Rapidly functional immobilization of immortalized human hepatocytes using cell adhesive GRGDS peptide-carrying cellulose microspheres

With the development of biotechnology, hepatic support by a hybrid artifici al liver (HAL) using hepatocytes has been given much attention. Because the availability of human livers is limited, we have established a tightly reg ulated immortal human hepatocyte cell line, NKNT-3, for developing HAL. Bec ause hi.-h-density cell culture allows the compactness of the HAL device an d its easy use under emergency circumstances, we have developed cell adhesi ve GRGDS peptide-containing cellulose microspheres (GRGDS/CMS). The GRGDS/C MS efficiently immobilized NKNT-3 cells within 24 h in a stirred suspension culture. Electron microscopic examinations demonstrated glycogen granules and well-developed endoplasmic reticulum and mitochondria in NKNT-3 cells a ttached to the GRGDS/CMS. The cells showed ammonia clearance activity, wher eas HepG2-transformed human liver cells did not remove the loaded ammonia. An efficient adenoviral delivery of the lacZ reporter gene was performed in GRGDS/CMS-immobilized NKNT-3 cells. In this study we present rapid immobil ization of NKNT-3 immortal human hepatocytes using cellulose microspheres c arrying GRGDS peptides. These microspheres satisfied immediate preparation of NKNT-3 cells in sufficient quantity and of adequate quality.