Hepatocyte purification column using thermoresponsive glycopolymer-modified silica beads

Abstract

Regenerative medicine involving hepatocytes holds promise as a therapeutic strategy for liver diseases. The clinical application of this approach requires a reliable technique for hepatocyte isolation. In this study, we developed a cell separation column that employs a thermoresponsive glycopolymer to isolate hepatocytes through temperature modulation. Silica beads were coated with poly(N-p-vinylbenzyl-O-β-D-galactopyranosyl-1→4-D-guliconamide)-b-poly(N-isopropylacrylamide) (PVLA-b-PNIPAAm) via atom transfer radical polymerization and were used as packing material for the column. The modified beads were characterized using scanning electron microscopy, CHN elemental analysis, Fourier transform infrared spectroscopy, and X-ray photoelectron spectroscopy. Nine columns were fabricated by altering the PNIPAAm and PVLA chain lengths of PVLA-b-PNIPAAm. Evaluation of the elution characteristics of HepG2 cells revealed that columns with intermediate PVLA and PNIPAAm chain lengths exhibited temperature-dependent cell retention and elution. This column enabled the separation of HepG2 cells from contaminating RAW264.7 cells by adjusting the temperature. The recovered HepG2 cells retained high levels of activity. Collectively, these findings indicate that the developed column enables efficient separation of hepatocytes solely through temperature modulation.