一种基于混流控制的多细胞三维受控组装系统

细胞三维受控组装技术为体外构建活体组织器官提供了理论依据和技术支持。功能性组织和器官是多种细胞的有序三维排列,具有复杂的空间结构,因而在体外构建具有生物活性的组织器官,就需要细胞三维受控组装技术能够在空间上实现多种细胞材料的精确定位组装。本研究面向多细胞组装的需求,在单喷头细胞组装系统基础上,设计了新的混流喷头系统以及脉冲切换控制电路,开发出多细胞三维受控组装系统,并基于该系统进行了多组分的类细胞外基质材料的堆积实验。研究结果证明该系统可以准确地进行多细胞三维受控组装,达到构建非均质结构体的目的。     

基于细胞三维受控组装技术的细胞芯片构建

如何将细胞准确装配于细胞芯片的设计位置,是制约细胞芯片向组织和生理系统细胞芯片方向发展的关键问题之一。本研究用细胞组装技术将细胞装配到细胞芯片的设计位置并检测的理论和方法。首先设计构建了含多组传感器阵列的细胞芯片;选取输送和固定细胞的明胶-海藻酸钠复合水凝胶材料,探讨用细胞组装技术在芯片上精确装配细胞的方法;测试复合材料和多种电解溶液对电阻抗的影响,并用芯片对细胞增殖进行检测。结果显示:复合材料能输送并将细胞固定在芯片上超过4周;细胞组装技术可精确将细胞装配到指定传感器,组装的细胞/材料微丝直径100～120μm。复合材料对芯片基础电阻抗影响小,在大于103Hz的高频段,基础阻抗为小于102.6Ω;PBS和DMEM溶液在103.5～105Hz高频段可替代氰铁化钾溶液作为电解液;芯片上的传感器在103.5Hz可准确测出Hela细胞增殖引起的电阻抗变化。研究证明了用细胞组装技术构建复杂细胞芯片的可行性。     

3D打印器官芯片研究进展

器官芯片是一种将生物体活细胞植入精准设计的微流体芯片内,可特定再现生物体组织器官功能的仿生的微生理系统,在疾病模拟、毒性检测、新药研发、精准医疗等许多方面具有独特应用前景。3D生物打印技术与器官芯片技术相结合制作3D打印器官芯片,可实现芯片制造工艺的简易化和低成本化,同时满足对复杂异质三维微环境的精细需求。综述3D打印器官芯片在打印方式、打印墨水等方面的研究进展,阐述其最新生物医学应用,总结器官芯片结构和功能单元的打印方式和打印墨水,探讨基于现有打印工艺实现器官芯片一体化制造的潜在可行性,概述3D打印器官芯片技术在心、肝、肺、肾、神经、肿瘤等组织和器官结构和功能仿生方面的最新进展,最后展望3D打印器官芯片技术领域的发展趋势和有待解决的关键问题。     

基于细胞3D打印技术的肿瘤药物筛选细胞芯片研究

现有的药物筛选评价技术中,动物筛选模型存在种属差异和周期长等缺点,高通量筛选和细胞筛选模型则与体内环境差异大,药物筛选准确率低。细胞3D打印技术为在体外构建仿真的组织器官模型提供了可能,当其与细胞芯片技术结合则为体外构建高效准确的药物筛选模型提供了新的技术空间。本研究构建了含有多个叉指电极（IDEs）阵列的细胞芯片,用细胞3D打印技术在芯片上组装了卵巢癌细胞HO-8910和人肝间充质干细胞HMSC-H组织模型,并通过对组织模型内细胞阻抗变化的检测,反映细胞生长、贴附、增殖、凋亡的过程及药物对细胞活性的影响等,最终基于该模型检测了抗癌药物顺铂和环磷酰胺对肿瘤细胞的杀伤和肝毒性。结果显示：支架微丝直径和孔径约为200～300 μm,肿瘤细胞和肝细胞在三维结构里生长良好;DMEM作为电解液,芯片在10.4 Hz可准确检测到三维结构中细胞增殖引起的阻抗变化,20 h后阻抗升高69.6%;基于该筛选模型,能同步检测到药物的抗肿瘤作用和肝毒性,并筛选出需要肝的二次代谢产物才能产生抗肿瘤性的药物环磷酰胺。     

血管组织工程支架的计算机辅助低温沉积制造研究

实现血管支架的个性化制造,对于组织工程血管移植物(tissue-engineered vascular graft,TEVG)进入临床应用具有重要意义。本研究利用低温沉积制造(LDM)技术制造了管状支架及结构较复杂的血管网支架。并控制螺杆转速、扫描速度和喷头温度等工艺参数调控支架特性,对其表观特征进行了评测。所得支架能准确再现复杂的计算机辅助设计(CAD)的三维血管模型结构;支架壁厚随着螺杆转速与扫描速度之比的增加而增加;微孔尺寸、壁面平整度及支架壁厚与喷头温度成正相关,但是孔隙率受喷头温度影响不大。利用LDM技术可以制作出具有特定结构及表观特征的血管支架,对血管支架的个性化制造具有极大推动作用。     

3D打印技术在医学领域中的应用

3D打印技术是新型的快速打印技术,它结合了三维扫描、计算机辅助数字化成型等技术,实现高精度、高效率、低费用地打印立体结构。在医学领域,3D打印通过对骨骼、肌肉、血管、神经等解剖结构进行高仿真地精确打印,用于辅助解剖学、病理学教学、临床实习生的手术模拟、医生术前讨论与手术方案制定,也被作为植入体用于患者缺损组织的修补。通过将活细胞与仿生材料相结合进行的3D生物打印,可用于药物筛选、药效评价,还可实现组织器官再造、器官移植等。     

人造胸腺能产生T细胞

美国的研究人员已经制造出一种人造胸腺 ,这种人造胸腺能够产生 T细胞并比现行使用的方法更为有效。这种技术将来可被用于制作胸腺类器官 ,将这类器官植入患者体内可重建或改善患者的免疫系统。现行技术主要是培养 T细胞并用细胞活素使T细胞成长和分化 ,但效果不是很好。有证据表明 T细胞在拟态的胸腺三维结构环境中成长比较好。研究人员采用了用于修复头的多孔材料。这种材料由覆盖有生物适合的金属钽的网状碳基质组成。研究人员用鼠胸腺细胞种基质 ,首先胸腺细胞进入支架 ,然后研究增加人类促细胞生成素原始细胞 ,发现 14天内系统能产生…     

基于MRI切片的人体复杂结构的三维建模方法

目的：探讨使用Mimics软件构建包含皮肤、肌肉、脂肪以及骨骼的复杂三维人体模型的方法及意义。方法：利用MRJ（MagneticResonanceImaging）切片通过MIMICS软件分别建立人体骨骼、肌肉、脂肪、皮肤等几何面模型，准确表达人体组织和器官的复杂几何结构；将组织和器官的几何面模型导入到MAGICS中进行面网格划分，通过ANSYS软件将面网格转化为体网格，得到人体各器官和组织的三维几何体模型；进而给骨骼、肌肉、脂肪、皮肤分别赋予材料特性（弹性模量和泊松比等），可以模拟真实人体组织和器官的物理属性。结果：利用MIMICS的布尔运算将各部分组织和器官进行组装，得到人体复杂结构的三维实体模型。结论：与目前常用的基于几何学的人体模型建模方法相比，使用该方法建立的三维实体模型可以完全模拟人体器官和组织的真实结构，等效程度高、建模速度快。所建立的数学模型可以直接用于3D打印、模具制造及等效假人的运动学和力学分析。广泛应用于医学、人机工程学等领域。     

生物三维打印细胞负载水凝胶类组织的精准优化

生物三维打印技术与水凝胶的完美结合,可为制造复杂结构功能的组织器官提供一种极具吸引力的解决方案。定制打印细胞负载水凝胶类组织的内部结构,可以更好地仿生真实组织器官的三维微环境,对打印后细胞生长、组织形成和功能再生至关重要。但水凝胶理化特性多变,精准打印与设计结构匹配的多孔结构仍然极具挑战。提出基于光学相干层析成像技术(OCT)的生物三维打印细胞负载水凝胶类组织的精准优化方法,通过自制的三维扫频OCT系统无损在线成像打印组织块和定量评价结构参数,迭代降低设计与打印间的结构差异,提高细胞负载水凝胶打印的精准性和稳定性。实验结果表明,基于OCT无损定量表征结果反馈优化打印参数设置,指导打印过程,使得细胞负载水凝胶类组织的结构形态参数与设计值的偏差从40%左右控制到7%以内,包括内部孔隙尺寸、支撑尺寸、孔隙率、表面积、体积五项关键参数;细胞培养两周后的存活率从80%左右显著提高到90%以上。研究表明OCT技术为批量定制细胞负载水凝胶类组织、生物三维打印组织和器官等提供了具有潜力的精准化工具。     

动态三维超声心动图研究

目前广泛使用的二维超声心动图技术仅能在二维平面上评价心脏的结构与功能,但心脏本身是一个具有复杂结构的立体器官,临床医生只能通过观察多帧不同方位上的二维切面后,再在头脑内想象出心脏本身的立体结构形态和病理改变,这一过程通常十分困难,且不准确,特别是在诊断复杂先天性心脏病时更是如此,故自七十年代开始,许多研究者都在致力于开发心脏三维结构的超声显示方法,近年来,随着计算机技术的迅速发展,动态三维超声心动图技术在其成像方法研究到临床应用的过程中日臻成熟.同二维超声心动图相比,动态三维超声心动图能获得心脏真实的三维解剖结构图像,并可进行准实时的动态观察,提供更准确的定量数据及获取新的定量信息和功能参数。     

Multinozzle low-temperature deposition system for construction of gradient tissue engineering scaffolds

Tissue engineering is a technology that enables us to construct complicated hominine organs composed of many different types of cells. One of the key points to achieve this goal is to control the material composition and porous structure of the scaffold accurately. A disposable syringe based volume-driven injecting (VDI) nozzle was proposed and designed to extrude both natural derived and synthetic polymers. A multinozzle low-temperature deposition and manufacturing (M-LDM) system is proposed to fabricate scaffolds with heterogeneous materials and gradient hierarchical porous structures. PLGA, collagen, gelatin, chitosan can be extruded without leaking to form hierarchical porous scaffolds for primary study. Composite scaffolds with two kinds of materials were fabricated via two different nozzles to get both hydrophilic and mechanical properties. The results from scanning electron microscopy (SEM) demonstrated that the natural-derived biomaterials were strongly absorbed onto the synthetic biomaterials to form a stable network. Several gradient PLGA/TCP scaffolds were also fabricated to supply several samples.     

Coaxial nozzle-assisted 3D bioprinting with built-in microchannels for nutrients delivery

This study offers a novel 3D bioprinting method based on hollow calcium alginate filaments by using a coaxial nozzle, in which high strength cell-laden hydrogel 3D structures with built-in microchannels can be fabricated by controlling the crosslinking time to realize fusion of adjacent hollow filaments. A 3D bioprinting system with a Z-shape platform was used to realize layer-by-layer fabrication of cell-laden hydrogel structures. Curving, straight, stretched or fractured filaments can be formed by changes to the filament extrusion speed or the platform movement speed. To print a 3D structure, we first adjusted the concentration and flow rate of the sodium alginate and calcium chloride solution in the crosslinking process to get partially crosslinked filaments. Next, a motorized XY stages with the coaxial nozzle attached was used to control adjacent hollow filament deposition in the precise location for fusion. Then the Z stage attached with a Z-shape platform moved down sequentially to print layers of structure. And the printing process always kept the top two layers fusing and the below layers solidifying. Finally, the Z stage moved down to keep the printed structure immersed in the CaCl₂ solution for complete crosslinking. The mechanical properties of the resulting fused structures were investigated. High-strength structures can be formed using higher concentrations of sodium alginate solution with smaller distance between adjacent hollow filaments. In addition, cell viability of this method was investigated, and the findings show that the viability of L929 mouse fibroblasts in the hollow constructs was higher than that in alginate structures without built-in microchannels. Compared with other bioprinting methods, this study is an important technique to allow easy fabrication of lager-scale organs with built-in microchannels.     

基于RFID的医疗器械库管信息系统的设计与实现

目的：设计符合医疗器械仓库管理工作要求的信息管理系统,从而实现医疗器械的科学有效管理。方法：基于无线射频识别技术、数据库技术、信息处理和传输技术对医疗器械仓库管理工作实行模块化、系统化和信息化的管理。结果：测试表明,所设计的系统符合医疗器械仓库管理工作流程的设计要求,稳定可靠,通过串口及读写标签子程序很好地实现了电子标签与数据库信息的交换,并且具有操作简单、使用方便、功能强大和完善全面的安全保护措施。结论：该系统大大地提高了医疗器械仓库管理工作的效率,使器械信息的管理更加准确、方便、规范和完善,满足了医疗器械高效得动态化管理的需要。     

Electrosonic ejector microarray for drug and gene delivery

We report on development and experimental characterization of a novel cell manipulation device—the electrosonic ejector microarray—which establishes a pathway for drug and/or gene delivery with control of biophysical action on the length scale of an individual cell. The device comprises a piezoelectric transducer for ultrasound wave generation, a reservoir for storing the sample mixture and a set of acoustic horn structures that form a nozzle array for focused application of mechanical energy. The nozzles are micromachined in silicon or plastic using simple and economical batch fabrication processes. When the device is driven at a particular resonant frequency of the acoustic horn structures, the sample mixture of cells and desired transfection agents/molecules suspended in culture medium is ejected from orifices located at the nozzle tips. During sample ejection, focused mechanical forces (pressure and shear) are generated on a microsecond time scale (dictated by nozzle size/geometry and ejection velocity) resulting in identical “active” microenvironments for each ejected cell. This process enables a number of cellular bioeffects, from uptake of small molecules and gene delivery/transfection to cell lysis. Specifically, we demonstrate successful calcein uptake and transfection of DNA plasmid encoding green fluorescent protein (GFP) into human malignant glioma cells (cell line LN443) using electrosonic microarrays with 36, 45 and 50 μm diameter nozzle orifices and operating at ultrasound frequencies between 0.91 and 0.98 MHz. Our results suggest that efficacy and the extent of bioeffects are mainly controlled by nozzle orifice size and the localized intensity of the applied acoustic field.     

Temperature limits of mitosis in mammalian cell cultures

To determine the temperature limits of mitosis 11 strains of mammalian cells of different origin were grown in culture between temperatures of 25 and 41°C. Different strains of mammalian cells were shown to respond differently to a rise and fall of the cultivation temperature. The upper temperature limit lies not more than 5° above the optimum and a decrease of temperature of more than 10°C does not stop cell division. In most cell populations there are three types of cells. Most cells can divide within a certain range of temperatures, a smaller group of cells can start mitosis but not complete it, the process being stopped in metaphase, and a very small number of cells can complete one division at below the threshold temperature. The temperature limits of mitosis are unconnected with the species or tissue origin of the cells.     

Vacuum pressure generation via microfabricated converging-diverging nozzles for operation of automated pneumatic logic

Microfluidic devices with integrated pneumatic logic enable automated fluid handling without requiring external control instruments. These chips offer the additional advantage that they may be powered by vacuum and do not require an electricity source. This work describes a microfluidic converging-diverging (CD) nozzle optimized to generate vacuum at low input pressures, making it suitable for microfluidic applications including powering integrated pneumatic logic. It was found that efficient vacuum pressure was generated for high aspect ratios of the CD nozzle constriction (or throat) width to height and diverging angle of 3.6^o. In specific, for an inlet pressure of 42.2 psia (290.8 kPa) and a volumetric flow rate of approximately 1700 sccm, a vacuum pressure of 8.03 psia (55.3 kPa) was generated. To demonstrate the capabilities of our converging - diverging nozzle device, we connected it to a vacuum powered peristaltic pump driven by integrated pneumatic logic and obtained tunable flow rates from 0 to 130 μL/min. Finally, we demonstrate a proof of concept system for use where electricity and vacuum pressure are not readily available by powering a CD nozzle with a bicycle tire pump and pressure regulator. This system is able to produce a stable vacuum sufficient to drive pneumatic logic, and could be applied to power automated microfluidics in limited resource settings.     

Layer-by-layer microfluidics for biomimetic three-dimensional structures

Due to the complex structures of living systems, with size scales spanning from the micron to millimeter range, the use of microtechnology to recreate in vivo-like architecture has exciting potential applications. However, most microscale systems are two-dimensional, and few three-dimensional (3-D) systems are being explored. We have developed a versatile technique, combining surface engineering with layer-by-layer microfluidics technology, to create a 3-D microscale hierarchical tissue-like structure. The process involves immobilization of a cell-matrix assembly, cell-matrix contraction, and pressure-driven microfluidic delivery. An aminopropyltriethoxysilane-glutaraldehyde activated chip is used to effectively immobilize the cell-matrix assemblies while maintaining cell viability. Pressure-driven microfluidics is applied to transport cells-matrices with controlled flow rates, determined from dynamic flow imaging. By taking advantage of the contraction of the biopolymer matrices by cells, layer-by-layer microfluidics can be used to build multilayers of cell-matrix inside a microchannel and the thickness of each layer can be controlled down to microscale dimensions. Confocal and electron microscopy images of the final structure show a hierarchical layered cellular configuration composed of heterogeneous biomimetic materials. For a model system, a biomimetic arterial structure is formed using three types of vascular cells to mimic the 3-tunic structure found in vivo. This approach provides solutions to fabricate hierarchical "neotissues" with controlled microarchitectures and 3-D configurations of multiple cell types.     

Mammalian cell delivery via aerosol deposition

The objective of this study was to test the hypothesis that bovine dermal fibroblasts can survive aerosol delivery via an airbrush with mean cell survival rates greater than 50%. This technology has great implications for burn and other wound therapies, for delivery of genetically altered cells in gene therapies, and for tissue engineering with tissue scaffolds. Bovine dermal fibroblasts were suspended at a concentration of 200,000 cells/mL in Hank's Balanced Salt Solution, and delivered into six-well tissue culture plates using a Badger 100G airbrush. Cells were delivered through three nozzle diameters (312, 484, and 746 microm) at five different air pressures (41, 55, 69, 96, and 124 kPa). Nine repetitions were performed for each treatment group, and cell viability was measured using trypan blue exclusion assay. Mean cell viability ranged from 37 to 94%, and depended on the combination of nozzle diameter and delivery pressure (p < 0.0001). Linear regression analysis was used to develop a stochastic model of cell delivery viability as a function of nozzle diameter and delivery air pressure. This study demonstrates the feasibility of using an airbrush to deliver viable cells in an aerosol to a substrate.     

Analytic estimates of secondary neutron dose in proton therapy

Proton beam losses in various components of a treatment nozzle generate secondary neutrons, which bring unwanted out of field dose during treatments. The purpose of this study was to develop an analytic method for estimating neutron dose to a distant organ at risk during proton therapy. Based on radiation shielding calculation methods proposed by Sullivan, we developed an analytical model for converting the proton beam losses in the nozzle components and in the treatment volume into the secondary neutron dose at a point of interest. Using the MCNPx Monte Carlo code, we benchmarked the neutron dose rates generated by the proton beam stopped at various media. The Monte Carlo calculations confirmed the validity of the analytical model for simple beam stop geometry. The analytical model was then applied to neutron dose equivalent measurements performed on double scattering and uniform scanning nozzles at the Midwest Proton Radiotherapy Institute (MPRI). Good agreement was obtained between the model predictions and the data measured at MPRI. This work provides a method for estimating analytically the neutron dose equivalent to a distant organ at risk. This method can be used as a tool for optimizing dose delivery techniques in proton therapy.