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首页> 外文期刊>Tissue engineering, Part C. Methods >Layer-by-layer tissue microfabrication supports cell proliferation in vitro and in vivo
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Layer-by-layer tissue microfabrication supports cell proliferation in vitro and in vivo

机译：逐层组织微细加工可在体外和体内支持细胞增殖

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Layer-by-layer biofabrication represents a novel strategy to create three-dimensional living structures with a controlled internal architecture, using cell micromanipulation technologies. Laser assisted bioprinting (LAB) is an effective printing method for patterning cells, biomolecules, and biomaterials in two dimensions. "Biopapers," made of thin polymer scaffolds, may be appropriate to achieve three-dimensional constructs and to reinforce mechanical properties of printed materials. The aim of this work was to evaluate the effect of the tridimensional organization of cells and biomaterials on cell proliferation in vitro and in vivo. The experimental LAB setup was comprised of an infrared laser, focused onto a glass ribbon coated with an absorbing layer of gold. The cell bioink was made of MG63 cells (50 millions cells/mL in culture medium and 1% alginate), transduced with Luciferase gene for tracking and quantification. The printing substrate was a 100-μm-thick polycaprolacton (PCL) electrospun scaffold. The building sequence comprised sequential layers of cells and PCL scaffolds stacked using two different tridimensional arrangements, which were compared in this study (layer-by-layer vs. seeding on a single locus of the scaffolds). Then the cell-seeded materials were cultured in vitro or implanted in vivo in NOD-SCID mice. The qualitative follow-up involved scanning electron microscopy (SEM) observations, live-dead assays, and histology. The cell amount was quantified by photon imager during 21 days in vitro and 2 months in vivo. Live-dead assay and SEM revealed that the cells survived after printing and spread onto PCL membranes. Circle-shaped patterns were maintained in vitro during the first week but they were no longer observable after 2 weeks, due to cell proliferation. Luciferase tracking displayed that the cell amount was increased in vitro and in vivo when the materials and the cells where stacked layer by layer. Histological sections of the in vivo samples revealed a thicker fibrous tissue in the layer-by-layer samples. We have demonstrated in this study that PCL electrospun biopapers can act as a shock-absorbing mattress for cell printing and could further support cell proliferation. The layer-by-layer printing provided an appropriate 3D environment for cell survival and enhanced cell proliferation in vitro and in vivo.
机译：逐层生物制造代表一种新颖的策略，即使用细胞微操纵技术创建具有受控内部结构的三维居住结构。激光辅助生物打印（LAB）是一种有效的打印方法，可以在两个方向上对细胞，生物分子和生物材料进行图案化。由薄的聚合物支架制成的“生物纸”可能适合实现三维结构并增强印刷材料的机械性能。这项工作的目的是评估细胞和生物材料的三维组织对体外和体内细胞增殖的影响。实验LAB装置由红外激光组成，聚焦在涂有金吸收层的玻璃带上。细胞生物墨水由MG63细胞（培养基中的5000万个细胞/ mL和1％海藻酸盐）制成，并通过荧光素酶基因进行转导以进行跟踪和定量。印刷基材为100μm厚的聚己内酰胺（PCL）电纺丝支架。构建顺序包括使用两个不同的三维排列堆叠的细胞和PCL支架的连续层，在本研究中进行了比较（逐层对比在单个位点上接种）。然后，将细胞接种的材料体外培养或体内植入NOD-SCID小鼠中。定性随访包括扫描电子显微镜（SEM）观察，活死分析和组织学检查。通过光子成像仪在体外21天和体内2个月期间定量细胞量。活死测定和SEM显示，细胞在印刷后存活并扩散到PCL膜上。第一周在体外维持圆形模式，但由于细胞增殖，两周后不再可见。荧光素酶追踪显示，当材料和细胞逐层堆叠时，体外和体内细胞数量增加。体内样品的组织学切片显示在逐层样品中较厚的纤维组织。在这项研究中，我们证明了PCL电纺生物纸可以充当细胞印刷的减震垫，并且可以进一步支持细胞增殖。逐层打印为体外和体内细胞存活和增强细胞增殖提供了合适的3D环境。

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