STABLE EXPRESSION OF HUMAN INDUCIBLE NITRIC-OXIDE SYNTHASE IN V79 CHINESE-HAMSTER CELLS

A recombinant expression vector containing the full-length cDNA for hu man inducible nitric oxide (NO) synthase was constructed for constitut ive expression in V79 Chinese hamster cells. Expression was followed b y Western analyses using three different NO synthase antisera. Activit y remained stable during 4 months of continued cultivation. Activities were 25 pmol min(-1) mg(-1) cytosolic protein with L-arginine and 47 pmol min(-1) mg(-1) cytosolic protein with N-G-hydroxy-L-arginine as s ubstrates. Activity was concentration-dependently inhibited by inhibit ors such as N-G-methyl-L-arginine, N-G-nitro-L-arginine, N-G-nitro L-a rginine methyl ester, aminoguanidine, and S-methyl-isothiourea. The ra nk order of inhibitor potencies was different from published results o btained with rodent inducible NOS. Parental V79 cells do not express a nd cannot be induced for NO synthase activity. Therefore, the generica lly engineered V79 cell line is defined for the cDNA-encoded human ind ucible NO synthase. The new cell line may serve as a useful tool to st udy human inducible NO synthase. Copyright (C) 1996 Elsevier Science I nc.