td>
			<td></td>
			<td>Veterinary grade</td>
		</tr>
		<tr>
			<td>Anesthesia inhalation system</td>
			<td>various</td>
			<td></td>
			<td>Veterinary grade</td>
		</tr>
		<tr>
			<td>Fine scissor</td>
			<td>FST</td>
			<td>14085-09</td>
			<td>Straight, sharp/sharp</td>
		</tr>
		<tr>
			<td>Fine scissor</td>
			<td>FST</td>
			<td>14558-09</td>
			<td>Straight, sharp/sharp</td>
		</tr>
		<tr>
			<td>Metzenbaum</td>
			<td>FST</td>
			<td>14018-13</td>
			<td>Straight, blunt/blunt</td>
		</tr>
		<tr>
			<td>Forcep</td>
			<td>FST</td>
			<td>11252-00</td>
			<td>Dumont #5</td>
		</tr>
		<tr>
			<td>Forcep</td>
			<td>FST</td>
			<td>11018-12</td>
			<td>Micro-Adson</td>
		</tr>
		<tr>
			<td>Tyr-CreER; LSL-BrafV600E; Pten-f/f</td>
			<td>Jackson Labs</td>
			<td>13590</td>
			<td></td>
		</tr>
		<tr>
			<td>LSL-tdTomato</td>
			<td>Jackson Labs</td>
			<td>007914</td>
			<td>ai14</td>
		</tr>
		<tr>
			<td>Cre-1</td>
			<td></td>
			<td>n/a</td>
			<td>GCATTACCGGTCGATGCAACGAGTGATGAG</td>
		</tr>
		<tr>
			<td>Cre-2</td>
			<td></td>
			<td>n/a</td>
			<td>GAGTGAACGAACCTGGTCGAAATCAGTGCG</td>
		</tr>
		<tr>
			<td>Braf-V600E-1</td>
			<td></td>
			<td>n/a</td>
			<td>TGAGTATTTTTGTGGCAACTGC</td>
		</tr>
		<tr>
			<td>Braf-V600E-2</td>
			<td></td>
			<td>n/a</td>
			<td>CTCTGCTGGGAAAGCGGC</td>
		</tr>
		<tr>
			<td>Kras-G12D-1</td>
			<td></td>
			<td>n/a</td>
			<td>AGCTAGCCACCATGGCTTGAGTAAGTCTGCA</td>
		</tr>
		<tr>
			<td>Kras-G12D-2</td>
			<td></td>
			<td>n/a</td>
			<td>CCTTTACAAGCGCACGCAGACTGTAGA</td>
		</tr>
		<tr>
			<td>Pten-1</td>
			<td></td>
			<td>n/a</td>
			<td>ACTCAAGGCAGGGATGAGC</td>
		</tr>
		<tr>
			<td>Pten-2</td>
			<td></td>
			<td>n/a</td>
			<td>AATCTAGGGCCTCTTGTGCC</td>
		</tr>
		<tr>
			<td>Pten-3</td>
			<td></td>
			<td>n/a</td>
			<td>GCTTGATATCGAATTCCTGCAGC</td>
		</tr>
		<tr>
			<td>tdTomato-1</td>
			<td></td>
			<td>n/a</td>
			<td>AAGGGAGCTGCAGTGGAGTA</td>
		</tr>
		<tr>
			<td>tdTomato-2</td>
			<td></td>
			<td>n/a</td>
			<td>CCGAAAATCTGTGGGAAGTC</td>
		</tr>
		<tr>
			<td>tdTomato-3</td>
			<td></td>
			<td>n/a</td>
			<td>GGCATTAAAGCAGCGTATCC</td>
		</tr>
		<tr>
			<td>tdTomato-4</td>
			<td></td>
			<td>n/a</td>
			<td>CTGTTCCTGTACGGCATGG</td>
		</tr>
	</tbody>
</table>

spatial and temporal control of murine melanoma initiation from mutant melanocyte stem cells

皮肤黑色素瘤是众所周知的最具攻击性的皮肤癌。虽然风险因素和主要基因改变继续被记录以越来越深的深度, 但近几十年来, 皮肤黑色素瘤的发病率迅速而持续地增加。为了找到有效的预防方法, 重要的是要了解黑色素瘤在皮肤中开始的早期步骤。先前的数据表明, 在成人皮肤组织中的滤泡黑素细胞干细胞 (Mcsc) 可以作为黑色素瘤细胞的起源, 当表达致癌突变和基因改变。当 Mcsc 从静止状态过渡到活动状态时, 可诱发黑色素瘤易发性 Mcsc 引起的肿瘤发生。这种易黑色素瘤的 Mcsc 的转变可以通过调节毛囊干细胞的活性状态或通过外源环境因素, 如紫外线-b (UV-B) 来促进。这些因素可以在实验室中通过化学脱毛进行人工操作, 从而导致毛囊干细胞和 Mcsc 从静止状态过渡到活动状态, 并通过使用台式光进行 UV-B 照射。这些方法为小鼠背侧皮肤皮肤黑色素瘤的开始提供了成功的空间和时间控制。因此, 这些体内模型系统将有助于确定皮肤黑色素瘤萌发的早期步骤, 并可用于测试肿瘤预防的潜在方法。

化学脱毛引起的皮肤黑色素瘤
化学脱毛的过程如图 2所示。当老鼠出生后7周时, 它们的背侧皮肤就会出现端粒。在端粒期间, 毛囊干细胞和 Mcsc 已知处于静止、静止状态。剃须后, 皮肤不应显示出明显的头发生长。另一方面, 化学脱毛可以诱导毛囊干细胞激活, 进而显示出感兴趣区域的显著头?...

Watch this Scientific Journal Video about Spatial and Temporal Control of Murine Melanoma Initiation from Mutant Melanocyte Stem Cells at JoVE.com

Spatial and Temporal Control of Murine Melanoma Initiation from Mutant Melanocyte Stem Cells

突变黑色素细胞干细胞引发小鼠黑色素瘤的时空控制

下面的过程描述了一种方法, 空间和时间控制黑素细胞肿瘤开始在小鼠背皮肤, 使用基因工程小鼠模型。该协议描述宏观以及微观皮肤黑色素瘤的启动。

Cutaneous melanoma is well known as the most aggressive skin cancer. Although the risk factors and major genetic alterations continue to be documented ...

黑色素瘤在全世界被广泛诊断，发病率不断增加。这些协议允许仔细控制黑色素瘤开始的早期阶段，促进研究其发展。通过控制黑色素细胞干细胞活化的时间和位置，我们有能力辨别干细胞种群内部的变化及其新菌株黑色素瘤的启动。演示程序将是大喜姆金和卢叶安，研究生从实验室。在开始手术前两到三天，使用电动修剪器从每只麻醉的七周大小鼠的后侧皮肤上剃光。在实验当天，使用棉签在裸露的皮肤上涂抹一层薄薄的脱毛霜。均匀涂面霜后，使用干净的棉签去除奶油，然后用湿布轻轻擦拭，去除残留的奶油。然后将每只鼠标返回到其主笼，并进行监控，直到完全恢复。对于 UV-B 辐照，用 UV-B 保护材料覆盖后层皮肤，以便仅暴露所需的区域，并用防紫外线盖覆盖 UV 室。然后打开 UV-B 灯，在适当的实验时间内照射鼠标，然后将鼠标返回到空笼，并进行监控，直到完全恢复。要分离皮肤样本，请使用电动修剪器从安乐死辐照小鼠的后皮区域去除任何头发，并使用无绒组织轻轻刷刷裸露的皮肤以清除该区域。用锋利的剪刀在尾巴的上方做一个小切口，并将大钝剪刀插入切口，以分离次皮结缔组织。使用锋利的剪刀，小心地沿着感兴趣的皮肤区域的边缘切割，以分离组织样本，并将切除的皮肤组织放在干净的纸巾上。然后用沉闷的钳子伸展皮肤，使皮肤粘附在毛巾上，并绷紧。所有样品都收获后，将皮肤样品和纸巾背放在紧挨着折痕的一块折叠滤纸上，注意样品光滑，不要折叠或皱褶。用订书针合上滤纸的两侧，然后修剪纸张。然后在室温下将样品完全淹没在10%中性缓冲的甲醛中，在室温下洗涤组织3至5小时，然后用两次5分钟的洗涤在去电化水中清洗。第二次洗涤后，小心地从皮肤组织中去除任何残留物质，并修剪样品的边缘，使它们不会锯齿状。接下来，在每个样品中进行三次下垂切口，使条带的宽度不超过 5 毫米，并进行横向切口，使样本宽度不超过 20 毫米。将含有最佳切割温度或 OCT 介质的低温模具放在纵向，并在 OCT 顶部放置四到五块皮肤。当所有碎片都到位时，使用两对细钳将每块皮肤的长边带到低温模具的底部，使样品垂直于 OCT 内，并垂直于模具底部。然后将模具填充到第二唇的附加 OCT，然后将模具放在一块干冰的平坦表面上，使用钳子调整在转移过程中可能转移的任何皮肤片，因为块开始根据需要冻结。当小鼠在产后七周时，它们的后皮是泰源。在泰洛根期间，毛囊和黑色素细胞干细胞已知处于静止休息状态，剃须后皮肤应无显著毛长。然而，化学脱毛可以诱导毛囊干细胞活化，在感兴趣的区域内诱导显著的头发生长。由于化学脱皮可以显著诱导毛囊和黑色素细胞干细胞的激活，黑色素瘤易发性黑色素细胞干细胞在活动状态可以形成肿瘤和显微黑色素瘤启动可以在化学脱皮后两周内观察到。UV-B辐照也可以诱导肿瘤从静态，肿瘤易发性黑色素细胞干细胞启动。重要的是，UV-B诱导黑色素细胞干细胞从卵泡干细胞利基直接移植到卵泡表皮间。此外，UV-B可在整个室间表皮中诱导黑色素细胞干细胞原源性皮肤黑色素瘤的形成。与后皮类似，UV-B 诱导黑色素瘤的起始可以在其他皮肤区域（如耳皮）明显观察到。事实上，与控制耳皮相比，在抗紫外线布覆盖下，暴露于UV-B的耳皮由于黑色素瘤的启动负担较高，表现出更高的色素沉着。不同剂量的UV-B对小鼠皮肤和黑色素细胞活性可能有不同的影响。因此，在开始实验之前确定和优化辐照剂量非常重要。

Research

JoVE Journal

Cancer Research

A Mouse Model of Fatigue Induced by Peripheral Irradiation

疲劳的小鼠模型的外围照射引起

Cancer-related fatigue (CRF) is a distressing and costly condition that often affects patients receiving cancer treatments, including radiation therapy. ...

科研

癌症研究

Harnessing the DNA Dye-triggered Side Population Phenotype to Detect and Purify Cancer Stem Cells from Biological Samples

利用DNA染色体引发的侧群体表型，从生物样品中检测和纯化癌细胞

Cancer is a stem cell-driven disease and eradication of these cells has become a major therapeutic goal. Deciphering vulnerabilities of Cancer Stem Cells ...

Using CRISPR/Cas9 Gene Editing to Investigate the Oncogenic Activity of Mutant Calreticulin in Cytokine Dependent Hematopoietic Cells

利用 CRISPR/Cas9 基因编辑研究细胞因子依赖性造血细胞中突变钙的致癌活性

Clustered regularly interspaced short palindromic repeats (CRISPR) is an adaptive immunity system in prokaryotes that has been repurposed by scientists to ...

Investigation of Genetic Dependencies Using CRISPR-Cas9-based Competition Assays

基于 crispr-cas9 的竞争检测对遗传依赖性的调查

Gene perturbation studies have been extensively used to investigate the role of individual genes in AML pathogenesis. For achieving complete gene ...

Mesenchymal Stem Cell Isolation from Pulp Tissue and Co-Culture with Cancer Cells to Study Their Interactions

骨髓间充质干细胞在浆组织中的分离及与癌细胞联合培养研究其相互作用

Cancer as a multistep process and complicated disease is not only regulated by individual cell proliferation and growth but also controlled by tumor ...

Rapid Generation of Primary Murine Melanocyte and Fibroblast Cultures

初级毛质素黑色素细胞和纤维细胞培养的快速生成

Defects in fibroblast or melanocyte function are associated with skin diseases, including poor barrier function, defective wound healing, pigmentation ...

Flow Cytometric Analysis of Mitochondrial Reactive Oxygen Species in Murine Hematopoietic Stem and Progenitor Cells and MLL-AF9 Driven Leukemia

木尿造血干细胞和祖细胞和MLL-AF9驱动白血病线粒体反应氧物种的流细胞学分析

We present a flow cytometric approach for analyzing mitochondrial ROS in various live bone marrow (BM)-derived stem and progenitor cell populations from ...

Isolation of Exosome-Enriched Extracellular Vesicles Carrying Granulocyte-Macrophage Colony-Stimulating Factor from Embryonic Stem Cells

分离外体富集的细胞外囊泡携带颗粒细胞-巨噬细胞-胚胎干细胞的细胞外刺激因子

Embryonic stem cells (ESCs) are pluripotent stem cells capable of self-renewal and differentiation into all types of embryonic cells. Like many other cell ...

Isolation of Stem-like Cells from 3-Dimensional Spheroid Cultures

从3维球体培养中分离出类似干细胞的细胞

Despite advances in adult stem cell research, identification and isolation of stem cells from tissue specimens remains a major challenge. While resident ...

Preparation of Mitochondria from Ovarian Cancer Tissues and Control Ovarian Tissues for Quantitative Proteomics Analysis

卵巢癌组织线粒体的制备与控制卵巢组织进行定量蛋白质组学分析

Ovarian cancer is a common gynecologic cancer with high mortality but unclear molecular mechanism. Most ovarian cancers are diagnosed in the advanced ...