Automated production of nerve repair constructs containing endothelial cell tube-like structures

The data that support the findings of this study, entitled Automated production of nerve repair constructs containing endothelial cell tube-like structuresVideos of the automated production of engineered neural tissue (EngNT) constructs using gel aspiration-ejection (GAE) were taken at UCL and McGill University. These data refer to the characterisation of two sizes of EngNT, 12G and 16G, produced using this technique containing human umbilical vein endothelial cells (HUVECs). Tensile testing data was collected from acellular constructs, performed using the Univert Mechanical Testing Instrument with a calibrated 10 N load cell and a controlled displacement rate of 0.1 mm/s. Viability data of engineered neural tissue containing HUVECs (EngNT-HUVEC) covers local viability, using LIVE/DEADTM Cell Imaging Kit (488/570), and whole construct viability, using Lactase Dehydrogenase (LDH) Assay Kit (Cytotoxicity) collected using a NanoZoomer S60 Digital Slide Scanner and a SpectraMAX M5 Multi-Mode Microplate Reader, respectively. CellTiter-Glo® 3D Cell Viability Assay data reports the ATP concentration in EngNT-HUVEC constructs relative to their starting hydrogels, measured using a SpectraMAX M5 Multi-Mode Microplate Reader. Alignment of cells within EngNT-HUVEC was captured by VolocityTM v6.5.1 analysis of 20 µm z-stacks taken using Zeiss LSM 710 confocal microscope at predetermined positions along the construct stained with Rhodamine 110 Phalloidin and Hoechst 33342. Viability, metabolic activity and alignment assays were performed 24 h after EngNT-HUVEC construct formation. Neurite length and alignment data were collected using ImageJ and VolocityTM v6.5.1, respectively, analysing the co-culture of neurons on the EngNT-HUVEC and acellular constructs imaged using Zeiss LSM 710 confocal microscope of samples stained with anti-betaIII tubulin and Hoechst 33342. EngNT-HUVEC made from starting hydrogels with two seeding densities (0.5 mill/mL and 2.0 mill/mL) were cultured for 2 or 4 days. These samples were stained with Rhodamine 110 Phalloidin and Hoechst 33342 and analysed for inner angiogenic features, using ImageJ plugin Angiogenesis Analyzer, and outer endothelialisation, using VolocityTM v6.5.1. 16G EngNT-HUVEC constructs were further analysed for lumen number from 10 µm sections stained with Rhodamine 110 Phalloidin and Hoechst 33342, imaged on Zeiss AxioLab A1 fluorescent microscope. RT-PCR data from EngNT-HUVEC were collected using GoScript Reverse Transcriptase Kit (Promega) and Power SYBRä Green PCR Master Mix using SimpliAmp Thermal Cycler and QuantStudio3 instrument, analysed by QuantStudio Design and Analysis Software v1.5.1 with hypoxanthine guanine phosphoribosyl transferase (HPRT1), ribosomal protein lateral stalk subunit P0 (RPLP0) and ribosomal protein S18 (RPS18) reference genes. Data collected between December 2022 and January 2024.

本研究所支持的数据集，名为《含内皮细胞管状结构的神经修复结构的自动化生产》，记录了在伦敦大学学院和麦吉尔大学使用凝胶吸射-喷射（GAE）技术自动化生产工程化神经组织（EngNT）结构的视频。这些数据涉及使用该技术制备的两种尺寸的EngNT，即12G和16G，其中包含人脐静脉内皮细胞（HUVECs）。通过Univert机械测试仪，配备校准的10N负荷细胞和0.1mm/s的受控位移速率，对去细胞结构进行了拉伸测试数据收集。包含HUVECs的工程化神经组织（EngNT-HUVEC）的存活数据包括局部存活率，采用LIVE/DEAD™细胞成像试剂盒（488/570）检测，以及整体结构存活率，采用乳糖脱氢酶（LDH）测定试剂盒（细胞毒性）检测，分别使用NanoZoomer S60数字切片扫描仪和SpectraMAX M5多模态微孔板阅读器收集。使用CellTiter-Glo® 3D细胞存活率测定数据报告了EngNT-HUVEC结构相对于其起始水凝胶的ATP浓度，该数据是通过SpectraMAX M5多模态微孔板阅读器测量的。通过Volocity™ v6.5.1分析，在预定位置沿结构用罗丹明110磷酰胺和 Hoechst 33342染色的20µm z堆栈图像中捕获了EngNT-HUVEC内细胞对齐情况。在EngNT-HUVEC结构形成后24小时，进行了存活率、代谢活性和对齐性检测。使用ImageJ和Volocity™ v6.5.1分别收集了神经突长度和对齐数据，分析了在EngNT-HUVEC和去细胞结构上共培养的神经元，这些样本使用Zeiss LSM 710共聚焦显微镜进行染色，分别使用抗-βIII微管蛋白和 Hoechst 33342。从起始水凝胶中制备的两种播种密度（0.5毫毫升和2.0毫毫升）的EngNT-HUVEC在2或4天后进行培养。这些样本使用罗丹明110磷酰胺和 Hoechst 33342染色，并使用ImageJ插件血管生成分析器和Volocity™ v6.5.1进行分析，以检测内部血管生成特征和外部内皮化。进一步对16G EngNT-HUVEC结构进行分析，以确定用罗丹明110磷酰胺和 Hoechst 33342染色的10µm切片中的管腔数量，并在Zeiss AxioLab A1荧光显微镜上进行成像。EngNT-HUVEC的RT-PCR数据使用Promega的GoScript逆转录酶试剂盒和Power SYBR® Green PCR Master Mix，通过SimpliAmp热循环仪和QuantStudio3仪器收集，并使用QuantStudio设计和分析软件v1.5.1进行分析，参考基因包括次黄嘌呤鸟嘌呤磷酸核糖转移酶（HPRT1）、核糖体蛋白侧臂亚基P0（RPLP0）和核糖体蛋白S18（RPS18）。收集数据的时间为2022年12月至2024年1月。