作者感谢于峰教授、詹娜·布里德尔博士、伊恩·伍德沃德博士和卢卡斯·阿提亚教授的有益讨论。

1. 准备3D打印实验组件
注:协议中使用的所有软件均在 材料表中注明。此外，所使用的切片软件是特定于 材料表中列出的 3D 打印机:但是，此协议可以扩展到各种立体石刻 （SLA） 3D 打印机。
<ol><li>将患者 CT 扫描转换为 3D 对象 （.stl 文件）。 注:要更详细地讨论这些研究中使用的特定肺模型的几何特征，请参阅冯等人5。<ol>
<li>使用 CT 扫描软件将 CT 扫描渲染为 3D 对象（参见 材料表）。打开 CT 扫描，使用 阈值 工具在空域上创建一个面罩，其设置范围为 -800 到 -1000。使用 3D 预览 工具查看 3D 渲染并导出对象（文件|导出）作为。stl文件。</li>
		<li>将文件导入网格编辑软件（参见 材料表），使用 选择 工具删除任何锯齿状的功能（雕刻|画笔:"收缩/平滑"|属性: 强度 （50）， 大小 （10）， 深度 （0）） 。平滑表面（ctrl+A |变形|平滑|平滑 （0.2）， 平滑量表 （1））.</li>
		<li>在网格编辑软件中，将这些对象的墙壁延长 2 毫米（Ctrl+A |编辑|偏移），并允许内部物体保持空心，使只有墙仍然存在。切片对象（选择|编辑|平面切割）在气管形成一个入口，并在第2代或第3代，对象分支到每个叶创建插座（图1A）。 注:2毫米的厚度是根据 材料表中列出的3D打印机制造商指定的可接受功能尺寸选择的。如果保持模型的内部几何形状，则可以根据可用的 3D 打印机的规格调整此厚度。</li>
	</ol></li>
	<li>修改患者肺模型插座几何形状，以与材料表中列出的先前设计的叶出口盖组件（图1B，C）兼容。<ol>
<li>导入3D对象，复制内部CT扫描，壁厚度为2毫米，在入口和插座打开，进入3D建模软件（见 材料表）作为固体体（打开|网格文件|选项|固体身体）。</li>
		<li>根据每个插座的面部创建平面（插入|参考几何学|飞机）。使用拼接工具，在平面上的草图中跟踪出口的内壁和外壁（素描|斯普林）。</li>
		<li>将气缸（OD 18.5 mm、ID 12.5 mm、H 15.15 mm）连接到模型的内墙和外壁，从而将出口扩展到每个叶的均匀（功能|阁楼老板/基地）。在插座边缘添加一个缺口，以配合盖子（功能|挤压切割|偏移）。 注:盖（图1D）是一个空心圆柱体，匹配插座的尺寸，并有一个架子，与模型插座的档次相互连接。盖子的一端被阻塞，使 ID 小于部分的其余部分，这确保了带刺管连接（图 1E）的紧密配合。带刺管连接是一个带刺锥形，使刺通过帽的开口适合，但其余部分不，允许管连接牢固地适合在帽。因此，帽紧密地贴合在带刺管连接和肺模型（图1F，G）周围。</li>
		<li>根据所需的实验条件修改肺模型的入口。喉咙和咽喉区域可以包括模仿病人，可以自己呼吸（图1B）。气管上方的区域可以使用挤压的切口来模拟呼吸机支架上的插管患者（功能|挤压切割）（图1C）。</li>
	</ol></li>
	<li>3D 打印机制造商提供的切片软件中的定向和支持实验组件。<ol>
<li>将 3D 部分文件导入 3D 打印机切片软件，并选择适当的树脂。使用硬树脂打印肺模型和带刺管连接，以及软树脂打印帽。 注意:用于打印盖子的树脂必须具有弹性特性，使其能够伸展到叶出口，并创建一个密封的密封。</li>
		<li>设置部分方向，以便将任何"岛屿"和未发明的卷最小化。肺模型的最佳方向是叶口面向远离打印平台。确保带刺管连接和盖子都具有面向打印平台的更宽部分。 注:可以查看单个切片，以检查"岛屿"的出现，该部分最初出现在切片中，而不连接到部分的主体。审查功能可用于检查未发明卷的切片，未治愈的树脂在打印过程中可能被困在部分内的区域。"岛屿"和未发明的卷都会降低打印质量，并可能导致打印失败。</li>
		<li>单独查看每片，为该部分中任何剩余的"岛屿"以及任何悬垂严重的区域添加支撑。导出并查看打印切片，以验证所有区域是否得到适当的支持。</li>
	</ol></li>
	<li>按照制造商的说明打印实验组件并完成后处理。 注:以下所有后处理步骤均特定于 材料表中列出的 3D 打印机。当使用替代打印机或材料时，调整这些步骤以反映制造商的说明。<ol>
<li>对于用软树脂打印的零件，根据制造商规格，用≥99%纯度异丙醇 （IPA） 清洗，以去除多余的未固化树脂，并根据制造商规格在对流烤箱中热固化 8 小时。 注意:在软树脂中打印的零件在打印后可以非常精细，因此在清洁步骤时应特别小心。接触 IPA 应保持在材料溶剂暴露限制以下，以防止部分降解。</li>
		<li>对于用硬树脂打印的零件，用 IPA 清洗以去除多余的未固化树脂，并在紫外线烤箱中固化（365 nm 光在 5-10 mW/cm2），每侧 1 分钟。 注意:为了评估 3D 打印副本的准确性，建议使用印刷部分和 CT 扫描软件的 μCT 扫描来定量地比较原始 3D 渲染和 3D 打印复制品之间的变化。</li>
	</ol></li>
</ol>2. 流量控制管系统组装
<ol><li>螺丝1/4"带刺管配件到歧管的一侧，有6个端口（图2A-6）和一个3/8"带刺管配件到剩余的端口。</li>
	<li>将 1/4" 管子切成所需的长度，并插入推接阀的每一端（图 2A-5）。将每个阀门连接到插入歧管中的 1/4" 配件之一。</li>
	<li>将流量计（图2A-4）连接到每个阀门的另一端。</li>
	<li>将管子系统放置在木板顶部，使多管的单个 3/8"配件延伸至木板边缘。为了固定到位，在木板的一侧添加两个螺丝，然后用电线将歧管连接到螺钉上。</li>
	<li>在每个阀门和流量计周围添加四个螺丝，然后使用电线将每个螺钉固定在木板上（图 2E）。</li>
	<li>使用大约 6" 的 3/8" ID 管，将歧管连接到在线 0.1 μm 孔径大小的真空级过滤器。使用另一个 6" 的 3/8" ID 管将过滤器的另一端连接到流量控制器 注:管子系统只需组装一次。</li>
</ol>3. 带患者肺模型的叶出口盖组装
注意:协议的这一部分必须在每次实验运行之前完成。
<ol><li>将带刺管连接插入盖中，喷嘴通过盖底的开口突出。首先，将椭圆形带刺管连接底座的一端插入盖中。然后，小心地将柔性盖伸到椭圆形底座的另一端，特别注意不要裂开薄底座。 注意:新打印的帽子可能比预期的更硬，并且可以通过沿着盖子内部运行两根手指来伸展。</li>
	<li>切割 10μm 滤纸，使其略大于出口区域。将滤纸折叠在叶出口上，用一只手保持原位。</li>
	<li>另一方面，使用钳子在插座上用带刺管连接将帽子拉伸。按下盖，直到盖的缺口与叶出口上的相应缺口匹配（图 2C）。 注意:在此步骤中撕开滤纸可能会使结果失效，因此在将盖压到插座上时应特别小心以避免过度用力。</li>
	<li>重复所有剩余的叶插座 （图 2D）。</li>
</ol>4. 生成临床相关气流剖面
注意:协议的这一部分必须在每次实验运行之前完成。
<ol><li>将每个肺模型叶出口连接到相应的流量计和阀门的管子，注意不要对带刺管连接施加过多的横向压力。将电子流量计连接到肺模型口入口，以测量肺模型的总气流速率。</li>
	<li>打开流量控制器（图2A-7）和真空泵（图2A-8）。选择流量控制器上的"测试设置"设置，并缓慢地增加流速，直到电子流量计显示所需的总流速率。</li>
	<li>使用阀门（图2E-5），通过五个肺叶中的每一个调整流速:右上（RU）、右中叶（RM）、右下（RL）、左上（LU）和左下（LL）。一旦流表（图2E-4）上显示的叶流速率稳定在所需值，请再次检查电子流量表上的整体流速，以验证系统中没有泄漏。<ol>
<li>如果总流速存在差异，则通过流量控制器降低流速，将所有阀门设置为完全打开的配置，并重复步骤 4.2 和 4.3。 注:此处介绍的结果是利用Sul等人报告的数据，利用全灵感和过期患者肺部的薄片计算断层扫描图像计算得出的，比较了每个肺叶体积的相对变化。结果针对两种不同的流量条件，两种条件的总体入口流量均为 1 L/min。健康的肺叶出口流向每个出口按入口流量的以下百分比分布:LL-23.7%、LU-23.7%、RL-18.7%、RM-14.0%、RU-20.3%。COPD 叶出口流介按入口流量的以下百分比分布于每个出口之间:LL-10.0%、LU-29.0%、RL-13.0%、RM-5.0%、RU-43.0%9、10。</li>
	</ol>
</li>
	<li>退出流量控制器的"测试设置"功能，但请打开真空泵。 注意:在设置流速和执行沉积实验之间关闭真空泵可能导致生成的流量配置文件不准确。建议在设定所需的流量后将真空泵留在真空泵上，以完成气溶胶沉积测试。</li>
</ol>5. 向肺模型输送气溶胶
注:必须在烟罩中进行实验，关闭窗框，以尽量减少暴露在雾化器产生的任何气溶胶中。
<ol><li>填充雾化器与所需的荧光颗粒溶液 （图2A-1），并连接到肺模型入口 （图 2B）. 注:此处介绍的结果是使用甲醇中 1:100 稀释 1μm 荧光聚苯乙烯颗粒的 30 mL 得出的。<ol>
<li>要验证实验设置，请将雾化器直接连接到肺模型入口，无需任何定位装置。</li>
		<li>为了测量靶向装置的功效，将雾化器连接到设备，并将设备插入肺模型。</li>
	</ol>
</li>
	<li>将压缩的气管连接到雾化器，并尽可能关闭烟罩窗框。</li>
	<li>设置流控制器运行一个 10s 试用版。在按下启动之前，稍微打开压缩空气阀，开始在雾化器内产生气溶胶。</li>
	<li>按动启动流量控制器，并立即完全打开压缩空气阀。一旦流量控制器达到约9s，开始关闭压缩空气阀。</li>
	<li>压缩空气阀完全关闭后，将雾化器与压缩气管分离，完全关闭烟罩窗框，关闭真空泵，让任何气溶胶从烟斗中清除约 10 分钟。 注意:在完成运行后关闭真空泵非常重要，以防止真空在管道系统内积木。</li>
	<li>等待足够时间后，将肺模型与管系统断开，特别注意不要破裂带刺管连接。</li>
	<li>通过在盖子边缘下运行一对钳子，并轻轻地将其从肺模型上抬起来，取下叶出口盖。</li>
	<li>将滤纸从盖子上取下，放入 24 个井板中，颗粒沉积在底部，朝向板的油井。重复为剩余的插座，并标记与每个叶对应的井。 注意:为了防止任何残余颗粒沉积影响后续实验，在运行之间用 IPA 或适当的溶剂冲洗肺模型和盖组件非常重要。这可以根据需要收集并包含在分析中。此外，还保留日志，以确保使用的所有副本都很少暴露在 IPA 中，以保持部分完整性，并建议在使用前进行视觉部分检查。</li>
</ol>6. 出口滤纸成像
<ol><li>将井板放入数字荧光显微镜中，并将显微镜设置为 4 倍放大和适当的荧光通道。</li>
	<li>目视识别哪个叶的滤纸有最高数量的颗粒沉积，并使用"自动暴露"功能。注意由此产生的曝光和集成时间值。</li>
	<li>将此曝光应用到运行的所有滤镜中，并评估设置是否为过滤器的所有高沉积区域生成满意的图像。 注意:对焦设置可以从筛选器更改为筛选器:但是，必须在相同的曝光设置下分析给定运行的所有滤镜。一次只能有一帧焦点，因此滤纸中的弯曲或撕裂可能会阻止视图中所有沉积的粒子聚焦。通过确保滤纸与井板底部平整，可以避免这种情况。</li>
	<li>在随机位置拍摄每个叶滤纸的至少三张图像，并保存为.tiff文件。</li>
</ol>7. 粒子沉积量化
<ol><li>导入所有滤纸图片，以在给定运行到图像J会话中。</li>
	<li>通过选择图像|将每个图像的类型更改为 8 位 类型|8 位。</li>
	<li>打开具有最高荧光的图片，并选择图像|调整|打开阈值窗口的阈值。调整阈值以最大限度地减少滤纸的背景信号，并清楚地定义粒子的边缘。有关优质和劣质阈值的描述，请参阅图 3。 注意:对于沉积水平高的过滤器，可以在大颗粒群周围观察到由滤纸纤维的光扩散引起的荧光"冠冕"。阈值这些图像时，太大的范围会显示这些组周围的小点或"羽毛状"形状，如 图 3中的"差"阈值图像中所示。这可以通过逐步提高阈值的下限来改进，直到过滤纸纤维发出的信号最小化，而不会遮挡颗粒本身的信号。</li>
	<li>将最高荧光图像的阈值设置传播到所有其他图像。</li>
	<li>通过选择分析|来量化颗粒数量和整个荧光区 分析粒子。 注:使用 Sidak 的多重比较测试和双向 ANOVA 比较数据集。此外，仅使用学生 T 测试假设相等的方差，比较了兴趣叶中的沉积。</li>
</ol>

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B., Katz, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Deposition+Patterns+of+Polydisperse+Aerosols+Within+Human+Lungs.">Deposition Patterns of Polydisperse Aerosols Within Human Lungs.</a> Journal of Aerosol Medicine. 6 (4), 251-274 (1993).</li><li>Nahar, 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=In+vitro,+in+vivo+and+ex+vivo+models+for+studying+particle+deposition+and+drug+absorption+of+inhaled+pharmaceuticals.">In vitro, in vivo and ex vivo models for studying particle deposition and drug absorption of inhaled pharmaceuticals.</a> European Journal of Pharmaceutical Sciences. 49 (5), 805-818 (2013).</li><li>Nichols, S. C., 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+Multi-laboratory+in+Vitro+Study+to+Compare+Data+from+Abbreviated+and+Pharmacopeial+Impactor+Measurements+for+Orally+Inhaled+Products:+a+Report+of+the+European+Aerosol+Group+(EPAG).">A Multi-laboratory in Vitro Study to Compare Data from Abbreviated and Pharmacopeial Impactor Measurements for Orally Inhaled Products: a Report of the European Aerosol Group (EPAG).</a> AAPS PharmSciTech. 17 (6), 1383-1392 (2016).</li><li>Yoshida, H., Kuwana, A., Shibata, H., Izutsu, K. I., Goda, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+Aerodynamic+Particle+Size+Distribution+Between+a+Next+Generation+Impactor+and+a+Cascade+Impactor+at+a+Range+of+Flow+Rates.">Comparison of Aerodynamic Particle Size Distribution Between a Next Generation Impactor and a Cascade Impactor at a Range of Flow Rates.</a> AAPS PharmSciTech. 18 (3), 646-653 (2017).</li><li>Feng, Y., 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=An+in+silico+inter-subject+variability+study+of+extra-thoracic+morphology+effects+on+inhaled+particle+transport+and+deposition.">An in silico inter-subject variability study of extra-thoracic morphology effects on inhaled particle transport and deposition.</a> Journal of Aerosol Science. 123, 185-207 (2018).</li><li>Kleinstreuer, C., Seelecke, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inhaler+system+for+targeted+maximum+drug-aerosol+delivery.">Inhaler system for targeted maximum drug-aerosol delivery.</a> United States patent. , (2005).</li><li>. How Medical 3D Printing is Gaining Ground in Top Hospitals Available from: <a target="_blank" href="https://www.materialise.com/en/blog/3d-printing-hospitals">https://www.materialise.com/en/blog/3d-printing-hospitals</a> (2019)</li><li>Weber, P. W., Price, O. T., McClellan, G. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Demographic+Variability+of+Inhalation+Mechanics:+A+Review.">Demographic Variability of Inhalation Mechanics: A Review.</a> Defense Threat Reduction Agency. , (2016).</li><li>Jiang, Y. Y., Xu, X., Su, H. L., Liu, D. X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gender-related+difference+in+the+upper+airway+dimensions+and+hyoid+bone+position+in+Chinese+Han+children+and+adolescents+aged+6-18+years+using+cone+beam+computed+tomography.">Gender-related difference in the upper airway dimensions and hyoid bone position in Chinese Han children and adolescents aged 6-18 years using cone beam computed tomography.</a> Acta Odontologica Scandinavica. 73 (5), 391-400 (2015).</li><li>Martin, S. E., Mathur, R., Marshall, I., Douglas, N. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effect+of+age,+sex,+obesity+and+posture+on+upper+airway+size.">The effect of age, sex, obesity and posture on upper airway size.</a> European Respiratory Journal. 10 (9), 2087 (1997).</li><li>Xi, J., Longest, P. W., Martonen, T. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+the+laryngeal+jet+on+nano-+and+microparticle+transport+and+deposition+in+an+approximate+model+of+the+upper+tracheobronchial+airways.">Effects of the laryngeal jet on nano- and microparticle transport and deposition in an approximate model of the upper tracheobronchial airways.</a> Journal of Applied Physiology. 104 (6), 1761-1777 (2008).</li><li>Zhao, J., Feng, Y., Fromen, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glottis+motion+effects+on+the+particle+transport+and+deposition+in+a+subject-specific+mouth-to-trachea+model:+A+CFPD+study.">Glottis motion effects on the particle transport and deposition in a subject-specific mouth-to-trachea model: A CFPD study.</a> Computers in Biology and Medicine. 116, 103532 (2020).</li><li>Kim, S. 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=Chronic+obstructive+pulmonary+disease:+lobe-based+visual+assessment+of+volumetric+CT+by+Using+standard+images--comparison+with+quantitative+CT+and+pulmonary+function+test+in+the+COPDGene+study.">Chronic obstructive pulmonary disease: lobe-based visual assessment of volumetric CT by Using standard images--comparison with quantitative CT and pulmonary function test in the COPDGene study.</a> Radiology. 266 (2), 626-635 (2013).</li><li>. The Cancer Imaging Archive Available from: <a target="_blank" href="https://www.cancerimagingarchive.net/">https://www.cancerimagingarchive.net/</a> (2020)</li><li>Li, A., Ahmadi, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Computer+Simulation+of+Deposition+of+Aerosols+in+a+Turbulent+Channel+Flow+with+Rough+Walls.">Computer Simulation of Deposition of Aerosols in a Turbulent Channel Flow with Rough Walls.</a> Aerosol Science and Technology. 18 (1), 11-24 (1993).</li><li>Khalili, S. F., Ghanbarzadeh, S., Nokhodchi, A., Hamishehkar, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effect+of+different+coating+materials+on+the+prevention+of+powder+bounce+in+the+next+generation+impactor.">The effect of different coating materials on the prevention of powder bounce in the next generation impactor.</a> Research in Pharmaceutical Sciences. 13 (3), 283-287 (2018).</li><li>Galliger, Z., Vogt, C. D., Panoskaltsis-Mortari, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=3D+bioprinting+for+lungs+and+hollow+organs.">3D bioprinting for lungs and hollow organs.</a> Translational Research. 211, 19-34 (2019).</li><li>Schwarz, K., Biller, H., Windt, H., Koch, W., Hohlfeld, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterization+of+exhaled+particles+from+the+healthy+human+lung--a+systematic+analysis+in+relation+to+pulmonary+function+variables.">Characterization of exhaled particles from the healthy human lung--a systematic analysis in relation to pulmonary function variables.</a> Journal of Aerosol Medicine and Pulmonary Drug Delivery. 23 (6), 371-379 (2010).</li><li>Patton, J. S., Byron, 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=Inhaling+medicines:+delivering+drugs+to+the+body+through+the+lungs.">Inhaling medicines: delivering drugs to the body through the lungs.</a> Nature Reviews Drug Discovery. 6 (1), 67-74 (2007).</li><li>Zhang, Z., Kleinstreuer, C., Kim, C. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cyclic+micron-size+particle+inhalation+and+deposition+in+a+triple+bifurcation+lung+airway+model.">Cyclic micron-size particle inhalation and deposition in a triple bifurcation lung airway model.</a> Journal of Aerosol Science. 33 (2), 257-281 (2002).</li><li>Ju, Y., 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=Engineering+of+Nebulized+Metal-Phenolic+Capsules+for+Controlled+Pulmonary+Deposition.">Engineering of Nebulized Metal-Phenolic Capsules for Controlled Pulmonary Deposition.</a> Advanced Science. 7 (6), 1902650 (2020).</li></ol>

作者没有什么可透露的。

目前用于完全吸入剂量肺药测试的先进设备是下一个发电机冲击器 （NGI），用于测量气溶胶4的空气动力学直径。然后，这个大小的数据用于预测肺生成，气溶胶将沉积基于为健康的成年男性11开发的相关性。不幸的是，这种方法在评估区域肺沉积差异、确定疾病条件对药物输送的影响以及预测不同年龄组、种族和性别12、13、14</sup...

肺癌、慢性阻塞性肺病等许多肺病在疾病特征上表现出区域差异:然而，缺乏治疗技术，以目标药物交付到只有肺1的病区。多个计算流体动态（CFD）模型已经证明，通过识别肺部2，3的特定流线，可以调节药物沉积图谱。我们的实验室正在开发具有区域定位能力的吸入器和内脏（ET）管适配器，以控制气溶胶向患病肺部区域的分布。这些原则推广到临床使用受当前临床前测试能力的限制。众所周知，药物沉积在肺部的确切位置是疗效的最佳预测器:然而，目前的药物评估的可吸入治疗最常预测使用体外体内的粒子大小相关性，只是近似沉积4。此技术不允许任何空间分析来确定不同气道几何形状对通过肺部不同叶的区域分布的影响。此外，这项测试缺乏解剖学上准确的肺几何学，研究人员已经表明，这可能对沉积图5产生重大影响。已作出一些努力，通过增加上呼吸道，将患者特异性肺几何学纳入测试方案:然而，这些方法大多样本气溶胶输送到不同代的肺，而不是每个肺叶6，7，8。下列协议提出了一种高通量的方法，以产生患者特定的肺模型，并能够量化肺9的五个叶中的每个叶的相对粒子沉积。
解剖学上准确的模型肺是由3D打印患者计算断层扫描（CT）扫描产生的。当与一个容易组装的流动系统一起使用时，可以通过每个模型肺叶的相对流率进行独立控制和定制，以模拟不同患者人口统计学和/或疾病状态的相对流率。通过这种方法，研究人员可以在相关的肺几何学中测试潜在治疗方法的有效性，并将每种方法的性能与疾病形态的进展联系起来。在这里，我们实验室开发的两种设备设计通过控制口腔或气管中气溶胶释放的位置来增强所需肺叶沉积的能力。该协议还有可能通过促进对患者CT扫描数据特定于模型肺的治疗效果的快速预测，显著影响患者个性化程序的发展。

<table>
	<tbody>
		<tr>
			<td>1/4&#34; Plastic Barbed Tube Fitting</td>
			<td>McMaster Carr</td>
			<td>5372K111</td>
			<td></td>
		</tr>
		<tr>
			<td>10 um Filter Paper</td>
			<td>Fisher</td>
			<td>1093-110</td>
			<td></td>
		</tr>
		<tr>
			<td>1um Fluorescent Polystyrene Particles</td>
			<td>Polysciences</td>
			<td>15702-10</td>
			<td></td>
		</tr>
		<tr>
			<td>1um Non-Fluorescent Polystyrene Particles</td>
			<td>Polysciences</td>
			<td>8226</td>
			<td></td>
		</tr>
		<tr>
			<td>2-Propanol</td>
			<td>Fisher</td>
			<td>A516-4</td>
			<td>Referred to in protocol as &#34;IPA&#34;</td>
		</tr>
		<tr>
			<td>3/8&#34; Plastic Barbed Tube Fitting</td>
			<td>McMaster Carr</td>
			<td>5372K117</td>
			<td></td>
		</tr>
		<tr>
			<td>Air Flow Meter (1 - 280 mL/min)</td>
			<td>McMaster Carr</td>
			<td>41695K32</td>
			<td>Referred to in protocol as &#34;flow meter&#34;</td>
		</tr>
		<tr>
			<td>Carbon M1 3D Printer</td>
			<td>Carbon 3D</td>
			<td></td>
			<td>https://www.carbon3d.com/, Associated software referred to in protocol as &#34;slicing software&#34;</td>
		</tr>
		<tr>
			<td>Collison Jet Nebulizer</td>
			<td>CH Technologies</td>
			<td>ARGCNB0008 (CN-25)</td>
			<td>6 Jet MRE style horizontal collision with glass jar, Referred to in protocol as &#34;nebulizer&#34;, http://chtechusa.com/Manuals/MRE_Collison_Manual.pdf</td>
		</tr>
		<tr>
			<td>Convection Oven</td>
			<td>Yamato</td>
			<td>DKN602</td>
			<td></td>
		</tr>
		<tr>
			<td>Copley Critical Flow Controller TPK2000 Reve 120V</td>
			<td>MSP Corp</td>
			<td>0001-01-9810</td>
			<td>Referred to in protocol as &#34;flow controller&#34;</td>
		</tr>
		<tr>
			<td>Copley High Capacity Pump Model HCP5</td>
			<td>MSP Corp</td>
			<td>0001-01-9982</td>
			<td>Referred to in protocol as &#34;vacuum pump&#34;</td>
		</tr>
		<tr>
			<td>Cytation</td>
			<td>BioTek</td>
			<td>CYT5MPV</td>
			<td>Multifunctional Spectrophotometer/Fluorescent imager equiped with 4x/20x/40x objectives and DAPI/GFP/TexasRed laser/filter cubes</td>
		</tr>
		<tr>
			<td>EPU40 Resin</td>
			<td>Carbon 3D</td>
			<td></td>
			<td>https://www.carbon3d.com/materials/epu-elastomeric-polyurethane/, Referred to in protocol as &#34;soft resin&#34;</td>
		</tr>
		<tr>
			<td>Filter for vacuum pump</td>
			<td>Whatman</td>
			<td>6722-5000</td>
			<td></td>
		</tr>
		<tr>
			<td>Flow Meter Model DFM 2000</td>
			<td>MSP Corp</td>
			<td>0001-01-8764</td>
			<td>Referred to in protocol as &#34;electronic flow meter&#34;</td>
		</tr>
		<tr>
			<td>ImageJ Software</td>
			<td>ImageJ</td>
			<td></td>
			<td>https://imagej.nih.gov/ij/download.html</td>
		</tr>
		<tr>
			<td>Inline Air Flow Control Valve (Push-to-Connect)</td>
			<td>McMaster Carr</td>
			<td>62005K333</td>
			<td>Referred to in protocol as &#34;valve&#34;</td>
		</tr>
		<tr>
			<td>Inline Filter Devices</td>
			<td>Whatman</td>
			<td>WHA67225000</td>
			<td></td>
		</tr>
		<tr>
			<td>Marine-Grade Plywood Sheet</td>
			<td>McMaster Carr</td>
			<td>62005K333</td>
			<td>Referred to in protocol as &#34;wooden board&#34;</td>
		</tr>
		<tr>
			<td>Materialise Mimics Software</td>
			<td>Materialise</td>
			<td></td>
			<td>https://www.materialise.com/en/medical/mimics-innovation-suite, Referred to in protocol as &#34;CT scan software&#34;</td>
		</tr>
		<tr>
			<td>Meshmixer Software</td>
			<td>Autodesk</td>
			<td></td>
			<td>http://www.meshmixer.com/, Referred to in protocol as &#34;mesh editing software&#34;</td>
		</tr>
		<tr>
			<td>Methanol</td>
			<td>Fisher</td>
			<td>A454-4</td>
			<td></td>
		</tr>
		<tr>
			<td>Opticure LED Cube</td>
			<td>APM Technica</td>
			<td>102843</td>
			<td>Referred to in protocol as &#34;UV oven&#34;</td>
		</tr>
		<tr>
			<td>PR25 Resin</td>
			<td>Carbon 3D</td>
			<td></td>
			<td>https://www.carbon3d.com/materials/uma-urethanemethacrylate, /Referred to in protocol as &#34;hard resin&#34;</td>
		</tr>
		<tr>
			<td>PVC Tube for Chemicals</td>
			<td>McMaster Carr</td>
			<td>5231K161</td>
			<td>1/4&#34; ID</td>
		</tr>
		<tr>
			<td>Screws</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>SolidWorks Software</td>
			<td>Dassault Syst&#232;mes&#160;SolidWorks Corporation</td>
			<td></td>
			<td>https://www.solidworks.com/, Referred to in protocol as &#34;3D modeling software&#34;</td>
		</tr>
		<tr>
			<td>Straight Flow Rectangular Manifold</td>
			<td>McMaster Carr</td>
			<td>1125T31</td>
			<td></td>
		</tr>
		<tr>
			<td>Tubing to Flow Controller</td>
			<td>McMaster Carr</td>
			<td>5233K65</td>
			<td>3/8&#34; ID</td>
		</tr>
		<tr>
			<td>Wire</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

evaluating regional pulmonary deposition using patient-specific 3d printed lung models

肺病靶向疗法的发展受到可预测区域气溶胶输送能力的药物前检测方法的制约。利用 3D 打印生成患者专用肺模型，我们概述了用于量化球状肺沉积的高通量体外实验设置的设计。该系统由市售和 3D 打印组件组合而成，可独立控制肺部每个叶的流速。用荧光显微镜测量荧光气溶胶对每个叶的输送。该协议具有促进呼吸系统疾病个性化药物增长的潜力，因为它能够模拟广泛的患者人口统计学和疾病状态。3D 打印肺模型的几何形状和气流轮廓设置都可以轻松调整，以反映不同年龄、种族和性别患者的临床数据。临床相关的药物输送设备，如此处显示的内分泌管，可以纳入测试设置，以更准确地预测设备将治疗交付目标定向到肺部疾病区域的能力。这种实验设置的多功能性使其能够定制，以反映多种吸入条件，从而提高前结节治疗测试的严格性。

此大小范围（1-5 μm）和流速条件（1-10 L/min）的粒子根据其理论斯托克斯数和体内数据遵循流体流线;因此，在没有定向输送装置的情况下，释放到肺模型中的颗粒会根据流向每个叶的总气流百分比沉积。然后，通过分析患者特异性高分辨率计算机断层扫描 （HRCT） 扫描（HRCT）扫描获得的临床叶流率数据，可以比较每个叶的相对粒子输送量。经验证的实验设置将产生非靶向粒子沉...

Watch this Scientific Journal Video about Evaluating Regional Pulmonary Deposition using Patient-Specific 3D Printed Lung Models at JoVE.com

Evaluating Regional Pulmonary Deposition using Patient-Specific 3D Printed Lung Models

我们采用CT扫描衍生的3D打印肺模型和可调谐气流剖面，提出一种高通量体外方法，用于在叶位水平上量化区域肺沉积。

使用患者专用的 3D 打印肺模型评估区域肺沉积

Development of targeted therapies for pulmonary diseases is limited by the availability of preclinical testing methods with the ability to predict ...

evaluating-regional-pulmonary-deposition-using-patient-specific-3d

Research

JoVE Journal

Bioengineering

4.2K 次观看.  University of Delaware. 我们采用CT扫描衍生的3D打印肺模型和可调谐气流剖面，提出一种高通量体外方法，用于在叶位水平上量化区域肺沉积。

视频: Evaluating Regional Pulmonary Deposition using Patient-Specific 3D Printed Lung Models

Patient-specific Modeling of the Heart: Estimation of Ventricular Fiber Orientations

Patient-specific simulations of heart (dys)function aimed at personalizing cardiac therapy are hampered by the absence of in vivo imaging technology for clinically acquiring myocardial fiber orientations. The objective of this project was to develop a methodology to estimate cardiac fiber orientations from in vivo images of patient heart geometries. An accurate representation of ventricular geometry and fiber orientations was reconstructed, respectively, from high-resolution ex vivo structural magnetic resonance (MR) and diffusion tensor (DT) MR images of a normal human heart, referred to as the atlas. Ventricular geometry of a patient heart was extracted, via semiautomatic segmentation, from an in vivo computed tomography (CT) image. Using image transformation algorithms, the atlas ventricular geometry was deformed to match that of the patient. Finally, the deformation field was applied to the atlas fiber orientations to obtain an estimate of patient fiber orientations. The accuracy of the fiber estimates was assessed using six normal and three failing canine hearts. The mean absolute difference between inclination angles of acquired and estimated fiber orientations was 15.4 &deg;. Computational simulations of ventricular activation maps and pseudo-ECGs in sinus rhythm and ventricular tachycardia indicated that there are no significant differences between estimated and acquired fiber orientations at a clinically observable level.The new insights obtained from the project will pave the way for the development of patient-specific models of the heart that can aid physicians in personalized diagnosis and decisions regarding electrophysiological interventions.

A methodology to estimate ventricular fiber orientations from in vivo images of patient heart geometries for personalized modeling is described. Validation of the methodology performed using normal and failing canine hearts demonstrate that that there are no significant differences between estimated and acquired fiber orientations at a clinically observable level.

病人的具体建模之心:的心室纤维方向估计

Patient-specific  simulations of heart (dys)function aimed at personalizing cardiac therapy are hampered  by the absence of in vivo imaging technology for ...

科研

生物工程

Longitudinal Measurement of Extracellular Matrix Rigidity in 3D Tumor Models Using Particle-tracking Microrheology

The mechanical microenvironment has been shown to act as a crucial regulator of tumor growth behavior and signaling, which is itself remodeled and modified as part of a set of complex, two-way mechanosensitive interactions. While the development of biologically-relevant 3D tumor models have facilitated mechanistic studies on the impact of matrix rheology on tumor growth, the inverse problem of mapping changes in the mechanical environment induced by tumors remains challenging. Here, we describe the implementation of particle-tracking microrheology (PTM) in conjunction with 3D models of pancreatic cancer as part of a robust and viable approach for longitudinally monitoring physical changes in the tumor microenvironment, in situ. The methodology described here integrates a system of preparing in vitro 3D models embedded in a model extracellular matrix (ECM) scaffold of Type I collagen with fluorescently labeled probes uniformly distributed for position- and time-dependent microrheology measurements throughout the specimen. In vitro tumors are plated and probed in parallel conditions using multiwell imaging plates. Drawing on established methods, videos of tracer probe movements are transformed via the Generalized Stokes Einstein Relation (GSER) to report the complex frequency-dependent viscoelastic shear modulus, G*(&#969;). Because this approach is imaging-based, mechanical characterization is also mapped onto large transmitted-light spatial fields to simultaneously report qualitative changes in 3D tumor size and phenotype. Representative results showing contrasting mechanical response in sub-regions associated with localized invasion-induced matrix degradation as well as system calibration, validation data are presented. Undesirable outcomes from common experimental errors and troubleshooting of these issues are also presented. The 96-well 3D culture plating format implemented in this protocol is conducive to correlation of microrheology measurements with therapeutic screening assays or molecular imaging to gain new insights into impact of treatments or biochemical stimuli on the mechanical microenvironment.

Particle-tracking microrheology can be used to non-destructively quantify and spatially map changes in extracellular matrix mechanical properties in 3D tumor models.

细胞外基质刚性三维肿瘤模型的纵向测量用粒子跟踪Microrheology

The mechanical microenvironment has been shown to act as a crucial regulator of tumor growth behavior and signaling, which is itself remodeled and ...

Rapid and Low-cost Prototyping of Medical Devices Using 3D Printed Molds for Liquid Injection Molding

Biologically inert elastomers such as silicone are favorable materials for medical device fabrication, but forming and curing these elastomers using traditional liquid injection molding processes can be an expensive process due to tooling and equipment costs. As a result, it has traditionally been impractical to use liquid injection molding for low-cost, rapid prototyping applications. We have devised a method for rapid and low-cost production of liquid elastomer injection molded devices that utilizes fused deposition modeling 3D printers for mold design and a modified desiccator as an injection system. Low costs and rapid turnaround time in this technique lower the barrier to iteratively designing and prototyping complex elastomer devices. Furthermore, CAD models developed in this process can be later adapted for metal mold tooling design, enabling an easy transition to a traditional injection molding process. We have used this technique to manufacture intravaginal probes involving complex geometries, as well as overmolding over metal parts, using tools commonly available within an academic research laboratory. However, this technique can be easily adapted to create liquid injection molded devices for many other applications.

We have devised a method for low-cost and rapid prototyping of liquid elastomer rubber injection molded devices by using fused deposition modeling 3D printers for mold design and a modified desiccator as a liquid injection system.

医疗器械的使用3D印刷模具的液体注射成型快速，低成本原型

Biologically inert elastomers such as silicone are favorable materials for medical device fabrication, but forming and curing these elastomers using ...

Improving 2D and 3D Skin In Vitro Models Using Macromolecular Crowding

The glycoprotein family of collagens represents the main structural proteins in the human body, and are key components of biomaterials used in modern tissue engineering. A technical bottleneck is the deposition of collagen in vitro, as it is notoriously slow, resulting in sub-optimal formation of connective tissue and subsequent tissue cohesion, particularly in skin models. Here, we describe a method which involves the addition of differentially-sized sucrose co-polymers to skin cultures to generate macromolecular crowding (MMC), which results in a dramatic enhancement of collagen deposition. Particularly, dermal fibroblasts deposited a significant amount of collagen I/IV/VII and fibronectin under MMC in comparison to controls.
The protocol also describes a method to decellularize crowded cell layers, exposing significant amounts of extracellular matrix (ECM) which were retained on the culture surface as evidenced by immunocytochemistry. Total matrix mass and distribution pattern was studied using interference reflection microscopy. Interestingly, fibroblasts, keratinocytes and co-cultures produced cell-derived matrices (CDM) of varying composition and morphology. CDM could be used as &#34;bio-scaffolds&#34; for secondary cell seeding, where the current use of coatings or scaffolds, typically from xenogenic animal sources, can be avoided, thus moving towards more clinically relevant applications.
In addition, this protocol describes the application of MMC during the submerged phase of a 3D-organotypic skin co-culture model which was sufficient to enhance ECM deposition in the dermo-epidermal junction (DEJ), in particular, collagen VII, the major component of anchoring fibrils. Electron microscopy confirmed the presence of anchoring fibrils in cultures developed with MMC, as compared to controls. This is significant as anchoring fibrils tether the dermis to the epidermis, hence, having a pre-formed mature DEJ may benefit skin graft recipients in terms of graft stability and overall wound healing. Furthermore, culture time was condensed from 5 weeks to 3 weeks to obtain a mature construct, when using MMC, reducing costs.

Improving 2D and 3D Skin In Vitro Models Using Macromolecular Crowding

We present a protocol to obtain cell-derived matrices rich in extracellular matrix proteins, using macromolecular crowders (MMC). In addition, we present a protocol which incorporates MMC in 3D organotypic skin co-culture generation, which reduces culture time while maintaining maturity of construct.

提高2D和3D皮肤<em&gt;体外</em&gt;模型使用大分子拥挤

The glycoprotein family of collagens represents the main structural proteins in the human body, and are key components of biomaterials used in modern ...

Tunable Hydrogels from Pulmonary Extracellular Matrix for 3D Cell Culture

Here we present a method for establishing multiple component cell culture hydrogels for in vitro lung cell culture. Beginning with healthy en bloc lung tissue from porcine, rat, or mouse, the tissue is perfused and submerged in subsequent chemical detergents to remove the cellular debris. Histological comparison of the tissue before and after processing confirms removal of over 95% of double stranded DNA and alpha galactosidase staining suggests the majority of cellular debris is removed. After decellularization, the tissue is lyophilized and then cryomilled into a powder. The matrix powder is digested for 48 hr in an acidic pepsin digestion solution and then neutralized to form the pregel solution. Gelation of the pregel solution can be induced by incubation at 37 &#176;C and can be used immediately following neutralization or stored at 4 &#176;C for up to two weeks. Coatings can be formed using the pregel solution on a non-treated plate for cell attachment. Cells can be suspended in the pregel prior to self-assembly to achieve a 3D culture, plated on the surface of a formed gel from which the cells can migrate through the scaffold, or plated on the coatings. Alterations to the strategy presented can impact gelation temperature, strength, or protein fragment sizes. Beyond hydrogel formation, the hydrogel stiffness may be increased using genipin.

This is a method to create a 3-dimensional cell culture scaffold from pulmonary extracellular matrix. Intact lung is processed into hydrogels that can support the growth of cells in three-dimensions.

可调水凝胶由肺细胞外基质的三维细胞培养

Here we present a method for establishing multiple component cell culture hydrogels for in vitro lung cell culture. Beginning with healthy en bloc lung ...

Scaling of Engineered Vascular Grafts Using 3D Printed Guides and the Ring Stacking Method

Coronary artery disease remains a leading cause of death, affecting millions of Americans. With the lack of autologous vascular grafts available, engineered grafts offer great potential for patient treatment. However, engineered vascular grafts are generally not easily scalable, requiring manufacture of custom molds or polymer tubes in order to customize to different sizes, constituting a time-consuming and costly practice. Human arteries range in lumen diameter from about 2.0-38 mm and in wall thickness from about 0.5-2.5 mm. We have created a method, termed the "Ring Stacking Method," in which variable size rings of tissue of the desired cell type, demonstrated here with vascular smooth muscle cells (SMCs), can be created using guides of center posts to control lumen diameter and outer shells to dictate vessel wall thickness. These tissue rings are then stacked to create a tubular construct, mimicking the natural form of a blood vessel. The vessel length can be tailored by simply stacking the number of rings required to constitute the length needed. With our technique, tissues of tubular forms, similar to a blood vessel, can be readily manufactured in a variety of dimensions and lengths to meet the needs of the clinic and patient.

Scalable engineered blood vessels would improve clinical applicability. Using easily sizable 3D-printed guides, rings of vascular smooth muscle were created and stacked into a tubular form, forming a vascular graft. Grafts can be sized to meet the range of human coronary artery dimensions by simply changing the 3D-printed guide size.

工程血管移植缩放使用3D打印指南和环堆叠方法

Coronary artery disease remains a leading cause of death, affecting millions of Americans. With the lack of autologous vascular grafts available, ...

Scaled Anatomical Model Creation of Biomedical Tomographic Imaging Data and Associated Labels for Subsequent Sub-surface Laser Engraving (SSLE) of Glass Crystals

Biomedical imaging modalities like computed tomography (CT) and magnetic resonance (MR) provide excellent platforms for collecting three-dimensional data sets of patient or specimen anatomy in clinical or preclinical settings. However, the use of a virtual, on-screen display limits the ability of these tomographic images to fully convey the anatomical information embedded within. One solution is to interface a biomedical imaging data set with 3D printing technology to generate a physical replica. Here we detail a complementary method to visualize tomographic imaging data with a hand-held model: Sub Surface Laser Engraving (SSLE) of crystal glass. SSLE offers several unique benefits including: the facile ability to include anatomical labels, as well as a scale bar; streamlined multipart assembly of complex structures in one medium; high resolution in the X, Y, and Z planes; and semi-transparent shells for visualization of internal anatomical substructures. Here we demonstrate the process of SSLE with CT data sets derived from pre-clinical and clinical sources. This protocol will serve as a powerful and inexpensive new tool with which to visualize complex anatomical structures for scientists and students in a number of educational and research settings.

Scaled Anatomical Model Creation of Biomedical Tomographic Imaging Data and Associated Labels for Subsequent Sub-surface Laser Engraving SSLE of Glass Crystals

A methodology is described herein for representing anatomical imaging data within crystals. We create scaled three-dimensional models of biomedical imaging data for use in Sub-Surface Laser Engraving (SSLE) of crystal glass. This tool offers a useful complement to computational display or three-dimensionally printed models used within clinical or educational settings.

缩放生物医学层析成像数据的解剖模型的创建和相关联的标签玻璃晶体的随后的子表面激光雕刻（SSLE）

Biomedical imaging modalities like computed tomography (CT) and magnetic resonance (MR) provide excellent platforms for collecting three-dimensional data ...

Fabrication of Custom Agarose Wells for Cell Seeding and Tissue Ring Self-assembly Using 3D-Printed Molds

Engineered tissues are being used clinically for tissue repair and replacement, and are being developed as tools for drug screening and human disease modeling. Self-assembled tissues offer advantages over scaffold-based tissue engineering, such as enhanced matrix deposition, strength, and function. However, there are few available methods for fabricating 3D tissues without seeding cells on or within a supporting scaffold. Previously, we developed a system for fabricating self-assembled tissue rings by seeding cells into non-adhesive agarose wells. A polydimethylsiloxane (PDMS) negative was first cast in a machined polycarbonate mold, and then agarose was gelled in the PDMS negative to create ring-shaped cell seeding wells. However, the versatility of this approach was limited by the resolution of the tools available for machining the polycarbonate mold. Here, we demonstrate that 3D-printed plastic can be used as an alternative to machined polycarbonate for fabricating PDMS negatives. The 3D-printed mold and revised mold design is simpler to use, inexpensive to produce, and requires significantly less agarose and PDMS per cell seeding well. We have demonstrated that the resulting agarose wells can be used to create self-assembled tissue rings with customized diameters from a variety of different cell types. Rings can then be used for mechanical, functional, and histological analysis, or for fabricating larger and more complex tubular tissues.

This protocol describes a platform for fabricating self-assembled tissue rings in variable sizes using a customized 3D-printed plastic mold. PDMS negatives are cured in the 3D-printed mold; then agarose is cast in the cured PDMS negatives. Cells are seeded into the resulting agarose wells where they aggregate into tissue rings.

用 3 d-印刷模具制作细胞播种和组织环自组装用的自定义琼脂糖井

Engineered tissues are being used clinically for tissue repair and replacement, and are being developed as tools for drug screening and human disease ...

Targeting Neuronal Fiber Tracts for Deep Brain Stimulation Therapy Using Interactive, Patient-Specific Models

Deep brain stimulation (DBS), which involves insertion of an electrode to deliver stimulation to a localized brain region, is an established therapy for movement disorders and is being applied to a growing number of disorders. Computational modeling has been successfully used to predict the clinical effects of DBS; however, there is a need for novel modeling techniques to keep pace with the growing complexity of DBS devices. These models also need to generate predictions quickly and accurately. The goal of this project is to develop an image processing pipeline to incorporate structural magnetic resonance imaging (MRI) and diffusion weighted imaging (DWI) into an interactive, patient specific model to simulate the effects of DBS. A virtual DBS lead can be placed inside of the patient model, along with active contacts and stimulation settings, where changes in lead position or orientation generate a new finite element mesh and solution of the bioelectric field problem in near real-time, a timespan of approximately 10 seconds. This system also enables the simulation of multiple leads in close proximity to allow for current steering by varying anodes and cathodes on different leads. The techniques presented in this paper reduce the burden of generating and using computational models while providing meaningful feedback about the effects of electrode position, electrode design, and stimulation configurations to researchers or clinicians who may not be modeling experts.

The goal of this project is to develop an interactive, patient-specific modeling pipeline to simulate the effects of deep brain stimulation in near real-time and provide meaningful feedback as to how these devices influence neural activity in the brain.

应用交互式、患者特异性模型对神经纤维束进行深部脑刺激治疗的靶向性

Deep brain stimulation (DBS), which involves insertion of an electrode to deliver stimulation to a localized brain region, is an established therapy for ...

High-resolution Patterned Biofilm Deposition Using pDawn-Ag43

Spatial structure and patterning play an important role in bacterial biofilms. Here we demonstrate an accessible method for culturing E. coli biofilms into arbitrary spatial patterns at high spatial resolution. The technique uses a genetically encoded optogenetic construct&#8212;pDawn-Ag43&#8212;that couples biofilm formation in E. coli to optical stimulation by blue light. We detail the process for transforming E. coli with&#160;pDawn-Ag43, preparing the required optical set-up, and the protocol for culturing patterned biofilms using pDawn-Ag43 bacteria. Using this protocol, biofilms with a spatial resolution below 25 &#956;m can be patterned on various surfaces and environments, including enclosed chambers, without requiring microfabrication, clean-room facilities, or surface pretreatment. The technique is convenient and appropriate for use in applications that investigate the effect of biofilm structure, providing tunable control over biofilm patterning. More broadly, it also has potential applications in biomaterials, education, and bio-art.

We demonstrate a method for depositing Escherichia coli bacterial biofilms in arbitrary spatial patterns with a high resolution using optical stimulation of a genetically encoded surface-adhesion construct.

使用 pDawn-Ag43 的高分辨率图案生物膜沉积

Spatial structure and patterning play an important role in bacterial biofilms. Here we demonstrate an accessible method for culturing E. coli biofilms ...