<Emphasis Type="Italic">Development of a Tissue Engineering Scaffold Structure Library for Rapid Prototyping. Part 1: Investigation and Classification</Emphasis>

In tissue engineering (TE), a porous scaffold structure may be required as a template to guide the proliferation, growth and development of cells appropriately in three dimensions. Although TE scaffolds can be created using one of many conventional techniques available, most will suffer from a lack of mechanical strength and/or uniformity in pore distribution and sizes. This study is focused on creating scaffolds using rapid prototyping (RP) techniques. Utilising these novel techniques, a computer-aided design (CAD) of the scaffold structure must first be modelled. The scaffold structure is then fabricated directly from CAD data using a RP system. The objective of this research is to (1) investigate and select various polyhedral shapes suitable for scaffold modelling, (2) classify the selected unit cells, (3) create a parametric library of scaffold structures and (4) verify by building the CAD models using the selective laser sintering process. The first two objectives are covered in Part 1 of this two-part paper. The remaining objectives will be described and discussed in Part 2. ID="A1"Correspondance and offprint requests to: Dr C. K. Chua, School of Mechanical and Production Engineering, Nanyang Technological University, 50 Nanyang Avenue, Singapore 639798. E-mail: mckchua@ntu.edu.sg     

Improvement of the representative volume element method for 3-D scaffold simulation

Predicting the mechanical properties of the 3-D scaffold using finite element method (FEM) simulation is important to the practical application of tissue engineering. However, the porous structure of the scaffold complicates computer simulations, and calculating scaffold models at the pore level is time-consuming. In some cases, the demands of the procedure are too high for a computer to run the standard code. To address this problem, the representative volume element (RVE) theory was introduced, but studies on RVE modeling applied to the 3-D scaffold model have not been focused. In this paper, we propose an improved FEM-based RVE modeling strategy to better predict the mechanical properties of the scaffold prior to fabrication. To improve the precision of RVE modeling, we evaluated various RVE models of newly designed 3-D scaffolds using FEM simulation. The scaffolds were then constructed using microstereolithography technology, and their mechanical properties were measured for comparison.     

Skin regeneration stimulation: the role of PCL‐platelet gel nanofibrous scaffold

Skin is the largest organ of the human body. Thus far, tissue engineering of skin has developed rapidly and has used many types of growth factors and nanofibrous scaffolds. In this study, we differentiated neonate keratinocytes for epithelialization on the polycaprolactone‐Platelet gel (PCL‐PG) scaffold. Fabricated PCL nanofibers prepared by electrospinning technology and coated by platelet gel. Subsequently, the structure of the scaffold was evaluated by SEM, FTIR‐ATR, contact angle and tensile test assays. After seeding the neonate keratinocytes on neat PCL and PCL‐PG scaffolds, the epidermal maturation was tested by detecting cytokeratin 10 and loricrin determinants by immunocytochemistry; moreover, keratinocyte genes such as keratin 14, keratin 10, and Involucrin were investigated by real‐time PCR. The results of MTT assay indicated an increase in cell viability and cell proliferation of neonate keratinocytes on PCL‐PG nanofiber scaffolds compared with PCL. RT‐PCR and immunocytochemical analysis showed better cell differentiation on the PCL‐PG scaffolds than neat PCL. Furthermore, SEM microscopy images demonstrated that neo‐keratinocytes enhance adhesion and proliferation on PCL‐PG nanofiber scaffolds. We found that PG increases biocompatibility and wettability of scaffold, cell adhesion, and expression of keratinocyte markers. Overall, this procedure is recommended to be employed in skin tissue engineering and wounds healing.     

<Emphasis Type="Italic">Development of a Tissue Engineering Scaffold Structure Library for Rapid Prototyping. Part 2: Parametric Library and Assembly Program</Emphasis>

Rapid prototyping (RP) techniques have been found to be advantageous for tissue engineering (TE) scaffold fabrication due to their ability to address and overcome the problems of uncontrollable microstructure and the feasibility issues of complex three-dimensional structures found in conventional processing techniques. This research proposes a novel approach for TE scaffold manufacture using RP techniques. The approach involves the integration of medical imaging devices (CT/MRI) for the acquisition of anatomic structural data, three-dimensional CAD modelling for designing and creating the digital scaffold models and RP for fabricating the physical scaffolds. To aid the user in CAD modelling, a standard parametric library of scaffold structures is designed and developed. With the library, a user can select the geometry of the scaffold unit cell and size it to suit the end application of the TE scaffold. A developed application program will then assemble the scaffold structure from the selected unit cell, following the surface profile of the anatomic structure to be replicated. A physical scaffold will then be built using an RP system. ID="A1"Correspondance and offprint requests to: Dr C. K. Chua, School of Mechanical and Production Engineering, Nanyang Technological University, 50 Nanyang Avenue, Singapore 639798. E-mail: mckchua@ntu.edu.sg     

The design of a semi-additive manufacturing shape using metal 3D printing for a partially strengthened mold based on a high-alloy tool steel powder

In this paper, describe the fabrication of high strength punch molds that can be applied to ultra-high strength sheet materials after processing. A method for improving the strength of the punching die by additive manufacturing (AM) of a high strength powder material using a metal 3D printer was proposed. Furthermore, a semi-additive technique was proposed to increase the punch strength through partial AM of specific parts of the punch that require high strength. A preprocessing process for predicting the semi-additive shape for the punch function portion is proposed for application of the AM technology of a metal 3D printer to this semi-additive technique. The preprocessing for determining the semi-additive shape consists of the predicting step of the punch strength based on the shear process of the sheet material, analyzing step the stress distribution of the punch, defining step the semi-additive range, designing step the semi-additive shape, and verifying step the additive interface strength. Based on this simulation, the range of shapes for the semi-additive was 1.21 mm and 2.62 mm for sheet material CP1180 and 1.3 mm and 3.2 mm for sheet material 22MnB5. The shape and range determined in the simulation process defines a semi-additive area (volume) for the 3D printing AM technique using a high-strength powder material, and a semi-additive punch was manufactured according to the defined area. The semi-additive punch (HWS powder material) fabricated in this study was performed a durability test for validity verification in the piercing process of high-strength sheet material (CR980). This validation test compared the state of the punch after 1000 piercing processes with a typical cold piercing punch (SKD11 solid material). From this test, the feasibility of the semi-additive punch was confirmed by showing a similar state of scratches and abrasion from the two punches. The simulation analysis processor for the additive shape and the additive range prediction for the semi-additive punch manufacturing presented in this paper can be useful for the additive manufacture of cutting and trimming punch mold.

     

Development of in-process visualization system for laser-assisted three-dimensional microfabrication using photocatalyst nanoparticles

Recently, microfabrication technology has been used to develop micro-electro-mechanical systems (MEMSs), micro-total analysis systems (μ-TASs), and photonic crystals. Various microfabrication techniques have been proposed; however, a technique that can be used to efficiently fabricate 3-D structures via a simple procedure has not been reported thus far. Because 3-D metal structures have not only mechanical functions but also electromagnetic functions, it is desirable to develop such a technique. Our research group is in the process of developing a new technique for 3-D microfabrication that involves the use of a lower power continuous wave laser. Our technique is characterized by the reduction of silver ions via the photocatalysis of titanium dioxide (TiO2) excited at the laser beam waist. For the analysis and development of our microfabrication technique, we developed a microscope system that enabled us to observe the microfabrication process along the fabrication beam optical axis and its radial direction. We successfully visualized the microfabrication process in 3-D. The visualization showed that when the beam waist was swept, the silver structure grew in 3-D following its path. The effect of the substrate on the deposition condition was examined.     

基于增量制造的再生骨支架制备路径规划方法

基于增量制造的再生骨支架制备方法在生物制造领域表现出巨大应用潜力,也越来越受到重视,其中制造路径规划方法直接影响到制备的支架的结构特点,在很大程度上决定了后期支架的细胞培养以及组织修复效果。目前典型的面向工业以及民用领域的增量制造工艺的路径规划方法不能完全适用于骨支架的制备,无法获得能够有效模拟细胞外基质结构特征的三维贯通结构。针对这一问题,提出一种面向生物增量挤出成形技术的再生骨支架制备路径规划方法,将材料成形过程中出现的尺寸变形的补偿融入到路径规划方法的设计中,能有效满足骨支架制备时对复杂外轮廓和梯度孔隙结构的需求。在此基础上,给出再生骨支架增量挤出成形系统的软件系统架构,实现方法及结果,并通过支架成形试验验证支架的性能,研究成果对自主开发生物增量制造装备具有重要价值和意义。     

Comparing different methods to fix and to dehydrate cells on alginate hydrogel scaffolds using scanning electron microscopy

Scanning electron microscopy (SEM) is commonly used in the analysis of scaffolds morphology, as well as cell attachment, morphology and spreading on to the scaffolds. However, so far a specific methodology to prepare the alginate hydrogel (AH) scaffolds for SEM analysis has not been evaluated. This study compared different methods to fix/dehydrate cells in AH scaffolds for SEM analysis. AH scaffolds were prepared and seeded with NIH/3T3 cell line; fixed with glutaraldehyde, osmium tetroxide, or the freeze drying method and analyzed by SEM. Results demonstrated that the freeze dried method interferes less with cell morphology and density, and preserves the scaffolds structure. The fixation with glutaraldehyde did not affect cells morphology and density; however, the scaffolds morphology was affected in some level. The fixation with osmium tetroxide interfered in the natural structure of cells and scaffold. In conclusion the freeze drying and glutaraldehyde are suitable methods for cell fixation in AH scaffold for SEM, although scaffolds structure seems to be affected by glutaraldehyde. Microsc. Res. Tech. 78:553–561, 2015. © 2015 Wiley Periodicals, Inc.     

二维直方图准分的Tsallis熵阈值分割及其快速实现

传统二维Tsallis熵阈值法主要由于对二维直方图采用近似假设等原因,导致分割结果不够准确,由此提出了基于二维直方图准分的Tsallis熵快速图像分割方法.首先,准确选择邻域模板构建二维直方图并将Tsallis熵法用于此直方图上以便提高分割性能;然后,舍弃二维直方图中关于主对角区域的概率和近似为1的假设而准确计算使阈值选取更准确;最后,结合Tsallis熵公式对二维直方图进行分析得到其特性和2个定理,利用此特性和2个定理导出新型、快速的递推算法来降低计算复杂度.实验结果表明,与传统二维Tsallis熵法相比,所提出的方法不仅分割更准确和抗噪性更强,而且占用的存储空间和运行时间都更少.     

Structural evaluation of scaffolds prototypes produced by three-dimensional printing

The fabrication of porous scaffolds with complex architectures represents a challenge in tissue engineering. Recent studies have shown that it is possible to construct tissue-engineered bone repair scaffolds with tight pore size distributions and controlled geometries using 3D printing techniques. In this context, this work aims to evaluate the 3D printing process in order to study its potential for scaffold fabrication. Despite the wide use of porous scaffolds, its design, geometry optimization and mechanical assessment, for successful integration in tissue engineering, require further developments and studies to help in its optimal design. In the present work, cubical scaffolds prototypes with different architectures were produced by 3D printing technology. Scaffolds dimensional accuracy, porosity and mechanical stiffness were comprehensive analysed by means of an experimental investigation. The microporosity, inherent to the fabrication process, and the mechanical characterization of the bulk material were also considered. This paper addresses methodologies to overcome some limitations of 3D printing technique to produce scaffolds for tissue engineering and proposes procedures for their suitable mechanical characterization. Results of this work indicate that 3D printing process has great potential for scaffold fabrication.     

Development of composite porous scaffolds based on poly(lactide-co-glycolide)/nano-hydroxyapatite via selective laser sintering

Poly(lactide-co-glycolide) (PLGA)/nano-hydroxyapatite (nano-HAP) composite porous scaffolds with well-controlled pore architectures as well as high exposure of the bioactive ceramics to the scaffold surface were fabricated via selective laser sintering. Neat PLGA and the composite of PLGA/nano-HAP were used to obtain suitable process parameters. The effects of nano-HAP content on the microstructure and mechanical properties were investigated. The testing results showed that the compressive strength and modulus of the scaffolds were highly enhanced when the nano-HAP content reached from 0 to 20 wt%, while the mechanical properties experienced a sharp dropped with the nano-HAP content further increased. This might be due to the large reduction in polymer which decreased the interface bond strength between particles. It suggests that the introduction of nano-HAP as a reinforcing phase can improve the mechanical properties of the polymer porous scaffolds. The novel developed scaffolds may serve as a three-dimensional bone substrate in tissue engineering.     

可控微结构支架光固化快速成形间接构造方法

提出一种以计算机辅助设计(CAD)和光固化快速成形为基础,精确设计和控制支架内部微观结构,实现支架外形与内部可控微结构一体化制造的方法。根据实际CT数据,应用三维CAD软件重构骨骼外形;依据利于细胞生长和促进成骨的原则,设计不同结构的支架内部微管道,控制微管道的尺寸、形状、走向、分支以及相互连通性。利用光固化快速成形技术构造相应的树脂模具,在模具中填充生物材料,待其固化后,通过热分解去除树脂模具,形成具有可控微结构的支架。光学显微镜下观察支架内部微管道结构,其结果与设计相符合。与传统支架构造方法相比,该方法在对支架外形重构和内部微结构制造的控制方面得到了改善。     

Mc3D: A three-dimensional Monte Carlo system simulating image contrast in surface analytical scanning electron microscopy I—Object-oriented software design and tests

The main features of a three-dimensional (3-D) Monte Carlo software system (Mc3D), designed for the simulation of electron scattering and image contrast in a scanning electron microscope, are reported. Before simulating electron trajectories in the sample, impingement of the incident electron beam is described by introducing the idea of a virtual scan path in 3-D space. A general and concise algorithm is given for simulating the intersection of electron trajectories leaving the sample onto multidetector entrance apertures distributed in 3-D space. By optimising the object-oriented design in conjunction with the use of a process-oriented and data-oriented code structure, Mc3D is capable of simulating microscopic analysis of a sample with a 3-D geometry or structure that can be expressed with formulae. Three examples of the use of Mc3D are given. The first is for linescans across a block of SiO₂ on top of a Si substrate; the second is for a stripe of SiO₂ embedded in a Si substrate. Finally, the simulation of Auger linescans across an Au overlay on Si is compared with experimental results. The relationships between experimental linescans and the true beam impact positions on the sample are revealed through the virtual scan path. An edge effect, parallel-edge enhancement, is predicted when the incident electron beam size, the distance of impact position to the terrace edge, and the inelastic mean free path of the Auger electron from a given element are comparable, and the linescan is parallel to the terrace edge. All three examples demonstrate the sensitivity of image contrast to the disposition of the sample with respect to the electron column and the detector position.     

Fabrication and characterization of three-dimensional poly(ether- ether- ketone)/-hydroxyapatite biocomposite scaffolds using laser sintering

The ability to have precise control over porosity, scaffold shape, and internal pore architecture is critical in tissue engineering. For anchorage-dependent cells, the presence of three-dimensional scaffolds with interconnected pore networks is crucial to aid in the proliferation and reorganization of cells. This research explored the potential of rapid prototyping techniques such as selective laser sintering to fabricate solvent-free porous composite polymeric scaffolds comprising of different blends of poly(ether-ether-ketone) (PEEK) and hydroxyapatite (HA). The architecture of the scaffolds was created with a scaffold library of cellular units and a corresponding algorithm to generate the structure. Test specimens were produced and characterized by varying the weight percentage, starting with 10 wt% HA to 40 wt% HA, of physically mixed PEEK-HA powder blends. Characterization analyses including porosity, microstructure, composition of the scaffolds, bioactivity, and in vitro cell viability of the scaffolds were conducted. The results obtained showed a promising approach in fabricating scaffolds which can produce controlled microarchitecture and higher consistency.     

空气孔硅光子晶体偏振无关3 dB分光器

提出并成功设计了基于自准直马赫-曾德尔干涉仪(SMZI)的空气孔硅光子晶体偏振无关3dB分光器。介绍了采用偏振透射谱匹配的方法,SMZI可以实现偏振无关分束的基础理论。通过连续改变SMZI两臂的光程差,基于匹配两个偏振的透射谱,成功实现了在归一化频率下将TE偏振和TM偏振的入射自准直光束进行1∶1分光。最后,利用时域有限差分数值模拟软件计算出来的TE偏振瞬时磁场分布图和TM偏振瞬时电场分布图验证了自准直光束的分光情况。结果显示:如果将工作波长定在1 550nm,该偏振无关3dB分光器大小仅为16.7μm×16.7μm。由于具有尺寸微小、结构简单并使用单一硅材料等特性,该分光器有望应用于集成光路中。     

组织工程支架三维结构点单元数据建模

组织工程支架所用材料和呈现结构是影响其性能的关键因素。以包含材料成分信息的点单元堆积所建立的三维离散模型,可以同时记录支架的几何结构信息和材料成分信息,为使用快速成形方法与数字微滴喷射技术制造具有更高精确的结构梯度和材料梯度的组织工程支架提供了数字模型。材料点单元堆积造型方法主要是基于模型分解的方式,将具有材料梯度和结构梯度的三维实体分割成一系列单元空间并制定单元值。主要分以下几个步骤：单元划分,所用单元形式、确定单元划分密度的法则;单元值量化方式以及量化值、从能量的角度进行局部误差分析。在理论研究的基础上,通过Matlab语言编写了功能强大的点单元数据建模软件,并应用该软件提供人体大段骨和颅部的点单元模型,从而证明此种方法的可行性。     

基于嵌入式Linux的桌面级DLP型3D打印机设计

针对现有DLP型3D打印机价格高、体积大、普及率低、打印时需要连接个人电脑等问题,详细研究了DLP型3D打印机打印原理及实现方式,使用廉价解决方案改进了原有硬件结构,利用Python语言重新编写控制软件,提出了一种基于嵌入式Linux的桌面级DLP型3D打印机设计方案。3D打印机采用Cortex-A7内核的树莓派为硬件核心,运用开源的计算机视觉库Open CV实现了图片解析与显示,通过控制步进电机带动丝杆螺母实现了打印平台精确升降,利用串口触摸屏代替电脑上位机实现了用户脱机打印。该DLP型3D打印机体积为240×220×500 mm(长*宽*高),最大成型体积达到120×120×160 mm(长*宽*高),打印层高最小为0.025 mm,图片显示分辨率达到1 080 P。实际打印结果表明,该DLP型3D打印机具有打印精度高,体积小,成本低,可实现脱机打印的优点。     

基于RP的骨组织工程支架构造及生物学特性分析

应用三维CAD软件设计支架和相应的模具结构,通过光固化快速成形技术制造出树脂模具,并在模具中填充磷酸钙(CPC)生物材料,然后通过热分解方法去掉树脂模具,得到具有可控微管道结构的骨组织工程支架。该方法克服了传统构造方法中支架内部微管道结构不可控的缺点,为制造出更有利于细胞/组织长入和成活的支架三维空间结构提供了一个更理想的方案。INSTRONMicrotester试验设备上测得支架的最大抗压强度为7.12MPa;测得其表面粗糙度Ra=2.16μm。扫描电镜观察支架表面微结构特征,能谱分析测出支架中所含钙元素和磷元素的摩尔比是1.59∶1。结果表明所构造的支架具有良好的生物学特性。     

基于STM32的微型热敏打印机的设计和实现

基于STM32的微型热敏打印机控制系统以STM32F103C8单片机为控制器,以存储器、步进电机和热敏打印头为控制对象,上位机和单片机之间进行串口通信,串口调试软件发送命令和字符,就可以在热敏纸上直接打印相应的字符。该文阐述了系统的硬件组成及软件的实现过程,并给出系统测试结果。测试结果表明,该微型热敏打印机控制系统稳定可靠,打印字迹清晰,不仅可以打印英文、汉字,还可以打印条形码和二维码等。     

Design of multi-scaffold fabrication system for various 3D scaffolds

Three-dimensional (3D) porous scaffolds have been fabricated recently for tissue engineering applications through solid free-form fabrication (SFF) technologies. A multi-scaffold fabrication system for the fabrication of scaffolds, such as polymer, polymer/ceramic, ceramic, and nanofiber, was designed in this study. The various components, including a dispenser with a maximum pressure of 750 kPa, a thermostat with a maximum temperature of 250°C, a high-voltage power supply with a maximum output of 60 kV, and a syringe pump with small flow control, play important roles in determining the process characteristics of scaffolds. The system can process applicable biomaterials with extremely high accuracy with a precision nozzle. Several 3D scaffolds, including PCL, PCL/PLGA/β-TCP, β-TCP, and PCL nanofibers, were fabricated. The morphology and pore size of fabricated scaffolds were observed through scanning electron microscopy. Results show that the scaffolds manufactured in this study can be effectively utilized as bone regeneration scaffolds.