Name: modeling brain metastasis via tail-vein injection of inflammatory breast cancer cells
Uploaded: 2021-02-04T15:00:00
Description: Watch this Scientific Journal Video about Modeling Brain Metastasis Via Tail-Vein Injection of Inflammatory Breast Cancer Cells Video at JoVE.com

我们感谢MD安德森放射肿瘤学部门的Christine F. Wogan，MS，ELS对手稿进行科学编辑，以及MD安德森外科组织学核心部门的Carol M. Johnston在苏木精和伊红染色方面的帮助。我们感谢MD安德森的兽医和外科核心对动物研究的支持。这项工作得到了以下资助的支持：Susan G. Komen Career Catalyst Research Grant（CCR16377813 to BGD），American Cancer Society Research Scholar Grant（RSG-19-126-01 to BGD）和德克萨斯州罕见和侵袭性乳腺癌研究计划。还得到了美国国立卫生研究院国家癌症研究所向德克萨斯大学MD安德森癌症中心的癌症中心支持（核心）拨款P30 CA016672的部分支持。

此处描述的方法已获得MD安德森癌症中心的机构动物护理和使用委员会（IACUC）的批准，并符合美国国立卫生研究院实验动物护理和使用指南。包括所有步骤在内的原理图工作流程如图 1所示。
1. 细胞制备
注意：在Woodward博士的实验室24中产生的MDA-IBC3（ER-/ PR-/ HER2 +）细胞系用荧光素酶绿色荧?...

<ol>
	<li>Achrol, A. S., 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=Brain+metastases.">Brain metastases.</a> Nature Reviews Disease Primers. 5 (1), 5 (2019).</li><li>Nayak, L., Lee, E. Q., Wen, P. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epidemiology+of+brain+metastases.">Epidemiology of brain metastases.</a> Current Oncology Report. 14 (1), 48-54 (2012).</li><li>Lowery, F. J., Yu, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Brain+metastasis:+Unique+challenges+and+open+opportunities.">Brain metastasis: Unique challenges and open opportunities.</a> Biochimica et Biophysica Acta Review Cancer. 1867 (1), 49-57 (2017).</li><li>Brufsky, A. 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=Central+nervous+system+metastases+in+patients+with+HER2-positive+metastatic+breast+cancer:+incidence,+treatment,+and+survival+in+patients+from+registHER.">Central nervous system metastases in patients with HER2-positive metastatic breast cancer: incidence, treatment, and survival in patients from registHER.</a> Clinical Cancer Research. 17 (14), 4834-4843 (2011).</li><li>Valiente, 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=The+evolving+landscape+of+brain+metastasis.">The evolving landscape of brain metastasis.</a> Trends in Cancer. 4 (3), 176-196 (2018).</li><li>Bos, P. D., 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=Genes+that+mediate+breast+cancer+metastasis+to+the+brain.">Genes that mediate breast cancer metastasis to the brain.</a> Nature. 459 (7249), 1005-1009 (2009).</li><li>Woditschka, S., 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=DNA+double-strand+break+repair+genes+and+oxidative+damage+in+brain+metastasis+of+breast+cancer.">DNA double-strand break repair genes and oxidative damage in brain metastasis of breast cancer.</a> Journal of the National Cancer Institute. 106 (7), (2014).</li><li>Palmieri, D., 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=Vorinostat+inhibits+brain+metastatic+colonization+in+a+model+of+triple-negative+breast+cancer+and+induces+DNA+double-strand+breaks.">Vorinostat inhibits brain metastatic colonization in a model of triple-negative breast cancer and induces DNA double-strand breaks.</a> Clinical Cancer Research. 15 (19), 6148-6157 (2009).</li><li>Kim, S. J., 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=Astrocytes+upregulate+survival+genes+in+tumor+cells+and+induce+protection+from+chemotherapy.">Astrocytes upregulate survival genes in tumor cells and induce protection from chemotherapy.</a> Neoplasia. 13 (3), 286-298 (2011).</li><li>Zhang, S., 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=SRC+family+kinases+as+novel+therapeutic+targets+to+treat+breast+cancer+brain+metastases.">SRC family kinases as novel therapeutic targets to treat breast cancer brain metastases.</a> Cancer Research. 73 (18), 5764-5774 (2013).</li><li>Valiente, 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=Serpins+promote+cancer+cell+survival+and+vascular+co-option+in+brain+metastasis.">Serpins promote cancer cell survival and vascular co-option in brain metastasis.</a> Cell. 156 (5), 1002-1016 (2014).</li><li>Gril, 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=Effect+of+lapatinib+on+the+outgrowth+of+metastatic+breast+cancer+cells+to+the+brain.">Effect of lapatinib on the outgrowth of metastatic breast cancer cells to the brain.</a> Journal of the National Cancer Institute. 100 (15), 1092-1103 (2008).</li><li>Gril, 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=Pazopanib+reveals+a+role+for+tumor+cell+B-Raf+in+the+prevention+of+HER2++breast+cancer+brain+metastasis.">Pazopanib reveals a role for tumor cell B-Raf in the prevention of HER2+ breast cancer brain metastasis.</a> Clinical Cancer Research. 17 (1), 142-153 (2011).</li><li>Palmieri, D., 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=Profound+prevention+of+experimental+brain+metastases+of+breast+cancer+by+temozolomide+in+an+MGMT-dependent+manner.">Profound prevention of experimental brain metastases of breast cancer by temozolomide in an MGMT-dependent manner.</a> Clinical Cancer Research. 20 (10), 2727-2739 (2014).</li><li>Priego, N., 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=STAT3+labels+a+subpopulation+of+reactive+astrocytes+required+for+brain+metastasis.">STAT3 labels a subpopulation of reactive astrocytes required for brain metastasis.</a> Nature Medicine. 24 (7), 1024-1035 (2018).</li><li>Debeb, B. G., 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=miR-141-mediated+regulation+of+brain+metastasis+from+breast+cancer.">miR-141-mediated regulation of brain metastasis from breast cancer.</a> Journal of the National Cancer Institute. 108 (8), (2016).</li><li>Chang, S., Parker, S. L., Pham, T., Buzdar, A. U., Hursting, S. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inflammatory+breast+carcinoma+incidence+and+survival:+the+surveillance,+epidemiology,+and+end+results+program+of+the+National+Cancer+Institute,+1975-1992.">Inflammatory breast carcinoma incidence and survival: the surveillance, epidemiology, and end results program of the National Cancer Institute, 1975-1992.</a> Cancer. 82 (12), 2366-2372 (1998).</li><li>Hance, K. W., Anderson, W. F., Devesa, S. S., Young, H. A., Levine, P. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trends+in+inflammatory+breast+carcinoma+incidence+and+survival:+the+surveillance,+epidemiology,+and+end+results+program+at+the+National+Cancer+Institute.">Trends in inflammatory breast carcinoma incidence and survival: the surveillance, epidemiology, and end results program at the National Cancer Institute.</a> Journal of National Cancer Institute. 97 (13), 966-975 (2005).</li><li>Dirix, L. Y., Van Dam, P., Prove, A., Vermeulen, P. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inflammatory+breast+cancer:+Current+understanding.">Inflammatory breast cancer: Current understanding.</a> Current Opinion in Oncology. 18 (6), 563-571 (2006).</li><li>Wang, Z., 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=Pattern+of+distant+metastases+in+inflammatory+breast+cancer+-+A+large-cohort+retrospective+study.">Pattern of distant metastases in inflammatory breast cancer - A large-cohort retrospective study.</a> Journal of Cancer. 11 (2), 292-300 (2020).</li><li>Uemura, M. I., 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=Development+of+CNS+metastases+and+survival+in+patients+with+inflammatory+breast+cancer.">Development of CNS metastases and survival in patients with inflammatory breast cancer.</a> Cancer. 124 (11), 2299-2305 (2018).</li><li>Smith, D. L., Debeb, B. G., Thames, H. D., Woodward, W. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Computational+modeling+of+micrometastatic+breast+cancer+radiation+dose+response.">Computational modeling of micrometastatic breast cancer radiation dose response.</a> International Journal of Radiation Oncology, Biology, Physics. 96 (1), 179-187 (2016).</li><li>Fukumura, K., 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=Multi-omic+molecular+profiling+reveals+potentially+targetable+abnormalities+shared+across+multiple+histologies+of+brain+metastasis.">Multi-omic molecular profiling reveals potentially targetable abnormalities shared across multiple histologies of brain metastasis.</a> Acta Neuropathol. , (2021).</li><li>Klopp, 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=Mesenchymal+stem+cells+promote+mammosphere+formation+and+decrease+E-cadherin+in+normal+and+malignant+breast+cells.">Mesenchymal stem cells promote mammosphere formation and decrease E-cadherin in normal and malignant breast cells.</a> PLoS One. 5 (8), 12180 (2010).</li><li>Villodre, E. S., 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=Abstract+P3-01-10:+Ndrg1-egfr+axis+in+inflammatory+breast+cancer+tumorigenesis+and+brain+metastasis.">Abstract P3-01-10: Ndrg1-egfr axis in inflammatory breast cancer tumorigenesis and brain metastasis.</a> Cancer Research. 80 (4), 10 (2020).</li></ol>

该协议包括几个关键步骤。细胞应在冰上保持不超过1小时以保持活力。注射前应使用酒精化妆棉擦拭小鼠的尾巴，注意不要擦得太用力或太频繁，以免损坏尾部皮肤。确保细胞悬液中不存在气泡，以防止小鼠死于血管栓塞。将注射角度保持在45°或更小，以避免刺穿尾部血管，并将至少1/3的针头插入尾静脉，以确保所有细胞注射成功。注射细胞的总体积可以根据小鼠的重量进行调整，但细胞总数?...

脑转移是系统性恶性肿瘤的常见和致命并发症。大多数脑转移瘤起源于肺、乳腺或皮肤的原发性肿瘤，合计占病例的67-80%1,2。脑转移发生率的估计在 100,000 至 240,000 例之间，这些数字可能被低估了，因为死于转移性癌症的患者很少进行尸检3.与无脑转移的患者相比，脑转移患者的预后更差，总生存期更低4.目前脑转移瘤的治疗方案主要是姑息性的，无法改善大多数患者的生存结果5。因此，脑转移仍然是一个挑战，并且仍然迫切需要更好地了解脑转移进展的机制，以开发更有效的治疗方法。
实验模型的使用为乳腺癌转移到大脑的特定机制提供了重要的见解，并允许评估各种治疗方法的疗效6,7,8,9,10,11,12,13,14,15,16 .然而，很少有模型能够准确和完整地概括脑转移发展的复杂性。通过不同的给药途径将癌细胞接种到小鼠体内，已经产生了几种实验性体内模型，包括原位、尾静脉、心内、颈内动脉和脑内注射。每种技术都有优点和缺点，详见其他专题3。然而，这些小鼠模型都不能完全复制脑转移的临床进展。
脑转移在炎症性乳腺癌（IBC）患者中特别常见，IBC是原发性乳腺癌的一种罕见但侵袭性的变体。IBC占乳腺癌病例的1%至4%，但它占美国乳腺癌相关死亡人数的10%不成比例17,18。已知 IBC 会迅速转移;事实上，三分之一的IBC患者在诊断时有远处转移19,20。特异性脑转移，IBC 患者的脑转移发生率高于非IBC 21 患者。最近，我们证明了MDA-IBC3细胞系来源于ER-/PR-/HER2 + IBC患者的恶性胸腔积液，概括了小鼠异种移植物中的IBC特征，当通过尾静脉注射时，小鼠发生脑转移而不是肺转移的倾向增强，使该细胞系成为研究脑转移发展的良好模型16。
在这里，我们描述了通过MDA-IBC3细胞的尾静脉注射产生脑转移并通过立体荧光显微镜和荧光素酶成像评估转移负荷的程序。该方法已被用于发现乳腺癌转移到大脑的关键介质，并测试治疗干预的疗效16,22,23。这种技术的缺点是它不能概括大脑转移过程中的所有步骤。然而，它的主要优点包括稳健性和可重复性，涉及内渗的相关转移生物学，穿越肺部和外渗进入大脑，以及其在技术方面的相对简单性。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Cell Culture</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>1000 &#181;L pipette tip filtered</td>
			<td>Genesee Scientific</td>
			<td>23430</td>
			<td></td>
		</tr>
		<tr>
			<td>10 mL Serological Pipets</td>
			<td>Genesee Scientific</td>
			<td>12-112</td>
			<td></td>
		</tr>
		<tr>
			<td>Antibiotic-antimycotic&#160;</td>
			<td>Thermo Fisher Scientific</td>
			<td>15240062</td>
			<td>1%</td>
		</tr>
		<tr>
			<td>Centrifuge tubes 15 mL bulk</td>
			<td>Genesee Scientific</td>
			<td>28103&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>Corning&#160; 500 mL Hams F-12 Medium [+] L-glutamine</td>
			<td>GIBICO Inc. USA</td>
			<td>MT10080CV</td>
			<td></td>
		</tr>
		<tr>
			<td>Countess II Automated Cell Counter (Invitrogen)</td>
			<td>Thermo Fisher Scientific</td>
			<td>AMQAX1000</td>
			<td></td>
		</tr>
		<tr>
			<td>1x DPBS</td>
			<td>Thermo Fisher Scientific</td>
			<td>21-031-CV</td>
			<td></td>
		</tr>
		<tr>
			<td>Eppendorf centufuge 5810R</td>
			<td>Eppendorf&#160;</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Fetal bovine serum (FBS)</td>
			<td>GIBICO Inc. USA</td>
			<td>16000044</td>
			<td>10%</td>
		</tr>
		<tr>
			<td>Fisherbrand &#160;Sterile Cell Strainers (40 &#956;m)</td>
			<td>Thermo Fisher Scientific</td>
			<td>22-363-547</td>
			<td></td>
		</tr>
		<tr>
			<td>Hydrocortisone</td>
			<td>Sigma-Aldrich</td>
			<td>H0888</td>
			<td>1 &#181;g/mL</td>
		</tr>
		<tr>
			<td>Insulin&#160;</td>
			<td>Thermo Fisher Scientific</td>
			<td>12585014</td>
			<td>5 &#181;g/mL</td>
		</tr>
		<tr>
			<td>Invitrogen&#160;Countess Cell Counting Chamber Slides</td>
			<td>Thermo Fisher Scientific</td>
			<td>C10228&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>MDA-IBC3 cell lines</td>
			<td>MD Anderson Cancer Center</td>
			<td></td>
			<td>Generated by Dr. Woodward&#39;s lab24</td>
		</tr>
		<tr>
			<td>Luciferase&#8211;green fluorescent protein (Luc&#8211;GFP) plasmid</td>
			<td>System Biosciences</td>
			<td>BLIV713PA-1</td>
			<td></td>
		</tr>
		<tr>
			<td>microtubes clear sterile 1.7 mL</td>
			<td>Genesee Scientific</td>
			<td>24282S</td>
			<td></td>
		</tr>
		<tr>
			<td>Olympus 10 &#181;L Reach Barrier Tip, Low Binding, Racked, Sterile</td>
			<td>Genesee Scientific</td>
			<td>23-401C&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>TC Treated Flasks (T75), 250mL, Vent</td>
			<td>Genesee Scientific</td>
			<td>25-209</td>
			<td></td>
		</tr>
		<tr>
			<td>Trypan Blue Stain (0.4%) for use with the Countess Automated Cell Counter</td>
			<td>Thermo Fisher Scientific</td>
			<td>T10282</td>
			<td></td>
		</tr>
		<tr>
			<td>Trypsin-EDTA (0.25%), phenol red</td>
			<td>Thermo Fisher Scientific</td>
			<td>25200114</td>
			<td></td>
		</tr>
		<tr>
			<td>Tail vein injection</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>C.B-17/IcrHsd-Prkdc scid Lyst bg-J - SCID/Beige</td>
			<td>Envigo</td>
			<td>SCID/beige mice</td>
			<td></td>
		</tr>
		<tr>
			<td>BD Insulin Syringe with the BD Ultra-Fine Needle 0.5mL 30Gx1/2&#34; (12.7mm)</td>
			<td>BD</td>
			<td>328466</td>
			<td></td>
		</tr>
		<tr>
			<td>Plas Labs &#160;Broome-Style Rodent Restrainers</td>
			<td>Plas Labs 551BSRR</td>
			<td>01-288-32A</td>
			<td>Order fromThermo Fisher Scientific</td>
		</tr>
		<tr>
			<td>Volu SolSupplier Diversity Partner Ethanol 95% SDA (190 Proof)</td>
			<td>Thermo Fisher Scientific</td>
			<td>50420872</td>
			<td>70 % used</td>
		</tr>
		<tr>
			<td>Imaging</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>BD&#160;Lo-Dose&#160; U-100 Insulin Syringes</td>
			<td>BD</td>
			<td>329461</td>
			<td></td>
		</tr>
		<tr>
			<td>Disposable PES Filter Units 0.45 &#181;m</td>
			<td>Fisherbrand</td>
			<td>FB12566501</td>
			<td>filter system to sterilize the D-luciferin</td>
		</tr>
		<tr>
			<td>D-Luciferin</td>
			<td>Biosynth</td>
			<td>L8220-1g</td>
			<td>stock concentration = 47.6 mM (15.15 mg/mL); use concentration = 1.515 mg/mL</td>
		</tr>
		<tr>
			<td>1.7 mL microtube amber</td>
			<td>Genesee Scientific</td>
			<td>24-282AM</td>
			<td></td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>Patterson Veterinary</td>
			<td>NDC-14043-704-06</td>
			<td>Liquid anesthetic for use in anesthetic vaporizer</td>
		</tr>
		<tr>
			<td>IVIS 200&#160;</td>
			<td>PerkinElmer</td>
			<td></td>
			<td>machine for luciferase imaging, up to 5 mice imaging at the same time, with anesthesia machine</td>
		</tr>
		<tr>
			<td>Plastic Containers with Lids&#160;</td>
			<td>Fisherbrand</td>
			<td>02-544-127</td>
			<td></td>
		</tr>
		<tr>
			<td>Tissue Cassettes</td>
			<td>Thermo Scientific</td>
			<td>1000957</td>
			<td></td>
		</tr>
		<tr>
			<td>Webcol Alcohol Prep&#160;</td>
			<td>Covidien</td>
			<td>6818</td>
			<td></td>
		</tr>
		<tr>
			<td>Stereomicroscope Imaging</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Stereomicroscope AZ100&#160;</td>
			<td>Nikon</td>
			<td>model AZ-STGE</td>
			<td>software NIS-ELEMENT</td>
		</tr>
		<tr>
			<td>Formalin 10%</td>
			<td>Fisher Chemical</td>
			<td>SF100-4</td>
			<td></td>
		</tr>
		<tr>
			<td>TC treated dishes 100x20 mm</td>
			<td>Genesee Scientific</td>
			<td>25202</td>
			<td></td>
		</tr>
	</tbody>
</table>

modeling brain metastasis via tail-vein injection of inflammatory breast cancer cells

转移到大脑是许多类型癌症的常见和破坏性表现。仅在美国，每年就有大约20万名患者被诊断出患有脑转移瘤。在改善原发性乳腺癌和系统性恶性肿瘤患者的生存结局方面取得了重大进展;然而，临床脑转移瘤患者的悲惨预后突出了开发针对这种致命疾病的新型治疗药物和策略的迫切需要。缺乏合适的实验模型一直是阻碍我们对脑转移生物学和治疗的理解的主要障碍之一。在本文中，我们描述了一种异种移植小鼠模型，该模型是通过尾静脉注射源自炎症性乳腺癌（IBC）的内源性HER2扩增细胞系产生的脑转移，这是一种罕见且侵袭性的乳腺癌形式。用萤火虫荧光素酶和绿色荧光蛋白标记细胞以监测脑转移，并通过生物发光成像、荧光立体显微镜和组织学评估量化转移负荷。小鼠稳健且一致地发展脑转移，允许研究转移过程中的关键介质并开发新治疗策略的临床前测试。

基于标记细胞有助于监测和可视化临床前小鼠模型中脑转移的基本原理，我们用Luc和GFP标记MDA-IBC3细胞，以监测脑转移并通过使用生物发光成像和荧光立体显微镜量化转移负荷。将标记的MDA-IBC3细胞注射到免疫功能低下的SCID / 米色小鼠的尾静脉中，导致高比例的小鼠发生脑转移（即66.7%至100%）16,23,25。早在注射后8周，通过荧光?...

Watch this Scientific Journal Video about Modeling Brain Metastasis Via Tail-Vein Injection of Inflammatory Breast Cancer Cells Video at JoVE.com

Modeling Brain Metastasis Via Tail-Vein Injection of Inflammatory Breast Cancer Cells Video (Video) | JoVE

我们描述了通过尾静脉注射内源性HER2扩增的炎性乳腺癌细胞系产生的乳腺癌脑转移的异种移植小鼠模型。

通过炎症性乳腺癌细胞的尾静脉注射模拟脑转移

Metastatic spread to the brain is a common and devastating manifestation of many types of cancer. In the United States alone, about 200,000 patients are ...

我们利用尾静脉方法在小鼠模型中使用来自炎症性乳腺癌（IBC，一种高度侵袭性的原发性乳腺癌）患者的细胞系产生脑转移。小鼠模型中脑转移的产生通常涉及心内或颈内动脉注射。然而，尾静脉方法比这些技术更具优势，因为它更好地模仿了脑转移定植的步骤，并且相对容易执行。医生Xiaoding Hu和Emilly Villodre都是我实验室的讲师，他们将演示该程序。为了在肿瘤细胞注射后8至10周通过GFP成像检测脑转移，用70%乙醇消毒小鼠并去除头部和耳朵上的皮毛。切开颈部的颈部区域。切除并切除头骨以收获大脑。将大脑放入组织盒中，并将盒式磁带放入冷DPBS容器中不超过一小时。对于立体显微镜成像，将大脑转移到配备紫外线的立体显微镜下的100厘米纸巾皿盖中，并选择0.5倍物镜以可视化整个大脑。按下实时取景并在大脑处于腹侧位置时聚焦视图。在软件和显微镜中为细胞中的荧光标记物选择合适的滤光片，然后拍照。在不移动样品的情况下，切换到明场成像并选择明场滤光片。拍照。然后如前所述，在背部位置对大脑进行成像。要分析全脑图像，首先将图像文件从TIFF转换为JPEG，以便可以使用任何没有相关软件的计算机打开它们。接下来，打开一对明场和荧光图像并选择文件和合并通道。选择适当的组件，单击“确定”，然后保存合并的图像。要量化荧光图像中的脑肿瘤区域，请选择自动选择。将箭头移动到 GFP 位置区域，然后单击以自动创建选区。然后再次单击以确认选择。将显示所有测量值。如果可以观察到多个GFP正区域，则对后续区域重复测量。脑转移性病变最早可在注射后八周通过荧光素酶成像和立体荧光显微镜检查检测到。成像后，可以对部分脑转移瘤进行福尔马林固定和处理，用于苏木精和伊红染色，以验证是否存在脑转移病变，并进行免疫组织化学染色以检测感兴趣的特定蛋白质标志物。尝试此协议时，重要的是通过将细胞保持在冰上不超过一小时来保持细胞活力，并避免细胞悬液中的气泡以防止栓子形成和小鼠死亡。尾静脉注射后，我们通过荧光素酶成像监测脑转移的生长和进展。安乐死后，立体荧光显微镜检查和组织学检查证实转移病变。

Research

JoVE Journal

Cancer Research

Modeling Brain Metastasis Via Tail-Vein Injection of Inflammatory Breast Cancer Cells

Intra-iliac Artery Injection for Efficient and Selective Modeling of Microscopic Bone Metastasis

Intra-iliac Artery Injection for Efficient and Selective Modeling of Microscopic Bone Metastasis Video (Video) | JoVE

内髂动脉注射显微骨转移的效率和选择性建模

Intra-iliac artery (IIA) injection is an efficient approach to introduce metastatic lesions of various cancer cells in animals. Compared to the widely ...

科研

癌症研究

Murine Experimental Model of Original Tumor Development and Peritoneal Metastasis via Orthotopic Inoculation with Ovarian Carcinoma Cells

Murine Experimental Model of Original Tumor Development and Peritoneal Metastasis via Orthotopic Inoculation with Ovarian Carcinoma Cells Video (Video) | JoVE

原发肿瘤发展小鼠实验模型和腹膜转移<em&gt;通过</em&gt;原位接种卵巢癌细胞

Epithelial ovarian carcinoma (EOC) is associated with a poor prognosis because it shows peritoneal dissemination. To improve the prognosis, it is ...

A Portal Vein Injection Model to Study Liver Metastasis of Breast Cancer

A Portal Vein Injection Model to Study Liver Metastasis of Breast Cancer Video (Video) | JoVE

门静脉注射模型来研究肝转移乳腺癌

Breast cancer is the leading cause of cancer-related mortality in women worldwide. Liver metastasis is involved in upwards of 30% of cases with breast ...

Intracarotid Cancer Cell Injection to Produce Mouse Models of Brain Metastasis

Intracarotid Cancer Cell Injection to Produce Mouse Models of Brain Metastasis Video (Video) | JoVE

颈动脉肿瘤细胞注射，产生脑转移瘤小鼠模型

Metastasis, the spread and growth of malignant cells at secondary sites within a patient&#39;s body, accounts for &#62; 90% of cancer-related mortality. ...

Orthotopic Injection of Breast Cancer Cells into the Mice Mammary Fat Pad

Orthotopic Injection of Breast Cancer Cells into the Mice Mammary Fat Pad Video (Video) | JoVE

乳腺癌细胞原位注射到小鼠乳房脂肪板

A proper animal model is crucial for a better understanding of diseases. Animal models established by different methods (subcutaneous injections, ...

Modeling Breast Cancer via an Intraductal Injection of Cre-expressing Adenovirus into the Mouse Mammary Gland

Modeling Breast Cancer via an Intraductal Injection of Cre-expressing Adenovirus into the Mouse Mammary Gland Video (Video) | JoVE

通过将表达的腺病毒注射到小鼠乳腺腺中,模拟乳腺癌

Breast cancer is a heterogeneous disease, possibly due to complex interactions between different cells of origins and oncogenic events. Mouse models are ...

Micromanipulation of Circulating Tumor Cells for Downstream Molecular Analysis and Metastatic Potential Assessment

Micromanipulation of Circulating Tumor Cells for Downstream Molecular Analysis and Metastatic Potential Assessment Video (Video) | JoVE

循环肿瘤细胞的微操作, 用于下游分子分析和转移电位评估

Blood-borne metastasis accounts for&#160;most cancer-related deaths and involves circulating tumor cells (CTCs) that are successful in establishing new ...

Combined Use of Tail Vein Metastasis Assays and Real-Time In Vivo Imaging to Quantify Breast Cancer Metastatic Colonization and Burden in the Lungs

Combined Use of Tail Vein Metastasis Assays and Real-Time In Vivo Imaging to Quantify Breast Cancer Metastatic Colonization and Burden in the Lungs Video (Video) | JoVE

结合使用尾静脉转移分析与实时 Vivo 成像，以量化乳腺癌转移殖民化和肺负担

Metastasis is the main cause of cancer-related deaths and there are limited therapeutic options for patients with metastatic disease. The identification ...

Pathological Analysis of Lung Metastasis Following Lateral Tail-Vein Injection of Tumor Cells

Pathological Analysis of Lung Metastasis Following Lateral Tail-Vein Injection of Tumor Cells Video (Video) | JoVE

肿瘤细胞外侧尾静脉注射后肺转移的病理分析

Metastasis, the primary cause of morbidity and mortality for most cancer patients, can be challenging to model preclinically in mice. Few spontaneous ...

Modeling Brain Metastases Through Intracranial Injection and Magnetic Resonance Imaging

Modeling Brain Metastases Through Intracranial Injection and Magnetic Resonance Imaging Video (Video) | JoVE

通过颅内注射和磁共振成像对大脑转移进行建模

Metastatic spread of cancer is an unfortunate consequence of disease progression, aggressive cancer subtypes, and/or late diagnosis. Brain metastases are ...