醛基化海藻多糖复合材料作为人工肝载体的可行性

背景：三维肝细胞体外培养中肝细胞可在支架孔隙中良好生长，但酶消化脱细胞过程严重影响了细胞获得率和活性。目的：制备大孔三维细胞支架N-聚异丙基丙烯酰胺-醛基化海藻多糖，探索其作为人工肝生物反应器细胞载体的可行性。方法：以海藻酸钠为致孔剂合成大孔N-聚异丙基丙烯酰胺，再交联醛基化海藻多糖，制备出大孔三维细胞支架N-聚异丙基丙烯酰胺-醛基化海藻多糖。测定其孔径及孔隙率。①细胞毒性实验：MTT法检测在新鲜细胞培养液、高密度聚乙烯浸提液、高密度聚氯乙烯浸提液及N-聚异丙基丙烯酰胺-醛基化海藻多糖浸提液中培养小鼠成纤维细胞的肖值。②非酶脱细胞性能实验：将第3代小鼠成纤维细胞接种到盛有N-聚异丙基丙烯酰胺一醛基化海藻多糖支架的6孔板中，实验组通过降温脱附回收细胞，对照组以胰蛋白酶消化回收细胞。结果与结论：大孔三维细胞支架N-聚异丙基丙烯酰胺-醛基化海藻多糖的孔径为（180．23±62．30）Um，孔隙率为（89．67±2．401％。细胞毒性实验证明N-聚异丙基丙烯酰胺-醛基化海藻多糖支架无细胞毒性；培养第15天实验组非酶温控脱细胞与对照组酶消化脱细胞相比细胞获得率增加26．24％。表明大孔三维细胞支架N-聚异丙基丙烯酰胺．醛基化海藻多糖具有适宜细胞生长的大孔结构；温度调控脱细胞性能克服了传统酶消化脱细胞对细胞的损伤，可提高细胞培养的数量和质量。

Aldehyde seaweed polysaccharide composites serve as artificial liver carriers

BACKGROUND： During the throe-dimensional culture in vitro, hepatocytes can grow well in the pores of the scaffold, but enzymatic digestion for deceUularization severely affects cel! survival rate and activity. OBJECTIVE： To prepare a macroporous scaffold, N-poly-isopropyl acrylamide-aldehyde seaweed polysaccharides, and to investigate the feasibility of this scaffold as a cell carrier for artificial liver bioroactor METHODS： Sodium alginate as the porogen was used to synthesize the macroporous N-poly-isopropyl acryiamide that was then cross-linked with aldehyde seaweed poiysaccharides to prepare the macreporous throe-dimensional cytoskeleton. Scaffold aperture and porosity were measured. （1） Cytotoxicity test： 3-（4,5-Dimethylthiazol-2-yl）-2,5-diphenyltetrazolium bromide assay was used to detect absorbance values of mouse fibroblasts cultured in fresh cell culture medium, high-density polyethylene extract, high-density polyvinyl chloride extract and N-poly-isopropyl acrylamide-aldehyde seaweed polyssccharide extract. （2） Non-enzymatic acellular performance tests： The third generation of mouse fibroblasts cell line was inoculated into 6-well plates containing N-poly isopropylacrylamide-aldehyde seaweed polysaccharide scaffolds. In the experimental group, cells were recovered by cooling desorption, while in the control group, the cells were recovered using trypsin digestion method. RESULTS AND CONCLUSION： The aperture and porosity of the macroporous scaffold were （180.23±62.30） pm and （89.67±2.40）%, respectively. Cytotoxicity test showed that the macroporous scaffold of N-poly-isopropyl acrylamide-aldehyde seaweed polysaccharide had no cytotoxicity. The cell survival rate in the experimental group was increased 26.24% than that in the control group. These findings indicate that the throe-dimensional macroporous cytoskeleton, N-poly-isopropyl acrylamide-atdehyde seaweed polysaccharide, has a macrooporous structure suitable for cell growth, and thermo-rosponsive acellular performance overcomes cellular damage resulting from traditional enzymatic digestion and promotes the number of quality of cultured cells.