This research was supported by the Technology Innovation Program (Contract grant No. 0037915, WPM Biomedical Materials-Implant Materials) and Industrial Strategic Technology Development Program (Contract grant No. 10045329, Development of customized implant with porous structure for bone replacement), funded by the Ministry of Trade, industry &#38; Energy (MI, Korea), and BK21 PLUS SNU Materials Division for Educating Creative Global Leaders (Contract grant No. 21A20131912052).

1.制备多孔金属支架的<ol><li>通过混合市售的Ti粉末，莰烯，和KD-4 如表1中描述的多孔钛支架具有四个初始孔隙率（40，50，60，和70）称量材料的适当的量后制备的Ti-莰泥浆。倾浆料到500ml聚乙烯（PE）瓶和旋转瓶在55℃下30分钟在球磨机烘箱在30rpm。 </li><li>倾从PE瓶浆料成圆柱形的铝（Al）的模具，其直径为60毫米和60毫米的高度。密封每个铝模具与相应的A1盖玻片并旋转在球磨机烘箱模具以30rpm的速度在55℃下进行10分钟。 <ol><li>随后，降低球磨机烘箱的温度升高到44℃，并连续旋转模具以30rpm的速度在44℃进行12小时的恒温。 </li></ol></li><li>取出模具从球 -磨炉之后附加地旋转所述模具在RT 1小时的冷却过程。从使用Al柱塞铝模具中取出固化的钛/莰生坯。 </li><li>将固化的钛/莰生坯在一橡胶袋通过手和完全由捆扎袋的口用字符串密封橡胶袋。将橡胶袋在冷等静压（CIP），机的水槽中，并应用​​为200MPa的等静压力10分钟。从橡胶袋中取出压缩坯体。 </li><li>上手工传送的Ti-莰生坯氧化铝坩埚放置在坩埚中的冷冻干燥机。冷冻干燥该生坯升华莰相在生坯在 - 40℃下放置24小时。 </li><li>接着，关闭该坩埚有氧化铝盖玻片并将封闭坩埚在真空炉中（低于10 -6托 ）于RT。然后，增加了炉的温度升高到1300℃，在一个加热ř吃了5℃/分钟并保持温度在1300℃进行2小时。 </li><li>热处理后，保持烧结多孔钛在炉为6-7小时，直至炉完全冷却至室温。 注意：在6小时的冷却过程中，400℃以上的熔炉的平均冷却速度为〜15℃/分钟和400℃以下的熔炉的平均冷却速度为2〜℃/分钟。 </li><li>如果需要的话，切割烧结的多孔钛的块到盘形样品，直径为通过放电加工16个毫米（EDM）。27 注意：根据所述Al模具的大小，需要通过加工过程（图2A）进行修改的烧结多孔的Ti的大小。 </li><li>放置在高压釜中的多孔钛样品的玻璃烧杯中，并灭菌样品在121℃下进行15分钟。从高压釜中取出样品。洗多孔钛样品用蒸馏水洗涤两次，然后用70％乙醇两次。最后，离开多孔钛到培养皿并空气干燥在RT样品上在UV光下净化台。 </li></ol>支架的2浸涂与生物活性剂<ol><li>通过混合1毫升的GFP用9ml的Dulbecco磷酸盐缓冲盐水（DPBS，pH7.4）中的溶液在10毫升灭菌的从1毫克/毫升，以在清洁台上100微克/毫升稀释商用绿色荧光蛋白（GFP）的聚苯乙烯（PS）管，如表1所示。 </li><li>通过将钛样品到PS管在RT将GFP溶液和放置在净化台浸没灭菌致密或多孔钛在10毫升稀释的GFP溶液（100微克/毫升）。 </li><li>放置在PS管在真空干燥器并撤离干燥器10分钟，以确保将GFP溶液更有效地渗入多孔钛的孔中。 </li><li>除去使用镊子将PS管上的多孔钛。将GFP-涂覆的多孔钛成直径为10cm的聚乙烯三道菜和风干O / N在室温在一个干净的长椅上。 </li><li>用在玻璃烧杯中10个​​ml的Dulbecco氏磷酸盐缓冲盐水（DPBS）中冲洗多孔钛两次，并使用镊子和风干移动多孔钛成直径为10cm的培养皿在RT上的净化台。 </li></ol> 3.致密多孔支架的<ol><li>放置在一个圆柱形钢模具的GFP-涂覆的多孔钛的样品不同的高度，并插入了一组冲头进入钢模具（图3A）的顶部和底部的孔。 </li><li>用压制机以0.05〜0.1秒的中间应变率压缩所述多孔钛在RT中的样品（图3A）的z方向上的钢模具组件内-1针对在表2所示的预定的施加应变。握持压力卸货前1分钟。 </li><li>从钢模具去除致密的Ti样品。用10毫升的DPBS洗涤两次致密样品在一个干净的长椅的烧杯中，空气干燥O / N在室温。 </li></ol>绿色荧光蛋白涂层支架4释放测试<ol><li>沉浸三种试样（GFP涂覆致密的Ti（后步2），绿色荧光蛋白涂覆的多孔钛（后步骤1和2）和GFP-涂覆致密多孔的Ti（后步骤1-3））的5ml DPBS（pH值7.4）在37℃下在清洁台上所载10ml的灭菌的PS管溶液。 </li><li>吸出所有来自每个PS管的DPBS溶液与GFP-涂覆的样品和补充与使用根据预定的次数的1，2，3，5，8，12的吸移管新5 ml的DPBS溶液（pH 7.4），浸泡后15，22和29天。 </li><li>取GFP涂覆样品的荧光图像浸渍（第0天）前，用共聚焦激光扫描光谱（CLSM）后22天，浸泡。 </li><li>测量所释放的GFP在1ml溶液中的荧光信号强度从总共5毫升DPBS溶液从每个试管的PS在节4.2绘制使用紫外光谱在215纳米的波长。的强度值转换成利用标准曲线将GFP溶液的浓度。 注意：在测量之前，通过测量在0纳克/毫升的浓度范围将GFP溶液的荧光信号强度绘制的GFP溶液的标准曲线 - 10微克/毫升。 </li></ol> 5.制造分级多孔钛支架的<ol><li>通过重复步骤1.1至步骤1.7产生烧结多孔Ti的块。 </li><li>机根据预定结构设计 （如， 图5a和 5d）通过电火花烧结多孔的Ti块。 </li><li>放置加工的Ti样品高度分布在一个钢模其中多孔钛的直径为〜小于模具的直径0.1毫米小，并插入了一组冲头进入钢模具的顶部和底部的孔。 </li><li>执行步骤3.2和3.3。 </li></ol> 6.孔隙率我钛支架的asurement <ol><li>衡量的Ti支架的质量（m 多个）。 </li><li>计算的表观容积（V S），通过测量钛支架长度，宽度和高度。 </li><li>使用下列公式计算孔隙率： <img alt="式（1）" src="/files/ftp_upload/53279/53279eq1.jpg" /> 其中P是总孔隙率，ρTi为钛和米S的理论密度/ V S是样品的测量的密度。 注意：钛样品的孔隙率可以从显微图像来直接检索后显微成像是使用微计算机断层扫描仪。 </li></ol>

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The authors declare that they have no competing financial interests.

而biometal系统已被广泛地用于生物医学应用，特别是，作为承重材料，高刚度和金属的低生物活性已被视为重大的挑战。在这项研究中，我们建立了一个新的金属系统，致密多孔金属支架具有仿生机械性能以及生物活性表面与可持续释放行为的制造方法。我们的制作方法的主要优点包括：1）在前面的动态冷冻的铸造方法没有变化，我们已经开发，28 2）中的一个参数度的控制致密化-实现机...

而金属生物材料已被广泛用作由于其优异的机械强度和韧性，1-3承重植入物和内固定装置，他们涉及两个关键的挑战：1）机械失配，因为金属是比生物体组织硬得多，从而导致不希望的损害到周围组织和2）低的生物活性，往往导致界面差与生物组织，往往挑起异物反应 （如炎症或血栓形成）。已经提出了促进骨向内生长的结构4-6多孔金属支架，改善骨-植入物接触，而应力屏蔽效果，因为它们降低的刚性抑制7-9此外，各种表面改性已经应用于提高金属植入物的生物活性;这些修改包括涂层将金属表面与生物活性分子（例如，生长的fac器）或药物 （如万古霉素，四环素）10-12但是，存在的问题，如多孔金属支架降低机械性能，降低的硬度和快速释放的生物活性涂层的层仍然没有得到解决。13-16 特别是，钛（Ti）和Ti的合金是一体的，因为它们的优异的机械性能，化学稳定性的最流行​​biometal系统，以及良好的生物相容性。13,17-19其泡沫形的应用也引起越来越多的关注，因为在3D多孔网络促进除了骨样的机械性能的骨向内生长。20-22已作出努力通过开发新的制造技术，包括聚合海绵的复制，金属粒子，快速成型（RP）的方法的烧结来提高机械性能，并为了控制孔的各种特征空间保持器的方法（例如，孔隙分数，形状，大小，分布，和连接）和材料性质（例如，金属相和杂质）23-25 ​​最近，水性金属浆料的冻结铸造已经获得了相当大的注意，以产生机械增强的Ti形式具有良好对准孔结构通过利用在凝固过程中的单向冰枝晶生长;然而，所造成的金属粉末与水接触氧污染需要特别小心，以尽量减少钛支架的脆化。14,15 因此，我们开发了对制造生物活性和机械可调多孔钛支架的新方法。25支架最初有超过50％的孔隙率多孔结构。所制造的多孔支架涂覆有生物活性分子，然后进行压缩使用期间的最后的孔隙率，机械性能和药物释放行为由APPLI分别控制一个机械压力机编辑应变。致密多孔钛种植体，尽管低刚度比得上骨（3-20 GPA）2由于被覆层的一者所示低孔隙率具有良好的强度，致密多孔钛的生物活性显著改善。因为引起的致密化过程中的独特扁平孔结构。此外，涂布的生物活性分子被视为被逐渐从支架释放时，保持其功效为长时间。 在这项研究中，我们介绍了我们建立的方法来制作致密多孔钛支架的生物医学应用的潜在用途。该协议包括动态凝固的铸造​​金属浆料和多孔支架的致密化。首先，为了制造多孔钛支架具有良好的延展性的动态冷冻铸造方法被引入，如图1A。钛粉末分散在液体莰;然后，通过降低温度，液相固化，从而在钛粉末网络和固体莰晶体之间的相分离。接着，将固化的Ti-莰生坯进行烧结，其中的Ti粉末缩合的连续钛支柱，和莰相完全除去，得到多孔结构。该涂层和致密化过程用所获得的多孔支架受雇于，变化的致密化和初始孔隙的程度。被覆层和它的释放行为进行观察和使用绿色荧光蛋白（GFP）包被的多孔钛具有和不致密化相比，GFP-涂覆致密的Ti定量。最后，提出并通过改变多孔支架的内层和外层部分的致密化的程度表现出功能梯度的Ti支架具有两个不同的多孔结构。

<table>
	<tbody>
		<tr>
			<td>Titanium powder</td>
			<td>Alfa Aesar</td>
			<td>#42624</td>
			<td>-325 mesh, 99.5% (metals basis)</td>
		</tr>
		<tr>
			<td>Camphene</td>
			<td>SigmaAldrich</td>
			<td>#456055</td>
			<td>95%, C10H16</td>
		</tr>
		<tr>
			<td>KD-4</td>
			<td>Croda</td>
			<td>&#173;</td>
			<td>Hypermer, polymeric dispersant</td>
		</tr>
		<tr>
			<td>Phosphate Buffer Solution (PBS)</td>
			<td>Welgene</td>
			<td>ML 008-01</td>
			<td>&#173;</td>
		</tr>
		<tr>
			<td>Green Fluorescent Protein (GFP)</td>
			<td>Genoss Co.</td>
			<td>-</td>
			<td>&#62;98% purity, 1mg/ml</td>
		</tr>
		<tr>
			<td>Ball mill oven</td>
			<td>SAMHENUG ENERGY</td>
			<td>SH-BDO150</td>
			<td>&#173;</td>
		</tr>
		<tr>
			<td>Freeze dryer</td>
			<td>Ilshin Lab.</td>
			<td>PVTFD50A</td>
			<td>&#173;</td>
		</tr>
		<tr>
			<td>Cold isostatic pressing (CIP) machine</td>
			<td>SONGWON SYSTEMS</td>
			<td>CIP 42260</td>
			<td>&#173;</td>
		</tr>
		<tr>
			<td>Vaccum furnace</td>
			<td>JEONG MIN INDUSTRIAL</td>
			<td>JM-HP20</td>
			<td>&#173;</td>
		</tr>
		<tr>
			<td>electical chaege machine</td>
			<td>FANUC robocut</td>
			<td>0iB</td>
			<td>External use</td>
		</tr>
		<tr>
			<td>Press machine</td>
			<td>CG&#38;S</td>
			<td>AJP-200</td>
			<td>&#173;</td>
		</tr>
		<tr>
			<td>Confocal laser scanning spectroscopy (CLSM)</td>
			<td>Olympus</td>
			<td>FluoView FV1000</td>
			<td>External use</td>
		</tr>
	</tbody>
</table>

fabrication of mechanically tunable and bioactive metal scaffolds for biomedical applications

Biometal systems have been widely used for biomedical applications, in particular, as load-bearing materials. However, major challenges are high stiffness and low bioactivity of metals. In this study, we have developed a new method towards fabricating a new type of bioactive and mechanically reliable porous metal scaffolds-densified porous Ti scaffolds. The method consists of two fabrication processes, 1) the fabrication of porous Ti scaffolds by dynamic freeze casting, and 2) coating and densification of the porous scaffolds. The dynamic freeze casting method to fabricate porous Ti scaffolds allowed the densification of porous scaffolds by minimizing the chemical contamination and structural defects. The densification process is distinctive for three reasons. First, the densification process is simple, because it requires a control of only one parameter (degree of densification). Second, it is effective, as it achieves mechanical enhancement and sustainable release of biomolecules from porous scaffolds. Third, it has broad applications, as it is also applicable to the fabrication of functionally graded porous scaffolds by spatially varied strain during densification.

用于生产多孔钛的支架的制造方法示于图1A。钛粉末被保持在莰均匀分散由容器的连续旋转在44℃下12小时和，而液体莰被完全固化，相对较重的Ti粉末的任何沉积物最小化。其结果是，将均匀的Ti-莰生坯使用动态冷冻浇铸方法生产，如图1B所示，在其中三维地相互连接的大莰孔隙由钛粉末相（图1C）所包围。然而，不正确的容器的转动...

Watch this Scientific Journal Video about Fabrication of Mechanically Tunable and Bioactive Metal Scaffolds for Biomedical Applications at JoVE.com

Fabrication of Mechanically Tunable and Bioactive Metal Scaffolds for Biomedical Applications

Bioactive and mechanically reliable metal scaffolds have been fabricated through a method which consists of two processes, dynamic freeze casting for the fabrication of porous Ti, and coating and densification of the Ti scaffolds. The densification process is simple, effective and applicable to the fabrication of functionally graded scaffolds.

加工机械可调和生物活性金属支架的生物医学应用

Biometal systems have been widely used for biomedical applications, in particular, as load-bearing materials. However, major challenges are high stiffness ...

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10.6K 次观看.  Korea Institute of Industrial Technology. Bioactive and mechanically reliable metal scaffolds have been fabricated through a method which consists of two processes, dynamic freeze casting for the fabrication of porous Ti, and coating and densification of the Ti scaffolds. The densification process is simple, effective and applicable to the fabrication of functionally graded scaffolds.