BACKGROUND:: During the three-dimensional culture in vitro, hepatocytes can grow well in the pores of the scaffold, but enzymatic digestion for decellularization severely affects cell 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 bioreactor. METHODS:: Sodium alginate as the porogen was used to synthesize the macroporous N-poly-isopropyl acrylamide that was then cross-linked with aldehyde seaweed polysaccharides to prepare the macroporous three-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 polysaccharide 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) μm 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 three-dimensional macroporous cytoskeleton, N-poly-isopropyl acrylamide-aldehyde seaweed polysaccharide, has a macroporous structure suitable for cell growth, and thermo-responsive acellular performance overcomes cellular damage resulting from traditional enzymatic digestion and promotes the number of quality of cultured cells.