A Novel Cell Co-Culture Method Utilizing Thermo-Responsive Surface

摘要

In this work, we developed a novel patterned co-culture method with thermo-responsive poly(N-isopropylacrylamide) (PIPAAm) and poly(N-ρ-vinylbenzyl-O-β-D-galactopyranosyl-(1→ 4)-D-gluconamide) (PVLA) inducing active hepatocyte attachment. Patterned graft of PIPAAm onto PS dishes was carried out by electron beam irradiation using cover-glass as a photomask. PVLA was only coated onto PIPAAm-ungrafted domain because of hydrated hydrophilic property of PIPAAm at below the LCST. Analysis by attenuated total reflection-Fourier transform infrared and electron spectroscopy for chemical analysis revealed that PIPAAm and PVLA were successfully grafted and coated on surfaces of PS dishes. PIPAAm-grafted surface exhibited decreasing contact angle by changing temperature from 37 to 20°C, while PVLA-coated PS and non-treated PS had negligible contact angle changes with temperature alternation. Atomic force microscopy (AFM) results showed that PIPAAm-grafted and PVLA-coated PS had smoother surfaces than that of ungrafted PS dishes. After culture for 12 hours, hepatocytes were well attached on PVLA-coated domain. Hepatocytes adherent on PIPAAm-grafted domain were detached by decreasing temperature. And then, fibroblasts were seeded onto PIPAAm pattern-grafted domain. Fibroblasts were only attached and spread onto PIPAAm-grafted domain. Co-cultured hepatocytes showed better differentiated function of albumin expression compared to homotypic hepatocyte culture