Name: magnetic resonance-guided high intensity focused ultrasound generated hyperthermia: a feasible treatment method in a murine rhabdomyosarcoma model
Uploaded: 2023-01-13T14:10:00
Description: Watch this Scientific Journal Video about Magnetic Resonance-Guided High Intensity Focused Ultrasound Generated Hyperthermia: A Feasible Treatment Method in a Murine Rhabdomyosarcoma Model at JoVE.com

我们要感谢该项目的资金来源和参与人员，包括：C17研究补助金，加拿大研究生奖学金，安大略省学生机会信托基金和詹姆斯J.哈蒙德基金。

研究是在表型基因组学中心（TCP）和大学健康网络（UHN）动物资源中心（ARC）动物研究设施的监督兽医的监督兽医的监督下按照动物护理委员会批准的动物使用方案进行的。所有涉及动物的程序，不包括MRgHIFU，都是在生物安全柜（BSC）中进行的，以尽量减少动物暴露于外部空气或易感染。
1.小鼠育种
注意：共有65只小鼠（菌株B6.129S2-Trp53tm1Tyj /...

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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=Human+cancer+growth+and+therapy+in+immunodeficient+mouse+models.">Human cancer growth and therapy in immunodeficient mouse models.</a> Cold Spring Harbor Protocols. 2014 (7), 694-708 (2014).</li><li>De Vita, A., 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=Deciphering+the+genomic+landscape+and+pharmacological+profile+of+uncommon+entities+of+adult+rhabdomyosarcomas.">Deciphering the genomic landscape and pharmacological profile of uncommon entities of adult rhabdomyosarcomas.</a> International Journal of Molecular Sciences. 22 (21), 11564 (2021).</li><li>McKinnon, T., 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=Functional+screening+of+FGFR4-driven+tumorigenesis+identifies+PI3K/mTOR+inhibition+as+a+therapeutic+strategy+in+rhabdomyosarcoma.">Functional screening of FGFR4-driven tumorigenesis identifies PI3K/mTOR inhibition as a therapeutic strategy in rhabdomyosarcoma.</a> Oncogene. 37 (20), 2630-2644 (2018).</li><li>Zaporzan, 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=MatMRI+and+MatHIFU:+software+toolboxes+for+real-time+monitoring+and+control+of+MR-guided+HIFU.">MatMRI and MatHIFU: software toolboxes for real-time monitoring and control of MR-guided HIFU.</a> Journal of Therapeutic Ultrasound. 1, (2013).</li><li>Dunne, 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=Heat-activated+drug+delivery+increases+tumor+accumulation+of+synergistic+chemotherapies.">Heat-activated drug delivery increases tumor accumulation of synergistic chemotherapies.</a> Journal of Controlled Release. 308, 197-208 (2019).</li><li>Zhao, Y. 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本文开发的方案用于使用MRgHIFU靶向后肢肿瘤进行轻度激素疗法治疗，并从体内脂质体中释放包封的药物 。 在试点研究期间，该协议中遇到了几个关键步骤，优化这些关键步骤可以提高试点研究的治疗成功率。首先是完全去除要超声处理的区域上的毛发。皮毛内的任何气体捕获都会阻止超声波束通过并阻止超声波进入目标组织1.其次，小鼠定位对于成功治疗至关重要;肿瘤?...

An erratum was issued for:&#160;<a href="https://www.jove.com/t/64544">Magnetic Resonance-Guided High Intensity Focused Ultrasound Generated Hyperthermia: A Feasible Treatment Method in a Murine Rhabdomyosarcoma Model </a>. The Authors section was updated from:
Claire Wunker1,2 
Karolina Piorkowska3 
Ben Keunen3 
Yael Babichev2 
Suzanne M. Wong3,4 
Maximilian Regenold5 
Michael Dunne5 
Julia Nomikos1,2 
Maryam Siddiqui6 
Samuel Pichardo6 
Warren Foltz7 
Adam C. Waspe3,8 
Justin T. Gerstle3,9 
Rebecca A. Gladdy1,2,10 
1 Institute of Medical Science, University of Toronto 
2 2Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital 
3 The Wilfred and Joyce Posluns Centre for Image-Guided Innovation and Therapeutic Intervention, The Hospital for Sick Children 
4 Institute of Biomedical Engineering, University of Toronto 
5 Leslie Dan Faculty of Pharmacy, University of Toronto 
6 Departments of Radiology and Clinical Neurosciences, University of Calgary 
7 Department of Radiation Oncology, University of Toronto 
8 Department of Medical Imaging, University of Toronto 
9 Department of Pediatric Surgery, University of Toronto 
10 Department of Surgery, University of Toronto
to:
Claire Wunker1,2 
Karolina Piorkowska3 
Ben Keunen3 
Yael Babichev2 
Suzanne M. Wong3,4 
Maximilian Regenold5 
Michael Dunne5 
Julia Nomikos1,2 
Maryam Siddiqui6 
Samuel Pichardo6 
Warren Foltz7 
Adam C. Waspe3,8 
Justin T. Gerstle3,9 
James M. Drake1,3,4,10 
Rebecca A. Gladdy1,2,10 
1 Institute of Medical Science, University of Toronto 
2 Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital 
3 The Wilfred and Joyce Posluns Centre for Image-Guided Innovation and Therapeutic Intervention, The Hospital for Sick Children 
4 Institute of Biomedical Engineering, University of Toronto 
5 Leslie Dan Faculty of Pharmacy, University of Toronto 
6 Departments of Radiology and Clinical Neurosciences, University of Calgary 
7 Department of Radiation Oncology, University of Toronto 
8 Department of Medical Imaging, University of Toronto 
9 Department of Pediatric Surgery, University of Toronto 
10 Department of Surgery, University of Toronto

An erratum was issued for:&#160;<a href="https://www.jove.com/t/64544">Magnetic Resonance-Guided High Intensity Focused Ultrasound Generated Hyperthermia: A Feasible Treatment Method in a Murine Rhabdomyosarcoma Model
</a>. The Authors section was updated.

Erratum: Magnetic Resonance-Guided High Intensity Focused Ultrasound Generated Hyperthermia: A Feasible Treatment Method in a Murine Rhabdomyosarcoma Model

横纹肌肉瘤 （RMS） 是一种骨骼肌肿瘤，最常见于儿童和年轻人1.局部疾病通常采用多模式治疗，包括化疗、电离放疗和手术。多药化疗方案的使用在儿科患者中更为普遍，与成人患者相比，结局有所改善2;然而，尽管正在进行研究努力，在最具侵略性的疾病中，5 年生存率仍保持在 30% 左右3，4。化疗标准治疗是一种多药方案，包括长春新碱、环磷酰胺和放线菌素 D。在疾病复发或复发的情况下，使用替代化疗，包括标准（游离）多柔比星 （FD） 和异环磷酰胺1。虽然所有这些化疗都具有全身毒性，但阿霉素的心脏毒性施加了终生剂量限制5-7。为了增加递送到肿瘤的药物量并尽量减少全身毒性，已经开发了替代制剂，包括脂质体包封。这些可以是非热敏性阿霉素，已被批准用于治疗乳腺癌和肝细胞癌，也可以是热敏性阿霉素，其临床试验正在进行中8，9，10，11，12，13。递送脂质体包封药物的替代方法，例如多泡脂质体和配体靶向脂质体已经得到评估，并显示出治疗肿瘤的希望9。在这项研究中，热量的增加具有多因素影响，包括药物释放14。热疗（HT）与磁共振引导的高强度聚焦超声（MRgHIFU）和热敏脂质体阿霉素（TLD）的组合是一种新颖的多模式治疗方法，用于使用这种有毒但有效的药物治疗RMS，同时最大限度地减少剂量限制毒性并可能增加对肿瘤的免疫反应。
阿霉素在>39°C的温度下从TLD迅速释放，远高于人体平均体温37°C，但不足以引起组织损伤或消融;这在43°C时开始发生，但随着温度接近60°C15，发生得更快。已经使用了多种方法在体内产生HT，包括激光，微波，射频消融和聚焦超声，其中许多是侵入性加热方法16。MRgHIFU是一种无创、非电离的加热方法，有助于在原位目标组织内精确设置温度。磁共振（MR）成像至关重要地提供了实时成像，其中可以使用计算机软件来计算整个治疗过程中组织的测温测量值;随后，该数据可用于实时控制超声治疗以达到并保持所需的温度设定点17。MRgHIFU已在各种组织类型中进行了测试，可用于从轻度激素疗法到消融术的各种温度治疗，以及临床上成功治疗疼痛的骨转移18。此外，HT已被证明可引起肿瘤细胞毒性，调节蛋白表达并改变肿瘤微环境中的免疫反应19，20，21，22。一项研究将轻度激素疗法与TLD相结合，然后用MRgHIFU消融，在协同R1大鼠模型23中，导致肿瘤核心坏死和药物输送到外围。传统上，放疗被用作辅助疗法，以破坏肿瘤细胞并减少局部疾病复发。然而，它的使用受到终生剂量和脱靶损伤的限制1.因此，HT的独特之处在于它可以引起一些相同的效果而没有相同的毒性或限制。
RMS的临床前动物模型包括免疫功能低下宿主中的同源免疫功能模型和患者来源的异种移植物（PDX）。虽然免疫功能低下的模型允许人类肿瘤的生长，但它们缺乏适当的肿瘤微环境，并且研究免疫反应的能力有限24。FGFR4 激活突变是预后不良的有希望的标志物，也是成人和儿童 RMS1，25 的潜在治疗靶点。在Gladdy实验室开发的同源RMS模型中，肿瘤能够在免疫功能正常的宿主中生长，该宿主对肿瘤产生先天性和适应性免疫反应26。由于HT影响免疫应答，观察小鼠免疫应答的变化是该肿瘤模型的宝贵优势。为了测试肿瘤对TLD的反应与FD相比，以及肿瘤对化疗和激素疗法的免疫反应的变化，开发并采用一种方案使用MRgHIFU和TLD治疗 体内 同系鼠RMS肿瘤，这是本研究的重点。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>1.5mL Eppendorf tubes</td>
			<td>Eppendorf</td>
			<td>22363204</td>
			<td></td>
		</tr>
		<tr>
			<td>1kb plus DNA Ladder</td>
			<td>Froggabio</td>
			<td>DM015-R500</td>
			<td></td>
		</tr>
		<tr>
			<td>2x HS-Red Taq (PCR mix)</td>
			<td>Wisent</td>
			<td>801-200-MM</td>
			<td></td>
		</tr>
		<tr>
			<td>7 Tesla MRI BioSpec</td>
			<td>Bruker</td>
			<td>T184931</td>
			<td>70/30 BioSpec, Bruker, Ettlingen, Germany</td>
		</tr>
		<tr>
			<td>C1000 Thermal cycler</td>
			<td>Biorad</td>
			<td>1851148</td>
			<td></td>
		</tr>
		<tr>
			<td>Clippers</td>
			<td>Whal Peanut</td>
			<td>8655</td>
			<td></td>
		</tr>
		<tr>
			<td>Compressed ultrasound gel</td>
			<td>Aquaflex</td>
			<td>HF54-004</td>
			<td></td>
		</tr>
		<tr>
			<td>Convection heating device</td>
			<td>3M Bair Hugger</td>
			<td>70200791401</td>
			<td></td>
		</tr>
		<tr>
			<td>Depiliatory cream</td>
			<td>Nair</td>
			<td>61700222611</td>
			<td>Shopper&#39;s Drug Mart</td>
		</tr>
		<tr>
			<td>DMEM</td>
			<td>Wisent</td>
			<td>219-065-LK</td>
			<td></td>
		</tr>
		<tr>
			<td>DNeasy extraction kit</td>
			<td>Qiagen&#160;</td>
			<td>69504</td>
			<td></td>
		</tr>
		<tr>
			<td>DPBS</td>
			<td>Wisent</td>
			<td>311-420-CL</td>
			<td></td>
		</tr>
		<tr>
			<td>Drug injection system</td>
			<td>Harvard Apparatus</td>
			<td>PY2 70-2131</td>
			<td>PHD 22/2200 MRI compatible Syringe Pump</td>
		</tr>
		<tr>
			<td>Eye lubricant</td>
			<td>Optixcare</td>
			<td>50-218-8442</td>
			<td></td>
		</tr>
		<tr>
			<td>F10 Media</td>
			<td>Wisent</td>
			<td>318-050-CL</td>
			<td></td>
		</tr>
		<tr>
			<td>FBS</td>
			<td>Wisent</td>
			<td>081-105</td>
			<td></td>
		</tr>
		<tr>
			<td>Froggarose</td>
			<td>FroggaBio</td>
			<td>A87</td>
			<td></td>
		</tr>
		<tr>
			<td>Gel Molecular Imager</td>
			<td>BioRad</td>
			<td>GelDocXR</td>
			<td></td>
		</tr>
		<tr>
			<td>Glutamax</td>
			<td>Wisent</td>
			<td>609-065-EL</td>
			<td></td>
		</tr>
		<tr>
			<td>Heat Lamp</td>
			<td>Morganville Scientific</td>
			<td>HL0100&#160;</td>
			<td>Similar to this product</td>
		</tr>
		<tr>
			<td>Intravascular Polyethylene tubing (0.015&#34; ID x 0.043&#34; OD, 20G)</td>
			<td>SAI infusion</td>
			<td>PE-20-100</td>
			<td></td>
		</tr>
		<tr>
			<td>Isoflurane</td>
			<td>Sigma</td>
			<td>792632</td>
			<td></td>
		</tr>
		<tr>
			<td>M25FV24C Cell line</td>
			<td>Gladdy Lab</td>
			<td>N/A</td>
			<td></td>
		</tr>
		<tr>
			<td>Microliter Syringe</td>
			<td>Hamilton</td>
			<td>01-01-7648</td>
			<td></td>
		</tr>
		<tr>
			<td>Molecular Imager Gel Doc XR</td>
			<td>Biorad</td>
			<td>170-8170</td>
			<td></td>
		</tr>
		<tr>
			<td>Mouse holder</td>
			<td>The 3D printing material used was ABS-M30i, and it was printed on FDM Fortus 380mc machine&#160;</td>
			<td>N/A</td>
			<td>Dimensions: length = 43 mm, outer radius = 15 mm, inner width (where the mouse would sit) = 20.7 mm.&#160;</td>
		</tr>
		<tr>
			<td>MyRun Machine</td>
			<td>Cosmo Bio Co Ltd</td>
			<td>CBJ-IMR-001-EX</td>
			<td></td>
		</tr>
		<tr>
			<td>Nanodrop 8000 Spectrophotometer</td>
			<td>Thermo Scientific</td>
			<td>ND-8000-GL</td>
			<td></td>
		</tr>
		<tr>
			<td>p53 primers</td>
			<td>Eurofins</td>
			<td>N/A</td>
			<td>Custom Primers</td>
		</tr>
		<tr>
			<td>PCR tubes</td>
			<td>Diamed</td>
			<td>SSI3131-06</td>
			<td></td>
		</tr>
		<tr>
			<td>Penicillin/Streptomycin</td>
			<td>Wisent</td>
			<td>450-200-EL</td>
			<td></td>
		</tr>
		<tr>
			<td>Proteus software&#160;</td>
			<td>Pichardo lab</td>
			<td>N/A</td>
			<td></td>
		</tr>
		<tr>
			<td>Respiratory monitoring system</td>
			<td>SAII</td>
			<td>Model 1030</td>
			<td>MR-compatible monitoring and gating system for small animals</td>
		</tr>
		<tr>
			<td>Small Bore HIFU device, LabFUS</td>
			<td>Image Guided Therapy</td>
			<td>N/A</td>
			<td>LabFUS, Image Guided Therapy, Pessac, France Number of elements 8 
			frequency 2.5 MHz 
			diameter&#160; 25 mm 
			radius of curvature 20 mm 
			Focal spot size 0.6 mm x 0.6 mm x 2.0 mm 
			 
			Motor: axes 2 
			 
			Generator: 
			Number of channels 8 
			Maximum electrical power/channel Wel 4 
			Maximum electrical power Wel 32 
			Bandwidth 0.5 - 5 MHz 
			Control per channel: Freq., Phase and. amplitude 
			Measurements per channel: Vrms, Irms, cos(theta) 
			Duty Cycle at 100% power % 100% for 1 min. 
			 
			Transducer: 
			Number of elements 8 
			frequency&#160; 2.5 MHz 
			diameter 25 mm 
			radius of curvature 20 mm 
			Focal spot size&#160; 0.6 mm x 0.6 mm x 2.0 mm</td>
		</tr>
		<tr>
			<td>SYBR Safe</td>
			<td>ThermoFisher Scientific</td>
			<td>S33102</td>
			<td></td>
		</tr>
		<tr>
			<td>TAE</td>
			<td>Wisent</td>
			<td>811-540-FL</td>
			<td></td>
		</tr>
		<tr>
			<td>Tail vein catheter (27G 0.5&#34; )</td>
			<td>Terumo Medical Corp</td>
			<td>15253</td>
			<td></td>
		</tr>
		<tr>
			<td>Thermal probes</td>
			<td>Rugged Monitoring</td>
			<td>L201-08</td>
			<td></td>
		</tr>
		<tr>
			<td>Trypan blue</td>
			<td>ThermoFisher Scientific</td>
			<td>15250061</td>
			<td></td>
		</tr>
		<tr>
			<td>Trypsin</td>
			<td>Wisent</td>
			<td>325-052-EL</td>
			<td></td>
		</tr>
		<tr>
			<td>Ultrasound Gel</td>
			<td>Aquasonic</td>
			<td>PLI 01-08</td>
			<td></td>
		</tr>
	</tbody>
</table>

magnetic resonance-guided high intensity focused ultrasound generated hyperthermia: a feasible treatment method in a murine rhabdomyosarcoma model

磁共振引导的高强度聚焦超声（MRgHIFU）是一种产生局部热疗的既定方法。考虑到实时成像和声能调制，这种模式可以在定义的区域内实现精确的温度控制。这种非侵入性、非电离技术正在探索许多热应用，例如热疗产生，以从热敏脂质体载体中释放药物。这些药物可以包括化疗，如阿霉素，由于剂量限制性全身副作用，即心脏毒性，需要靶向释放。多柔比星是治疗多种恶性肿瘤的主要药物，通常用于复发或复发性横纹肌肉瘤 （RMS）。RMS是儿童和年轻人中最常见的实体性软组织颅外肿瘤。尽管进行了积极的多模式治疗，但RMS存活率在过去30年中保持不变。为了探索解决这一未满足需求的解决方案，开发了一种实验方案，以评估使用MRgHIFU作为药物释放热疗源的免疫功能正常的同系RMS小鼠模型中热敏脂质体阿霉素（TLD）的释放。

使用MRgHIFU生成的热疗方案，后肢肿瘤能够在治疗期间持续加热到所需的设定温度（图4 显示了代表性治疗，10或20分钟，n = 65）。为了认为治疗成功，整个治疗过程中的ROI必须保持在39°C以上，在整个治疗过程中变化<6°C，并且不加热脱靶组织。此外，根据直肠探头或起始直肠温度加上食管探头温度的变化，核心温度必须保持在39°C以下（补充图2）。一旦MRgHIFU的?...

Watch this Scientific Journal Video about Magnetic Resonance-Guided High Intensity Focused Ultrasound Generated Hyperthermia: A Feasible Treatment Method in a Murine Rhabdomyosarcoma Model at JoVE.com

Magnetic Resonance-Guided High Intensity Focused Ultrasound Generated Hyperthermia: A Feasible Treatment Method in a Murine Rhabdomyosarcoma Model

这里介绍的是一种协议，使用由磁共振引导的高强度聚焦超声产生的受控热疗来触发横纹肌肉瘤小鼠模型中温度敏感脂质体的药物释放。

磁共振引导的高强度聚焦超声产生热疗：小鼠横纹肌肉瘤模型中的可行治疗方法

Magnetic resonance-guided high intensity focused ultrasound (MRgHIFU) is an established method for producing localized hyperthermia. Given the real-time ...

该方案允许肿瘤靶向热疗与热敏药物联合进行新的临床前测试，可用于提高肉瘤患者的生存率并降低全身毒性。该技术的主要优点是化疗释放仅限于加热区域，最大化肿瘤内的药物释放，同时最小化药物在健康组织中的全身释放。我们的方法可以在临床上转化为治疗肉瘤。此外，我们的工作是了解局部低温如何激发免疫系统，帮助身体在标准化疗的同时靶向癌症。展示该程序的将是Star创新中心的首席MRI物理学家Warren Foltz博士，病童医院的首席HIFU物理学家Adam Waspe博士和多伦多病童医院德雷克实验室应用科学硕士Suzanne Wong。开始使用剪刀剃除被约束的鼠标的后腿，并用70%乙醇擦拭。使用微升注射器在10微升细胞悬液中吸出约10， 000个细胞。伸展鼠标的右后腿，并使用带有26号针头的微升注射器以稳定的运动将细胞悬液注射到大腿肌肉组织中。针头应平行于股骨朝膝盖插入，注意不要撞到坐骨神经。取下针头并将鼠标放回第二个笼子。每天评估动物，并通过触诊监测它们的后肢肿瘤生长。使用MRI扫描仪对麻醉小鼠进行成像。在 T2 加权图像上，注意平坦尺寸和肿瘤出现的轴向切片数。注意肿瘤相对于股骨和超声波进入的大腿侧表面的位置。要准备小口径HIFU系统，请打开发生器，将膜放在换能器上，并在换能器中填充足够的去离子水，直到膜在换能器下方膨胀。但不要太硬，以至于它会压缩鼠标。对换能器电路中的水进行脱气 30 分钟，以去除介质中的溶解氧。麻醉小鼠并将其转移到鼻锥后，在眼睛上涂抹角膜润滑剂，以防止由于麻醉下缺乏眨眼反射而造成的损害。剃掉鼠标的右后肢。在剃光区域（包括整个右后肢）涂抹脱毛膏。在生物安全柜中将鼠标放在加热灯下，以帮助进行热调节。并在使用后一分钟去除头发和脱毛膏。将鼠标移动到MRI雪橇上的MRI兼容鼻锥上，并将鼠标置于非肿瘤轴承面朝下，并将肿瘤上部置于雪橇上的3D打印鼠标支架内。在鼠标附近放置加热灯以保持温暖。如果需要，切成压缩的超声凝胶垫段，将鼠标衬里放在支架底部的底部，厚度使肿瘤水平到支架的顶部。将未受累的腿从肿瘤腿上塞开，要么在小鼠下方，要么在肿瘤腿弯曲的情况下伸展。确保脚不在肿瘤和超声束路径的近场或远场。要插入食管温度探头，请将食管探头穿过鼻锥并擦拭小鼠的颈部。通过伸展头部，将鼠标的鼻子向上倾斜，从嘴巴直接到胃部形成一条线。将舌头上方的热探头滑入小鼠食道约 0.5 厘米，并更换小鼠鼻子周围的鼻锥。将食管探头和鼻锥固定在雪橇顶部。接下来，插入直肠温度探头并用胶带固定。如果需要，插入另一个凝胶垫，并在右后肢周围涂抹眼睛润滑剂或超声波凝胶以填充任何间隙。将 27 号蝶针尾静脉导管插入连接到带有 40 微升死区并牢固胶带的微管的侧尾静脉中。将带有连接电缆的呼吸监测器朝向鼠标头部，以免干扰超声换能器的放置，并用胶带固定。用两个人将准备好的鼠标、鼠标雪橇、麻醉线、呼吸线、尾静脉导管和热探头线带入 MRI 扫描仪，并将它们放入 MRI 雪橇支架中。每个人都经过了筛选，没有任何东西被带入MR套件，这是铁磁性的，或者在系统的强磁场中可能被损坏。HIFU软件操作员将换能器的半月板直接移动到肿瘤上进行初始对齐后，将眼部润滑剂或脱气超声凝胶涂在肿瘤上方的无毛皮肤上，并将HIFU换能器耦合到肿瘤区域。将自动泵的给药管线连接到尾静脉导管，并计算尾静脉线和连接线中的死区量。将MRI导轨上的鼠标HIFU滑板滑入MRI的中心，并将空气对流加热装置置于最温暖的设置上。将管子吹向MRI孔中心的鼠标，并用胶带固定。加热设备稍后将转到其最低设置，以防止在超声处理过程中鼠标过热。获取轴向和矢状面的调查MR图像，以确定超声靶向的肿瘤位置，包括深度。使用HIFU软件相应地调整换能器位置，方法是在图像上插入测量的所需移动距离，然后单击箭头方向进行移动。另外，请注意漂移管的位置。根据需要重复此操作。在测试注射测温采集期间，通过执行简短的50毫伏连续测试超声处理5秒钟来确定探头在角平面中焦点的位置。将 MR 测量图像与 HIFU 软件中焦点的视图对齐。查看图像中肿瘤相对于骨结构和直肠的位置，并根据需要修改换能器位置。在九次重复热成像期间重复测试射击超声处理，以确认肿瘤体积中是否存在均匀准确的加热，并且脱靶加热最小。调整切片位置、传感器位置和转向深度，并在必要时通过重复测试镜头来确认加热性能。使用HIFU处理监测软件，通过测量移动距离，然后在程序中改变网格坐标，定义最终加热曲线内温度监测的感兴趣区域或ROI。在漂移管的流体信号内设置ROI，以便对测温输出进行漂移校正。如果在肿瘤水平的轴向规划图像中没有很好地定义该流体信号，则漂移管内光纤温度探头的直接输出用于漂移校正。根据直肠探头温度输入基线温度以进行测温。在软件中打开20分钟的热疗规格，并在收集参考MR图像并开始测温后开始超声处理。在热成像过程中，使用内置的比例积分微分（PID）控制器软件进行20分钟的处理。在ROI中的温度升温至所需温度后1.5分钟注射所选药物。如果在治疗过程中直肠温度迅速升高并超过 40 摄氏度，请停止超声处理并让直肠温度下降，然后再重新开始。治疗完成后，将小鼠从MRI孔中取出，确保尾静脉导管插入部位止血并将其转移到生物安全柜中。用水洗掉后腿上的超声波凝胶和润滑剂。将鼠标转移到笼子上并监控其恢复情况。将加热灯对准笼子的一端，以帮助在恢复过程中进行热调节。使用这种热疗方案，后肢的肿瘤能够在治疗期间持续加热到所需的设定温度。代表性治疗期间监测的温度如图所示。此处显示了 ROI 中所有体素的平均前 10 个百分位数和前 90 个百分位数的温度。在优化阶段以标准偏差测试每只小鼠ROI内处理期间的平均温度。还显示了整体平均温度。随着时间的推移，热疗的成功率也有所提高。治疗成功与否取决于纳入标准。即全身温度，肿瘤温度和随ROI的变化，没有远端加热。在这里，蓝线表示治疗成功的小鼠百分比，橙色条表示治疗的小鼠数量。每种处理都是指进行实验的单独日期。为了确定进一步研究的最佳低温治疗时间，测试了两种治疗持续时间，10分钟和20分钟。高效液相色谱和质谱（HPLC MS）用于评估肿瘤中阿霉素的量，并量化测试持续时间之间阿霉素积累的差异。结果表明，与正常热疗对照相比，20分钟热敏脂质体阿霉素加热疗组肿瘤中阿霉素的含量具有显着意义。游离阿霉素组的肿瘤没有差异。在准备和定位砌块时，HIFU光束必须畅通无阻。必须避免凝胶中的气泡或光束路径中的骨头。我们收集组织和血液以分析各种器官的药代动力学和毒性水平，并检查免疫系统对治疗的即时和延迟反应。

Research

JoVE Journal

Cancer Research

Endobronchial Ultrasound-guided Intratumoral Injection of Cisplatin for the Treatment of Isolated Mediastinal Recurrence of Lung Cancer

支气管内超声引导下顺铂瘤内注射对肺癌纵隔孤立复发的治疗

Isolated hilar and mediastinal recurrence (IMHR) following external beam radiation therapy (EBRT) in patients with lung cancer is common. These patients ...

科研

癌症研究

A Murine Orthotopic Bladder Tumor Model and Tumor Detection System

小鼠原位膀胱肿瘤模型和肿瘤检测系统

This protocol describes the generation of bladder tumors in female C57BL/6J mice using the murine bladder cancer cell line MB49, which has been modified ...

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. ...

In Vivo EPR Assessment of pH, pO2, Redox Status, and Concentrations of Phosphate and Glutathione in the Tumor Microenvironment

In Vivo EPR Assessment of pH, pO2, Redox Status, and Concentrations of Phosphate and Glutathione in the Tumor Microenvironment

体内在肿瘤微环境中 pH 值、 pO2、氧化还原状态、磷酸盐和谷胱甘肽浓度的 EPR 评价

This protocol demonstrates the capability of low-field electron paramagnetic resonance (EPR)-based techniques in combination with functional paramagnetic ...

A Method to Define the Effects of Environmental Enrichment on Colon Microbiome Biodiversity in a Mouse Colon Tumor Model

一种确定环境富集对小鼠结肠肿瘤模型结肠菌群生物多样性影响的方法

Several recent studies have illustrated the beneficial effects of living in an enriched environment on improving human disease. In mice, environmental ...

High-sensitivity Detection of Micrometastases Generated by GFP Lentivirus-transduced Organoids Cultured from a Patient-derived Colon Tumor

淋巴结慢病毒北疆-转基因 Organoids 培养的大鼠结肠肿瘤的高灵敏度检测

Despite current advances in human colorectal cancer (CRC) treatment, few radical therapies are effective for the late stages of CRC. To overcome this ...

Evaluation of the Feasibility, Safety, and Accuracy of an Intraoperative High-intensity Focused Ultrasound Device for Treating Liver Metastases

术中高强度聚焦超声装置治疗肝转移的可行性、安全性和准确性评价

Today, the only potentially curative option in patients with colorectal liver metastases is surgery. However, liver resection is feasible in less than 20% ...

Characterize Disease-related Mutants of RAF Family Kinases by Using a Set of Practical and Feasible Methods

使用一套实用可行的方法,描述英国皇家空军家族激酶与疾病相关的突变体

The rapidly accelerated fibrosarcoma (RAF) family kinases play a central role in cell biology and their dysfunction leads to cancers and developmental ...

Ultrasound-Guided Orthotopic Implantation of Murine Pancreatic Ductal Adenocarcinoma

毛尿胰腺腺腺腺癌超声引导正射植入

The recent success of immune checkpoint blockade in melanoma and lung adenocarcinoma has galvanized the field of immuno-oncology as well as revealed the ...

Modeling Brain Metastases Through Intracranial Injection and Magnetic Resonance Imaging

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 ...