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 artificial liver (HAL) using hepatocytes has been given much attention. Because the availability of human livers is limited, we have established a tightly regulated immortal human hepatocyte cell line, NKNT-3, for developing HAL. Because high-density cell culture allows the compactness of the HAL device and its easy use under emergency circumstances, we have developed cell adhesive GRGDS peptide-containing cellulose microspheres (GRGDS/CMS). The GRGDS/CMS 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 attached to the GRGDS/CMS. The cells showed ammonia clearance activity, whereas 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 immobilization of NKNT-3 immortal human hepatocytes using cellulose microspheres carrying GRGDS peptides. These microspheres satisfied immediate preparation of NKNT-3 cells in sufficient quantity and of adequate quality.
Kobayashi
N
N
First Department of Surgery, Okayama University Medical School, Japan. immortal@md.okayama-u.ac.jp
Taguchi
T
T
Noguchi
H
H
Okitsu
T
T
Totsugawa
T
T
Watanabe
T
T
Matsumura
T
T
Fujiwara
T
T
Urata
H
H
Kishimoto
N
N
Hayashi
N
N
Nakaji
S
S
Murakami
T
T
Tanaka
N
N
D000641:Ammonia / Q000378:metabolism; D002448:Cell Adhesion / Q000502:physiology*; D018929:Cell Culture Techniques / Q000379:methods; D002460:Cell Line; D048429:Cell Size; D018914:Cells, Immobilized; D002482:Cellulose / Q000378:metabolism; D018014:Gene Transfer Techniques; D022781:Hepatocytes / Q000502:physiology* / Q000648:ultrastructure; D006801:Humans; D019164:Liver, Artificial; D008863:Microspheres; D009842:Oligopeptides / Q000378:metabolism; D013997:Time Factors
D009842:Oligopeptides; D000641:Ammonia; D002482:Cellulose; C049554:glycyl-arginyl-glycyl-aspartyl-serine