A novel large-scale production system for modified basement membrane matrices using gene-swapped parietal endoderm cells.
Parietal endoderm-like cells, including Engelbreth-Holm-Swarm tumor and differentiated F9 embryonal carcinoma cells, produce huge amounts of basement membrane components, including laminin-1 (alpha1beta1gamma1). We employed a double-lox system-based gene-swapping strategy in F9 cells to replace the laminin alpha1 gene with a laminin alpha5 minigene. The gene-swapped F9 cells secreted laminin-10 (alpha5beta1gamma1) consisting of the exogenous alpha5 subunit and endogenous beta1 and gamma1 subunits on differentiation. The laminin-10 concentration in the conditioned medium exceeded 10 mg/l, which is 10-fold higher than the concentrations achieved by conventional recombinant expression systems. The gene-swapped F9 cells deposited basement membrane-like matrices containing laminin-10 on culture dishes, offering a novel microenvironment for in vitro cell manipulation.
Hayashi
Yoshitaka
Y
Sekiguchi Biomatrix Signaling Project, ERATO, Japanese Science and Technology Agency (JST), Aichi Medical University, 21 Karimata, Aichi-gun, Aichi 480-1195, Japan. hayashiy@riem.nagoya-u.ac.jp
Weber
Charles N
CN
Emoto
Tomomi
T
Fujiwara
Hironobu
H
Sanzen
Noriko
N
Futaki
Sugiko
S
Sekiguchi
Kiyotoshi
K
D000818:Animals; D001485:Basement Membrane / Q000196:embryology / Q000502:physiology* / Q000648:ultrastructure; D045744:Cell Line, Tumor; D018507:Gene Expression Regulation, Developmental; D005822:Genetic Vectors; D007797:Laminin / Q000235:genetics*
D007797:Laminin